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Sample records for vascular smooth-muscle cells

  1. Vinpocetine Attenuates the Osteoblastic Differentiation of Vascular Smooth Muscle Cells.

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    Yun-Yun Ma

    Full Text Available Vascular calcification is an active process of osteoblastic differentiation of vascular smooth muscle cells; however, its definite mechanism remains unknown. Vinpocetine, a derivative of the alkaloid vincamine, has been demonstrated to inhibit the high glucose-induced proliferation of vascular smooth muscle cells; however, it remains unknown whether vinpocetine can affect the osteoblastic differentiation of vascular smooth muscle cells. We hereby investigated the effect of vinpocetine on vascular calcification using a beta-glycerophosphate-induced cell model. Our results showed that vinpocetine significantly reduced the osteoblast-like phenotypes of vascular smooth muscle cells including ALP activity, osteocalcin, collagen type I, Runx2 and BMP-2 expression as well as the formation of mineralized nodule. Vinpocetine, binding to translocation protein, induced phosphorylation of extracellular signal-related kinase and Akt and thus inhibited the translocation of nuclear factor-kappa B into the nucleus. Silencing of translocator protein significantly attenuated the inhibitory effect of vinpocetine on osteoblastic differentiation of vascular smooth muscle cells. Taken together, vinpocetine may be a promising candidate for the clinical therapy of vascular calcification.

  2. Calcium dynamics in vascular smooth muscle

    OpenAIRE

    Amberg, Gregory C.; Navedo, Manuel F.

    2013-01-01

    Smooth muscle cells are ultimately responsible for determining vascular luminal diameter and blood flow. Dynamic changes in intracellular calcium are a critical mechanism regulating vascular smooth muscle contractility. Processes influencing intracellular calcium are therefore important regulators of vascular function with physiological and pathophysiological consequences. In this review we discuss the major dynamic calcium signals identified and characterized in vascular smooth muscle cells....

  3. An α-smooth muscle actin (acta2/αsma zebrafish transgenic line marking vascular mural cells and visceral smooth muscle cells.

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    Thomas R Whitesell

    Full Text Available Mural cells of the vascular system include vascular smooth muscle cells (SMCs and pericytes whose role is to stabilize and/or provide contractility to blood vessels. One of the earliest markers of mural cell development in vertebrates is α smooth muscle actin (acta2; αsma, which is expressed by pericytes and SMCs. In vivo models of vascular mural cell development in zebrafish are currently lacking, therefore we developed two transgenic zebrafish lines driving expression of GFP or mCherry in acta2-expressing cells. These transgenic fish were used to trace the live development of mural cells in embryonic and larval transgenic zebrafish. acta2:EGFP transgenic animals show expression that largely mirrors native acta2 expression, with early pan-muscle expression starting at 24 hpf in the heart muscle, followed by skeletal and visceral muscle. At 3.5 dpf, expression in the bulbus arteriosus and ventral aorta marks the first expression in vascular smooth muscle. Over the next 10 days of development, the number of acta2:EGFP positive cells and the number of types of blood vessels associated with mural cells increases. Interestingly, the mural cells are not motile and remain in the same position once they express the acta2:EGFP transgene. Taken together, our data suggests that zebrafish mural cells develop relatively late, and have little mobility once they associate with vessels.

  4. Electron histochemical and autoradiographic studies of vascular smooth muscle cell

    International Nuclear Information System (INIS)

    Kameyama, Kohji; Aida, Takeo; Asano, Goro

    1982-01-01

    The authors have studied the vascular smooth muscle cell in the aorta and the arteries of brain, heart in autopsied cases, cholesterol fed rabbits and canine through electron histochemical and autoradiographic methods, using 3 H-proline and 3 H-thymidine. The vascular changes are variable presumably due to the functional and morphological difference of vessels. Aging, pathological condition and physiological requirement induce the disturbances of vascular functions as contractility. According to various pathological conditions, the smooth muscle cell altered their shape, surface properties and arrangement of subcellular organelles including changes in number. The morphological features of arteries during aging is characterized by the thickening of endothelium and media. Decreasing cellularity and increasing collagen contents in media. The autoradiographic and histochemical observations using periodic acid methenamine silver (PAM) and ruthenium red stains demonstrated that the smooth muscle cell is a connective tissue synthetic cell. The PAM impregnation have proved that the small bundle of microfilaments become associated with small conglomerate of collagen and elastic fibers. Cytochemical examination will provide sufficient evidence to establish the contribution of subcellular structure. The acid phosphatase play an important role in vascular disease and they are directly involved in cellular lipid metabolism in cholesterol fed animals, and the activity of Na-K ATPase on the plasma membrane may contribute to the regulation of vascular blood flow and vasospasms. Direct injury and subsequent abnormal contraction of smooth muscle cell may initiate increased permeability of plasma protein and lipid in the media layer and eventually may developed and enhance arteriosclerosis. (author)

  5. Notch signal reception is required in vascular smooth muscle cells for ductus arteriosus closure

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    Krebs, Luke T.; Norton, Christine R.; Gridley, Thomas

    2017-01-01

    Summary The ductus arteriosus is an arterial vessel that shunts blood flow away from the lungs during fetal life, but normally occludes after birth to establish the adult circulation pattern. Failure of the ductus arteriosus to close after birth is termed patent ductus arteriosus, and is one of the most common congenital heart defects. Our previous work demonstrated that vascular smooth muscle cell expression of the Jag1 gene, which encodes a ligand for Notch family receptors, is essential for postnatal closure of the ductus arteriosus in mice. However, it was not known what cell population was responsible for receiving the Jag1-mediated signal. Here we show, using smooth muscle cell-specific deletion of the Rbpj gene, which encodes a transcription factor that mediates all canonical Notch signaling, that Notch signal reception in the vascular smooth muscle cell compartment is required for ductus arteriosus closure. These data indicate that homotypic vascular smooth muscle cell interactions are required for proper contractile smooth muscle cell differentiation and postnatal closure of the ductus arteriosus in mice. PMID:26742650

  6. Augmented vascular smooth muscle cell stiffness and adhesion when hypertension is superimposed on aging.

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    Sehgel, Nancy L; Sun, Zhe; Hong, Zhongkui; Hunter, William C; Hill, Michael A; Vatner, Dorothy E; Vatner, Stephen F; Meininger, Gerald A

    2015-02-01

    Hypertension and aging are both recognized to increase aortic stiffness, but their interactions are not completely understood. Most previous studies have attributed increased aortic stiffness to changes in extracellular matrix proteins that alter the mechanical properties of the vascular wall. Alternatively, we hypothesized that a significant component of increased vascular stiffness in hypertension is due to changes in the mechanical and adhesive properties of vascular smooth muscle cells, and that aging would augment the contribution from vascular smooth muscle cells when compared with the extracellular matrix. Accordingly, we studied aortic stiffness in young (16-week-old) and old (64-week-old) spontaneously hypertensive rats and Wistar-Kyoto wild-type controls. Systolic and pulse pressures were significantly increased in young spontaneously hypertensive rats when compared with young Wistar-Kyoto rats, and these continued to rise in old spontaneously hypertensive rats when compared with age-matched controls. Excised aortic ring segments exhibited significantly greater elastic moduli in both young and old spontaneously hypertensive rats versus Wistar-Kyoto rats. were isolated from the thoracic aorta, and stiffness and adhesion to fibronectin were measured by atomic force microscopy. Hypertension increased both vascular smooth muscle cell stiffness and vascular smooth muscle cell adhesion, and these increases were both augmented with aging. By contrast, hypertension did not affect histological measures of aortic collagen and elastin, which were predominantly changed by aging. These findings support the concept that stiffness and adhesive properties of vascular smooth muscle cells are novel mechanisms contributing to the increased aortic stiffness occurring with hypertension superimposed on aging. © 2014 American Heart Association, Inc.

  7. Suppression of vascular smooth muscle cells' proliferation and ...

    African Journals Online (AJOL)

    This study aimed to determine the effects of valsartan on the proliferation and migration of isolated rat vascular smooth muscle cells (VSMCs) and the expression of phospho-p42/44 mitogen-activated protein kinase (MAPK) promoted by angiotensin II (Ang II). VSMCs from the rat thoracic aorta were cultured by ...

  8. Cholesterol is necessary both for the toxic effect of Abeta peptides on vascular smooth muscle cells and for Abeta binding to vascular smooth muscle cell membranes.

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    Subasinghe, Supundi; Unabia, Sharon; Barrow, Colin J; Mok, Su San; Aguilar, Marie-Isabel; Small, David H

    2003-02-01

    Accumulation of beta amyloid (Abeta) in the brain is central to the pathogenesis of Alzheimer's disease. Abeta can bind to membrane lipids and this binding may have detrimental effects on cell function. In this study, surface plasmon resonance technology was used to study Abeta binding to membranes. Abeta peptides bound to synthetic lipid mixtures and to an intact plasma membrane preparation isolated from vascular smooth muscle cells. Abeta peptides were also toxic to vascular smooth muscle cells. There was a good correlation between the toxic effect of Abeta peptides and their membrane binding. 'Ageing' the Abeta peptides by incubation for 5 days increased the proportion of oligomeric species, and also increased toxicity and the amount of binding to lipids. The toxicities of various Abeta analogs correlated with their lipid binding. Significantly, binding was influenced by the concentration of cholesterol in the lipid mixture. Reduction of cholesterol in vascular smooth muscle cells not only reduced the binding of Abeta to purified plasma membrane preparations but also reduced Abeta toxicity. The results support the view that Abeta toxicity is a direct consequence of binding to lipids in the membrane. Reduction of membrane cholesterol using cholesterol-lowering drugs may be of therapeutic benefit because it reduces Abeta-membrane binding.

  9. Biophysical induction of vascular smooth muscle cell podosomes.

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    Na Young Kim

    Full Text Available Vascular smooth muscle cell (VSMC migration and matrix degradation occurs with intimal hyperplasia associated with atherosclerosis, vascular injury, and restenosis. One proposed mechanism by which VSMCs degrade matrix is through the use of podosomes, transient actin-based structures that are thought to play a role in extracellular matrix degradation by creating localized sites of matrix metalloproteinase (MMP secretion. To date, podosomes in VSMCs have largely been studied by stimulating cells with phorbol esters, such as phorbol 12,13-dibutyrate (PDBu, however little is known about the physiological cues that drive podosome formation. We present the first evidence that physiological, physical stimuli mimicking cues present within the microenvironment of diseased arteries can induce podosome formation in VSMCs. Both microtopographical cues and imposed pressure mimicking stage II hypertension induce podosome formation in A7R5 rat aortic smooth muscle cells. Moreover, wounding using a scratch assay induces podosomes at the leading edge of VSMCs. Notably the effect of each of these biophysical stimuli on podosome stimulation can be inhibited using a Src inhibitor. Together, these data indicate that physical cues can induce podosome formation in VSMCs.

  10. Epigenetic regulation of vascular smooth muscle cell function in atherosclerosis.

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    Findeisen, Hannes M; Kahles, Florian K; Bruemmer, Dennis

    2013-04-01

    Epigenetics involve heritable and acquired changes in gene transcription that occur independently of the DNA sequence. Epigenetic mechanisms constitute a hierarchic upper-level of transcriptional control through complex modifications of chromosomal components and nuclear structures. These modifications include, for example, DNA methylation or post-translational modifications of core histones; they are mediated by various chromatin-modifying enzymes; and ultimately they define the accessibility of a transcriptional complex to its target DNA. Integrating epigenetic mechanisms into the pathophysiologic concept of complex and multifactorial diseases such as atherosclerosis may significantly enhance our understanding of related mechanisms and provide promising therapeutic approaches. Although still in its infancy, intriguing scientific progress has begun to elucidate the role of epigenetic mechanisms in vascular biology, particularly in the control of smooth muscle cell phenotypes. In this review, we will summarize epigenetic pathways in smooth muscle cells, focusing on mechanisms involved in the regulation of vascular remodeling.

  11. Human induced pluripotent stem cell-derived vascular smooth muscle cells

    DEFF Research Database (Denmark)

    Ayoubi, Sohrab; Sheikh, Søren P; Eskildsen, Tilde V

    2017-01-01

    . To this end, human induced pluripotent stem cells (hiPSCs) have generated great enthusiasm, and have been a driving force for development of novel strategies in drug discovery and regenerative cell-therapy for the last decade. Hence, investigating the mechanisms underlying the differentiation of hi......PSCs into specialized cell types such as cardiomyocytes, endothelial cells, and vascular smooth muscle cells (VSMCs) may lead to a better understanding of developmental cardiovascular processes and potentiate progress of safe autologous regenerative therapies in pathological conditions. In this review, we summarize...

  12. Enhanced elastin synthesis and maturation in human vascular smooth muscle tissue derived from induced-pluripotent stem cells.

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    Eoh, Joon H; Shen, Nian; Burke, Jacqueline A; Hinderer, Svenja; Xia, Zhiyong; Schenke-Layland, Katja; Gerecht, Sharon

    2017-04-01

    Obtaining vascular smooth muscle tissue with mature, functional elastic fibers is a key obstacle in tissue-engineered blood vessels. Poor elastin secretion and organization leads to a loss of specialization in contractile smooth muscle cells, resulting in over proliferation and graft failure. In this study, human induced-pluripotent stem cells (hiPSCs) were differentiated into early smooth muscle cells, seeded onto a hybrid poly(ethylene glycol) dimethacrylate/poly (l-lactide) (PEGdma-PLA) scaffold and cultured in a bioreactor while exposed to pulsatile flow, towards maturation into contractile smooth muscle tissue. We evaluated the effects of pulsatile flow on cellular organization as well as elastin expression and assembly in the engineered tissue compared to a static control through immunohistochemistry, gene expression and functionality assays. We show that culturing under pulsatile flow resulted in organized and functional hiPSC derived smooth muscle tissue. Immunohistochemistry analysis revealed hiPSC-smooth muscle tissue with robust, well-organized cells and elastic fibers and the supporting microfibril proteins necessary for elastic fiber assembly. Through qRT-PCR analysis, we found significantly increased expression of elastin, fibronectin, and collagen I, indicating the synthesis of necessary extracellular matrix components. Functionality assays revealed that hiPSC-smooth muscle tissue cultured in the bioreactor had an increased calcium signaling and contraction in response to a cholinergic agonist, significantly higher mature elastin content and improved mechanical properties in comparison to the static control. The findings presented here detail an effective approach to engineering elastic human vascular smooth muscle tissue with the functionality necessary for tissue engineering and regenerative medicine applications. Obtaining robust, mature elastic fibers is a key obstacle in tissue-engineered blood vessels. Human induced-pluripotent stem cells have

  13. KCl cotransport regulation and protein kinase G in cultured vascular smooth muscle cells.

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    Adragna, N C; Zhang, J; Di Fulvio, M; Lincoln, T M; Lauf, P K

    2002-05-15

    K-Cl cotransport is activated by vasodilators in erythrocytes and vascular smooth muscle cells and its regulation involves putative kinase/phosphatase cascades. N-ethylmaleimide (NEM) activates the system presumably by inhibiting a protein kinase. Nitrovasodilators relax smooth muscle via cGMP-dependent activation of protein kinase G (PKG), a regulator of membrane channels and transporters. We investigated whether PKG regulates K-Cl cotransport activity or mRNA expression in normal, PKG-deficient-vector-only-transfected (PKG-) and PKG-catalytic-domain-transfected (PKG+) rat aortic smooth muscle cells. K-Cl cotransport was calculated as the Cl-dependent Rb influx, and mRNA was determined by semiquantitative RT-PCR. Baseline K-Cl cotransport was higher in PKG+ than in PKG- cells (p <0.01). At 0.5 mM, NEM stimulated K-Cl cotransport by 5-fold in PKG- but not in PKG+ cells. However, NEM was more potent although less effective to activate K-Cl cotransport in normal (passage 1-3) and PKG+ than in PKG- cells. In PKG- cells, [(dihydroindenyl) oxy] alkanoic acid (300 mM) but not furosemide (1 mM) inhibited K-Cl cotransport. Furthermore, no difference in K-Cl cotransport mRNA expression was observed between these cells. In conclusion, this study shows that manipulation of PKG expression in vascular smooth muscle cells affects K-Cl cotransport activity and its activation by NEM.

  14. LRP1 in brain vascular smooth muscle cells mediates local clearance of Alzheimer's amyloid-β.

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    Kanekiyo, Takahisa; Liu, Chia-Chen; Shinohara, Mitsuru; Li, Jie; Bu, Guojun

    2012-11-14

    Impaired clearance of amyloid-β (Aβ) is a major pathogenic event for Alzheimer's disease (AD). Aβ depositions in brain parenchyma as senile plaques and along cerebrovasculature as cerebral amyloid angiopathy (CAA) are hallmarks of AD. A major pathway that mediates brain Aβ clearance is the cerebrovascular system where Aβ is eliminated through the blood-brain barrier (BBB) and/or degraded by cerebrovascular cells along the interstitial fluid drainage pathway. An Aβ clearance receptor, the low-density lipoprotein receptor-related protein 1 (LRP1), is abundantly expressed in cerebrovasculature, in particular in vascular smooth muscle cells. Previous studies have indicated a role of LRP1 in endothelial cells in transcytosing Aβ out of the brain across the BBB; however, whether this represents a significant pathway for brain Aβ clearance remains controversial. Here, we demonstrate that Aβ can be cleared locally in the cerebrovasculature by an LRP1-dependent endocytic pathway in smooth muscle cells. The uptake and degradation of both endogenous and exogenous Aβ were significantly reduced in LRP1-suppressed human brain vascular smooth muscle cells. Conditional deletion of Lrp1 in vascular smooth muscle cell in amyloid model APP/PS1 mice accelerated brain Aβ accumulation and exacerbated Aβ deposition as amyloid plaques and CAA without affecting Aβ production. Our results demonstrate that LRP1 is a major Aβ clearance receptor in cerebral vascular smooth muscle cell and a disturbance of this pathway contributes to Aβ accumulation. These studies establish critical functions of the cerebrovasculature system in Aβ metabolism and identify a new pathway involved in the pathogenesis of both AD and CAA.

  15. Dipeptidyl peptidase-4 inhibitor gemigliptin protects against vascular calcification in an experimental chronic kidney disease and vascular smooth muscle cells.

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    Soon-Youn Choi

    Full Text Available Although dipeptidyl peptidase-4 inhibitors, a class of antidiabetic drugs, have various pleiotropic effects, it remains undetermined whether gemigliptin has a beneficial effect on vascular calcification. Therefore, this study was performed to evaluate the effect of gemigliptin on vascular calcification in a rat model of adenine-induced chronic kidney disease and in cultured vascular smooth muscle cells. Gemigliptin attenuated calcification of abdominal aorta and expression of RUNX2 in adenine-induced chronic kidney disease rats. In cultured vascular smooth muscle cells, phosphate-induced increase in calcium content was reduced by gemigliptin. Gemigliptin reduced phosphate-induced PiT-1 mRNA expression, reactive oxygen species generation, and NADPH oxidase mRNA expression (p22phox and NOX4. The reduction of oxidative stress by gemigliptin was associated with the downregulation of phospho-PI3K/AKT expression. High phosphate increased the expression of frizzled-3 (FDZ3 and decreased the expression of dickkopf-related protein-1 (DKK-1 in the Wnt pathway. These changes were attenuated by gemigliptin treatment. Gemigliptin restored the decreased expression of vascular smooth muscle cells markers (α-SMA and SM22α and increased expression of osteogenic makers (CBFA1, OSX, E11, and SOST induced by phosphate. In conclusion, gemigliptin attenuated vascular calcification and osteogenic trans-differentiation in vascular smooth muscle cells via multiple steps including downregulation of PiT-1 expression and suppression of reactive oxygen species generation, phospho-PI3K/AKT, and the Wnt signaling pathway.

  16. Expression of smooth muscle and non-muscle myosin heavy chain isoforms in cultured vascular smooth muscle cells

    International Nuclear Information System (INIS)

    Rovner, A.S.; Murphy, R.A.; Owens, G.K.

    1986-01-01

    Immunocytochemical studies of cultured smooth muscle cells (SMCs) have disagreed on the nature of myosin expression. This investigation was undertaken to test for the presence of heterogeneous myosin heavy chain (MHC) isoforms in cell culture as a possible explanation for these results. Previously, Rovner et al. detected two MHCs in intact smooth muscles which differed in molecular weight by ca. 4000 daltons (SM1 and SM2) using a 3-4% acrylamide gradient SDS gel system. When sub-confluent primary cultures of rat aorta SMCs were assayed by this system, SM1 and SM2 were seen, along with large amounts of a third, unique MHC, NM, which closely resembled the MHC from human platelet in size and antigenicity. Data from 35 S-methionine autoradiograms showed that the log growth phase SMC cultures were producing almost exclusively NM, but the growth arrest, post-confluent cultures synthesized increased relative amounts of the SM MHC forms and contained comparable amounts of SM1, SM2, and NM. The same patterns of MHC synthesis were seen in sub-passaged SMCs. The expression of the SM-specific forms of myosin in quiescent, post-confluent cultures parallels that of smooth muscle actin suggesting that density induced growth arrest promotes cytodifferentiation in cultured vascular SMCs

  17. Experimental study on effect of arsenic trioxide on vascular smooth muscle cells

    International Nuclear Information System (INIS)

    Lu Qin; An Yanli; Niu Huanzhang; Teng Gaojun; Wang Zihao; Zhang Dongsheng; Fang Juanjuan

    2007-01-01

    Objective: To investigate the effect of arsenic trioxide (As 2 O 3 ) nanoparticles on rabbit vascular smooth muscle cells in vitro in comparison with normal form As 2 O 3 . Methods: The rabbit vascular smooth muscle cells were cultured in vitro. Nano and normal forms of As 2 O 3 with drug concentrations of 3 μmol/L were added into the cells. Cell proliferation curve was drawn according to the light absorption values of MTT test. Flow cytometry was applied to observe the apoptosis. DNA was extracted and underwent electrophoresis. Results: Cell proliferation treated with the 3 μmol/L concentration of As 2 O 3 was inhibited. Cell growth was inhibited markedly with increased treatment time, and the inhibition effect of nano drug form seemed stronger than that of normal form. MTT light absorption values of cells treated at 24, 48 and 72 h showed statistically significant difference (H=10.934, 15.039, 15.539, P 2 O 3 , normal drug form of As 2 O 3 and control group of cells without As 2 O 3 were 44.97%, 58.54%, 74.02% respectively. The early apoptosis rates were 16.89%, 11.27%, 11.20%, late apoptosis rates were 26.56%, 23.60%, 12.46%, and necrosis rates were 11.58%, 6.59%, 2.32% respectively. Agarose gel electrophoresis showed 'ladder' strand of DNA, with more strands and obscurity for nano drug form treated cells. Conclusion: Arsenic trioxide may inhibit the growth of rabbit vascular smooth muscle cells. The nano drug form showed stronger inhibition effect than that of the normal drug form. (authors)

  18. Anti-atherosclerotic plants which modulate the phenotype of vascular smooth muscle cells.

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    Saleh Al-Shehabi, Tuqa; Iratni, Rabah; Eid, Ali H

    2016-10-15

    Cardiovascular disease (CVD) remains the leading cause of global death, with atherosclerosis being a major contributor to this mortality. Several mechanisms are implicated in the pathogenesis of this disease. A key element in the development and progression of atherosclerotic lesions is the phenotype of vascular smooth muscle cells. Under pathophysiologic conditions such as injury, these cells switch from a contractile to a synthetic phenotype that often possesses high proliferative and migratory capacities. Despite major advances made in the management and treatment of atherosclerosis, mortality associated with this disease remains high. This mandates that other approaches be sought. Herbal medicine, especially for the treatment of CVD, has been gaining more attention in recent years. This is in no small part due to the evidence-based values associated with the consumption of many plants as well as the relatively cheaper prices, easier access and conventional folk medicine "inherited" over generations. Sections: In this review, we provide a brief introduction about the pathogenesis of atherosclerosis then we highlight the role of vascular smooth muscle cells in this disease, especially when a phenotypic switch of these cells arises. We then thoroughly discuss the various plants that show potentially beneficial effects as anti-atherosclerotic, with prime attention given to herbs and plants that inhibit the phenotypic switch of vascular smooth muscle cells. Accumulating evidence provides the justification for the use of botanicals in the treatment or prevention of atherosclerosis. However, further studies, especially clinical ones, are warranted to better define several pharmacological parameters of these herbs, such as toxicity, tolerability, and efficacy. Copyright © 2015 Elsevier Ltd. All rights reserved.

  19. In-depth evaluation of commercially available human vascular smooth muscle cells phenotype: Implications for vascular tissue engineering

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    Timraz, Sara B.H., E-mail: sara.timraz@kustar.ac.ae [Department of Biomedical Engineering, Khalifa University, PO Box 127788, Abu Dhabi (United Arab Emirates); Farhat, Ilyas A.H., E-mail: ilyas.farhat@outlook.com [Department of Applied Mathematics and Sciences, Khalifa University, PO Box 127788, Abu Dhabi (United Arab Emirates); Alhussein, Ghada, E-mail: ghada.alhussein@kustar.ac.ae [Department of Biomedical Engineering, Khalifa University, PO Box 127788, Abu Dhabi (United Arab Emirates); Christoforou, Nicolas, E-mail: nicolas.christoforou@kustar.ac.ae [Department of Biomedical Engineering, Khalifa University, PO Box 127788, Abu Dhabi (United Arab Emirates); Department of Biomedical Engineering, Duke University, Durham, NC 27708 (United States); Teo, Jeremy C.M., E-mail: jeremy.teo@kustar.ac.ae [Department of Biomedical Engineering, Khalifa University, PO Box 127788, Abu Dhabi (United Arab Emirates)

    2016-05-01

    In vitro research on vascular tissue engineering has extensively used isolated primary human or animal smooth muscle cells (SMC). Research programs that lack such facilities tend towards commercially available primary cells sources. Here, we aim to evaluate the capacity of commercially available human SMC to maintain their contractile phenotype, and determine if dedifferentiation towards the synthetic phenotype occurs in response to conventional cell culture and passaging without any external biochemical or mechanical stimuli. Lower passage SMC adopted a contractile phenotype marked by a relatively slower proliferation rate, higher expression of proteins of the contractile apparatus and smoothelin, elongated morphology, and reduced deposition of collagen types I and III. As the passage number increased, migratory capacity was enhanced, average cell speed, total distance and net distance travelled increased up to passage 8. Through the various assays, corroborative evidence pinpoints SMC at passage 7 as the transition point between the contractile and synthetic phenotypes, while passage 8 distinctly and consistently exhibited characteristics of synthetic phenotype. This knowledge is particularly useful in selecting SMC of appropriate passage number for the target vascular tissue engineering application, for example, a homeostatic vascular graft for blood vessel replacement versus recreating atherosclerotic blood vessel model in vitro. - Highlights: • Ability of human smooth muscle cells to alter phenotype in culture is evaluated. • Examined the effect of passaging human smooth muscle cells on phenotype. • Phenotype is assessed based on morphology, proliferation, markers, and migration. • Multi-resolution assessment methodology, single-cell and cell-population. • Lower and higher passages than P7 adopted a contractile and synthetic phenotype respectively.

  20. In-depth evaluation of commercially available human vascular smooth muscle cells phenotype: Implications for vascular tissue engineering

    International Nuclear Information System (INIS)

    Timraz, Sara B.H.; Farhat, Ilyas A.H.; Alhussein, Ghada; Christoforou, Nicolas; Teo, Jeremy C.M.

    2016-01-01

    In vitro research on vascular tissue engineering has extensively used isolated primary human or animal smooth muscle cells (SMC). Research programs that lack such facilities tend towards commercially available primary cells sources. Here, we aim to evaluate the capacity of commercially available human SMC to maintain their contractile phenotype, and determine if dedifferentiation towards the synthetic phenotype occurs in response to conventional cell culture and passaging without any external biochemical or mechanical stimuli. Lower passage SMC adopted a contractile phenotype marked by a relatively slower proliferation rate, higher expression of proteins of the contractile apparatus and smoothelin, elongated morphology, and reduced deposition of collagen types I and III. As the passage number increased, migratory capacity was enhanced, average cell speed, total distance and net distance travelled increased up to passage 8. Through the various assays, corroborative evidence pinpoints SMC at passage 7 as the transition point between the contractile and synthetic phenotypes, while passage 8 distinctly and consistently exhibited characteristics of synthetic phenotype. This knowledge is particularly useful in selecting SMC of appropriate passage number for the target vascular tissue engineering application, for example, a homeostatic vascular graft for blood vessel replacement versus recreating atherosclerotic blood vessel model in vitro. - Highlights: • Ability of human smooth muscle cells to alter phenotype in culture is evaluated. • Examined the effect of passaging human smooth muscle cells on phenotype. • Phenotype is assessed based on morphology, proliferation, markers, and migration. • Multi-resolution assessment methodology, single-cell and cell-population. • Lower and higher passages than P7 adopted a contractile and synthetic phenotype respectively.

  1. Hypoxic contraction of cultured pulmonary vascular smooth muscle cells

    International Nuclear Information System (INIS)

    Murray, T.R.; Chen, L.; Marshall, B.E.; Macarak, E.J.

    1990-01-01

    The cellular events involved in generating the hypoxic pulmonary vasoconstriction response are not clearly understood, in part because of the multitude of factors that alter pulmonary vascular tone. The goal of the present studies was to determine if a cell culture preparation containing vascular smooth muscle (VSM) cells could be made to contract when exposed to a hypoxic atmosphere. Cultures containing only fetal bovine pulmonary artery VSM cells were assessed for contractile responses to hypoxic stimuli by two methods. In the first, tension forces generated by cells grown on a flexible growth surface (polymerized polydimethyl siloxane) were manifested as wrinkles and distortions of the surface under the cells. Wrinkling of the surface was noted to progressively increase with time as the culture medium bathing the cells was made hypoxic (PO2 approximately 25 mmHg). The changes were sometimes reversible upon return to normoxic conditions and appeared to be enhanced in cells already exhibiting evidence of some baseline tone. Repeated passage in culture did not diminish the hypoxic response. Evidence for contractile responses to hypoxia was also obtained from measurements of myosin light chain (MLC) phosphorylation. Conversion of MLC to the phosphorylated species is an early step in the activation of smooth muscle contraction. Lowering the PO2 in the culture medium to 59 mmHg caused a 45% increase in the proportion of MLC in the phosphorylated form as determined by two-dimensional gel electrophoresis. Similarly, cultures preincubated for 4 h with 32P and then exposed to normoxia or hypoxia for a 5-min experimental period showed more than twice as much of the label in MLCs of the hypoxic cells

  2. Tetraspanin CD9 regulates cell contraction and actin arrangement via RhoA in human vascular smooth muscle cells.

    Directory of Open Access Journals (Sweden)

    Michael J Herr

    Full Text Available The most prevalent cardiovascular diseases arise from alterations in vascular smooth muscle cell (VSMC morphology and function. Tetraspanin CD9 has been previously implicated in regulating vascular pathologies; however, insight into how CD9 may regulate adverse VSMC phenotypes has not been provided. We utilized a human model of aortic smooth muscle cells to understand the consequences of CD9 deficiency on VSMC phenotypes. Upon knocking down CD9, the cells developed an abnormally small and rounded morphology. We determined that this morphological change was due to a lack of typical parallel actin arrangement. We also found similar total RhoA but decreased GTP-bound (active RhoA levels in CD9 deficient cells. As a result, cells lacking a full complement of CD9 were less contractile than their control treated counterparts. Upon restoration of RhoA activity in the CD9 deficient cells, the phenotype was reversed and cell contraction was restored. Conversely, inhibition of RhoA activity in the control cells mimicked the CD9-deficient cell phenotype. Thus, alteration in CD9 expression was sufficient to profoundly disrupt cellular actin arrangement and endogenous cell contraction by interfering with RhoA signaling. This study provides insight into how CD9 may regulate previously described vascular smooth muscle cell pathophysiology.

  3. Hepatocyte growth factor triggers signaling cascades mediating vascular smooth muscle cell migration

    NARCIS (Netherlands)

    Taher, Taher E. I.; Derksen, Patrick W. B.; de Boer, Onno J.; Spaargaren, Marcel; Teeling, Peter; van der Wal, Allard C.; Pals, Steven T.

    2002-01-01

    A key event in neointima formation and atherogenesis is the migration of vascular smooth muscle cells (VSMCs) into the intima. This is controlled by cytokines and extracellular matix (ECM) components within the microenvironment of the diseased vessel wall. At present, these signals have only been

  4. Akt1/PKB upregulation leads to vascular smooth muscle cell hypertrophy and polyploidization

    Science.gov (United States)

    Hixon, Mary L.; Muro-Cacho, Carlos; Wagner, Mark W.; Obejero-Paz, Carlos; Millie, Elise; Fujio, Yasushi; Kureishi, Yasuko; Hassold, Terry; Walsh, Kenneth; Gualberto, Antonio

    2000-01-01

    Vascular smooth muscle cells (VSMCs) at capacitance arteries of hypertensive individuals and animals undergo marked age- and blood pressure–dependent polyploidization and hypertrophy. We show here that VSMCs at capacitance arteries of rat models of hypertension display high levels of Akt1/PKB protein and activity. Gene transfer of Akt1 to VSMCs isolated from a normotensive rat strain was sufficient to abrogate the activity of the mitotic spindle cell–cycle checkpoint, promoting polyploidization and hypertrophy. Furthermore, the hypertrophic agent angiotensin II induced VSMC polyploidization in an Akt1-dependent manner. These results demonstrate that Akt1 regulates ploidy levels in VSMCs and contributes to vascular smooth muscle polyploidization and hypertrophy during hypertension. PMID:11032861

  5. Identification and characterization of novel smoothelin isoforms in vascular smooth muscle.

    Science.gov (United States)

    Krämer, J; Quensel, C; Meding, J; Cardoso, M C; Leonhardt, H

    2001-01-01

    Smoothelin is a cytoskeletal protein specifically expressed in differentiated smooth muscle cells and has been shown to colocalize with smooth muscle alpha actin. In addition to the small smoothelin isoform of 59 kD, we recently identified a large smoothelin isoform of 117 kD. The aim of this study was to identify and characterize novel smoothelin isoforms. The genomic structure and sequence of the smoothelin gene were determined by genomic PCR, RT-PCR and DNA sequencing. Comparison of the cDNA and genomic sequences shows that the small smoothelin isoform is generated by transcription initiation 10 kb downstream of the start site of the large isoform. In addition to the known smoothelin cDNA (c1 isoform) we identified two novel cDNA variants (c2 and c3 isoform) that are generated by alternative splicing within a region, which shows similarity to the spectrin family of F-actin cross-linking proteins. Visceral organs express the c1 form, while the c2 form prevails in well-vascularized tissue as analyzed by RT-PCR. We then generated specific antibodies against the major smoothelin isoforms and could show by Western blotting and immunohistochemistry that the large isoform is specifically expressed in vascular smooth muscle cells, while the small isoform is abundant in visceral smooth muscle. These results strongly suggest that the smoothelin gene contains a vascular and a visceral smooth muscle promoter. The cell-type-specific expression of smoothelin isoforms that are associated with actin filaments may play a role in the modulation of the contractile properties of different smooth muscle cell types. Copyright 2001 S. Karger AG, Basel

  6. Notch signaling regulates platelet-derived growth factor receptor-β expression in vascular smooth muscle cells

    NARCIS (Netherlands)

    Jin, S.; Hansson, E.M.; Tikka, S.; Lanner, F.; Sahlgren, C.; Farnebo, F.; Baumann, M.; Kalimo, H.; Lendahl, U.

    2008-01-01

    Notch signaling is critically important for proper architecture of the vascular system, and mutations in NOTCH3 are associated with CADASIL, a stroke and dementia syndrome with vascular smooth muscle cell (VSMC) dysfunction. In this report, we link Notch signaling to platelet-derived growth factor

  7. Vascular wall-resident CD44+ multipotent stem cells give rise to pericytes and smooth muscle cells and contribute to new vessel maturation.

    Directory of Open Access Journals (Sweden)

    Diana Klein

    Full Text Available Here, we identify CD44(+CD90(+CD73(+CD34(-CD45(- cells within the adult human arterial adventitia with properties of multipotency which were named vascular wall-resident multipotent stem cells (VW-MPSCs. VW-MPSCs exhibit typical mesenchymal stem cell characteristics including cell surface markers in immunostaining and flow cytometric analyses, and differentiation into adipocytes, chondrocytes and osteocytes under culture conditions. Particularly, TGFß1 stimulation up-regulates smooth muscle cell markers in VW-MPSCs. Using fluorescent cell labelling and co-localisation studies we show that VW-MPSCs differentiate to pericytes/smooth muscle cells which cover the wall of newly formed endothelial capillary-like structures in vitro. Co-implantation of EGFP-labelled VW-MPSCs and human umbilical vein endothelial cells into SCID mice subcutaneously via Matrigel results in new vessels formation which were covered by pericyte- or smooth muscle-like cells generated from implanted VW-MPSCs. Our results suggest that VW-MPSCs are of relevance for vascular morphogenesis, repair and self-renewal of vascular wall cells and for local capacity of neovascularization in disease processes.

  8. The neuropeptide catestatin promotes vascular smooth muscle cell proliferation through the Ca{sup 2+}-calcineurin-NFAT signaling pathway

    Energy Technology Data Exchange (ETDEWEB)

    Guo, Xiaoxia [Department of Cardiology, People' s Hospital, Peking University, No. 11 South Avenue, Xi Zhi Men Xicheng District, Beijing 100044 (China); Zhou, Chunyan, E-mail: chunyanzhou@bjmu.edu.cn [Department of Biochemistry and Molecular Biology, School of Basic Medical Sciences, Peking University, 38 Xueyuan Road, Haidian District, Beijing 100191 (China); Sun, Ningling, E-mail: nlsun@263.net [Department of Cardiology, People' s Hospital, Peking University, No. 11 South Avenue, Xi Zhi Men Xicheng District, Beijing 100044 (China)

    2011-04-22

    Highlights: {yields} Catestatin stimulates proliferation of vascular smooth muscle cells in a dose-dependent manner. {yields} Catestatin provokes sustained increase in intracellular Ca{sup 2+}. {yields} Catestatin produces increased activation of calcineurin and promotes NFATc1 translocation into the nucleus. -- Abstract: The Chromogranin A-derived neuropeptide catestatin is an endogenous nicotinic cholinergic antagonist that acts as a pleiotropic hormone. Since catestatin shares several functions with other members derived from the chromogranin/secretogranin protein family and other neuropeptides which exert proliferative effects on vascular smooth muscle cells (VSMCs), we therefore hypothesized that catestatin would regulate VSMC proliferation. The present study demonstrates that catestatin caused a dose-dependent induction of proliferation in rat aortic smooth muscle cells and furthermore evoked a sustained increase in intracellular calcium. This subsequently leaded to enhanced activation of the Ca{sup 2+}/calmodulin-dependent phosphatase, calcineurin and resulted in an activation of the Ca{sup 2+}-dependent transcription factor, nuclear factor of activated T cells (NFAT), initiating transcription of proliferative genes. In addition, cyclosporin A (CsA), a potent inhibitor of calcineurin, abrogated catestatin-mediated effect on VSMCs, indicating that the calcineurin-NFAT signaling is strongly required for catestatin-induced growth of VSMCs. The present study establishes catestatin as a novel proliferative cytokine on vascular smooth muscle cells and this effect is mediated by the Ca{sup 2+}-calcineurin-NFAT signaling pathway.

  9. Phenotypic modulation of smooth muscle cells during formation of neointimal thickenings following vascular injury.

    Science.gov (United States)

    Thyberg, J

    1998-07-01

    Smooth muscle cells build up the media of mammalian arteries and constitute one of the principal cell types in atherosclerotic and restenotic lesions. Accordingly, they show a high degree of plasticity and are able to shift from a differentiated, contractile phenotype to a less differentiated, synthetic phenotype, and then back again. This modulation occurs as a response to vascular injury and includes a prominent structural reorganization with loss of myofilaments and formation of an extensive endoplasmic reticulum and a large Golgi complex. At the same time, the expression of cytoskeletal proteins and other gene products is altered. As a result, the cells lose their contractility and become able to migrate from the media to the intima, proliferate, and secrete extracellular matrix components, thereby contributing to the formation of intimal thickenings. The mechanisms behind this change in morphology and function of the smooth muscle cells are still incompletely understood. A crucial role has been ascribed to basement membrane proteins such as laminin and collagen type IV and adhesive proteins such as fibronectin. A significant role is also played by mitogenic proteins such as platelet-derived growth factor (PDGF) and basic fibroblast growth factor (bFGF). An improved knowledge of the regulation of smooth muscle differentiated properties represents an important part in the search for new methods of prevention and treatment of vascular disease.

  10. Experimental studies of mitochondrial function in CADASIL vascular smooth muscle cells

    International Nuclear Information System (INIS)

    Viitanen, Matti; Sundström, Erik; Baumann, Marc; Poyhonen, Minna; Tikka, Saara; Behbahani, Homira

    2013-01-01

    Cerebral Autosomal Dominant Arteriopathy with Subcortical Infarcts and Leukoencephalopathy (CADASIL) is a familiar fatal progressive degenerative disorder characterized by cognitive decline, and recurrent stroke in young adults. Pathological features include a dramatic reduction of brain vascular smooth muscle cells and severe arteriopathy with the presence of granular osmophilic material in the arterial walls. Here we have investigated the cellular and mitochondrial function in vascular smooth muscle cell lines (VSMCs) established from CADASIL mutation carriers (R133C) and healthy controls. We found significantly lower proliferation rates in CADASIL VSMC as compared to VSMC from controls. Cultured CADASIL VSMCs were not more vulnerable than control cells to a number of toxic substances. Morphological studies showed reduced mitochondrial connectivity and increased number of mitochondria in CADASIL VSMCs. Transmission electron microscopy analysis demonstrated increased irregular and abnormal mitochondria in CADASIL VSMCs. Measurements of mitochondrial membrane potential (Δψ m ) showed a lower percentage of fully functional mitochondria in CADASIL VSMCs. For a number of genes previously reported to be changed in CADASIL VSMCs, immunoblotting analysis demonstrated a significantly reduced SOD1 expression. These findings suggest that alteration of proliferation and mitochondrial function in CADASIL VSMCs might have an effect on vital cellular functions important for CADASIL pathology. -- Highlights: ► CADASIL is an inherited disease of cerebral vascular cells. ► Mitochondrial dysfunction has been implicated in the pathogenesis of CADASIL. ► Lower proliferation rates in CADASIL VSMC. ► Increased irregular and abnormal mitochondria and lower mitochondrial membrane potential in CADASIL VSMCs. ► Reduced mitochondrial connectivity and increased number of mitochondria in CADASIL VSMCs.

  11. Experimental studies of mitochondrial function in CADASIL vascular smooth muscle cells

    Energy Technology Data Exchange (ETDEWEB)

    Viitanen, Matti [Division of Clinical Geriatrics, Department of Neurobiology, Care Sciences and Society, Karolinska Institutet, Stockholm (Sweden); Department of Geriatrics, Turku City Hospital and University of Turku, Turku (Finland); Sundström, Erik [Division of Neurodegeneration, Department of Neurobiology, Care Sciences and Society, Karolinska Institutet, Stockholm (Sweden); Baumann, Marc [Protein Chemistry Unit, Institute of Biomedicine/Anatomy, University of Helsinki, Helsinki (Finland); Poyhonen, Minna [Department of Clinical Genetics, Helsinki University Hospital, HUSLAB, Helsinki (Finland); Tikka, Saara [Protein Chemistry Unit, Institute of Biomedicine/Anatomy, University of Helsinki, Helsinki (Finland); Behbahani, Homira, E-mail: homira.behbahani@ki.se [Division of Clinical Geriatrics, Department of Neurobiology, Care Sciences and Society, Karolinska Institutet, Stockholm (Sweden); Karolinska Institutet Alzheimer' s Disease Research Center, Department of Neurobiology, Care Sciences and Society, Karolinska Institutet, Stockholm (Sweden)

    2013-02-01

    Cerebral Autosomal Dominant Arteriopathy with Subcortical Infarcts and Leukoencephalopathy (CADASIL) is a familiar fatal progressive degenerative disorder characterized by cognitive decline, and recurrent stroke in young adults. Pathological features include a dramatic reduction of brain vascular smooth muscle cells and severe arteriopathy with the presence of granular osmophilic material in the arterial walls. Here we have investigated the cellular and mitochondrial function in vascular smooth muscle cell lines (VSMCs) established from CADASIL mutation carriers (R133C) and healthy controls. We found significantly lower proliferation rates in CADASIL VSMC as compared to VSMC from controls. Cultured CADASIL VSMCs were not more vulnerable than control cells to a number of toxic substances. Morphological studies showed reduced mitochondrial connectivity and increased number of mitochondria in CADASIL VSMCs. Transmission electron microscopy analysis demonstrated increased irregular and abnormal mitochondria in CADASIL VSMCs. Measurements of mitochondrial membrane potential (Δψ{sub m}) showed a lower percentage of fully functional mitochondria in CADASIL VSMCs. For a number of genes previously reported to be changed in CADASIL VSMCs, immunoblotting analysis demonstrated a significantly reduced SOD1 expression. These findings suggest that alteration of proliferation and mitochondrial function in CADASIL VSMCs might have an effect on vital cellular functions important for CADASIL pathology. -- Highlights: ► CADASIL is an inherited disease of cerebral vascular cells. ► Mitochondrial dysfunction has been implicated in the pathogenesis of CADASIL. ► Lower proliferation rates in CADASIL VSMC. ► Increased irregular and abnormal mitochondria and lower mitochondrial membrane potential in CADASIL VSMCs. ► Reduced mitochondrial connectivity and increased number of mitochondria in CADASIL VSMCs.

  12. Towards the therapeutic use of vascular smooth muscle progenitor cells.

    Science.gov (United States)

    Merkulova-Rainon, Tatyana; Broquères-You, Dong; Kubis, Nathalie; Silvestre, Jean-Sébastien; Lévy, Bernard I

    2012-07-15

    Recent advances in the development of alternative proangiogenic and revascularization processes, including recombinant protein delivery, gene therapy, and cell therapy, hold the promise of greater efficacy in the management of cardiovascular disease in the coming years. In particular, vascular progenitor cell-based strategies have emerged as an efficient treatment approach to promote vessel formation and repair and to improve tissue perfusion. During the past decade, considerable progress has been achieved in understanding therapeutic properties of endothelial progenitor cells, while the therapeutic potential of vascular smooth muscle progenitor cells (SMPC) has only recently been explored; the number of the circulating SMPC being correlated with cardiovascular health. Several endogenous SMPC populations with varying phenotypes have been identified and characterized in the peripheral blood, bone marrow, and vascular wall. While the phenotypic entity of vascular SMPC is not fully defined and remains an evolving area of research, SMPC are increasingly recognized to play a special role in cardiovascular biology. In this review, we describe the current approaches used to define vascular SMPC. We further summarize the data on phenotype and functional properties of SMPC from various sources in adults. Finally, we discuss the role of SMPC in cardiovascular disease, including the contribution of SMPC to intimal proliferation, angiogenesis, and atherosclerotic plaque instability as well as the benefits resulting from the therapeutic use of SMPC.

  13. Functional vascular smooth muscle cells derived from human induced pluripotent stem cells via mesenchymal stem cell intermediates

    Science.gov (United States)

    Bajpai, Vivek K.; Mistriotis, Panagiotis; Loh, Yuin-Han; Daley, George Q.; Andreadis, Stelios T.

    2012-01-01

    Aims Smooth muscle cells (SMC) play an important role in vascular homeostasis and disease. Although adult mesenchymal stem cells (MSC) have been used as a source of contractile SMC, they suffer from limited proliferation potential and culture senescence, particularly when originating from older donors. By comparison, human induced pluripotent stem cells (hiPSC) can provide an unlimited source of functional SMC for autologous cell-based therapies and for creating models of vascular disease. Our goal was to develop an efficient strategy to derive functional, contractile SMC from hiPSC. Methods and results We developed a robust, stage-wise, feeder-free strategy for hiPSC differentiation into functional SMC through an intermediate stage of multipotent MSC, which could be coaxed to differentiate into fat, bone, cartilage, and muscle. At this stage, the cells were highly proliferative and displayed higher clonogenic potential and reduced senescence when compared with parental hair follicle mesenchymal stem cells. In addition, when exposed to differentiation medium, the myogenic proteins such as α-smooth muscle actin, calponin, and myosin heavy chain were significantly upregulated and displayed robust fibrillar organization, suggesting the development of a contractile phenotype. Indeed, tissue constructs prepared from these cells exhibited high levels of contractility in response to receptor- and non-receptor-mediated agonists. Conclusion We developed an efficient stage-wise strategy that enabled hiPSC differentiation into contractile SMC through an intermediate population of clonogenic and multipotent MSC. The high yield of MSC and SMC derivation suggests that our strategy may facilitate an acquisition of the large numbers of cells required for regenerative medicine or for studying vascular disease pathophysiology. PMID:22941255

  14. Epidermal growth factor-mediated effects on equine vascular smooth muscle cells

    International Nuclear Information System (INIS)

    Grosenbaugh, D.A.; Amoss, M.S.; Hood, D.M.; Morgan, S.J.; Williams, J.D.

    1988-01-01

    Epidermal growth factor (EGF) receptor binding kinetics and EGF-mediated stimulation of DNA synthesis and cellular proliferation were studied in cultured vascular smooth muscle cells (VSMC) from the equine thoracic aorta. Binding studies, using murine 125 I-labeled EGF, indicate the presence of a single class of high-affinity binding sites, with an estimated maximal binding capacity of 5,800 sites/cells. EGF stimulated [ 3 H]thymidine uptake in confluent quiescent monolayers in a dose-dependent fashion, half-maximal stimulation occurring at 7.5 x 10 -11 M. Likewise, EGF-mediated cellular proliferation was dose dependent under reduced serum concentrations. Equine VSMC contain specific receptors for EGF, and EGF can stimulate DNA synthesis and proliferation in these cultured cells, which suggests that EGF may participate in the proliferative changes observed in equine distal digital peripheral vascular disease

  15. Tissue-Engineered Vascular Rings from Human iPSC-Derived Smooth Muscle Cells

    Directory of Open Access Journals (Sweden)

    Biraja C. Dash

    2016-07-01

    Full Text Available There is an urgent need for an efficient approach to obtain a large-scale and renewable source of functional human vascular smooth muscle cells (VSMCs to establish robust, patient-specific tissue model systems for studying the pathogenesis of vascular disease, and for developing novel therapeutic interventions. Here, we have derived a large quantity of highly enriched functional VSMCs from human induced pluripotent stem cells (hiPSC-VSMCs. Furthermore, we have engineered 3D tissue rings from hiPSC-VSMCs using a facile one-step cellular self-assembly approach. The tissue rings are mechanically robust and can be used for vascular tissue engineering and disease modeling of supravalvular aortic stenosis syndrome. Our method may serve as a model system, extendable to study other vascular proliferative diseases for drug screening. Thus, this report describes an exciting platform technology with broad utility for manufacturing cell-based tissues and materials for various biomedical applications.

  16. Dynamin-related protein inhibitor downregulates reactive oxygen species levels to indirectly suppress high glucose-induced hyperproliferation of vascular smooth muscle cells

    Energy Technology Data Exchange (ETDEWEB)

    Maimaitijiang, Alimujiang; Zhuang, Xinyu; Jiang, Xiaofei; Li, Yong, E-mail: 11211220031@fudan.edu.cn

    2016-03-18

    Hyperproliferation of vascular smooth muscle cells is a pathogenic mechanism common in diabetic vascular complications and is a putatively important therapeutic target. This study investigated multiple levels of biology, including cellular and organellar changes, as well as perturbations in protein synthesis and morphology. Quantitative and qualitative analysis was utilized to assess the effect of mitochondrial dynamic changes and reactive oxygen species(ROS) levels on high-glucose-induced hyperproliferation of vascular smooth muscle cells. The data demonstrated that the mitochondrial fission inhibitor Mdivi-1 and downregulation of ROS levels both effectively inhibited the high-glucose-induced hyperproliferation of vascular smooth muscle cells. Downregulation of ROS levels played a more direct role and ROS levels were also regulated by mitochondrial dynamics. Increased ROS levels induced excessive mitochondrial fission through dynamin-related protein (Drp 1), while Mdivi-1 suppressed the sensitivity of Drp1 to ROS levels, thus inhibiting excessive mitochondrial fission under high-glucose conditions. This study is the first to propose that mitochondrial dynamic changes and ROS levels interact with each other and regulate high-glucose-induced hyperproliferation of vascular smooth muscle cells. This finding provides novel ideas in understanding the pathogenesis of diabetic vascular remodeling and intervention. - Highlights: • Mdivi-1 inhibits VSMC proliferation by lowering ROS level in high-glucose condition. • ROS may be able to induce mitochondrial fission through Drp1 regulation. • Mdivi-1 can suppress the sensitivity of Drp1 to ROS.

  17. Disruption of TGF-β signaling in smooth muscle cell prevents flow-induced vascular remodeling

    Energy Technology Data Exchange (ETDEWEB)

    Gao, Fu [Department of Vascular Surgery, Peking University People’s Hospital, Beijing (China); Chambon, Pierre [Institut de Génétique et de Biologie Moléculaire et Cellulaire (CNRS UMR7104, INSERM U596, ULP, Collége de France) and Institut Clinique de la Souris, ILLKIRCH, Strasbourg (France); Tellides, George [Department of Surgery, Interdepartmental Program in Vascular Biology and Therapeutics, Yale University School of Medicine, New Haven, CT (United States); Kong, Wei [Department of Physiology and Pathophysiology, Basic Medical College of Peking University, Beijing (China); Zhang, Xiaoming, E-mail: rmygxgwk@163.com [Department of Vascular Surgery, Peking University People’s Hospital, Beijing (China); Li, Wei [Department of Vascular Surgery, Peking University People’s Hospital, Beijing (China)

    2014-11-07

    Highlights: • TGF-β signaling in SMC contributes to the flow-induced vascular remodeling. • Disruption of TGF-β signaling in SMC can prevent this process. • Targeting SM-specific Tgfbr2 could be a novel therapeutic strategy for vascular remodeling. - Abstract: Transforming growth factor-β (TGF-β) signaling has been prominently implicated in the pathogenesis of vascular remodeling, especially the initiation and progression of flow-induced vascular remodeling. Smooth muscle cells (SMCs) are the principal resident cells in arterial wall and are critical for arterial remodeling. However, the role of TGF-β signaling in SMC for flow-induced vascular remodeling remains unknown. Therefore, the goal of our study was to determine the effect of TGF-β pathway in SMC for vascular remodeling, by using a genetical smooth muscle-specific (SM-specific) TGF-β type II receptor (Tgfbr2) deletion mice model. Mice deficient in the expression of Tgfbr2 (MyhCre.Tgfbr2{sup f/f}) and their corresponding wild-type background mice (MyhCre.Tgfbr2{sup WT/WT}) underwent partial ligation of left common carotid artery for 1, 2, or 4 weeks. Then the carotid arteries were harvested and indicated that the disruption of Tgfbr2 in SMC provided prominent inhibition of vascular remodeling. And the thickening of carotid media, proliferation of SMC, infiltration of macrophage, and expression of matrix metalloproteinase (MMP) were all significantly attenuated in Tgfbr2 disruption mice. Our study demonstrated, for the first time, that the TGF-β signaling in SMC plays an essential role in flow-induced vascular remodeling and disruption can prevent this process.

  18. Fibroblast growth factor regulates insulin-like growth factor-binding protein production by vascular smooth muscle cells.

    Science.gov (United States)

    Ververis, J; Ku, L; Delafontaine, P

    1994-02-01

    Insulin-like growth factor I is an important mitogen for vascular smooth muscle cells, and its effects are regulated by several binding proteins. Western ligand blotting of conditioned medium from rat aortic smooth muscle cells detected a 24 kDa binding protein and a 28 kDa glycosylated variant of this protein, consistent with insulin-like growth factor binding protein-4 by size. Low amounts of a glycosylated 38 to 42 kDa doublet (consistent with binding protein-3) and a 31 kDa non-glycosylated protein also were present. Basic fibroblast growth factor markedly increased secretion of the 24 kDa binding protein and its 28 kDa glycosylated variant. This effect was dose- and time-dependent and was inhibited by co-incubation with cycloheximide. Crosslinking of [125I]-insulin-like growth factor I to cell monolayers revealed no surface-associated binding proteins, either basally or after agonist treatment. Induction of binding protein production by fibroblast growth factor at sites of vascular injury may be important in vascular proliferative responses in vivo.

  19. Calcification of human vascular smooth muscle cells: associations with osteoprotegerin expression and acceleration by high-dose insulin

    DEFF Research Database (Denmark)

    Olesen, Ping; Knudsen, Kirsten Quyen Nguyen; Wogensen, Lise

    2007-01-01

    Arterial medial calcifications occur often in diabetic individuals as part of the diabetic macroangiopathy. The pathogenesis is unknown, but the presence of calcifications predicts risk of cardiovascular events. We examined the effects of insulin on calcifying smooth muscle cells in vitro...... and measured the expression of the bone-related molecule osteoprotegerin (OPG). Human vascular smooth muscle cells (VSMCs) were grown from aorta from kidney donors. Induction of calcification was performed with beta-glycerophosphate. The influence of insulin (200 microU/ml or 1,000 microU/ml) on calcification...... calcification in human smooth muscle cells from a series of donors after variable time in culture. Decreased OPG amounts were observed from the cells during the accelerated calcification phase. High dose of insulin (1,000 microU/ml) accelerated the calcification, whereas lower concentrations (200 microU/ml) did...

  20. Upregulation of decorin by FXR in vascular smooth muscle cells

    International Nuclear Information System (INIS)

    He Fengtian; Zhang Qiuhong; Kuruba, Ramalinga; Gao Xiang; Li Jiang; Li Yong; Gong Wei; Jiang, Yu; Xie Wen; Li Song

    2008-01-01

    Decorin is a member of the family of small leucine-rich proteoglycans that are present in blood vessels and synthesized by vascular smooth muscle cells (VSMCs). Decorin plays complex roles in both normal vascular physiology and the pathogenesis of various types of vascular disorders. However, the mechanisms of regulation of decorin expression in vasculature are not clearly understood. Particularly little information is available about a role of nuclear receptors in the regulation of decorin expression. In the present study, we report that activation of vascular FXR by a specific ligand resulted in upregulation of decorin at the levels of both mRNA and protein. FXR appears to induce decorin expression at a transcriptional level because (1) upregulation of decorin mRNA expression was abolished by the treatment of a transcription inhibitor, actinomycin D; and (2) decorin promoter activity was significantly increased by activation of FXR. Functional analysis of human decorin promoter identified an imperfect inverted repeat DNA motif, IR8 (-2313TGGTCAtagtgtcaTGACCT-2294), as a likely FXR-responsive element that is involved in decorin regulation

  1. IGF-1 has plaque-stabilizing effects in atherosclerosis by altering vascular smooth muscle cell phenotype

    NARCIS (Netherlands)

    von der Thüsen, Jan H.; Borensztajn, Keren S.; Moimas, Silvia; van Heiningen, Sandra; Teeling, Peter; van Berkel, Theo J. C.; Biessen, Erik A. L.

    2011-01-01

    Insulin-like growth factor-1 (IGF-1) signaling is important for the maintenance of plaque stability in atherosclerosis due to its effects on vascular smooth muscle cell (vSMC) phenotype. To investigate this hypothesis, we studied the effects of the highly inflammatory milieu of the atherosclerotic

  2. CCN5 modulates the antiproliferative effect of heparin and regulates cell motility in vascular smooth muscle cells

    Directory of Open Access Journals (Sweden)

    Castellot John J

    2003-11-01

    Full Text Available Abstract Background Vascular smooth muscle cell (VSMC hyperplasia plays an important role in both chronic and acute vascular pathologies including atherosclerosis and restenosis. Considerable work has focused on the mechanisms regulating VSMC proliferation and motility. Earlier work in our lab revealed a novel growth arrest-specific (gas gene induced in VSMC exposed to the antiproliferative agent heparin. This gene is a member of the CCN family and has been given the name CCN5. The objective of the present study is to elucidate the function of CCN5 protein and to explore its mechanism of action in VSMC. Results Using RNA interference (RNAi, we first demonstrate that CCN5 is required for the antiproliferative effect of heparin in VSMC. We also use this gene knockdown approach to show that CCN5 is an important negative regulator of motility. To explore the mechanism of action of CCN5 on VSMC motility, we use RNAi to demonstrate that knock down of CCN5 up regulates expression of matrix metalloproteinase-2 (MMP-2, an important stimulator of motility in VSMC. In addition, forced expression of CCN5 via adenovirus results in reduced MMP-2 activity, this also corroborates the gene knock down results. Finally, we show that loss of CCN5 expression in VSMC causes changes in VSMC morphology and cytoskeletal organization, including a reduction in the amount and macromolecular assembly of smooth muscle cell α-actin. Conclusions This work provides important new insights into the regulation of smooth muscle cell proliferation and motility by CCN5 and may aid the development of therapies for vascular diseases.

  3. Influence of 103Pd radioactive stent on apoptosis of vascular smooth muscle cells

    International Nuclear Information System (INIS)

    Liu Yingmei; Wu Wei; Chen Xiaochao; Zhang Xuming; Wang Jingfeng; Wei Yulin; Yang Li

    2003-01-01

    Objective: To evaluate the influence of 103 Pd radioactive stent on apoptosis and its relative genes bcl-2 and bax in injured vascular media smooth muscle cells of rabbit abdominal arteries and to investigate the mechanism of 103 Pd radioactive stent for preventing restenosis after angioplasty. Methods: Fifty male New Zealand rabbits were randomized into stent group and 103 Pd stent group. Each group was subdivided into 5 sub-groups. Control group was set up. The study arteries were harvested at 3, 7, 14, 28 and 56 d after stenting and the pathomorphology, apoptosis analysis and in situ hybridization were performed to evaluate the expression of bcl-2 and bax mRNA. Results: The severity of the restenosis in 103 Pd stent group was less than that of stent group. It was most obvious at the 56th day (P 103 Pd stent group had much more apoptosis of vascular smooth muscle cells than stent group did and reached the peak at the 7th day, (14.72±0.53)% vs (12.42±1.13)% (P 103 Pd stent group was much lower than that of stent group at 3 to 28 d. The difference was most obvious at the 28th day after stenting, (18.43± 0.67)% vs (21.55±0.93)% (P 103 Pd stent group was higher than that of stent group, the peak was at the 7th day, (11.17±0.94)% vs (9.30±1.01)%. The ratio of bcl-2/bax in 103 Pd stent group was much lower than that of stent group at 3 to 28 d. Linear correlation analysis showed that there was significant negative correlation between bcl-2 mRNA and apoptosis. Between bax mRNA and apoptosis, the positive correlation was found (P 103 Pd radioactive stent induced more significant apoptosis in vascular media smooth muscle cells by promoting the expression of apoptosis related genes and relieved the expanding of restenosis

  4. S1P receptor signalling and RGS proteins; expression and function in vascular smooth muscle cells and transfected CHO cells

    NARCIS (Netherlands)

    Hendriks-Balk, Mariëlle C.; van Loenen, Pieter B.; Hajji, Najat; Michel, Martin C.; Peters, Stephan L. M.; Alewijnse, Astrid E.

    2009-01-01

    Sphingosine-1-phosphate (S1P) signalling via G protein-coupled receptors is important for the regulation of cell function and differentiation. Specific Regulators of G protein Signalling (RGS) proteins modulate the function of these receptors in many cell types including vascular smooth muscle cells

  5. Oxytocin receptors expressed and coupled to Ca2+ signalling in a human vascular smooth muscle cell line.

    Science.gov (United States)

    Yazawa, H; Hirasawa, A; Horie, K; Saita, Y; Iida, E; Honda, K; Tsujimoto, G

    1996-03-01

    1. In a human vascular smooth muscle cell line (HVSMC), binding experiments with [3H]-arginine8-vasopressin (AVP) have shown the existence of a homogeneous population of binding sites with affinity (Kd value) of 0.65 nM and a maximum number of binding sites (Bmax) of 122 fmol mg-1 protein. 2. Nonlabelled compounds compete for [3H]-AVP binding in the HVSMC membrane with an order of potency of oxytocin > lyspressin > or = AVP > Thr4, Gly7-oxytocin > (beta-mercapto-beta-beta-cyclopentamethylenepropionyl-O-Me Tyr2, Arg8) vasopressin > desmopressin > OPC21268 > OPC31260. This order was markedly different from that observed in rat vascular smooth muscle cells (A10), a well-established V1A receptor system. 3. In HVSMC both oxytocin and AVP increased inositol 1,4,5-trisphosphate (IP3) production and [Ca2+]i response, but the efficacy of the responses was greater for oxytocin than AVP. 4. Reverse transcription-polymerase chain reaction (RT-PCR) assay detected only oxytocin receptor but not V1A or V2 receptors in HVSMC, whereas only V1A receptors were found in A10 cells. 5. In conclusion, in HVSMC only oxytocin receptors are expressed among the vasopressin receptor family, and they coupled to phosphatidyl inositol (PI) turnover/Ca2+ signalling. This unexpected observation should provide new insight into the functional role of the oxytocin receptor in a human vascular smooth muscle cell line.

  6. [Vascular Calcification - Pathological Mechanism and Clinical Application - . Role of vascular smooth muscle cells in vascular calcification].

    Science.gov (United States)

    Kurabayashi, Masahiko

    2015-05-01

    Vascular calcification is commonly seen with aging, chronic kidney disese (CKD), diabetes, and atherosclerosis, and is closely associated with cardiovascular morbidity and mortality. Vascular calcification has long been regarded as the final stage of degeneration and necrosis of arterial wall and a passive, unregulated process. However, it is now known to be an active and tightly regulated process involved with phenotypic transition of vascular smooth muscle cells (VSMC) that resembles bone mineralization. Briefly, calcium deposits of atherosclerotic plaque consist of hydroxyapatite and may appear identical to fully formed lamellar bone. By using a genetic fate mapping strategy, VSMC of the vascular media give rise to the majority of the osteochondrogenic precursor- and chondrocyte-like cells observed in the calcified arterial media of MGP (- / -) mice. Osteogenic differentiation of VSMC is characterized by the expression of bone-related molecules including bone morphogenetic protein (BMP) -2, Msx2 and osteopontin, which are produced by osteoblasts and chondrocytes. Our recent findings are that (i) Runx2 and Notch1 induce osteogenic differentiation, and (ii) advanced glycation end-product (AGE) /receptor for AGE (RAGE) and palmitic acid promote osteogenic differentiation of VSMC. To understand of the molecular mechanisms of vascular calcification is now under intensive research area.

  7. Effects of the dual TP receptor antagonist and thromboxane synthase inhibitor EV-077 on human endothelial and vascular smooth muscle cells

    International Nuclear Information System (INIS)

    Petri, Marcelo H.; Tellier, Céline; Michiels, Carine; Ellertsen, Ingvill; Dogné, Jean-Michel; Bäck, Magnus

    2013-01-01

    Highlights: •EV-077 reduced TNF-α induced inflammation in endothelial cells. •The thromboxane mimetic U69915 enhanced vascular smooth muscle cell proliferation. •EV-077 inhibited smooth muscle cell proliferation. -- Abstract: The prothrombotic mediator thromboxane A 2 is derived from arachidonic acid metabolism through the cyclooxygenase and thromboxane synthase pathways, and transduces its effect through the thromboxane prostanoid (TP) receptor. The aim of this study was to determine the effect of the TP receptor antagonist and thromboxane synthase inhibitor EV-077 on inflammatory markers in human umbilical vein endothelial cells and on human coronary artery smooth muscle cell proliferation. To this end, mRNA levels of different proinflammatory mediators were studied by real time quantitative PCR, supernatants were analyzed by enzyme immune assay, and cell proliferation was assessed using WST-1. EV-077 significantly decreased mRNA levels of ICAM-1 and PTX3 after TNFα incubation, whereas concentrations of 6-keto PGF1α in supernatants of endothelial cells incubated with TNFα were significantly increased after EV-077 treatment. Although U46619 did not alter coronary artery smooth muscle cell proliferation, this thromboxane mimetic enhanced the proliferation induced by serum, insulin and growth factors, which was significantly inhibited by EV-077. In conclusion, EV-077 inhibited TNFα-induced endothelial inflammation and reduced the enhancement of smooth muscle cell proliferation induced by a thromboxane mimetic, supporting that the thromboxane pathway may be associated with early atherosclerosis in terms of endothelial dysfunction and vascular hypertrophy

  8. Effects of the dual TP receptor antagonist and thromboxane synthase inhibitor EV-077 on human endothelial and vascular smooth muscle cells

    Energy Technology Data Exchange (ETDEWEB)

    Petri, Marcelo H. [Department of Medicine, Karolinska Institutet and Center for Molecular Medicine, Karolinska University Hospital, Stockholm (Sweden); Tellier, Céline; Michiels, Carine [NARILIS, URBC, University of Namur, Namur (Belgium); Ellertsen, Ingvill [Department of Medicine, Karolinska Institutet and Center for Molecular Medicine, Karolinska University Hospital, Stockholm (Sweden); Dogné, Jean-Michel [Department of Pharmacy, Namur Thrombosis and Hemostasis Center, University of Namur, Namur (Belgium); Bäck, Magnus, E-mail: Magnus.Back@ki.se [Department of Medicine, Karolinska Institutet and Center for Molecular Medicine, Karolinska University Hospital, Stockholm (Sweden)

    2013-11-15

    Highlights: •EV-077 reduced TNF-α induced inflammation in endothelial cells. •The thromboxane mimetic U69915 enhanced vascular smooth muscle cell proliferation. •EV-077 inhibited smooth muscle cell proliferation. -- Abstract: The prothrombotic mediator thromboxane A{sub 2} is derived from arachidonic acid metabolism through the cyclooxygenase and thromboxane synthase pathways, and transduces its effect through the thromboxane prostanoid (TP) receptor. The aim of this study was to determine the effect of the TP receptor antagonist and thromboxane synthase inhibitor EV-077 on inflammatory markers in human umbilical vein endothelial cells and on human coronary artery smooth muscle cell proliferation. To this end, mRNA levels of different proinflammatory mediators were studied by real time quantitative PCR, supernatants were analyzed by enzyme immune assay, and cell proliferation was assessed using WST-1. EV-077 significantly decreased mRNA levels of ICAM-1 and PTX3 after TNFα incubation, whereas concentrations of 6-keto PGF1α in supernatants of endothelial cells incubated with TNFα were significantly increased after EV-077 treatment. Although U46619 did not alter coronary artery smooth muscle cell proliferation, this thromboxane mimetic enhanced the proliferation induced by serum, insulin and growth factors, which was significantly inhibited by EV-077. In conclusion, EV-077 inhibited TNFα-induced endothelial inflammation and reduced the enhancement of smooth muscle cell proliferation induced by a thromboxane mimetic, supporting that the thromboxane pathway may be associated with early atherosclerosis in terms of endothelial dysfunction and vascular hypertrophy.

  9. Akt1/PKB upregulation leads to vascular smooth muscle cell hypertrophy and polyploidization

    OpenAIRE

    Hixon, Mary L.; Muro-Cacho, Carlos; Wagner, Mark W.; Obejero-Paz, Carlos; Millie, Elise; Fujio, Yasushi; Kureishi, Yasuko; Hassold, Terry; Walsh, Kenneth; Gualberto, Antonio

    2000-01-01

    Vascular smooth muscle cells (VSMCs) at capacitance arteries of hypertensive individuals and animals undergo marked age- and blood pressure–dependent polyploidization and hypertrophy. We show here that VSMCs at capacitance arteries of rat models of hypertension display high levels of Akt1/PKB protein and activity. Gene transfer of Akt1 to VSMCs isolated from a normotensive rat strain was sufficient to abrogate the activity of the mitotic spindle cell–cycle checkpoint, promoting polyploidizati...

  10. Redundant control of migration and adhesion by ERM proteins in vascular smooth muscle cells

    International Nuclear Information System (INIS)

    Baeyens, Nicolas; Latrache, Iman; Yerna, Xavier; Noppe, Gauthier; Horman, Sandrine; Morel, Nicole

    2013-01-01

    Highlights: •The three ERM proteins are expressed in vascular smooth muscle cell. •ERM depletion inhibited PDGF-evoked migration redundantly. •ERM depletion increased cell adhesion redundantly. •ERM depletion did not affect PDGF-evoked Ca signal, Rac1 activation, proliferation. •ERM proteins control PDGF-induced migration by regulating adhesion. -- Abstract: Ezrin, radixin, and moesin possess a very similar structure with a C-terminal actin-binding domain and a N-terminal FERM interacting domain. They are known to be involved in cytoskeleton organization in several cell types but their function in vascular smooth muscle cells (VSMC) is still unknown. The aim of this study was to investigate the role of ERM proteins in cell migration induced by PDGF, a growth factor involved in pathophysiological processes like angiogenesis or atherosclerosis. We used primary cultured VSMC obtained from rat aorta, which express the three ERM proteins. Simultaneous depletion of the three ERM proteins with specific siRNAs abolished the effects of PDGF on cell architecture and migration and markedly increased cell adhesion and focal adhesion size, while these parameters were only slightly affected by depletion of ezrin, radixin or moesin alone. Rac1 activation, cell proliferation, and Ca 2+ signal in response to PDGF were unaffected by ERM depletion. These results indicate that ERM proteins exert a redundant control on PDGF-induced VSMC migration by regulating focal adhesion turn-over and cell adhesion to substrate

  11. Redundant control of migration and adhesion by ERM proteins in vascular smooth muscle cells

    Energy Technology Data Exchange (ETDEWEB)

    Baeyens, Nicolas; Latrache, Iman; Yerna, Xavier [Laboratory of Cell Physiology, IoNS, Université Catholique de Louvain (Belgium); Noppe, Gauthier; Horman, Sandrine [Pôle de Recherche Cardiovasculaire, IREC, Université Catholique de Louvain (Belgium); Morel, Nicole, E-mail: nicole.morel@uclouvain.be [Laboratory of Cell Physiology, IoNS, Université Catholique de Louvain (Belgium)

    2013-11-22

    Highlights: •The three ERM proteins are expressed in vascular smooth muscle cell. •ERM depletion inhibited PDGF-evoked migration redundantly. •ERM depletion increased cell adhesion redundantly. •ERM depletion did not affect PDGF-evoked Ca signal, Rac1 activation, proliferation. •ERM proteins control PDGF-induced migration by regulating adhesion. -- Abstract: Ezrin, radixin, and moesin possess a very similar structure with a C-terminal actin-binding domain and a N-terminal FERM interacting domain. They are known to be involved in cytoskeleton organization in several cell types but their function in vascular smooth muscle cells (VSMC) is still unknown. The aim of this study was to investigate the role of ERM proteins in cell migration induced by PDGF, a growth factor involved in pathophysiological processes like angiogenesis or atherosclerosis. We used primary cultured VSMC obtained from rat aorta, which express the three ERM proteins. Simultaneous depletion of the three ERM proteins with specific siRNAs abolished the effects of PDGF on cell architecture and migration and markedly increased cell adhesion and focal adhesion size, while these parameters were only slightly affected by depletion of ezrin, radixin or moesin alone. Rac1 activation, cell proliferation, and Ca{sup 2+} signal in response to PDGF were unaffected by ERM depletion. These results indicate that ERM proteins exert a redundant control on PDGF-induced VSMC migration by regulating focal adhesion turn-over and cell adhesion to substrate.

  12. Regulation of insulin-like growth factor I receptors on vascular smooth muscle cells by growth factors and phorbol esters.

    Science.gov (United States)

    Ververis, J J; Ku, L; Delafontaine, P

    1993-06-01

    Insulin-like growth factor I (IGF I) is an important mitogen for vascular smooth muscle cells. To characterize regulation of vascular IGF I receptors, we performed radioligand displacement experiments using rat aortic smooth muscle cells (RASMs). Serum deprivation for 48 hours caused a 40% decrease in IGF I receptor number. Exposure of quiescent RASMs to platelet-derived growth factor (PDGF), fibroblast growth factor (FGF), or angiotensin II (Ang II) caused a 1.5-2.0-fold increase in IGF I receptors per cell. After FGF exposure, there was a marked increase in the mitogenic response to IGF I. IGF I downregulated its receptors in the presence of platelet-poor plasma. Stimulation of protein kinase C (PKC) by exposure of quiescent RASMs to phorbol 12-myristate 13-acetate caused a biphasic response in IGF I binding; there was a 42% decrease in receptor number at 45 minutes and a 238% increase at 24 hours. To determine the role of PKC in growth factor-induced regulation of IGF I receptors, we downregulated PKC by exposing RASMs to phorbol 12,13-dibutyrate (PDBu) for 48 hours. PDGF- and FGF- but not Ang II-mediated upregulation of IGF I receptors was completely inhibited in PDBu-treated cells. Thus, acute PKC activation by phorbol esters inhibits IGF I binding, whereas chronic PKC activation increases IGF I binding. PDGF and FGF but not Ang II regulate vascular IGF I receptors through a PKC-dependent pathway. These data provide new insights into the regulation of vascular smooth muscle cell IGF I receptors in vitro and are of potential importance in characterizing vascular proliferative responses in vivo.

  13. Mitochondrial metabolism and the control of vascular smooth muscle cell proliferation

    Directory of Open Access Journals (Sweden)

    Mario eChiong

    2014-12-01

    Full Text Available Differentiation and dedifferentiation of vascular smooth muscle cells (VSMCs are essential processes of vascular development. VSMCs have biosynthetic, proliferative and contractile roles in the vessel wall. Alterations in the differentiated state of the VSMCs play a critical role in the pathogenesis of a variety of cardiovascular diseases, including atherosclerosis, hypertension and vascular stenosis. This review provides an overview of the current state of knowledge of molecular mechanisms involved in the control of VSMC proliferation, with particular focus on mitochondrial metabolism. Mitochondrial activity can be controlled by regulating mitochondrial dynamics, i.e. mitochondrial fusion and fission, and by regulating mitochondrial calcium handling through the interaction with the endoplasmic reticulum (ER. Alterations in both VSMC proliferation and mitochondrial function can be triggered by dysregulation of mitofusin-2, a small GTPase associated with mitochondrial fusion and mitochondrial-ER interaction. Several lines of evidence highlight the relevance of mitochondrial metabolism in the control of VSMC proliferation, indicating a new area to be explored in the treatment of vascular diseases.

  14. Adhesion, Growth, and Maturation of Vascular Smooth Muscle Cells on Low-Density Polyethylene Grafted with Bioactive Substances

    Czech Academy of Sciences Publication Activity Database

    Pařízek, Martin; Kasálková-Slepičková, N.; Bačáková, Lucie; Švindrych, Zdeněk; Slepička, P.; Bačáková, Markéta; Lisá, Věra; Švorčík, V.

    2013-01-01

    Roč. 2013, č. 2013 (2013), s. 371430 ISSN 2314-6133 R&D Projects: GA ČR(CZ) GBP108/12/G108 Institutional support: RVO:67985823 Keywords : biotechnology * tissue replacements * vascular smooth muscle cells * adhesion * modification Subject RIV: JJ - Other Materials

  15. Effects of x-irradiation on growth of vascular smooth muscle cells

    International Nuclear Information System (INIS)

    Dzoga, K.F.; Dimitrievich, G.S.; Sutton, H.G.; Griem, M.L.

    1984-01-01

    Effects of x-irradiation doses ranging from 0-2000 rads on vascular smooth muscle cells were measured. Explant cultures were from the medial layers of aortas from New Zealand rabbits. X-irradiation was delivered to narrow mediastinal port using a 250 kV Maxitron at a rate of 80 rads/min. and a S-C distance of 60 cm. Explantation was done either immediately following radiation or five days later. Two parameters were used to determine post-irradiation growth potential of these cells: number of outgrowing cells per seeded explant and size and number of cells/culture. Results were expressed as fraction of control. Irradiation immediately before explantation reduced number of cells/ explant 10% for 250 rads and over 50% for 500 rads. Doses of 1000 rads and over resulted in reductions of over 70% in number of growing explants and culture size. When five days were allowed to elapse between x-irradiation and explantation the same parameters were not significantly affected for doses of 500 rads or less. Doses of 1000 rads resulted in a reduction in number of cells of 40% and 2000 rads of over 80%. These results suggest the presence of a population of vascular repair cells five days following irradiation treatment. The nature of these cells is discussed

  16. Differentiation and Application of Induced Pluripotent Stem Cell-Derived Vascular Smooth Muscle Cells.

    Science.gov (United States)

    Maguire, Eithne Margaret; Xiao, Qingzhong; Xu, Qingbo

    2017-11-01

    Vascular smooth muscle cells (VSMCs) play a role in the development of vascular disease, for example, neointimal formation, arterial aneurysm, and Marfan syndrome caused by genetic mutations in VSMCs, but little is known about the mechanisms of the disease process. Advances in induced pluripotent stem cell technology have now made it possible to derive VSMCs from several different somatic cells using a selection of protocols. As such, researchers have set out to delineate key signaling processes involved in triggering VSMC gene expression to grasp the extent of gene regulatory networks involved in phenotype commitment. This technology has also paved the way for investigations into diseases affecting VSMC behavior and function, which may be treatable once an identifiable culprit molecule or gene has been repaired. Moreover, induced pluripotent stem cell-derived VSMCs are also being considered for their use in tissue-engineered blood vessels as they may prove more beneficial than using autologous vessels. Finally, while several issues remains to be clarified before induced pluripotent stem cell-derived VSMCs can become used in regenerative medicine, they do offer both clinicians and researchers hope for both treating and understanding vascular disease. In this review, we aim to update the recent progress on VSMC generation from stem cells and the underlying molecular mechanisms of VSMC differentiation. We will also explore how the use of induced pluripotent stem cell-derived VSMCs has changed the game for regenerative medicine by offering new therapeutic avenues to clinicians, as well as providing researchers with a new platform for modeling of vascular disease. © 2017 American Heart Association, Inc.

  17. Sphingosine-1-phosphate regulates RGS2 and RGS16 mRNA expression in vascular smooth muscle cells

    NARCIS (Netherlands)

    Hendriks-Balk, Mariëlle C.; Hajji, Najat; van Loenen, Pieter B.; Michel, Martin C.; Peters, Stephan L. M.; Alewijnse, Astrid E.

    2009-01-01

    Regulator of G protein signalling (RGS) protein expression is altered under growth promoting conditions in vascular smooth muscle cells (VSMCs). Since sphingosine-1-phosphate (S1P) is an important growth stimulatory factor, we investigated whether stimulation of VSMCs with S1P results in alterations

  18. Ageing induced vascular smooth muscle cell senescence in atherosclerosis.

    Science.gov (United States)

    Uryga, Anna K; Bennett, Martin R

    2016-04-15

    Atherosclerosis is a disease of ageing in that its incidence and prevalence increase with age. However, atherosclerosis is also associated with biological ageing, manifest by a number of typical hallmarks of ageing in the atherosclerotic plaque. Thus, accelerated biological ageing may be superimposed on the effects of chronological ageing in atherosclerosis. Tissue ageing is seen in all cells that comprise the plaque, but particularly in vascular smooth muscle cells (VSMCs). Hallmarks of ageing include evidence of cell senescence, DNA damage (including telomere attrition), mitochondrial dysfunction, a pro-inflammatory secretory phenotype, defects in proteostasis, epigenetic changes, deregulated nutrient sensing, and exhaustion of progenitor cells. In this model, initial damage to DNA (genomic, telomeric, mitochondrial and epigenetic changes) results in a number of cellular responses (cellular senescence, deregulated nutrient sensing and defects in proteostasis). Ultimately, ongoing damage and attempts at repair by continued proliferation overwhelm reparative capacity, causing loss of specialised cell functions, cell death and inflammation. This review summarises the evidence for accelerated biological ageing in atherosclerosis, the functional consequences of cell ageing on cells comprising the plaque, and the causal role that VSMC senescence plays in atherogenesis. © 2015 The Authors. The Journal of Physiology © 2015 The Physiological Society.

  19. Vascular smooth muscle cells exhibit a progressive loss of rigidity with serial culture passaging.

    Science.gov (United States)

    Dinardo, Carla Luana; Venturini, Gabriela; Omae, Samantha Vieira; Zhou, Enhua H; da Motta-Leal-Filho, Joaquim Maurício; Dariolli, Rafael; Krieger, José Eduardo; Alencar, Adriano Mesquita; Costa Pereira, Alexandre

    2012-01-01

    One drawback of in vitro cell culturing is the dedifferentiation process that cells experience. Smooth muscle cells (SMC) also change molecularly and morphologically with long term culture. The main objective of this study was to evaluate if culture passages interfere in vascular SMC mechanical behavior. SMC were obtained from five different porcine arterial beds. Optical magnetic twisting cytometry (OMTC) was used to characterize mechanically vascular SMC from different cultures in distinct passages and confocal microscopy/western blotting, to evaluate cytoskeleton and extracellular matrix proteins. We found that vascular SMC rigidity or viscoelastic complex modulus (G) decreases with progression of passages. A statistically significant negative correlation between G and passage was found in four of our five cultures studied. Phalloidin-stained SMC from higher passages exhibited lower mean signal intensity per cell (confocal microscopy) and quantitative western blotting analysis showed a decrease in collagen I content throughout passages. We concluded that vascular SMC progressively lose their stiffness with serial culture passaging. Thus, limiting the number of passages is essential for any experiment measuring viscoelastic properties of SMC in culture.

  20. A Dominant-Negative PPARγ Mutant Promotes Cell Cycle Progression and Cell Growth in Vascular Smooth Muscle Cells

    Directory of Open Access Journals (Sweden)

    Joey Z. Liu

    2009-01-01

    Full Text Available PPARγ ligands have been shown to have antiproliferative effects on many cell types. We herein report that a synthetic dominant-negative (DN PPARγ mutant functions like a growth factor to promote cell cycle progression and cell proliferation in human coronary artery smooth muscle cells (CASMCs. In quiescent CASMCs, adenovirus-expressed DN-PPARγ promoted G1→S cell cycle progression, enhanced BrdU incorporation, and increased cell proliferation. DN-PPARγ expression also markedly enhanced positive regulators of the cell cycle, increasing Rb and CDC2 phosphorylation and the expression of cyclin A, B1, D1, and MCM7. Conversely, overexpression of wild-type (WT or constitutively-active (CA PPARγ inhibited cell cycle progression and the activity and expression of positive regulators of the cell cycle. DN-PPARγ expression, however, did not up-regulate positive cell cycle regulators in PPARγ-deficient cells, strongly suggesting that DN-PPARγ effects on cell cycle result from blocking the function of endogenous wild-type PPARγ. DN-PPARγ expression enhanced phosphorylation of ERK MAPKs. Furthermore, the ERK specific-inhibitor PD98059 blocked DN-PPARγ-induced phosphorylation of Rb and expression of cyclin A and MCM7. Our data thus suggest that DN-PPARγ promotes cell cycle progression and cell growth in CASMCs by modulating fundamental cell cycle regulatory proteins and MAPK mitogenic signaling pathways in vascular smooth muscle cells (VSMCs.

  1. Luteolin Ameliorates Hypertensive Vascular Remodeling through Inhibiting the Proliferation and Migration of Vascular Smooth Muscle Cells

    Directory of Open Access Journals (Sweden)

    Jie Su

    2015-01-01

    Full Text Available Objectives. Preliminary researches showed that luteolin was used to treat hypertension. However, it is still unclear whether luteolin has effect on the hypertensive complication such as vascular remodeling. The present study was designed to investigate the effect of luteolin on the hypertensive vascular remodeling and its molecular mechanism. Method and Results. We evaluated the effect of luteolin on aorta thickening of hypertension in spontaneous hypertensive rats (SHRs and found that luteolin could significantly decrease the blood pressure and media thickness of aorta in vivo. Luteolin could inhibit angiotensin II- (Ang II- induced proliferation and migration of vascular smooth muscle cells (VSMCs. Dichlorofluorescein diacetate (DCFH-DA staining result showed that luteolin reduced Ang II-stimulated ROS production in VSMCs. Furthermore, western blot and gelatin zymography results showed that luteolin treatment leaded to a decrease in ERK1/2, p-ERK1/2, p-p38, MMP2, and proliferating cell nuclear antigen (PCNA protein level. Conclusion. These data support that luteolin can ameliorate hypertensive vascular remodeling by inhibiting the proliferation and migration of Ang II-induced VSMCs. Its mechanism is mediated by the regulation of MAPK signaling pathway and the production of ROS.

  2. Smooth muscle LDL receptor-related protein-1 deletion induces aortic insufficiency and promotes vascular cardiomyopathy in mice.

    Directory of Open Access Journals (Sweden)

    Joshua E Basford

    Full Text Available Valvular disease is common in patients with Marfan syndrome and can lead to cardiomyopathy. However, some patients develop cardiomyopathy in the absence of hemodynamically significant valve dysfunction, suggesting alternative mechanisms of disease progression. Disruption of LDL receptor-related protein-1 (Lrp1 in smooth muscle cells has been shown to cause vascular pathologies similar to Marfan syndrome, with activation of smooth muscle cells, vascular dysfunction and aortic aneurysms. This study used echocardiography and blood pressure monitoring in mouse models to determine whether inactivation of Lrp1 in vascular smooth muscle leads to cardiomyopathy, and if so, whether the mechanism is a consequence of valvular disease. Hemodynamic changes during treatment with captopril were also assessed. Dilation of aortic roots was observed in young Lrp1-knockout mice and progressed as they aged, whereas no significant aortic dilation was detected in wild type littermates. Diastolic blood pressure was lower and pulse pressure higher in Lrp1-knockout mice, which was normalized by treatment with captopril. Aortic dilation was followed by development of aortic insufficiency and subsequent dilated cardiomyopathy due to valvular disease. Thus, smooth muscle cell Lrp1 deficiency results in aortic dilation and insufficiency that causes secondary cardiomyopathy that can be improved by captopril. These findings provide novel insights into mechanisms of cardiomyopathy associated with vascular activation and offer a new model of valvular cardiomyopathy.

  3. Differential regulation of protease activated receptor-1 and tissue plasminogen activator expression by shear stress in vascular smooth muscle cells

    Science.gov (United States)

    Papadaki, M.; Ruef, J.; Nguyen, K. T.; Li, F.; Patterson, C.; Eskin, S. G.; McIntire, L. V.; Runge, M. S.

    1998-01-01

    Recent studies have demonstrated that vascular smooth muscle cells are responsive to changes in their local hemodynamic environment. The effects of shear stress on the expression of human protease activated receptor-1 (PAR-1) and tissue plasminogen activator (tPA) mRNA and protein were investigated in human aortic smooth muscle cells (HASMCs). Under conditions of low shear stress (5 dyn/cm2), PAR-1 mRNA expression was increased transiently at 2 hours compared with stationary control values, whereas at high shear stress (25 dyn/cm2), mRNA expression was decreased (to 29% of stationary control; Pmuscle cells, indicating that the effects of shear stress on human PAR-1 were not species-specific. Flow cytometry and ELISA techniques using rat smooth muscle cells and HASMCs, respectively, provided evidence that shear stress exerted similar effects on cell surface-associated PAR-1 and tPA protein released into the conditioned media. The decrease in PAR-1 mRNA and protein had functional consequences for HASMCs, such as inhibition of [Ca2+] mobilization in response to thrombin stimulation. These data indicate that human PAR-1 and tPA gene expression are regulated differentially by shear stress, in a pattern consistent with their putative roles in several arterial vascular pathologies.

  4. Vascular smooth muscle cell phenotypic changes in patients with Marfan syndrome.

    Science.gov (United States)

    Crosas-Molist, Eva; Meirelles, Thayna; López-Luque, Judit; Serra-Peinado, Carla; Selva, Javier; Caja, Laia; Gorbenko Del Blanco, Darya; Uriarte, Juan José; Bertran, Esther; Mendizábal, Yolanda; Hernández, Vanessa; García-Calero, Carolina; Busnadiego, Oscar; Condom, Enric; Toral, David; Castellà, Manel; Forteza, Alberto; Navajas, Daniel; Sarri, Elisabet; Rodríguez-Pascual, Fernando; Dietz, Harry C; Fabregat, Isabel; Egea, Gustavo

    2015-04-01

    Marfan's syndrome is characterized by the formation of ascending aortic aneurysms resulting from altered assembly of extracellular matrix microfibrils and chronic tissue growth factor (TGF)-β signaling. TGF-β is a potent regulator of the vascular smooth muscle cell (VSMC) phenotype. We hypothesized that as a result of the chronic TGF-β signaling, VSMC would alter their basal differentiation phenotype, which could facilitate the formation of aneurysms. This study explores whether Marfan's syndrome entails phenotypic alterations of VSMC and possible mechanisms at the subcellular level. Immunohistochemical and Western blotting analyses of dilated aortas from Marfan patients showed overexpression of contractile protein markers (α-smooth muscle actin, smoothelin, smooth muscle protein 22 alpha, and calponin-1) and collagen I in comparison with healthy aortas. VSMC explanted from Marfan aortic aneurysms showed increased in vitro expression of these phenotypic markers and also of myocardin, a transcription factor essential for VSMC-specific differentiation. These alterations were generally reduced after pharmacological inhibition of the TGF-β pathway. Marfan VSMC in culture showed more robust actin stress fibers and enhanced RhoA-GTP levels, which was accompanied by increased focal adhesion components and higher nuclear localization of myosin-related transcription factor A. Marfan VSMC and extracellular matrix measured by atomic force microscopy were both stiffer than their respective controls. In Marfan VSMC, both in tissue and in culture, there are variable TGF-β-dependent phenotypic changes affecting contractile proteins and collagen I, leading to greater cellular and extracellular matrix stiffness. Altogether, these alterations may contribute to the known aortic rigidity that precedes or accompanies Marfan's syndrome aneurysm formation. © 2015 American Heart Association, Inc.

  5. Effect of lovastatin on rabbit vascular smooth muscle cells

    International Nuclear Information System (INIS)

    Luan Zhaoxia; Pei Zhuguo

    2003-01-01

    Objective: To investigate the effect of lovastatin on binding activity of nuclear factor activator protein-1 (AP-1) to NF-κB and the expression of matrix metalloproteinase-9 (MMP-9) in rabbit vascular smooth muscle cells (VSMCs). Methods: The oligonucleotide corresponding to the consensus NF-κB element or the consensus AP-1 element was labeled by [γ- 32 P]-ATP. AP-1 and NF-κB binding activity was detected by electrophoretic mobility shift assay (EMSA), expression of MMP-9 was detected by zymography. Results: Lovastatin inhibited the expression of MMP-9 in a dose-dependent manner, this effect was reversed by mevalonate and GGPP but not by squalene; lovastatin significantly decreased AP-1 and NF-κB binding activity. Conclusion: Lovastatin decreased AP-1 and NF-κB binding activity and inhibited MMP-9 expression in rabbit VSMCs by the way of inhibiting prenylation of protein but not by cholestrol-lowering, and this might be the mechanism of its arteriosclerostic plaque stabilizing effects

  6. A role for mitochondrial oxidants in stress-induced premature senescence of human vascular smooth muscle cells

    Directory of Open Access Journals (Sweden)

    Yogita Mistry

    2013-01-01

    Full Text Available Mitochondria are a major source of cellular oxidants and have been implicated in aging and associated pathologies, notably cardiovascular diseases. Vascular cell senescence is observed in experimental and human cardiovascular pathologies. Our previous data highlighted a role for angiotensin II in the induction of telomere-dependent and -independent premature senescence of human vascular smooth muscle cells and suggested this was due to production of superoxide by NADPH oxidase. However, since a role for mitochondrial oxidants was not ruled out we hypothesise that angiotensin II mediates senescence by mitochondrial superoxide generation and suggest that inhibition of superoxide may prevent vascular smooth muscle cell aging in vitro. Cellular senescence was induced using a stress-induced premature senescence protocol consisting of three successive once-daily exposure of cells to 1×10−8 mol/L angiotensin II and was dependent upon the type-1 angiotensin II receptor. Angiotensin stimulated NADPH-dependent superoxide production as estimated using lucigenin chemiluminescence in cell lysates and this was attenuated by the mitochondrial electron transport chain inhibitor, rotenone. Angiotensin also resulted in an increase in mitoSOX fluorescence indicating stimulation of mitochondrial superoxide. Significantly, the induction of senescence by angiotensin II was abrogated by rotenone and by the mitochondria-targeted superoxide dismutase mimetic, mitoTEMPO. These data suggest that mitochondrial superoxide is necessary for the induction of stress-induced premature senescence by angiotensin II and taken together with other data suggest that mitochondrial cross-talk with NADPH oxidases, via as yet unidentified signalling pathways, is likely to play a key role.

  7. Contact-mediated and humoral communication between vascular endothelial and smooth muscle cells in vitro

    International Nuclear Information System (INIS)

    Davies, P.F.

    1986-01-01

    Vascular endothelial cells (EC) and smooth muscle cells (SMC) co-exist in close apposition to each other in all blood vessels except capillaries. Investigations of the metabolic interactions that may occur between these cells are essential to an understanding of vascular homeostasis and the pathogenesis of atherosclerosis. The authors have developed two in vitro models of co-temporal vascular cell communication. The first facilitates reversible microcarrier-mediated gap junctional communication between EC and SMC monolayers. When either EC or SMC were prelabelled with 3 H-uridine, intracellular nucleotide rapidly transferred across the region of heterocellular attachment to the complementary cell population. Cytoplasmic continuity between EC and SMC allowed metabolic cooperation via ions and small molecules (<1.5 KD). Thus, vascular reactivity, particularly in the microcirculation where myoendothelial gap junctions have been observed, may involve cytoplasmic second messengers transported from EC to SMC. In the second model, humoral communication was established between separated cultures of EC and SMC which shared the same culture medium. Endothelial-specific stimulation of SMC growth and lipoprotein metabolism via soluble factors was demonstrated. Two mechanisms of stimulation of SMC lipoprotein metabolism were identified; one endothelial derived mitogen-dependent, the other mitogen-independent which was mediated via low molecular weight endothelial cell products

  8. Vascular Smooth Muscle Cells From Hypertensive Patient-Derived Induced Pluripotent Stem Cells to Advance Hypertension Pharmacogenomics.

    Science.gov (United States)

    Biel, Nikolett M; Santostefano, Katherine E; DiVita, Bayli B; El Rouby, Nihal; Carrasquilla, Santiago D; Simmons, Chelsey; Nakanishi, Mahito; Cooper-DeHoff, Rhonda M; Johnson, Julie A; Terada, Naohiro

    2015-12-01

    Studies in hypertension (HTN) pharmacogenomics seek to identify genetic sources of variable antihypertensive drug response. Genetic association studies have detected single-nucleotide polymorphisms (SNPs) that link to drug responses; however, to understand mechanisms underlying how genetic traits alter drug responses, a biological interface is needed. Patient-derived induced pluripotent stem cells (iPSCs) provide a potential source for studying otherwise inaccessible tissues that may be important to antihypertensive drug response. The present study established multiple iPSC lines from an HTN pharmacogenomics cohort. We demonstrated that established HTN iPSCs can robustly and reproducibly differentiate into functional vascular smooth muscle cells (VSMCs), a cell type most relevant to vasculature tone control. Moreover, a sensitive traction force microscopy assay demonstrated that iPSC-derived VSMCs show a quantitative contractile response on physiological stimulus of endothelin-1. Furthermore, the inflammatory chemokine tumor necrosis factor α induced a typical VSMC response in iPSC-derived VSMCs. These studies pave the way for a large research initiative to decode biological significance of identified SNPs in hypertension pharmacogenomics. Treatment of hypertension remains suboptimal, and a pharmacogenomics approach seeks to identify genetic biomarkers that could be used to guide treatment decisions; however, it is important to understand the biological underpinnings of genetic associations. Mouse models do not accurately recapitulate individual patient responses based on their genetics, and hypertension-relevant cells are difficult to obtain from patients. Induced pluripotent stem cell (iPSC) technology provides a great interface to bring patient cells with their genomic data into the laboratory and to study hypertensive responses. As an initial step, the present study established an iPSC bank from patients with primary hypertension and demonstrated an effective

  9. Verapamil stereoisomers induce antiproliferative effects in vascular smooth muscle cells via autophagy

    Energy Technology Data Exchange (ETDEWEB)

    Salabei, Joshua K. [Diabetes and Obesity Center, University of Louisville, Louisville, KY 40202 (United States); Department of Biochemistry and Molecular Biology, University of Louisville, Louisville, KY 40202 (United States); Balakumaran, Arun [Department of Pathology, University of Texas Medical Branch, Galveston, TX 77555‐0438 (United States); Frey, Justin C. [Department of Biology, University of Wisconsin-Eau Claire, Eau Claire, WI 54702 (United States); Boor, Paul J. [Department of Pathology, University of Texas Medical Branch, Galveston, TX 77555‐0438 (United States); Treinen-Moslen, Mary [Department of Internal Medicine, University of Texas Medical Branch, Galveston, TX 77555‐0609 (United States); Department of Pathology, University of Texas Medical Branch, Galveston, TX 77555‐0438 (United States); Conklin, Daniel J., E-mail: dj.conklin@louisville.edu [Diabetes and Obesity Center, University of Louisville, Louisville, KY 40202 (United States); Division of Cardiovascular Medicine, University of Louisville, Louisville, KY 40202 (United States); Department of Biology, University of Wisconsin-Eau Claire, Eau Claire, WI 54702 (United States); Department of Pathology, University of Texas Medical Branch, Galveston, TX 77555‐0438 (United States)

    2012-08-01

    Calcium channel blockers (CCBs) are important in the management of hypertension and limit restenosis. Although CCB efficacy could derive from decreased blood pressure, other mechanisms independent of CCB activity also can contribute to antiproliferative action. To understand mechanisms of CCB-mediated antiproliferation, we studied two structurally dissimilar CCBs, diltiazem and verapamil, in cultured rat vascular smooth muscle cells (VSMC). To elucidate CCB-independent effects, pure stereoisomers of verapamil (R-verapamil, inactive VR; S-verapamil, active, VS) were used. The effects of CCB exposure on cell viability (MTT reduction), cell proliferation ({sup 3}H-thymidine incorporation), VSMC morphology by light and transmission electron microscopy (TEM) and autophagy (LC3I/II, ATG5) were measured. In general, verapamil, VR or VS treatment alone (80 μM) appreciably enhanced MTT absorbance although higher concentrations (VR or VS) slightly decreased MTT absorbance. Diltiazem (140 μM) markedly decreased MTT absorbance (40%) at 120 h. VR or VS treatment inhibited {sup 3}H-thymidine incorporation (24 h) and induced cytological alterations (i.e., karyokinesis, enhanced perinuclear MTT deposition, accumulated perinuclear “vacuoles”). TEM revealed perinuclear “vacuoles” to be aggregates of highly laminated and electron-dense vesicles resembling autophagosomes and lysosomes, respectively. Increased autophagosome activity was confirmed by a concentration-dependent increase in LC3-II formation by Western blotting and by increased perinuclear LC3-GFP{sup +} puncta in verapamil-treated VSMC. Verapamil stereoisomers appeared to decrease perinuclear mitochondrial density. These observations indicate that antiproliferative effects of verapamil stereoisomers are produced by enhanced mitochondrial damage and upregulated autophagy in VSMC. These effects are independent of CCB activity indicating a distinct mechanism of action that could be targeted for more efficacious anti

  10. Redox signaling in cardiovascular pathophysiology: A focus on hydrogen peroxide and vascular smooth muscle cells

    Directory of Open Access Journals (Sweden)

    Chang Hyun Byon

    2016-10-01

    Full Text Available Oxidative stress represents excessive intracellular levels of reactive oxygen species (ROS, which plays a major role in the pathogenesis of cardiovascular disease. Besides having a critical impact on the development and progression of vascular pathologies including atherosclerosis and diabetic vasculopathy, oxidative stress also regulates physiological signaling processes. As a cell permeable ROS generated by cellular metabolism involved in intracellular signaling, hydrogen peroxide (H2O2 exerts tremendous impact on cardiovascular pathophysiology. Under pathological conditions, increased oxidase activities and/or impaired antioxidant systems results in uncontrolled production of ROS. In a pro-oxidant environment, vascular smooth muscle cells (VSMC undergo phenotypic changes which can lead to the development of vascular dysfunction such as vascular inflammation and calcification. Investigations are ongoing to elucidate the mechanisms for cardiovascular disorders induced by oxidative stress. This review mainly focuses on the role of H2O2 in regulating physiological and pathological signals in VSMC.

  11. IGF-1 Has Plaque-Stabilizing Effects in Atherosclerosis by Altering Vascular Smooth Muscle Cell Phenotype

    Science.gov (United States)

    von der Thüsen, Jan H.; Borensztajn, Keren S.; Moimas, Silvia; van Heiningen, Sandra; Teeling, Peter; van Berkel, Theo J.C.; Biessen, Erik A.L.

    2011-01-01

    Insulin-like growth factor-1 (IGF-1) signaling is important for the maintenance of plaque stability in atherosclerosis due to its effects on vascular smooth muscle cell (vSMC) phenotype. To investigate this hypothesis, we studied the effects of the highly inflammatory milieu of the atherosclerotic plaque on IGF-1 signaling and stability-related phenotypic parameters of murine vSMCs in vitro, and the effects of IGF-1 supplementation on plaque phenotype in an atherosclerotic mouse model. M1-polarized, macrophage-conditioned medium inhibited IGF-1 signaling by ablating IGF-1 and increasing IGF-binding protein 3, increased vSMC apoptosis, and decreased proliferation. Expression of α-actin and col3a1 genes was strongly attenuated by macrophage-conditioned medium, whereas expression of matrix-degrading enzymes was increased. Importantly, all of these effects could be corrected by supplementation with IGF-1. In vivo, treatment with the stable IGF-1 analog Long R3 IGF-1 in apolipoprotein E knockout mice reduced stenosis and core size, and doubled cap/core ratio in early atherosclerosis. In advanced plaques, Long R3 IGF-1 increased the vSMC content of the plaque by more than twofold and significantly reduced the rate of intraplaque hemorrhage. We believe that IGF-1 in atherosclerotic plaques may have a role in preventing plaque instability, not only by modulating smooth muscle cell turnover, but also by altering smooth muscle cell phenotype. PMID:21281823

  12. Tungstate-Targeting of BKαβ1 Channels Tunes ERK Phosphorylation and Cell Proliferation in Human Vascular Smooth Muscle

    OpenAIRE

    López López, José Ramón; Fernández Mariño, Ana Isabel; Cidad, Pilar; Zafra, Delia; Nocito, Laura; Domínguez, Jorge; Oliván Viguera, Aida; Köhler, Ralf; Pérez García, María Teresa; Valverde, Miguel Ángel; Guinovart, Joan J.; Fernández Fernández, José Manuel

    2015-01-01

    Producción Científica Despite the substantial knowledge on the antidiabetic, antiobesity and antihypertensive actions of tungstate, information on its primary target/s is scarce. Tungstate activates both the ERK1/2 pathway and the vascular voltage- and Ca2+-dependent large-conductance BKαβ1 potassium channel, which modulates vascular smooth muscle cell (VSMC) proliferation and function, respectively. Here, we have assessed the possible involvement of BKαβ1 channels in the tungstate-induced...

  13. Thrombospondin-1, -2 and -5 have differential effects on vascular smooth muscle cell physiology

    Energy Technology Data Exchange (ETDEWEB)

    Helkin, Alex; Maier, Kristopher G. [SUNY Upstate Medical University, Division of Vascular Surgery and Endovascular Services, Syracuse, NY (United States); Department of Veterans Affairs VA Healthcare Network Upstate New York at Syracuse, Syracuse, NY (United States); Gahtan, Vivian, E-mail: gahtanv@upstate.edu [SUNY Upstate Medical University, Division of Vascular Surgery and Endovascular Services, Syracuse, NY (United States); Department of Veterans Affairs VA Healthcare Network Upstate New York at Syracuse, Syracuse, NY (United States)

    2015-09-04

    Introduction: The thrombospondins (TSPs) are matricellular proteins that exert multifunctional effects by binding cytokines, cell-surface receptors and other proteins. TSPs play important roles in vascular pathobiology and are all expressed in arterial lesions. The differential effects of TSP-1, -2, and -5 represent a gap in knowledge in vascular smooth muscle cell (VSMC) physiology. Our objective is to determine if structural differences of the TSPs imparted different effects on VSMC functions critical to the formation of neointimal hyperplasia. We hypothesize that TSP-1 and -2 induce similar patterns of migration, proliferation and gene expression, while the effects of TSP-5 are different. Methods: Human aortic VSMC chemotaxis was tested for TSP-2 and TSP-5 (1–40 μg/mL), and compared to TSP-1 and serum-free media (SFM) using a modified Boyden chamber. Next, VSMCs were exposed to TSP-1, TSP-2 or TSP-5 (0.2–40 μg/mL). Proliferation was assessed by MTS assay. Finally, VSMCs were exposed to TSP-1, TSP-2, TSP-5 or SFM for 3, 6 or 24 h. Quantitative real-time PCR was performed on 96 genes using a microfluidic card. Statistical analysis was performed by ANOVA or t-test, with p < 0.05 being significant. Results: TSP-1, TSP-2 and TSP-5 at 20 μg/mL all induce chemotaxis 3.1 fold compared to serum-free media. TSP-1 and TSP-2 induced proliferation 53% and 54% respectively, whereas TSP-5 did not. In the gene analysis, overall, cardiovascular system development and function is the canonical pathway most influenced by TSP treatment, and includes multiple growth factors, cytokines and proteases implicated in cellular migration, proliferation, vasculogenesis, apoptosis and inflammation pathways. Conclusions and relevance: The results of this study indicate TSP-1, -2, and -5 play active roles in VSMC physiology and gene expression. Similarly to TSP-1, VSMC chemotaxis to TSP-2 and -5 is dose-dependent. TSP-1 and -2 induces VSMC proliferation, but TSP-5 does not, likely

  14. Establishment of artery smooth muscle cell proliferation model after subarachnoid hemorrhage in rats

    Directory of Open Access Journals (Sweden)

    Yu-jie CHEN

    2011-12-01

    Full Text Available Objective The current paper aims to simulate the effects of hemolytic products on intracranial vascular smooth muscle cell after subarachnoid hemorrhage(SAH,and probe into the molecular mechanism and strategy for the prevention and cure of vascular proliferation after SAH.Methods Thirty Sprague-Dawley rats were randomly divided into three groups,including sham-operated,24 h after SAH,and 72 h after SAH groups.The artificial hemorrhage model around the common carotid artery was established for the latter two groups.The animals were put to death after 24 h and 72 h to take the common carotid artery,and to measure the expression level of PCNA,SM-α-actin protein,and mRNA in the smooth muscle cell.Results The PCNA mRNA expression was significantly up-regulated in the 24-h group(P < 0.01.The expression in the 72-h group was lower than that of the 24-h group(P < 0.01,whereas it was still remarkably higher than that of the sham group(P < 0.01.The SM-α-actin mRNA expression in the smooth muscle cell in the 24-h and 72-h groups decreased compared with that of the Sham group(P < 0.05,whereas the 72-h group was significantly lower than that of the 24-h group(P < 0.05.The protein expression of PCNA and SM-α-actin showed a similar trend.Conclusion The current experiment simulates better effects of the hemolytic products on vascular smooth muscle cell after SAH.It also shows that artificial hemorrhage around the common carotid artery could stimulate vascular smooth muscle cell to change from contractile phenotype into synthetic phenotype,and improve it to proliferate.

  15. Tyk2 mediates effects of urokinase on human vascular smooth muscle cell growth

    International Nuclear Information System (INIS)

    Patecki, Margret; Schaewen, Markus von; Tkachuk, Sergey; Jerke, Uwe; Dietz, Rainer; Dumler, Inna; Kusch, Angelika

    2007-01-01

    The urokinase (uPA)/uPA receptor (uPAR) system plays a role in the response of the vessel wall to injury, presumably by modulating vascular smooth muscle cell (VSMC) functional behaviour. The Jak/Stat signaling pathway has been implicated to mediate the uPA/uPAR-directed cell migration and proliferation in VSMC. We have therefore investigated the underlying molecular mechanisms, which remained not completely understood. In particular, we aimed at identification of the kinase involved in the signaling cascade leading to Stat1 phosphorylation by uPA and its impact on VSMC growth. We performed expression in VSMC of kinase-deficient mutant forms of the Janus kinases Jak1 and Tyk2 and used different cell culture models imitating the response to vascular injury. We provide evidence that Tyk2, but not Jak1, mediates uPA-induced Stat1 phosphorylation and VSMC growth inhibition and suggest a novel function for Tyk2 as an important modulator of the uPA-directed VSMC functional behaviour at the place of injury

  16. Nuclear import mechanism for myocardin family members and their correlation with vascular smooth muscle cell phenotype.

    Science.gov (United States)

    Nakamura, Seiji; Hayashi, Ken'ichiro; Iwasaki, Kazuhiro; Fujioka, Tomoaki; Egusa, Hiroshi; Yatani, Hirofumi; Sobue, Kenji

    2010-11-26

    Myocardin (Mycd), which is essential for the differentiation of the smooth muscle cell lineage, is constitutively located in the nucleus, although its family members, myocardin-related transcription factors A and B (MRTF-A/B), mostly reside in the cytoplasm and translocate to the nucleus in response to Rho signaling. The mechanism for their nuclear import is unclear. Here we investigated the mechanism for the nuclear import of Mycd family members and demonstrated any correlation between such mechanism and the phenotype of vascular smooth muscle cells (VSMCs). In cultured VSMCs, the knockdown of importin β1 inhibited the nuclear import of Mycd and MRTF-A/B. Their NH(2)-terminal basic domain was identified as a binding site for importin α/β1 by in vitro analyses. However, Mycd had a higher affinity for importin α/β1 than did MRTF-A/B, even in the absence of G-actin, and Mycd affinity for importin α1/β1 was stronger than for any other importin α/β1 heterodimers. The binding of Mycd to importin α/β1 was insensitive to G-actin, whereas that of MRTF-A/B was differently inhibited by G-actin. In dedifferentiated VSMCs, the levels of importins α1 and β1 were reduced concomitant with down-regulation of Mycd, serum response factor, and smooth muscle cell markers. By contrast, in differentiated VSMCs, their expressions were up-regulated. Thus, the nuclear import of Mycd family members in VSMCs depends on importin α/β1, and their relative affinities for importin α/β1 heterodimers determine Mycd nuclear import. The expression of Mycd nuclear import machineries is related to the expression levels of VSMC phenotype-dependent smooth muscle cell markers.

  17. Recipient origin of neointimal vascular smooth muscle cells in cardiac allografts with transplant arteriosclerosis

    NARCIS (Netherlands)

    Hillebrands, JL; van den Hurk, BMH; Klatter, FA; Popa, ER; Nieuwenhuis, P; Rozing, J

    2000-01-01

    Background: Coronary artery disease is today's most important post-heart transplantation problem after the first perioperative year. Histologically, coronary artery disease is characterized by transplant arteriosclerosis. The current view on this vasculopathy is that vascular smooth muscle (VSM)

  18. Gene expression profiling of resting and activated vascular smooth muscle cells by serial analysis of gene expression and clustering analysis

    NARCIS (Netherlands)

    Beauchamp, Nicholas J.; van Achterberg, Tanja A. E.; Engelse, Marten A.; Pannekoek, Hans; de Vries, Carlie J. M.

    2003-01-01

    Migration and proliferation of vascular smooth muscle cells (SMCs) are key events in atherosclerosis. However, little is known about alterations in gene expression upon transition of the quiescent, contractile SMC to the proliferative SMC. We performed serial analysis of gene expression (SAGE) of

  19. Statins activate GATA-6 and induce differentiated vascular smooth muscle cells

    International Nuclear Information System (INIS)

    Wada, Hiromichi; Abe, Mitsuru; Ono, Koh; Morimoto, Tatsuya; Kawamura, Teruhisa; Takaya, Tomohide; Satoh, Noriko; Fujita, Masatoshi; Kita, Toru; Shimatsu, Akira; Hasegawa, Koji

    2008-01-01

    The beneficial effects of 3-hydroxy-3-methylglutaryl coenzyme A reductase inhibitors (statins) beyond cholesterol lowering involve their direct actions on vascular smooth muscle cells (VSMCs). However, the effects of statins on phenotypic modulation of VSMCs are unknown. We herein show that simvastatin (Sm) and atorvastatin (At) inhibited DNA synthesis in human aortic VSMCs dose-dependently, while cell toxicity was not observed below the concentration of 1 μM of Sm or 100 nM of At. Stimulating proliferative VSMCs with Sm or At induced the expression of SM-α-actin and SM-MHC, highly specific markers of differentiated phenotype. Sm up-regulated the binding activity of GATA-6 to SM-MHC GATA site and activated the transfected SM-MHC promoter in proliferative VSMCs, while mutating the GATA-6 binding site abolished this activation. Geranylgeranylpyrophosphate (10 μM), an inhibitor of Rho family proteins, abolished the statin-mediated induction of the differentiated phenotype in VSMCs. These findings suggest that statins activate GATA-6 and induce differentiated VSMCs

  20. Statins activate GATA-6 and induce differentiated vascular smooth muscle cells

    Energy Technology Data Exchange (ETDEWEB)

    Wada, Hiromichi [Division of Translational Research, National Hospital Organization Kyoto Medical Center, 1-1 Mukaihata-cho, Fukakusa, Fushimi-ku, Kyoto 612-8555 (Japan); Abe, Mitsuru; Ono, Koh [Department of Cardiovascular Medicine, Graduate School of Medicine, Kyoto University, Kyoto (Japan); Morimoto, Tatsuya; Kawamura, Teruhisa; Takaya, Tomohide [Division of Translational Research, National Hospital Organization Kyoto Medical Center, 1-1 Mukaihata-cho, Fukakusa, Fushimi-ku, Kyoto 612-8555 (Japan); Satoh, Noriko [Division of Metabolic Research, National Hospital Organization Kyoto Medical Center, Kyoto (Japan); Fujita, Masatoshi [Department of Human Health Sciences, Graduate School of Medicine, Kyoto University, Kyoto (Japan); Kita, Toru [Department of Cardiovascular Medicine, Graduate School of Medicine, Kyoto University, Kyoto (Japan); Shimatsu, Akira [Clinical Research Institute, National Hospital Organization Kyoto Medical Center, Kyoto (Japan); Hasegawa, Koji [Division of Translational Research, National Hospital Organization Kyoto Medical Center, 1-1 Mukaihata-cho, Fukakusa, Fushimi-ku, Kyoto 612-8555 (Japan)

    2008-10-03

    The beneficial effects of 3-hydroxy-3-methylglutaryl coenzyme A reductase inhibitors (statins) beyond cholesterol lowering involve their direct actions on vascular smooth muscle cells (VSMCs). However, the effects of statins on phenotypic modulation of VSMCs are unknown. We herein show that simvastatin (Sm) and atorvastatin (At) inhibited DNA synthesis in human aortic VSMCs dose-dependently, while cell toxicity was not observed below the concentration of 1 {mu}M of Sm or 100 nM of At. Stimulating proliferative VSMCs with Sm or At induced the expression of SM-{alpha}-actin and SM-MHC, highly specific markers of differentiated phenotype. Sm up-regulated the binding activity of GATA-6 to SM-MHC GATA site and activated the transfected SM-MHC promoter in proliferative VSMCs, while mutating the GATA-6 binding site abolished this activation. Geranylgeranylpyrophosphate (10 {mu}M), an inhibitor of Rho family proteins, abolished the statin-mediated induction of the differentiated phenotype in VSMCs. These findings suggest that statins activate GATA-6 and induce differentiated VSMCs.

  1. Down-regulation of endothelin binding sites in rat vascular smooth muscle cells

    International Nuclear Information System (INIS)

    Roubert, P.; Gillard, V.; Plas, P.; Chabrier, P.E.; Braquet, P.

    1990-01-01

    In cultured rat aortic smooth muscle cells, [ 125 I]endothelin (ET-1) bound to an apparent single class of high affinity recognition sites with a dissociation constant of 1.84 +/- 0.29 nmol/L and a maximum binding of 62 +/- 10.5 fmol/10(6) cells. The binding was not affected by calcium antagonists or vasoactive substances, including angiotensin II, arginine vasopressin, atrial natriuretic factor and bradykinin. Exposure of the cells to ET-1 (0.01 nmol/L to 10 nmol/L) resulted in an apparent dose-dependent reduction of the number of endothelin binding sites with no significant modification of its binding affinity. The time course of the down-regulation of ET-1 binding sites showed that this effect was present after 30 min incubation and persisted after 18 h. This indicates that down-regulation of ET-1 binding sites can modulate the activity of ET-1 and suggests a rapid internalization of ET-1 in vascular cells

  2. Hydroxyapatite and Calcified Elastin Induce Osteoblast-like Differentiation in Rat Aortic Smooth Muscle Cells

    Science.gov (United States)

    Lei, Yang; Sinha, Aditi; Nosoudi, Nasim; Grover, Ankit; Vyavahare, Naren

    2014-01-01

    Vascular calcification can be categorized into two different types. Intimal calcification related to atherosclerosis and elastin-specific medial arterial calcification (MAC). Osteoblast-like differentiation of vascular smooth muscle cells (VSMCs) has been shown in both types; however, how this relates to initiation of vascular calcification is unclear. We hypothesize that the initial deposition of hydroxyapatite-like mineral in MAC occurs on degraded elastin first and that causes osteogenic transformation of VSMCs. To test this, rat aortic smooth muscle cells (RASMCs) were cultured on hydroxyapatite crystals and calcified aortic elastin. Using RT-PCR and specific protein assays, we demonstrate that RASMCs lose their smooth muscle lineage markers like alpha smooth muscle actin (SMA) and myosin heavy chain (MHC) and undergo chondrogenic/osteogenic transformation. This is indicated by an increase in the expression of typical chondrogenic proteins such as aggrecan, collagen type II alpha 1(Col2a1) and bone proteins such as runt-related transcription factor 2 (RUNX2), alkaline phosphatase (ALP) and osteocalcin (OCN). Furthermore, when calcified conditions are removed, cells return to their original phenotype. Our data supports the hypothesis that elastin degradation and calcification precedes VSMCs' osteoblast-like differentiation. PMID:24447384

  3. Selective Inhibitory Effect of Epigallocatechin-3-gallate on Migration of Vascular Smooth Muscle Cells

    Directory of Open Access Journals (Sweden)

    Jong-Chul Park

    2010-11-01

    Full Text Available In order to prevent restenosis after angioplasty or stenting, one of the most popular targets is suppression of the abnormal growth and excess migration of vascular smooth muscle cells (VSMCs with drugs. However, the drugs also adversely affect vascular endothelial cells (VECs, leading to the induction of late thrombosis. We have investigated the effect of epigallocatechin-3-gallate (EGCG on the proliferation and migration of VECs and VSMCs. Both cells showed dose-dependent decrease of viability in response to EGCG while they have different IC50 values of EGCG (VECs, 150 mM and VSMCs, 1050 mM. Incubating both cells with EGCG resulted in significant reduction in cell proliferation irrespective of cell type. The proliferation of VECs were greater affected than that of VSMCs at the same concentrations of EGCG. EGCG exerted differential migration-inhibitory activity in VECs vs. VSMCs. The migration of VECs was not attenuated by 200 mM EGCG, but that of VSMCs was significantly inhibited at the same concentration of EGCG. It is suggested that that EGCG can be effectively used as an efficient drug for vascular diseases or stents due to its selective activity, completely suppressing the proliferation and migration of VSMCs, but not adversely affecting VECs migration in blood vessels.

  4. Msx1 and Msx2 are expressed in sub-populations of vascular smooth muscle cells.

    Science.gov (United States)

    Goupille, Olivier; Saint Cloment, Cécile; Lopes, Miguel; Montarras, Didier; Robert, Benoît

    2008-08-01

    Using an nlacZ reporter gene inserted at the Msx1 and Msx2 loci, we could analyze the expression of these homeogenes in the adult mouse. We observed that Msx genes are prominently expressed in a subset of blood vessels. The Msx2nlacZ allele is mainly expressed in a restricted population of mural cells in peripheral arteries and veins. Msx1nlacZ is expressed to a lesser extent by vascular smooth muscle cells of peripheral arteries, but is highly expressed in arterioles and capillaries, making Msx1 a novel marker for a subpopulation of pericytes. Expression is set up early in developing vessels and maintained throughout life. In addition, expression of both genes is observed in a few endothelial cells of the aorta at fetal stages, and only Msx2 continues to be expressed in this layer at the adult stage. These results suggest major functions for Msx genes in vascular mural cell formation and remodeling. Copyright (c) 2008 Wiley-Liss, Inc.

  5. Triiodothyronine Potentiates Vasorelaxation via PKG/VASP Signaling in Vascular Smooth Muscle Cells

    Directory of Open Access Journals (Sweden)

    Sherin Samuel

    2017-04-01

    Full Text Available Background/Aims: Vascular relaxation caused by Triiodothyronine (T3 involves direct activation of endothelial cells (EC and vascular smooth muscle cells (VSMC. Activation of protein kinase G (PKG has risen as a novel contributor to the vasorelaxation mechanism triggered by numerous stimuli. We hypothesize that T3-induced vasorelaxation involves PKG/vasodilator-stimulated phosphoprotein (VASP signaling pathway in VSMC. Methods: Human aortic endothelial cells (HAEC and VSMC were treated with T3 for short (2 to 60 minutes and long term (24 hours. Nitric oxide (NO production was measured using DAF-FM. Expression of protein targets was determined using western blot. For functional studies, rat aortas were isolated and treated with T3 for 20 minutes and mounted in a wire myograph. Relaxation was measured by a concentration-dependent response to acetylcholine (ACh and sodium nitroprusside (SNP. Results: Aortas stimulated with T3 exhibited augmented sensitivity to ACh and SNP-induced relaxation, endothelium-dependent and endothelium-independent responses, respectively. T3 directly increased vasorelaxation, which was abolished in the presence of a PKG inhibitor. T3 markedly induced phosphorylation of Akt, eNOS and consequently increased NO production in EC. Likewise, T3 induced phosphorylation of VASP at serine 239 via the PKG pathway in VSMC. Conclusion: Our findings have uncovered a PKG/VASP signaling pathway in VSMC as a key molecular mechanism underlying T3-induced vascular relaxation.

  6. Triiodothyronine Potentiates Vasorelaxation via PKG/VASP Signaling in Vascular Smooth Muscle Cells.

    Science.gov (United States)

    Samuel, Sherin; Zhang, Kuo; Tang, Yi-Da; Gerdes, A Martin; Carrillo-Sepulveda, Maria Alicia

    2017-01-01

    Vascular relaxation caused by Triiodothyronine (T3) involves direct activation of endothelial cells (EC) and vascular smooth muscle cells (VSMC). Activation of protein kinase G (PKG) has risen as a novel contributor to the vasorelaxation mechanism triggered by numerous stimuli. We hypothesize that T3-induced vasorelaxation involves PKG/vasodilator-stimulated phosphoprotein (VASP) signaling pathway in VSMC. Human aortic endothelial cells (HAEC) and VSMC were treated with T3 for short (2 to 60 minutes) and long term (24 hours). Nitric oxide (NO) production was measured using DAF-FM. Expression of protein targets was determined using western blot. For functional studies, rat aortas were isolated and treated with T3 for 20 minutes and mounted in a wire myograph. Relaxation was measured by a concentration-dependent response to acetylcholine (ACh) and sodium nitroprusside (SNP). Aortas stimulated with T3 exhibited augmented sensitivity to ACh and SNP-induced relaxation, endothelium-dependent and endothelium-independent responses, respectively. T3 directly increased vasorelaxation, which was abolished in the presence of a PKG inhibitor. T3 markedly induced phosphorylation of Akt, eNOS and consequently increased NO production in EC. Likewise, T3 induced phosphorylation of VASP at serine 239 via the PKG pathway in VSMC. Our findings have uncovered a PKG/VASP signaling pathway in VSMC as a key molecular mechanism underlying T3-induced vascular relaxation. © 2017 The Author(s)Published by S. Karger AG, Basel.

  7. Novel Mechanism of Plasma Prekallikrein (PK) Activation by Vascular Smooth Muscle Cells: Evidence of the presence of PK Activator

    OpenAIRE

    Keum, Joo-Seob; Jaffa, Miran A; Luttrell, Louis M; Jaffa, Ayad A.

    2014-01-01

    The contribution of plasma prekallikrein (PK) to vascular remodeling is becoming increasingly recognized. Plasma PK is activated when the zymogen PK is digested to an active enzyme by activated factor XII (FXII). Here, we present our findings that vascular smooth muscle cells (VSMC) activate plasma PK in the absence of FXII. Extracted plasma membrane and cytosolic fractions of VSMCs activate PK, but the rate of PK activation was greater by the membrane fraction. FXII neutralizing antibody did...

  8. LPS, but not Angiotensin ll, lnduces Direct Pro-lnflammatory Effects in Cultured Mouse Arteries and Human Endothelial and Vascular Smooth Muscle Cells

    DEFF Research Database (Denmark)

    Outzen, Emilie M; Zaki, Marina; Mehryar, Rahila

    2017-01-01

    resistance-sized arteries (MRA) supported by experiments in cultured human primary endothelial and vascular smooth muscle cells. Results showed that 24-hr organ culture of mouse MRA with 10 nM Ang II had, unlike 100 ng/mL LPS, no effects on IL-6 or MCP-1 secretion, VCAM1 mRNA expression or endothelial......]-Ang II had no concentration- or time-dependent effects on IL-6 and MCP-1 secretion in human umbilical vein endothelial cells (HUVEC) and human aortic smooth muscle cells (HASMC). AGTR1 or AGTR2 mRNA expression were undetectable in HUVEC, whereas HASMC expressed only AGTR1 mRNA. In summary, contrary...... rights reserved....

  9. Key role of microRNA-15a in the KLF4 suppressions of proliferation and angiogenesis in endothelial and vascular smooth muscle cells

    International Nuclear Information System (INIS)

    Zheng, Xuemei; Li, Aiqin; Zhao, Liang; Zhou, Tengfei; Shen, Qiang; Cui, Qinghua; Qin, Xiaomei

    2013-01-01

    Highlights: •This is the first demonstration that miR-15a is a novel target gene of KLF4. •A novel finding that KLF4 increases the expression of miR-15a in ECs and VSMCs. •The novel mechanism is that KLF4 inhibits the proliferation of ECs via miR-15a. •The novel mechanism is that KLF4 inhibits the proliferation of VSMCs via miR-15. •miR-15a mediates the anti-angiogenic activity of KLF4. -- Abstract: While recent insights indicate that the transcription factor Krüppel-like factor 4 (KLF4) is indispensable for vascular homeostasis, its exact role in proliferation and angiogenesis and how it functions remain unresolved. Thus, the aim of the present study was to evaluate the role of KLF4 in the proliferations of endothelial and vascular smooth muscle cells, as well as the angiogenesis. The overexpression of KLF4 in endothelial cells significantly impaired tube formation. KLF4 inhibited the formation of a vascular network in implanted Matrigel plugs in nude mice. Importantly, we found that KLF4 significantly upregulated the miR-15a expression in endothelial cells and vascular smooth muscle cells, and conversely, KLF4 depletion reduced the amount of miR-15a. Furthermore, KLF4 blocked cell cycle progression and decreased cyclin D1 expression in endothelial cells and vascular smooth muscle cells through the induction of miR-15a. Intriguingly, the delivery of a miR-15a antagomir to nude mice resulted in marked attenuation of the anti-angiogenic effect of KLF4. Collectively, our present study provide the first evidence that miR-15a as a direct transcriptional target of KLF4 that mediates the anti-proliferative and anti-angiogenic actions of KLF4, which indicates that KLF4 upregulation of miR-15a may represent a therapeutic option to suppress proliferative vascular disorders

  10. Key role of microRNA-15a in the KLF4 suppressions of proliferation and angiogenesis in endothelial and vascular smooth muscle cells

    Energy Technology Data Exchange (ETDEWEB)

    Zheng, Xuemei; Li, Aiqin; Zhao, Liang; Zhou, Tengfei; Shen, Qiang [Institute of Cardiovascular Science, Peking University Health Science Center, Beijing 100191 (China); Key Laboratory of Molecular Cardiovascular Science of Ministry of Education, Peking University Health Science Center, Beijing 100191 (China); Cui, Qinghua [Department of Biomedical Informatics, Peking University Health Science Center, Beijing 100191 (China); Key Laboratory of Molecular Cardiovascular Science of Ministry of Education, Peking University Health Science Center, Beijing 100191 (China); Qin, Xiaomei, E-mail: xmqin@bjmu.edu.cn [Institute of Cardiovascular Science, Peking University Health Science Center, Beijing 100191 (China); Key Laboratory of Molecular Cardiovascular Science of Ministry of Education, Peking University Health Science Center, Beijing 100191 (China)

    2013-08-09

    Highlights: •This is the first demonstration that miR-15a is a novel target gene of KLF4. •A novel finding that KLF4 increases the expression of miR-15a in ECs and VSMCs. •The novel mechanism is that KLF4 inhibits the proliferation of ECs via miR-15a. •The novel mechanism is that KLF4 inhibits the proliferation of VSMCs via miR-15. •miR-15a mediates the anti-angiogenic activity of KLF4. -- Abstract: While recent insights indicate that the transcription factor Krüppel-like factor 4 (KLF4) is indispensable for vascular homeostasis, its exact role in proliferation and angiogenesis and how it functions remain unresolved. Thus, the aim of the present study was to evaluate the role of KLF4 in the proliferations of endothelial and vascular smooth muscle cells, as well as the angiogenesis. The overexpression of KLF4 in endothelial cells significantly impaired tube formation. KLF4 inhibited the formation of a vascular network in implanted Matrigel plugs in nude mice. Importantly, we found that KLF4 significantly upregulated the miR-15a expression in endothelial cells and vascular smooth muscle cells, and conversely, KLF4 depletion reduced the amount of miR-15a. Furthermore, KLF4 blocked cell cycle progression and decreased cyclin D1 expression in endothelial cells and vascular smooth muscle cells through the induction of miR-15a. Intriguingly, the delivery of a miR-15a antagomir to nude mice resulted in marked attenuation of the anti-angiogenic effect of KLF4. Collectively, our present study provide the first evidence that miR-15a as a direct transcriptional target of KLF4 that mediates the anti-proliferative and anti-angiogenic actions of KLF4, which indicates that KLF4 upregulation of miR-15a may represent a therapeutic option to suppress proliferative vascular disorders.

  11. A Robust Method to Generate Mechanically Anisotropic Vascular Smooth Muscle Cell Sheets for Vascular Tissue Engineering.

    Science.gov (United States)

    Backman, Daniel E; LeSavage, Bauer L; Shah, Shivem B; Wong, Joyce Y

    2017-06-01

    In arterial tissue engineering, mimicking native structure and mechanical properties is essential because compliance mismatch can lead to graft failure and further disease. With bottom-up tissue engineering approaches, designing tissue components with proper microscale mechanical properties is crucial to achieve the necessary macroscale properties in the final implant. This study develops a thermoresponsive cell culture platform for growing aligned vascular smooth muscle cell (VSMC) sheets by photografting N-isopropylacrylamide (NIPAAm) onto micropatterned poly(dimethysiloxane) (PDMS). The grafting process is experimentally and computationally optimized to produce PNIPAAm-PDMS substrates optimal for VSMC attachment. To allow long-term VSMC sheet culture and increase the rate of VSMC sheet formation, PNIPAAm-PDMS surfaces were further modified with 3-aminopropyltriethoxysilane yielding a robust, thermoresponsive cell culture platform for culturing VSMC sheets. VSMC cell sheets cultured on patterned thermoresponsive substrates exhibit cellular and collagen alignment in the direction of the micropattern. Mechanical characterization of patterned, single-layer VSMC sheets reveals increased stiffness in the aligned direction compared to the perpendicular direction whereas nonpatterned cell sheets exhibit no directional dependence. Structural and mechanical anisotropy of aligned, single-layer VSMC sheets makes this platform an attractive microstructural building block for engineering a vascular graft to match the in vivo mechanical properties of native arterial tissue. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  12. ASIC PROTEINS REGULATE SMOOTH MUSCLE CELL MIGRATION

    OpenAIRE

    Grifoni, Samira C.; Jernigan, Nikki L.; Hamilton, Gina; Drummond, Heather A.

    2007-01-01

    The purpose of the present study was to investigate Acid Sensing Ion Channel (ASIC) protein expression and importance in cellular migration. We recently demonstrated Epithelial Na+ Channel (ENaC) proteins are required for vascular smooth muscle cell (VSMC) migration, however the role of the closely related ASIC proteins has not been addressed. We used RT-PCR and immunolabeling to determine expression of ASIC1, ASIC2, ASIC3 and ASIC4 in A10 cells. We used small interference RNA to silence indi...

  13. Advanced Glycation End-Products Induce Apoptosis of Vascular Smooth Muscle Cells: A Mechanism for Vascular Calcification

    Directory of Open Access Journals (Sweden)

    Sayo Koike

    2016-09-01

    Full Text Available Vascular calcification, especially medial artery calcification, is associated with cardiovascular death in patients with diabetes mellitus and chronic kidney disease (CKD. To determine the underlying mechanism of vascular calcification, we have demonstrated in our previous report that advanced glycation end-products (AGEs stimulated calcium deposition in vascular smooth muscle cells (VSMCs through excessive oxidative stress and phenotypic transition into osteoblastic cells. Since AGEs can induce apoptosis, in this study we investigated its role on VSMC apoptosis, focusing mainly on the underlying mechanisms. A rat VSMC line (A7r5 was cultured, and treated with glycolaldehyde-derived AGE-bovine serum albumin (AGE3-BSA. Apoptotic cells were identified by Terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL staining. To quantify apoptosis, an enzyme-linked immunosorbent assay (ELISA for histone-complexed DNA fragments was employed. Real-time PCR was performed to determine the mRNA levels. Treatment of A7r5 cells with AGE3-BSA from 100 µg/mL concentration markedly increased apoptosis, which was suppressed by Nox inhibitors. AGE3-BSA significantly increased the mRNA expression of NAD(PH oxidase components including Nox4 and p22phox, and these findings were confirmed by protein levels using immunofluorescence. Dihydroethidisum assay showed that compared with cBSA, AGE3-BSA increased reactive oxygen species level in A7r5 cells. Furthermore, AGE3-induced apoptosis was significantly inhibited by siRNA-mediated knockdown of Nox4 or p22phox. Double knockdown of Nox4 and p22phox showed a similar inhibitory effect on apoptosis as single gene silencing. Thus, our results demonstrated that NAD(PH oxidase-derived oxidative stress are involved in AGEs-induced apoptosis of VSMCs. These findings might be important to understand the pathogenesis of vascular calcification in diabetes and CKD.

  14. Preliminary investigations on the effects of a Strongylus vulgaris larval extract, mononuclear factors and platelet factors on equine smooth muscle cells in vitro.

    Science.gov (United States)

    Morgan, S J; Storts, R W; Stromberg, P C; Sowa, B A; Lay, J C

    1989-01-01

    Factors involved in the proliferation of equine vascular smooth muscle cells were studied in vitro. The most prominent proliferative responses in cultured vascular smooth muscle cells were induced by Strongylus vulgaris larval antigen extract (LAE) and platelet-derived factors. Less significant proliferative responses were obtained with conditioned media from S. vulgaris LAE stimulated and from unstimulated equine mononuclear leukocytes. Additionally, vascular smooth muscle cells exposed to S. vulgaris LAE developed numerous perinuclear vacuoles and were more spindle-shaped than control or smooth muscle cells exposed to other factors. Equine mononuclear leukocytes exposed to LAE developed prominent morphological changes, including enlargement, clumping and increased numbers of mitotic figures.

  15. Bone morphogenetic proteins regulate osteoprotegerin and its ligands in human vascular smooth muscle cells

    DEFF Research Database (Denmark)

    Knudsen, Kirsten Quyen Nguyen; Olesen, Ping; Ledet, Thomas

    2007-01-01

    The bone-related protein osteoprotegerin (OPG) may be involved in the development of vascular calcifications, especially in diabetes, where it has been found in increased amounts in the arterial wall. Experimental studies suggest that members of the TGF-superfamily are involved in the transformat......The bone-related protein osteoprotegerin (OPG) may be involved in the development of vascular calcifications, especially in diabetes, where it has been found in increased amounts in the arterial wall. Experimental studies suggest that members of the TGF-superfamily are involved...... in the transformation of human vascular smooth muscle cells (HVSMC) to osteoblast-like cells. In this study, we evaluated the effect of BMP-2, BMP-7 and transforming growth factor beta (TGF-beta1) on the secretion and mRNA expression of OPG and its ligands receptor activator of nuclear factor-kappabeta ligand (RANKL......) and TNF-related apoptosis-inducing ligand (TRAIL) in HVSMC. All three growth factors decreased OPG protein production significantly; these results were paralleled by reduced OPG mRNA expression. TRAIL mRNA levels were also decreased. RANKL mRNA expression declined when treated with TGF-beta1 but were...

  16. Neuroblast survival depends on mature vascular network formation after mouse stroke: role of endothelial and smooth muscle progenitor cell co-administration.

    Science.gov (United States)

    Nih, Lina R; Deroide, Nicolas; Leré-Déan, Carole; Lerouet, Dominique; Soustrat, Mathieu; Levy, Bernard I; Silvestre, Jean-Sébastien; Merkulova-Rainon, Tatiana; Pocard, Marc; Margaill, Isabelle; Kubis, Nathalie

    2012-04-01

    Pro-angiogenic cell-based therapies constitute an interesting and attractive approach to enhancing post-stroke neurogenesis and decreasing neurological deficit. However, most new stroke-induced neurons die during the first few weeks after ischemia, thus impairing total recovery. Although the neovascularization process involves different cell types and various growth factors, most cell therapy protocols are based on the biological effects of single-cell-type populations or on the administration of heterogeneous populations of progenitors, namely human cord blood-derived CD34(+) cells, with scarce vascular progenitor cells. Tight cooperation between endothelial cells and smooth muscle cells/pericytes is critical for the development of functional neovessels. We hypothesized that neuroblast survival in stroke brain depends on mature vascular network formation. In this study, we injected a combination of endothelial progenitor cells (EPCs) and smooth muscle progenitor cells (SMPCs), isolated from human umbilical cord blood, into a murine model of permanent focal ischemia induced by middle cerebral artery occlusion. The co-administration of SMPCs and EPCs induced enhanced angiogenesis and vascular remodeling in the peri-infarct and infarct areas, where vessels exhibited a more mature phenotype. This activation of vessel growth resulted in the maintenance of neurogenesis and neuroblast migration to the peri-ischemic cortex. Our data suggest that a mature vascular network is essential for neuroblast survival after cerebral ischemia, and that co-administration of EPCs and SMPCs may constitute a novel therapeutic strategy for improving the treatment of stroke. © 2012 The Authors. European Journal of Neuroscience © 2012 Federation of European Neuroscience Societies and Blackwell Publishing Ltd.

  17. Tungstate-targeting of BKαβ1 channels tunes ERK phosphorylation and cell proliferation in human vascular smooth muscle.

    Directory of Open Access Journals (Sweden)

    Ana Isabel Fernández-Mariño

    Full Text Available Despite the substantial knowledge on the antidiabetic, antiobesity and antihypertensive actions of tungstate, information on its primary target/s is scarce. Tungstate activates both the ERK1/2 pathway and the vascular voltage- and Ca2+-dependent large-conductance BKαβ1 potassium channel, which modulates vascular smooth muscle cell (VSMC proliferation and function, respectively. Here, we have assessed the possible involvement of BKαβ1 channels in the tungstate-induced ERK phosphorylation and its relevance for VSMC proliferation. Western blot analysis in HEK cell lines showed that expression of vascular BKαβ1 channels potentiates the tungstate-induced ERK1/2 phosphorylation in a Gi/o protein-dependent manner. Tungstate activated BKαβ1 channels upstream of G proteins as channel activation was not altered by the inhibition of G proteins with GDPβS or pertussis toxin. Moreover, analysis of Gi/o protein activation measuring the FRET among heterologously expressed Gi protein subunits suggested that tungstate-targeting of BKαβ1 channels promotes G protein activation. Single channel recordings on VSMCs from wild-type and β1-knockout mice indicated that the presence of the regulatory β1 subunit was essential for the tungstate-mediated activation of BK channels in VSMCs. Moreover, the specific BK channel blocker iberiotoxin lowered tungstate-induced ERK phosphorylation by 55% and partially reverted (by 51% the tungstate-produced reduction of platelet-derived growth factor (PDGF-induced proliferation in human VSMCs. Our observations indicate that tungstate-targeting of BKαβ1 channels promotes activation of PTX-sensitive Gi proteins to enhance the tungstate-induced phosphorylation of ERK, and inhibits PDGF-stimulated cell proliferation in human vascular smooth muscle.

  18. Overexpression of Mitofusin 2 inhibited oxidized low-density lipoprotein induced vascular smooth muscle cell proliferation and reduced atherosclerotic lesion formation in rabbit

    International Nuclear Information System (INIS)

    Guo Yanhong; Chen Kuanghueih; Gao Wei; Li Qian; Chen Li; Wang Guisong; Tang Jian

    2007-01-01

    Our previous studies have implies that Mitofusin 2 (Mfn2), which was progressively reduced in arteries from ApoE -/- mice during the development of atherosclerosis, may take part in pathogenesis of atherosclerosis. In this study, we found that overexpression of Mfn2 inhibited oxidized low-density lipoprotein or serum induced vascular smooth muscle cell proliferation by down-regulation of Akt and ERK phosphorylation. Then we investigated the in vivo role of Mfn2 on the development of atherosclerosis in rabbits using adenovirus expressing Mitofusin 2 gene (AdMfn2). By morphometric analysis we found overexpression of Mfn2 inhibited atherosclerotic lesion formation and intima/media ratio by 66.7% and 74.6%, respectively, compared with control group. These results suggest that local Mfn2 treatment suppresses the development of atherosclerosis in vivo in part by attenuating the smooth muscle cell proliferation induced by lipid deposition and vascular injury

  19. 3,3'Diindolylmethane suppresses vascular smooth muscle cell phenotypic modulation and inhibits neointima formation after carotid injury.

    Directory of Open Access Journals (Sweden)

    Hongjing Guan

    Full Text Available 3,3'Diindolylmethane (DIM, a natural phytochemical, has shown inhibitory effects on the growth and migration of a variety of cancer cells; however, whether DIM has similar effects on vascular smooth muscle cells (VSMCs remains unknown. The purpose of this study was to assess the effects of DIM on the proliferation and migration of cultured VSMCs and neointima formation in a carotid injury model, as well as the related cell signaling mechanisms.DIM dose-dependently inhibited the platelet-derived growth factor (PDGF-BB-induced proliferation of VSMCs without cell cytotoxicity. This inhibition was caused by a G0/G1 phase cell cycle arrest demonstrated by fluorescence-activated cell-sorting analysis. We also showed that DIM-induced growth inhibition was associated with the inhibition of the expression of cyclin D1 and cyclin-dependent kinase (CDK 4/6 as well as an increase in p27(Kip1 levels in PDGF-stimulated VSMCs. Moreover, DIM was also found to modulate migration of VSMCs and smooth muscle-specific contractile marker expression. Mechanistically, DIM negatively modulated PDGF-BB-induced phosphorylation of PDGF-recptorβ (PDGF-Rβ and the activities of downstream signaling molecules including Akt/glycogen synthase kinase(GSK3β, extracellular signal-regulated kinase1/2 (ERK1/2, and signal transducers and activators of transcription 3 (STAT3. Our in vivo studies using a mouse carotid arterial injury model revealed that treatment with 150 mg/kg DIM resulted in significant reduction of the neointima/media ratio and proliferating cell nuclear antigen (PCNA-positive cells, without affecting apoptosis of vascular cells and reendothelialization. Infiltration of inflammatory cells was also inhibited by DIM administration.These results demonstrate that DIM can suppress the phenotypic modulation of VSMCs and neointima hyperplasia after vascular injury. These beneficial effects on VSMCs were at least partly mediated by the inhibition of PDGF-Rβ and the

  20. Effects of (-)-desmethoxyverapamil on heart and vascular smooth muscle

    International Nuclear Information System (INIS)

    Nawrath, H.; Raschack, M.

    1987-01-01

    (-)-Desmethoxyverapamil [also known as (-)-devapamil or (-)-D888] has been developed as a verapamil type radioligand for the study of calcium channels. In the present investigation, the effects of (-)-desmethoxyverapamil on action potential (AP) and force of contraction in heart muscle preparations and on tension and 45 Ca influx in vascular smooth muscle are described. In part, the effects were compared with the (+)-isomer of desmethoxyverapamil and the isomers of both verapamil and methoxyverapamil. In atrial and/or ventricular heart muscle preparations from guinea pigs, cats and man, (-)-desmethoxyverapamil decreased the force of contraction and shortened the AP duration. Slow response APs were depressed, whereas dV/dtmax of phase 0 of the AP remained unchanged. The rank order of potency of the (-)-isomers was as follows: desmethoxyverapamil greater than methoxyverapamil greater than verapamil. Potassium-induced contractures and 45 Ca influx were depressed by the (-)-isomers of desmethoxyverapamil, methoxyverapamil and verapamil in the same potency rank order as observed in heart muscle. The (+)-isomers exerted qualitatively similar effects at about 10 to 200 times higher concentrations. Correspondingly, the increase in potency of the racemic mixtures of the drugs was accompanied by increases in stereoselectivity. It is concluded that (-)-desmethoxyverapamil is the most potent stereoselective calcium antagonist of the verapamil type with respect to its effects on heart and vascular smooth muscle

  1. Effects of (-)-desmethoxyverapamil on heart and vascular smooth muscle

    Energy Technology Data Exchange (ETDEWEB)

    Nawrath, H.; Raschack, M.

    1987-09-01

    (-)-Desmethoxyverapamil (also known as (-)-devapamil or (-)-D888) has been developed as a verapamil type radioligand for the study of calcium channels. In the present investigation, the effects of (-)-desmethoxyverapamil on action potential (AP) and force of contraction in heart muscle preparations and on tension and /sup 45/Ca influx in vascular smooth muscle are described. In part, the effects were compared with the (+)-isomer of desmethoxyverapamil and the isomers of both verapamil and methoxyverapamil. In atrial and/or ventricular heart muscle preparations from guinea pigs, cats and man, (-)-desmethoxyverapamil decreased the force of contraction and shortened the AP duration. Slow response APs were depressed, whereas dV/dtmax of phase 0 of the AP remained unchanged. The rank order of potency of the (-)-isomers was as follows: desmethoxyverapamil greater than methoxyverapamil greater than verapamil. Potassium-induced contractures and /sup 45/Ca influx were depressed by the (-)-isomers of desmethoxyverapamil, methoxyverapamil and verapamil in the same potency rank order as observed in heart muscle. The (+)-isomers exerted qualitatively similar effects at about 10 to 200 times higher concentrations. Correspondingly, the increase in potency of the racemic mixtures of the drugs was accompanied by increases in stereoselectivity. It is concluded that (-)-desmethoxyverapamil is the most potent stereoselective calcium antagonist of the verapamil type with respect to its effects on heart and vascular smooth muscle.

  2. Fibulin-2 is present in murine vascular lesions and is important for smooth muscle cell migration

    DEFF Research Database (Denmark)

    Ström, A.; Olin, A. I.; Aspberg, A.

    2006-01-01

    /hyaluronan complexes, an ECM network that has been suggested to be important during tissue repair. In this study we have analysed the presence of fibulin-2 in two different models of murine vascular lesions. We have also examined how the fibulin-2/versican network influences SMC migration. Methods: Presence of fibulin......Objective: The vascular extracellular matrix (ECM) can affect smooth muscle cell (SMC) adhesion, migration and proliferation-events that are important during the atherosclerotic process. Fibulin-2 is a member of the ECM protein family of fibulins and has been found to cross-link versican...... and is upregulated during SMC phenotypic modulation in cell culture. Moreover, treatments with peptides that block the interaction between versican and fibulin-2 inhibit SMC migration in vitro. Conclusions: Fibulin-2 can be produced by SMC as a response to injury and may participate in the ECM organisation...

  3. Functional role of stromal interaction molecule 1 (STIM1) in vascular smooth muscle cells

    International Nuclear Information System (INIS)

    Takahashi, Yoichiro; Watanabe, Hiroyuki; Murakami, Manabu; Ono, Kyoichi; Munehisa, Yoshiko; Koyama, Takashi; Nobori, Kiyoshi; Iijima, Toshihiko; Ito, Hiroshi

    2007-01-01

    We investigated the functional role of STIM1, a Ca 2+ sensor in the endoplasmic reticulum (ER) that regulates store-operated Ca 2+ entry (SOCE), in vascular smooth muscle cells (VSMCs). STIM1 was mainly localized at the ER and plasma membrane. The knockdown of STIM1 expression by small interfering (si) RNA drastically decreased SOCE. In contrast, an EF-hand mutant of STIM1, STIM1 E87A , produced a marked increase in SOCE, which was abolished by co-transfection with siRNA to transient receptor potential canonical 1 (TRPC1). In addition, transfection with siRNA against STIM1 suppressed phosphorylation of cAMP-responsive element binding protein (CREB) and cell growth. These results suggest that STIM1 is an essential component of SOCE and that it is involved in VSMC proliferation

  4. Micropatterned coculture of vascular endothelial and smooth muscle cells on layered electrospun fibrous mats toward blood vessel engineering.

    Science.gov (United States)

    Li, Huinan; Liu, Yaowen; Lu, Jinfu; Wei, Jiaojun; Li, Xiaohong

    2015-06-01

    A major challenge in vascular engineering is the establishment of proper microenvironment to guide the spatial organization, growth, and extracellular matrix (ECM) productions of cells found in blood vessels. In the current study, micropatterned fibrous mats with distinct ridges and grooves of different width were created to load smooth muscle cells (SMCs), which were assembled by stacking on vascular endothelial cell (EC)-loaded flat fibrous mats to mimic the in vivo-like organized structure of blood vessels. SMCs were mainly distributed in the ridges, and aligned fibers in the patterned regions led to the formation of elongated cell bodies, intense actin filaments, and expressions of collagen I and α-smooth muscle actin in a parallel direction with fibers. ECs spread over the flat fibrous mats and expressed collagen IV and laminin with a cobblestone-like feature. A z-stack scanning of fluorescently stained fibrous mats indicated that SMCs effectively infiltrated into fibrous scaffolds at the depth of around 200 μm. Compared with SMCs cultured alone, the coculture with ECs enhanced the proliferation, infiltration, and cytoskeleton elongation of SMCs on patterned fibrous mats. Although the coculture of SMCs made no significant difference in the EC growth, the coculture system on patterned fibrous scaffolds promoted ECM productions of both ECs and SMCs. Thus, this patterned fibrous configuration not only offers a promising technology in the design of tissue engineering scaffolds to construct blood vessels with durable mechanical properties, but also provides a platform for patterned coculture to investigate cell-matrix and cell-cell interactions in highly organized tissues. © 2014 Wiley Periodicals, Inc.

  5. 3D Reconstruction of Coronary Artery Vascular Smooth Muscle Cells.

    Directory of Open Access Journals (Sweden)

    Tong Luo

    Full Text Available The 3D geometry of individual vascular smooth muscle cells (VSMCs, which are essential for understanding the mechanical function of blood vessels, are currently not available. This paper introduces a new 3D segmentation algorithm to determine VSMC morphology and orientation.A total of 112 VSMCs from six porcine coronary arteries were used in the analysis. A 3D semi-automatic segmentation method was developed to reconstruct individual VSMCs from cell clumps as well as to extract the 3D geometry of VSMCs. A new edge blocking model was introduced to recognize cell boundary while an edge growing was developed for optimal interpolation and edge verification. The proposed methods were designed based on Region of Interest (ROI selected by user and interactive responses of limited key edges. Enhanced cell boundary features were used to construct the cell's initial boundary for further edge growing. A unified framework of morphological parameters (dimensions and orientations was proposed for the 3D volume data. Virtual phantom was designed to validate the tilt angle measurements, while other parameters extracted from 3D segmentations were compared with manual measurements to assess the accuracy of the algorithm. The length, width and thickness of VSMCs were 62.9±14.9 μm, 4.6±0.6 μm and 6.2±1.8 μm (mean±SD. In longitudinal-circumferential plane of blood vessel, VSMCs align off the circumferential direction with two mean angles of -19.4±9.3° and 10.9±4.7°, while an out-of-plane angle (i.e., radial tilt angle was found to be 8±7.6° with median as 5.7°.A 3D segmentation algorithm was developed to reconstruct individual VSMCs of blood vessel walls based on optical image stacks. The results were validated by a virtual phantom and manual measurement. The obtained 3D geometries can be utilized in mathematical models and leads a better understanding of vascular mechanical properties and function.

  6. NF-kappaB signaling mediates vascular smooth muscle endothelin type B receptor expression in resistance arteries

    DEFF Research Database (Denmark)

    Zheng, Jian-Pu; Zhang, Yaping; Edvinsson, Lars

    2010-01-01

    Vascular smooth muscle cells (SMC) endothelin type B (ET(B)) receptor upregulation results in strong vasoconstriction and reduction of local blood flow. We hypothesizes that the underlying molecular mechanisms involve transcriptional factor nuclear factor-kappaB (NF-kappaB) pathway. ET(B) recepto...

  7. Lipid Metabolism in Vascular Smooth Muscle Cells Infuenced by HCMV Infection

    Directory of Open Access Journals (Sweden)

    Lingfang Li

    2016-10-01

    Full Text Available Background: The present study was designed to observe the infection of human cytomegalovirus (HCMV to human vascular smooth muscle cells (VSMCs, and the effect of viral infection on lipid metabolism in VSMCs. Methods: The cytopathic effects were observed by inverted microscopy and viral infection were examined by electron microscopy and RT-PCR. The lipid metabolism related gene profiling of VSMCs after HCMV infection was assayed by cDNA assay and the abnormal expression of genes were validated by quantitative RT-PCR. The content of cholesterol in VSMCs after HCMV infection was assayed by cholesterol detection kit. Results: VSMCs showed obvious cytopathic effects after HCMV infection. Intact viral particles could be detected in VSMCs using electron microscope. By use of RT-PCR technology, IE gene of HCMV could be amplified from VSMCs. The expression of cell lipid metabolism related gene profiling showed obvious disorders. The expression levels of HMG-CoA synthase and HMG-CoA reductase after infection increased significantly. The cellular cholesterol content (µmol/106 cells was significantly higher than that of mock infected group at 72h post infection. Conclusion: HCMV can infect VSMCs and the infection can affect cellular lipid metabolism related gene expression, which get involved in the occurrence and development of atherosclerosis (AS.

  8. Polysaccharide from Fuzi protects against Ox-LDL-induced calcification of human vascular smooth muscle cells by increasing autophagic activity

    Science.gov (United States)

    Liao, Lizhen; Zhuang, Xiaodong; Li, Weidong; Su, Qibiao; Zhao, Jie; Liu, Ying

    2018-01-01

    Polysaccharide from Fuzi (FPS) is a water-soluble polysaccharide isolated from the traditional Chinese herbal medicine Fuzi. It has been demonstrated to protect hepatocytes against ischemia-reperfusion injury through its potent antioxidant effects, and to attenuate starvation-induced cytotoxicity in H9c2 cells by increasing autophagic activity. In the present study, Alizarin Red S staining was used to detect mineral deposition and reverse transcription-quantitative polymerase chain reaction was used to detect the core binding factor α1 and smooth muscle 22α mRNA expression. To analyze autophagic activity, western blotting was used to detect microtubule-associated protein 1A/1B light chain 3 and nucleoporin P62 expression. In addition, green fluorescent protein-LC3 dots-per-cell was observed by fluorescence microscopy. It was demonstrated that oxidized low-density lipoprotein (Ox-LDL) could increase the calcification of human vascular smooth muscle cells (VSMCs) in a concentration-dependent manner, and that FPS treatment had a significant protective effect against Ox-LDL-induced calcification of human VSMCs. Furthermore, FPS treatment alleviated the Ox-LDL-induced downregulation of autophagic activity, and the protective effect of FPS on Ox-LDL-induced calcification was attenuated by the autophagy inhibitor 3-methyladenine. In conclusion, the present study demonstrated for the first time to the best of the authors' knowledge that FPS can protect against Ox-LDL-induced vascular calcification in human VSMCs, and that this likely occurs via the activation of autophagy. This supports the hypothesis that autophagy may be an endogenous protective mechanism counteracting vascular calcification, and that FPS may be used as a potential therapeutic for vascular calcification. PMID:29393437

  9. Kaempferol inhibits vascular smooth muscle cell migration by modulating BMP-mediated miR-21 expression.

    Science.gov (United States)

    Kim, Kwangho; Kim, Sunghwan; Moh, Sang Hyun; Kang, Hara

    2015-09-01

    Bioflavonoids are known to induce cardioprotective effects by inhibiting vascular smooth muscle cell (VSMC) proliferation and migration. Kaempferol has been shown to inhibit VSMC proliferation. However, little is known about the effect of kaempferol on VSMC migration and the underlying molecular mechanisms. Our studies provide the first evidence that kaempferol inhibits VSMC migration by modulating the BMP4 signaling pathway and microRNA expression levels. Kaempferol activates the BMP signaling pathway, induces miR-21 expression and downregulates DOCK4, 5, and 7, leading to inhibition of cell migration. Moreover, kaempferol antagonizes the PDGF-mediated pro-migratory effect. Therefore, our study uncovers a novel regulatory mechanism of VSMC migration by kaempferol and suggests that miRNA modulation by kaempferol is a potential therapy for cardiovascular diseases.

  10. Constituents of Mediterranean Spices Counteracting Vascular Smooth Muscle Cell Proliferation: Identification and Characterization of Rosmarinic Acid Methyl Ester as a Novel Inhibitor

    Czech Academy of Sciences Publication Activity Database

    Liu, R.; Heiss, E.H.; Waltenberger, B.; Blažević, T.; Schachner, B.; Jiang, B.; Kryštof, Vladimír; Liu, W.; Schwaiger, S.; Peña-Rodríguez, L. M.; Breuss, J.; Stuppner, H.; Dirsch, V.M.; Atanasov, A. G.

    2018-01-01

    Roč. 62, č. 7 (2018), č. článku 1700860. ISSN 1613-4125 Institutional support: RVO:61389030 Keywords : Mediterranean spices * neointima formation * rosmarinic acid * rosmarinic acid methyl ester * vascular smooth muscle cells Subject RIV: CE - Biochemistry OBOR OECD: Biochemical research methods Impact factor: 4.323, year: 2016

  11. Oxytocin receptors expressed and coupled to Ca2+ signalling in a human vascular smooth muscle cell line.

    OpenAIRE

    Yazawa, H.; Hirasawa, A.; Horie, K.; Saita, Y.; Iida, E.; Honda, K.; Tsujimoto, G.

    1996-01-01

    1. In a human vascular smooth muscle cell line (HVSMC), binding experiments with [3H]-arginine8-vasopressin (AVP) have shown the existence of a homogeneous population of binding sites with affinity (Kd value) of 0.65 nM and a maximum number of binding sites (Bmax) of 122 fmol mg-1 protein. 2. Nonlabelled compounds compete for [3H]-AVP binding in the HVSMC membrane with an order of potency of oxytocin > lyspressin > or = AVP > Thr4, Gly7-oxytocin > (beta-mercapto-beta-beta-cyclopentamethylenep...

  12. Nuclear fusion-independent smooth muscle differentiation of human adipose-derived stem cells induced by a smooth muscle environment.

    Science.gov (United States)

    Zhang, Rong; Jack, Gregory S; Rao, Nagesh; Zuk, Patricia; Ignarro, Louis J; Wu, Benjamin; Rodríguez, Larissa V

    2012-03-01

    Human adipose-derived stem cells hASC have been isolated and were shown to have multilineage differentiation capacity. Although both plasticity and cell fusion have been suggested as mechanisms for cell differentiation in vivo, the effect of the local in vivo environment on the differentiation of adipose-derived stem cells has not been evaluated. We previously reported the in vitro capacity of smooth muscle differentiation of these cells. In this study, we evaluate the effect of an in vivo smooth muscle environment in the differentiation of hASC. We studied this by two experimental designs: (a) in vivo evaluation of smooth muscle differentiation of hASC injected into a smooth muscle environment and (b) in vitro evaluation of smooth muscle differentiation capacity of hASC exposed to bladder smooth muscle cells. Our results indicate a time-dependent differentiation of hASC into mature smooth muscle cells when these cells are injected into the smooth musculature of the urinary bladder. Similar findings were seen when the cells were cocultured in vitro with primary bladder smooth muscle cells. Chromosomal analysis demonstrated that microenvironment cues rather than nuclear fusion are responsible for this differentiation. We conclude that cell plasticity is present in hASCs, and their differentiation is accomplished in the absence of nuclear fusion. Copyright © 2011 AlphaMed Press.

  13. UAP56 is an important mediator of Angiotensin II/platelet derived growth factor induced vascular smooth muscle cell DNA synthesis and proliferation

    International Nuclear Information System (INIS)

    Sahni, Abha; Wang, Nadan; Alexis, Jeffrey

    2013-01-01

    Highlights: ► Knockdown of UAP56 inhibits Angiotensin II/PDGF induced vascular smooth muscle cell proliferation. ► UAP56 is a positive regulator of E2F transcriptional activation. ► UAP56 is present in the vessel wall of low flow carotid arteries. -- Abstract: Angiotensin (Ang) II and platelet-derived growth factor (PDGF) are important mediators of pathologic vascular smooth muscle cell (VSMC) proliferation. Identifying downstream mediators of Ang II and PDGF signaling may provide insights for therapies to improve vascular proliferative diseases. We have previously demonstrated that breakpoint cluster region (Bcr) is an important mediator of Ang II/PDGF signaling in VSMC. We have recently reported that the DExD/H box protein UAP56 is an interacting partner of Bcr in regulating VSMC DNA synthesis. We hypothesized that UAP56 itself is an important regulator of VSMC proliferation. In this report we demonstrate that knockdown of UAP56 inhibits Ang II/PDGF induced VSMC DNA synthesis and proliferation, and inhibits E2F transcriptional activity. In addition, we demonstrate that UAP56 is present in the vessel wall of low-flow carotid arteries. These findings suggest that UAP56 is a regulator of VSMC proliferation and identify UAP56 as a target for preventing vascular proliferative disease

  14. Kir2.1 regulates rat smooth muscle cell proliferation, migration, and post-injury carotid neointimal formation

    International Nuclear Information System (INIS)

    Qiao, Yong; Tang, Chengchun; Wang, Qingjie; Wang, Dong; Yan, Gaoliang; Zhu, Boqian

    2016-01-01

    Phenotype switching of vascular smooth muscle cells (VSMC) from the contractile type to the synthetic type is a hallmark of vascular disorders such as atherosclerosis and restenosis after angioplasty. Inward rectifier K"+ channel 2.1 (Kir2.1) has been identified in VSMC. However, whether it plays a functional role in regulating cellular transformation remains obscure. In this study, we evaluated the role of Kir2.1 on VSMC proliferation, migration, phenotype switching, and post-injury carotid neointimal formation. Kir2.1 knockdown significantly suppressed platelet-derived growth factor BB-stimulated rat vascular smooth muscle cells (rat-VSMC) proliferation and migration. Deficiency in Kir2.1 contributed to the restoration of smooth muscle α-actin, smooth muscle 22α, and calponin and to a reduction in osteopontin expression in rat-VSMC. Moreover, the in vivo study showed that rat-VSMC switched to proliferative phenotypes and that knockdown of Kir2.1 significantly inhibited neointimal formation after rat carotid injury. Kir2.1 may be a potential therapeutic target in the treatment of cardiovascular diseases, such as atherosclerosis and restenosis following percutaneous coronary intervention.

  15. Kir2.1 regulates rat smooth muscle cell proliferation, migration, and post-injury carotid neointimal formation

    Energy Technology Data Exchange (ETDEWEB)

    Qiao, Yong; Tang, Chengchun, E-mail: tangchengchun@medmail.com.cn; Wang, Qingjie; Wang, Dong; Yan, Gaoliang; Zhu, Boqian

    2016-09-02

    Phenotype switching of vascular smooth muscle cells (VSMC) from the contractile type to the synthetic type is a hallmark of vascular disorders such as atherosclerosis and restenosis after angioplasty. Inward rectifier K{sup +} channel 2.1 (Kir2.1) has been identified in VSMC. However, whether it plays a functional role in regulating cellular transformation remains obscure. In this study, we evaluated the role of Kir2.1 on VSMC proliferation, migration, phenotype switching, and post-injury carotid neointimal formation. Kir2.1 knockdown significantly suppressed platelet-derived growth factor BB-stimulated rat vascular smooth muscle cells (rat-VSMC) proliferation and migration. Deficiency in Kir2.1 contributed to the restoration of smooth muscle α-actin, smooth muscle 22α, and calponin and to a reduction in osteopontin expression in rat-VSMC. Moreover, the in vivo study showed that rat-VSMC switched to proliferative phenotypes and that knockdown of Kir2.1 significantly inhibited neointimal formation after rat carotid injury. Kir2.1 may be a potential therapeutic target in the treatment of cardiovascular diseases, such as atherosclerosis and restenosis following percutaneous coronary intervention.

  16. Effect of Oxysterol-Induced Apoptosis of Vascular Smooth Muscle Cells on Experimental Hypercholesterolemia

    Science.gov (United States)

    Perales, Sonia; Alejandre, M. José; Palomino-Morales, Rogelio; Torres, Carolina; Iglesias, Jose; Linares, Ana

    2009-01-01

    Smooth muscle cells (SMCs) undergo changes related to proliferation and apoptosis in the physiological remodeling of vessels and in diseases such as atherosclerosis and restenosis. Recent studies also have demonstrated the vascular cell proliferation and programmed cell death contribute to changes in vascular architecture in normal development and in disease. The present study was designed to investigate the apoptotic pathways induced by 25-hydroxycholesterol in SMCs cultures, using an in vivo/in vitro cell model in which SMCs were isolated and culture from chicken exposed to an atherogenic cholesterol-rich diet (SMC-Ch) and/or an antiatherogenic fish oil-rich diet (SMC-Ch-FO). Cells were exposed in vitro to 25-hydroxycholesterol to study levels of apoptosis and apoptotic proteins Bcl-2, Bcl-XL and Bax and the expression of bcl-2 and bcl-xL, genes. The quantitative real-time reverse transcriptase-polymerase chain reaction and the Immunoblotting western blot analysis showed that 25-hydroxycholesterol produces apoptosis in SMCs, mediated by a high increase in Bax protein and Bax gene expression. These changes were more marked in SMC-Ch than in SMC-Ch-FO, indicating that dietary cholesterol produces changes in SMCs that make them more susceptible to 25-hydroxycholesterol-mediated apoptosis. Our results suggest that the replacement of a cholesterol-rich diet with a fish oil-rich diet produces some reversal of cholesterol-induced changes in the apoptotic pathways induced by 25-hydroxycholesterol in SMCs cultures, making SMCs more resistant to apoptosis. PMID:19727411

  17. Effect of Oxysterol-Induced Apoptosis of Vascular Smooth Muscle Cells on Experimental Hypercholesterolemia

    Directory of Open Access Journals (Sweden)

    Sonia Perales

    2009-01-01

    Full Text Available Smooth muscle cells (SMCs undergo changes related to proliferation and apoptosis in the physiological remodeling of vessels and in diseases such as atherosclerosis and restenosis. Recent studies also have demonstrated the vascular cell proliferation and programmed cell death contribute to changes in vascular architecture in normal development and in disease. The present study was designed to investigate the apoptotic pathways induced by 25-hydroxycholesterol in SMCs cultures, using an in vivo/in vitro cell model in which SMCs were isolated and culture from chicken exposed to an atherogenic cholesterol-rich diet (SMC-Ch and/or an antiatherogenic fish oil-rich diet (SMC-Ch-FO. Cells were exposed in vitro to 25-hydroxycholesterol to study levels of apoptosis and apoptotic proteins Bcl-2, Bcl-XL and Bax and the expression of bcl-2 and bcl-xL, genes. The quantitative real-time reverse transcriptase-polymerase chain reaction and the Immunoblotting western blot analysis showed that 25-hydroxycholesterol produces apoptosis in SMCs, mediated by a high increase in Bax protein and Bax gene expression. These changes were more marked in SMC-Ch than in SMC-Ch-FO, indicating that dietary cholesterol produces changes in SMCs that make them more susceptible to 25-hydroxycholesterol-mediated apoptosis. Our results suggest that the replacement of a cholesterol-rich diet with a fish oil-rich diet produces some reversal of cholesterol-induced changes in the apoptotic pathways induced by 25-hydroxycholesterol in SMCs cultures, making SMCs more resistant to apoptosis.

  18. Heterogeneity of smooth muscle cells in tunica media of aorta in ...

    African Journals Online (AJOL)

    ... of the tunica media of goat aorta are phenotypically heterogeneous and run in multiple directions. These characteristics probably confer mechanical strength and functional plasticity to the aortic wall. Designers of aortic substitutes should bear this in mind. Keywords: Vascular, Smooth Muscle Cells, Heterogeneity, Aorta ...

  19. Smooth Muscle Endothelin B Receptors Regulate Blood Pressure but Not Vascular Function or Neointimal Remodeling.

    Science.gov (United States)

    Miller, Eileen; Czopek, Alicja; Duthie, Karolina M; Kirkby, Nicholas S; van de Putte, Elisabeth E Fransen; Christen, Sibylle; Kimmitt, Robert A; Moorhouse, Rebecca; Castellan, Raphael F P; Kotelevtsev, Yuri V; Kuc, Rhoda E; Davenport, Anthony P; Dhaun, Neeraj; Webb, David J; Hadoke, Patrick W F

    2017-02-01

    The role of smooth muscle endothelin B (ET B ) receptors in regulating vascular function, blood pressure (BP), and neointimal remodeling has not been established. Selective knockout mice were generated to address the hypothesis that loss of smooth muscle ET B receptors would reduce BP, alter vascular contractility, and inhibit neointimal remodeling. ET B receptors were selectively deleted from smooth muscle by crossing floxed ET B mice with those expressing cre-recombinase controlled by the transgelin promoter. Functional consequences of ET B deletion were assessed using myography. BP was measured by telemetry, and neointimal lesion formation induced by femoral artery injury. Lesion size and composition (day 28) were analyzed using optical projection tomography, histology, and immunohistochemistry. Selective deletion of ET B was confirmed by genotyping, autoradiography, polymerase chain reaction, and immunohistochemistry. ET B -mediated contraction was reduced in trachea, but abolished from mesenteric veins, of knockout mice. Induction of ET B -mediated contraction in mesenteric arteries was also abolished in these mice. Femoral artery function was unaltered, and baseline BP modestly elevated in smooth muscle ET B knockout compared with controls (+4.2±0.2 mm Hg; P<0.0001), but salt-induced and ET B blockade-mediated hypertension were unaltered. Circulating endothelin-1 was not altered in knockout mice. ET B -mediated contraction was not induced in femoral arteries by incubation in culture medium or lesion formation, and lesion size was not altered in smooth muscle ET B knockout mice. In the absence of other pathology, ET B receptors in vascular smooth muscle make a small but significant contribution to ET B -dependent regulation of BP. These ET B receptors have no effect on vascular contraction or neointimal remodeling. © 2016 The Authors.

  20. Brain cytoplasmic RNA 1 suppresses smooth muscle differentiation and vascular development in mice.

    Science.gov (United States)

    Wang, Yung-Chun; Chuang, Ya-Hui; Shao, Qiang; Chen, Jian-Fu; Chen, Shi-You

    2018-04-13

    The cardiovascular system develops during the early stages of embryogenesis, and differentiation of smooth muscle cells (SMCs) is essential for that process. SMC differentiation is critically regulated by transforming growth factor (TGF)-β/SMAD family member 3 (SMAD3) signaling, but other regulators may also play a role. For example, long noncoding RNAs (lncRNAs) regulate various cellular activities and events, such as proliferation, differentiation, and apoptosis. However, whether long noncoding RNAs also regulate SMC differentiation remains largely unknown. Here, using the murine cell line C3H10T1/2, we found that brain cytoplasmic RNA 1 (BC1) is an important regulator of SMC differentiation. BC1 overexpression suppressed, whereas BC1 knockdown promoted, TGF-β-induced SMC differentiation, as indicated by altered cell morphology and expression of multiple SMC markers, including smooth muscle α-actin (αSMA), calponin, and smooth muscle 22α (SM22α). BC1 appeared to block SMAD3 activity and inhibit SMC marker gene transcription. Mechanistically, BC1 bound to SMAD3 via RNA SMAD-binding elements (rSBEs) and thus impeded TGF-β-induced SMAD3 translocation to the nucleus. This prevented SMAD3 from binding to SBEs in SMC marker gene promoters, an essential event in SMC marker transcription. In vivo , BC1 overexpression in mouse embryos impaired vascular SMC differentiation, leading to structural defects in the artery wall, such as random breaks in the elastic lamina, abnormal collagen deposition on SM fibers, and disorganized extracellular matrix proteins in the media of the neonatal aorta. Our results suggest that BC1 is a suppressor of SMC differentiation during vascular development. © 2018 by The American Society for Biochemistry and Molecular Biology, Inc.

  1. Vascular smooth muscle cell stiffness and adhesion to collagen I modified by vasoactive agonists.

    Directory of Open Access Journals (Sweden)

    Zhongkui Hong

    Full Text Available In vascular smooth muscle cells (VSMCs integrin-mediated adhesion to extracellular matrix (ECM proteins play important roles in sustaining vascular tone and resistance. The main goal of this study was to determine whether VSMCs adhesion to type I collagen (COL-I was altered in parallel with the changes in the VSMCs contractile state induced by vasoconstrictors and vasodilators. VSMCs were isolated from rat cremaster skeletal muscle arterioles and maintained in primary culture without passage. Cell adhesion and cell E-modulus were assessed using atomic force microscopy (AFM by repetitive nano-indentation of the AFM probe on the cell surface at 0.1 Hz sampling frequency and 3200 nm Z-piezo travelling distance (approach and retraction. AFM probes were tipped with a 5 μm diameter microbead functionalized with COL-I (1 mg\\ml. Results showed that the vasoconstrictor angiotensin II (ANG-II; 10-6 significantly increased (p<0.05 VSMC E-modulus and adhesion probability to COL-I by approximately 35% and 33%, respectively. In contrast, the vasodilator adenosine (ADO; 10-4 significantly decreased (p<0.05 VSMC E-modulus and adhesion probability by approximately -33% and -17%, respectively. Similarly, the NO donor (PANOate, 10-6 M, a potent vasodilator, also significantly decreased (p<0.05 the VSMC E-modulus and COL-I adhesion probability by -38% and -35%, respectively. These observations support the hypothesis that integrin-mediated VSMC adhesion to the ECM protein COL-I is dynamically regulated in parallel with VSMC contractile activation. These data suggest that the signal transduction pathways modulating VSMC contractile activation and relaxation, in addition to ECM adhesion, interact during regulation of contractile state.

  2. Vascular endothelial dysfunction in β-thalassemia occurs despite increased eNOS expression and preserved vascular smooth muscle cell reactivity to NO.

    Directory of Open Access Journals (Sweden)

    Ekatherina Stoyanova

    Full Text Available The hereditary β-thalassemia major condition requires regular lifelong blood transfusions. Transfusion-related iron overloading has been associated with the onset of cardiovascular complications, including cardiac dysfunction and vascular anomalies. By using an untransfused murine model of β-thalassemia major, we tested the hypothesis that vascular endothelial dysfunction, alterations of arterial structure and of its mechanical properties would occur despite the absence of treatments.Vascular function and structure were evaluated ex vivo. Compared to the controls, endothelium-dependent vasodilation with acetylcholine was blunted in mesenteric resistance arteries of β-thalassemic mice while the endothelium-independent vasodilator (sodium nitroprusside produced comparable vessel dilation, indicating endothelial cell impairment with preserved smooth muscle cell reactivity to nitric oxide (NO. While these findings suggest a decrease in NO bioavailability, Western blotting showed heightened expression of aortic endothelial NO synthase (eNOS in β-thalassemia. Vascular remodeling of the common carotid arteries revealed increased medial elastin content. Under isobaric conditions, the carotid arteries of β-thalassemic mice exhibited decreased wall stress and softening due to structural changes of the vessel wall.A complex vasculopathy was identified in untransfused β-thalassemic mice characterized by altered carotid artery structure and endothelial dysfunction of resistance arterioles, likely attributable to reduced NO bioavailability despite enhanced vascular eNOS expression.

  3. Arterial wall mechanics as a function of heart rate: role of vascular smooth muscle

    International Nuclear Information System (INIS)

    Salvucci, Fernando Pablo; Schiavone, Jonathan; Craiem, Damian; Barra, Juan Gabriel

    2007-01-01

    Vascular wall viscoelasticity can be evaluated using a first-order lumped model. This model consists of a spring with elastic constant E and a dashpot with viscous constant η. More importantly, this viscoelastic model can be fitted in-vivo measuring arterial pressure and diameter. The aim of this work is to analyze the influence of heart rate over E and η. In two anesthetized sheep, diameter in thoracic aorta and intravascular pressure has been registered. The right atrium was connected to a programmable stimulator through a pair of pace-maker wires to produce changes in stimulation heart rate (HR) from 80 to 160 bpm. Additionally, local activation of vascular smooth muscle was induced with phenylephrine. After converting pressure and diameter signals into stress and strain respectively, E y η were calculated in control state and during muscle activation. The elastic modulus E did not present significant changes with heart rate. The viscous modulus η decreased 49% with a two-fold acceleration in heart rate from 80 to 160 bpm. However, the product η HR remained stable. The viscous modulus η increased 39% with smooth muscle activation. No significant pressure changes were registered during the experiment. The contractile action of vascular smooth muscle could contribute to increasing arterial wall viscosity. The decrease of η when HR increased might be related to smooth muscle relaxation mediated by endothelium activity, which was stimulated by flow increase. We conclude that HR can modulate arterial wall viscoelasticity through endothelium-dependent mechanisms

  4. Substance-specific importance of EGFR for vascular smooth muscle cells motility in primary culture.

    Science.gov (United States)

    Schreier, Barbara; Schwerdt, Gerald; Heise, Christian; Bethmann, Daniel; Rabe, Sindy; Mildenberger, Sigrid; Gekle, Michael

    2016-07-01

    Besides their importance for the vascular tone, vascular smooth muscle cells (VSMC) also contribute to pathophysiological vessel alterations. Various G-protein coupled receptor ligands involved in vascular dysfunction and remodeling can transactivate the epidermal growth factor receptor (EGFR) of VSMC, yet the importance of EGFR transactivation for the VSMC phenotype is incompletely understood. The aims of this study were (i) to characterize further the importance of the VSMC-EGFR for proliferation, migration and marker gene expression for inflammation, fibrosis and reactive oxygen species (ROS) homeostasis and (ii) to test the hypothesis that vasoactive substances (endothelin-1, phenylephrine, thrombin, vasopressin and ATP) rely differentially on the EGFR with respect to the abovementioned phenotypic alterations. In primary, aortic VSMC from mice without conditional deletion of the EGFR, proliferation, migration, marker gene expression (inflammation, fibrosis and ROS homeostasis) and cell signaling (ERK 1/2, intracellular calcium) were analyzed. VSMC-EGFR loss reduced collective cell migration and single cell migration probability, while no difference between the genotypes in single cell velocity, chemotaxis or marker gene expression could be observed under control conditions. EGF promoted proliferation, collective cell migration, chemokinesis and chemotaxis and leads to a proinflammatory gene expression profile in wildtype but not in knockout VSMC. Comparing the impact of five vasoactive substances (all reported to transactivate EGFR and all leading to an EGFR dependent increase in ERK1/2 phosphorylation), we demonstrate that the importance of EGFR for their action is substance-dependent and most apparent for crowd migration but plays a minor role for gene expression regulation. Copyright © 2016 Elsevier B.V. All rights reserved.

  5. Piperlongumine inhibits atherosclerotic plaque formation and vascular smooth muscle cell proliferation by suppressing PDGF receptor signaling

    Energy Technology Data Exchange (ETDEWEB)

    Son, Dong Ju [Wallace H. Coulter Department of Biomedical Engineering, Georgia Institute of Technology and Emory University, Atlanta, GA (United States); Division of Cardiology, Department of Medicine, Emory University, Atlanta, GA (United States); Kim, Soo Yeon [Division of Life Science, Korea Basic Science Institute, Daejeon (Korea, Republic of); Han, Seong Su [University of Iowa Carver College of Medicine, Department of Pathology, Iowa City, IA (United States); Kim, Chan Woo [Wallace H. Coulter Department of Biomedical Engineering, Georgia Institute of Technology and Emory University, Atlanta, GA (United States); Division of Cardiology, Department of Medicine, Emory University, Atlanta, GA (United States); Department of Bioinspired Science, Ehwa Womans University, Seoul (Korea, Republic of); Kumar, Sandeep [Wallace H. Coulter Department of Biomedical Engineering, Georgia Institute of Technology and Emory University, Atlanta, GA (United States); Division of Cardiology, Department of Medicine, Emory University, Atlanta, GA (United States); Park, Byeoung Soo [Nanotoxtech Co., Ansan (Korea, Republic of); Lee, Sung Eun [Division of Applied Biology and Chemistry, Kyungpook National University, Daegu (Korea, Republic of); Yun, Yeo Pyo [College of Pharmacy, Chungbuk National University, Cheongju (Korea, Republic of); Jo, Hanjoong, E-mail: hjo@emory.edu [Wallace H. Coulter Department of Biomedical Engineering, Georgia Institute of Technology and Emory University, Atlanta, GA (United States); Division of Cardiology, Department of Medicine, Emory University, Atlanta, GA (United States); Department of Bioinspired Science, Ehwa Womans University, Seoul (Korea, Republic of); Park, Young Hyun, E-mail: pyh012@sch.ac.kr [Department of Food Science and Nutrition, College of Natural Sciences, Soonchunhyang University, Asan (Korea, Republic of)

    2012-10-19

    Highlights: Black-Right-Pointing-Pointer Anti-atherogenic effect of PL was examined using partial carotid ligation model in ApoE KO mice. Black-Right-Pointing-Pointer PL prevented atherosclerotic plaque development, VSMCs proliferation, and NF-{kappa}B activation. Black-Right-Pointing-Pointer Piperlongumine reduced vascular smooth muscle cell activation through PDGF-R{beta} and NF-{kappa}B-signaling. Black-Right-Pointing-Pointer PL may serve as a new therapeutic molecule for atherosclerosis treatment. -- Abstract: Piperlongumine (piplartine, PL) is an alkaloid found in the long pepper (Piper longum L.) and has well-documented anti-platelet aggregation, anti-inflammatory, and anti-cancer properties; however, the role of PL in prevention of atherosclerosis is unknown. We evaluated the anti-atherosclerotic potential of PL in an in vivo murine model of accelerated atherosclerosis and defined its mechanism of action in aortic vascular smooth muscle cells (VSMCs) in vitro. Local treatment with PL significantly reduced atherosclerotic plaque formation as well as proliferation and nuclear factor-kappa B (NF-{kappa}B) activation in an in vivo setting. PL treatment in VSMCs in vitro showed inhibition of migration and platelet-derived growth factor BB (PDGF-BB)-induced proliferation to the in vivo findings. We further identified that PL inhibited PDGF-BB-induced PDGF receptor beta activation and suppressed downstream signaling molecules such as phospholipase C{gamma}1, extracellular signal-regulated kinases 1 and 2 and Akt. Lastly, PL significantly attenuated activation of NF-{kappa}B-a downstream transcriptional regulator in PDGF receptor signaling, in response to PDGF-BB stimulation. In conclusion, our findings demonstrate a novel, therapeutic mechanism by which PL suppresses atherosclerosis plaque formation in vivo.

  6. Hypoxia-inducible factor-1 plays a role in phosphate-induced vascular smooth muscle cell calcification.

    Science.gov (United States)

    Mokas, Sophie; Larivière, Richard; Lamalice, Laurent; Gobeil, Stéphane; Cornfield, David N; Agharazii, Mohsen; Richard, Darren E

    2016-09-01

    Medial vascular calcification is a common complication of chronic kidney disease (CKD). Although elevated inorganic phosphate stimulates vascular smooth muscle cell (VSMC) osteogenic transdifferentiation and calcification, the mechanisms involved in their calcification during CKD are not fully defined. Because hypoxic gene activation is linked to CKD and stimulates bone cell osteogenic differentiation, we used in vivo and in vitro rodent models to define the role of hypoxic signaling during elevated inorganic phosphate-induced VSMC calcification. Cell mineralization studies showed that elevated inorganic phosphate rapidly induced VSMC calcification. Hypoxia strongly enhanced elevated inorganic phosphate-induced VSMC calcification and osteogenic transdifferentiation, as seen by osteogenic marker expression. Hypoxia-inducible factor-1 (HIF-1), the key hypoxic transcription factor, was essential for enhanced VSMC calcification. Targeting HIF-1 expression in murine VSMC blocked calcification in hypoxia with elevated inorganic phosphate while HIF-1 activators, including clinically used FG-4592/Roxadustat, recreated a procalcifying environment. Elevated inorganic phosphate rapidly activated HIF-1, even in normal oxygenation; an effect mediated by HIF-1α subunit stabilization. Thus, hypoxia synergizes with elevated inorganic phosphate to enhance VSMC osteogenic transdifferentiation. Our work identifies HIF-1 as an early CKD-related pathological event, prospective marker, and potential target against vascular calcification in CKD-relevant conditions. Copyright © 2016 International Society of Nephrology. Published by Elsevier Inc. All rights reserved.

  7. Bone marrow stromal and vascular smooth muscle cells have chemosensory capacity via bitter taste receptor expression.

    Directory of Open Access Journals (Sweden)

    Troy C Lund

    Full Text Available The ability of cells to detect changes in the microenvironment is important in cell signaling and responsiveness to environmental fluctuations. Our interest is in understanding how human bone marrow stromal-derived cells (MSC and their relatives, vascular smooth muscle cells (VSMC, interact with their environment through novel receptors. We found, through a proteomics screen, that MSC express the bitter taste receptor, TAS2R46, a protein more typically localized to the taste bud. Expression was also confirmed in VSMCs. A prototypical bitter compound that binds to the bitter taste receptor class, denatonium, increased intracellular calcium release and decreased cAMP levels as well as increased the extracellular release of ATP in human MSC. Denatonium also bound and activated rodent VSMC with a change in morphology upon compound exposure. Finally, rodents given denatonium in vivo had a significant drop in blood pressure indicating a vasodilator response. This is the first description of chemosensory detection by MSC and VSMCs via a taste receptor. These data open a new avenue of research into discovering novel compounds that operate through taste receptors expressed by cells in the marrow and vascular microenvironments.

  8. Mercury induces proliferation and reduces cell size in vascular smooth muscle cells through MAPK, oxidative stress and cyclooxygenase-2 pathways

    Energy Technology Data Exchange (ETDEWEB)

    Aguado, Andrea; Galán, María; Zhenyukh, Olha; Wiggers, Giulia A.; Roque, Fernanda R. [Departamento de Farmacología, Facultad de Medicina, Universidad Autónoma de Madrid, Instituto de Investigación Hospital Universitario La Paz (IdiPAZ), 28029, Madrid (Spain); Redondo, Santiago [Departamento de Farmacología, Facultad de Medicina, Universidad Complutense, 28040, Madrid (Spain); Peçanha, Franck [Departamento de Farmacología, Facultad de Medicina, Universidad Autónoma de Madrid, Instituto de Investigación Hospital Universitario La Paz (IdiPAZ), 28029, Madrid (Spain); Martín, Angela [Departamento de Bioquímica, Fisiología y Genética Molecular, Universidad Rey Juan Carlos, 28922, Alcorcón (Spain); Fortuño, Ana [Área de Ciencias Cardiovasculares, Centro de Investigación Médica Aplicada, Universidad de Navarra, 31008, Pamplona (Spain); Cachofeiro, Victoria [Departamento de Fisiología, Facultad de Medicina, Universidad Complutense, 28040, Madrid (Spain); Tejerina, Teresa [Departamento de Farmacología, Facultad de Medicina, Universidad Complutense, 28040, Madrid (Spain); Salaices, Mercedes, E-mail: mercedes.salaices@uam.es [Departamento de Farmacología, Facultad de Medicina, Universidad Autónoma de Madrid, Instituto de Investigación Hospital Universitario La Paz (IdiPAZ), 28029, Madrid (Spain); and others

    2013-04-15

    Mercury exposure is known to increase cardiovascular risk but the underlying cellular mechanisms remain undetermined. We analyzed whether chronic exposure to HgCl{sub 2} affects vascular structure and the functional properties of vascular smooth muscle cells (VSMC) through oxidative stress/cyclooxygenase-2 dependent pathways. Mesenteric resistance arteries and aortas from Wistar rats treated with HgCl{sub 2} (first dose 4.6 mg kg{sup −1}, subsequent doses 0.07 mg kg{sup −1} day{sup −1}, 30 days) and cultured aortic VSMC stimulated with HgCl{sub 2} (0.05–5 μg/ml) were used. Treatment of rats with HgCl{sub 2} decreased wall thickness of the resistance and conductance vasculature, increased the number of SMC within the media and decreased SMC nucleus size. In VSMCs, exposure to HgCl{sub 2}: 1) induced a proliferative response and a reduction in cell size; 2) increased superoxide anion production, NADPH oxidase activity, gene and/or protein levels of the NADPH oxidase subunit NOX-1, the EC- and Mn-superoxide dismutases and cyclooxygenase-2 (COX-2); 3) induced activation of ERK1/2 and p38 MAPK. Both antioxidants and COX-2 inhibitors normalized the proliferative response and the altered cell size induced by HgCl{sub 2}. Blockade of ERK1/2 and p38 signaling pathways abolished the HgCl{sub 2}-induced Nox1 and COX-2 expression and normalized the alterations induced by mercury in cell proliferation and size. In conclusion, long exposure of VSMC to low doses of mercury activates MAPK signaling pathways that result in activation of inflammatory proteins such as NADPH oxidase and COX-2 that in turn induce proliferation of VSMC and changes in cell size. These findings offer further evidence that mercury might be considered an environmental risk factor for cardiovascular disease. - Highlights: ► Chronic HgCl{sub 2} exposure induces vascular remodeling. ► HgCl{sub 2} induces proliferation and decreased cell size in vascular smooth muscle cells. ► HgCl{sub 2} induces

  9. Inhibition of Proliferation of Vascular Smooth Muscle Cells by Cucurbitanes from Momordica charantia.

    Science.gov (United States)

    Tuan, Nguyen Quoc; Lee, Do-Hyung; Oh, Joonseok; Kim, Chung Sub; Heo, Kyung-Sun; Myung, Chang-Seon; Na, MinKyun

    2017-07-28

    The cucurbitaceous plant Momordica charantia L., named "bitter melon", inhabits Asia, Africa, and South America and has been used as a traditional medicine. The atypical proliferation of vascular smooth muscle cells (VSMCs) plays an important role in triggering the pathogenesis of cardiovascular diseases. Platelet-derived growth factor (PDGF) is regarded as the most powerful growth factor in promoting the intimal accumulation of VSMCs. The current study features the identification of six new cucurbitane-type triterpenoids (1-6) from the fruits of M.  charantia, utilizing diverse chromatographic and spectroscopic techniques. In particular, the 2D structure of 1 was confirmed utilizing the long-range HSQMBC NMR pulse, capable of measuring heteronuclear long-range correlations ( 4-6 J CH ). The cucurbitanes were also assessed for their inhibitory activity against PDGF-induced VSMC proliferation. This current study may constitute a basis for developing those chemotypes into sensible pharmacophores alleviating cardiovascular disorders.

  10. 31P-nuclear magnetic resonance analysis of extracts of vascular smooth muscle

    International Nuclear Information System (INIS)

    Barron, J.T.; Messer, J.V.; Glonek, Thomas

    1986-01-01

    31 P-nuclear magnetic resonance spectroscopy was used to assess phosphate metabolites in perchloric acid extracts of rabbit aorta. In addition to the high energy phosphates, several other phosphorus compounds were detected and quantified. Most notable was the presence of a prominent phosphomonoester compound appearing at a chemical shift of 3.86 delta. This compound constituted 26% of the total extractable tissue phosphorus and is tentatively identified as ribose-5-phosphate, a pentose phosphate pathway intermediate. While ATP and phosphocreatine did not change during glucose and oxygen deprivation or during prolonged muscle contraction, the 3.86delta phosphate decreased significantly. Furthermore, theophylline, an agent that increases intracellular cAMP, also decreased the level of the 3.86 delta phosphate. These results are consistent with the concept that intermediate metabolism sustains high energy phosphate pools in vascular smooth muscle in the steady state under various conditions. The pentose phosphate pathway may play an important role in vascular smooth muscle metabolism. (author)

  11. Patent ductus arteriosus in mice with smooth muscle-specific Jag1 deletion

    Science.gov (United States)

    Feng, Xuesong; Krebs, Luke T.; Gridley, Thomas

    2010-01-01

    The ductus arteriosus is an arterial vessel that shunts blood flow away from the lungs during fetal life, but normally occludes after birth to establish the adult circulation pattern. Failure of the ductus arteriosus to close after birth is termed patent ductus arteriosus and is one of the most common congenital heart defects. Mice with smooth muscle cell-specific deletion of Jag1, which encodes a Notch ligand, die postnatally from patent ductus arteriosus. These mice exhibit defects in contractile smooth muscle cell differentiation in the vascular wall of the ductus arteriosus and adjacent descending aorta. These defects arise through an inability to propagate the JAG1-Notch signal via lateral induction throughout the width of the vascular wall. Both heterotypic endothelial smooth muscle cell interactions and homotypic vascular smooth muscle cell interactions are required for normal patterning and differentiation of the ductus arteriosus and adjacent descending aorta. This new model for a common congenital heart defect provides novel insights into the genetic programs that underlie ductus arteriosus development and closure. PMID:21068062

  12. The flavonoid quercetin induces apoptosis and inhibits JNK activation in intimal vascular smooth muscle cells

    International Nuclear Information System (INIS)

    Perez-Vizcaino, Francisco; Bishop-Bailley, David; Lodi, Federica; Duarte, Juan; Cogolludo, Angel; Moreno, Laura; Bosca, Lisardo; Mitchell, Jane A.; Warner, Timothy D.

    2006-01-01

    Quercetin, the most abundant dietary flavonol, exerts vasodilator, anti-hypertensive, and anti-atherogenic effects and reduces the vascular remodelling associated with elevated blood pressure. Here, we have compared the effects of quercetin in intimal- and medial-type rat vascular smooth muscle cells (VSMC) in culture. After 48 h, quercetin reduced the viability of a polyclonal intimal-type cell line derived from neonatal aorta but not of a medial-type cell line derived from adult aorta. These differential effects were similar in both proliferating and quiescent VSMC. Quercetin also preferentially reduced the viability of intimal-type over medial-type VSMC in primary cultures derived from balloon-injured carotid arteries. The effects of quercetin on cell viability were mainly dependent upon induction of apoptosis, as demonstrated by nuclear condensation and fragmentation, and were unrelated to PPARγ, pro-oxidant effects or nitric oxide. The expression of MAPKs (ERK, p38, and JNK) and ERK phosphorylation were not different between intimal- and medial-type VSMC. p38 phosphorylation was negligible in both cell types. Medial-type showed a weak JNK phosphorylation while this was markedly increased in intimal-type cells. Quercetin reduced JNK phosphorylation but had no consistent effect on ERK phosphorylation. In conclusion, quercetin preferentially produced apoptosis in intimal-type compared to medial-type VSMC. This might play a role in the anti-atherogenic and anti-hypertensive effects of quercetin

  13. Pituitary adenylate cyclase activating polypeptide induces vascular relaxation and inhibits non-vascular smooth muscle activity in the rabbit female genital tract

    DEFF Research Database (Denmark)

    Steenstrup, B R; Ottesen, B; Jørgensen, M

    1994-01-01

    In vitro effects of two bioactive forms of pituitary adenylate cyclase activating polypeptide (PACAP): PACAP-38 and PACAP-27 were studied on rabbit vascular and non-vascular smooth muscle. Segments of the ovarian artery and muscle strips from the fallopian tube were used. Two series of experiment...

  14. Hydroxysafflor yellow A suppresses oxidized low density lipoprotein induced proliferation of vascular smooth muscle cells

    Directory of Open Access Journals (Sweden)

    Lin Sheng

    2012-06-01

    Full Text Available To investigate the relationship between the suppression of Hydroxysafflor yellow A (HSYA on the oxidized low density lipoprotein (ox-LDL induced proliferation of vascular smooth muscle cells (VSMCs and the mRNA and protein expression of extracellular signal-regulated protein kinase 1/2 (ERK1/2 and mitogen activated protein kinase phospholipase-1 (MAKP-1, VSMCs were treated with HSYA at 10 ?mol/L and/or ox-LDL at 35 mg/L for 48 h. MTT assay was done to measure cell survival rate, flow cytometry to detect cell cycle, reverse transcription PCR and Western blot to detect the expression of ERK1/2 and MAKP-1. When compared to cells treated with ox-LDL alone, the survival rate of cells treated with two reagents was reduced and the proportion of cells in G0/G1 phase significantly increased, with increased MKP-1 expression. The study suggests HSYA can inhibit VSMC proliferation via increasing MKP-1 expression, reducing p-ERK1/2 activity and suppressing cell cycle.

  15. Buddleja officinalis suppresses high glucose-induced vascular smooth muscle cell proliferation: role of mitogen-activated protein kinases, nuclear factor-kappaB and matrix metalloproteinases.

    Science.gov (United States)

    Lee, Yun Jung; Kim, Jin Sook; Kang, Dae Gill; Lee, Ho Sub

    2010-02-01

    Diabetes mellitus is a well-established risk factor for vascular diseases caused by atherosclerosis. In the development of diabetic atherogenesis, vascular smooth muscle cell proliferation is recognized as a key event. Thus, we aimed to investigate whether an ethanol extract of Buddleja officinalis (EBO) suppresses high glucose-induced proliferation in primary cultured human aortic smooth muscle cells (HASMC). [(3)H]-thymidine incorporation revealed that incubation of HASMC with a high concentration of glucose (25 mmol/L) increased cell proliferation. The expression levels of cell cycle protein were also increased by treatment with high glucose concentration. Pretreatment of HASMC with EBO significantly attenuated the increase of high glucose-induced cell proliferation as well as p38 mitogen-activated protein kinases (MAPK) and JNK phosphorylation. EBO suppressed high glucose-induced matrix metalloproteinase (MMP)-9 activity in a dose-dependent manner. In addition, EBO suppressed nuclear factor-kappaB (NF-kappaB) nuclear translocation and transcriptional activity in high glucose conditions. Taken together, the present data suggest that EBO could suppress high glucose-induced atherosclerotic processes through inhibition of p38, JNK, NF-kappaB and MMP signal pathways in HASMC.

  16. Role of Cell-Cell bond for the viability and the function of vascular smooth muscle cells

    Directory of Open Access Journals (Sweden)

    M. Mura

    2010-01-01

    Full Text Available Vascular smooth muscle cell (VSMC viability and homeostasis is regulated by cell-matrix and cell-cell contact: disruption of these interactions are responsible of a switch from a mature to a high proliferative phenotype. VSMCs migration, rate of growth and apoptosis, and the extent of their extracellular matrix (ECM deposition can be also modulated by proatherogenic peptides. Among them, ATII induces the transactivation of IGF I R, which, together with the binding protein IGFBP3, represents a determinant of cell survival, growth and proliferation. Aim of our in vitro study was to verify the role of elective cell-cell bond in moulating the response to ATII. Thus, we evaluated viability, proliferation, IGFIR, IGFBP3 expression and the long term survival and production of ECM in a provisional tissue. A7r5 cell-line was used in adherent cultures or incubated in agarose-coated culture plates to inhibit cell-matrix interactions. Cells, treated or not with ATII 100 nM, were evaluated for apoptosis rate, cell cycle, IGFIR and IGFBP3 protei expression. Fibrin provisional tissue was developed polymerizing a fibrin solution. cantaining A7r5 cells with thrombin. Histological stainings for ECM components were performed on sections of prvisional tissue. An exclusive cell-cell contact resulted to monolayer cell cultures. ATII did not affect the cell survival in both culture conditions, but promoted a 10% decrease in "S" phase and an increases IGFIR expression only in adherent cells. while suspended cell aggregates were resistant to ATII administration; IGFBP3 was reduced both in ATII treated adherent cells and in floating clustered cells, irrespective of the treatmentn. VSMC conditioning in agarose-coated plates before seeding in fibrin provisional matrix reduced, but not abolished, the cell ability to colonize the clot and to produce ECM. This study demonstrates that the elective cell-cell contact induces a quiescent status in cells lacking of cell

  17. Antibodies against AT1 receptors are associated with vascular endothelial and smooth muscle function impairment: protective effects of hydroxysafflor yellow A.

    Directory of Open Access Journals (Sweden)

    Zhu Jin

    Full Text Available Ample evidence has shown that autoantibodies against AT1 receptors (AT1-AA are closely associated with human cardiovascular disease. The aim of this study was to investigate mechanisms underlying AT1-AA-induced vascular structural and functional impairments in the formation of hypertension, and explore ways for preventive treatment. We used synthetic peptide corresponding to the sequence of the second extracellular loop of the AT1 receptor (165-191 to immunize rats and establish an active immunization model. Part of the model received preventive therapy by losartan (20 mg/kg/day and hyroxysafflor yellow A (HSYA (10 mg/kg/day. The result show that systolic blood pressure (SBP and heart rate (HR of immunized rats was significantly higher, and closely correlated with the plasma AT1-Ab titer. The systolic response of thoracic aortic was increased, but diastolic effects were attenuated markedly. Histological observation showed that the thoracic aortic endothelium of the immunized rats became thinner or ruptured, inflammatory cell infiltration, medial smooth muscle cell proliferation and migration, the vascular wall became thicker. There was no significant difference in serum antibody titer between losartan and HSYA groups and the immunized group. The vascular structure and function were reversed, and plasma biochemical parameters were also improved significantly in the two treatment groups. These results suggest that AT1-Ab could induce injury to vascular endothelial cells, and proliferation of smooth muscle cells. These changes were involved in the formation of hypertension. Treatment with AT1 receptor antagonists and anti oxidative therapy could block the pathogenic effect of AT1-Ab on vascular endothelial and smooth muscle cells.

  18. Angiotensin II modulates interleukin-1β-induced inflammatory gene expression in vascular smooth muscle cells via interfering with ERK-NF-κB crosstalk

    International Nuclear Information System (INIS)

    Xu, Shanqin; Zhi, Hui; Hou, Xiuyun; Jiang, Bingbing

    2011-01-01

    Highlights: → We examine how angiotensin II modulates ERK-NF-κB crosstalk and gene expression. → Angiotensin II suppresses IL-1β-induced prolonged ERK and NF-κB activation. → ERK-RSK1 signaling is required for IL-1β-induced prolonged NF-κB activation. → Angiotensin II modulates NF-κB responsive genes via regulating ERK-NF-κB crosstalk. → ERK-NF-κB crosstalk is a novel mechanism regulating inflammatory gene expression. -- Abstract: Angiotensin II is implicated in cardiovascular diseases, which is associated with a role in increasing vascular inflammation. The present study investigated how angiotensin II modulates vascular inflammatory signaling and expression of inducible nitric oxide synthase (iNOS) and vascular cell adhesion molecule (VCAM)-1. In cultured rat aortic vascular smooth muscle cells (VSMCs), angiotensin II suppressed interleukin-1β-induced prolonged phosphorylation of extracellular signal-regulated kinase (ERK) and ribosomal S6 kinase (RSK)-1, and nuclear translocation of nuclear factor (NF)-κB, leading to decreased iNOS but enhanced VCAM-1 expression, associated with an up-regulation of mitogen-activated protein kinase phosphatase-1 expression. Knock-down of RSK1 selectively down regulated interleukin-1β-induced iNOS expression without influencing VCAM-1 expression. In vivo experiments showed that interleukin-1β, iNOS, and VCAM-1 expression were detectable in the aortic arches of both wild-type and apolipoprotein E-deficient (ApoE -/- ) mice. VCAM-1 and iNOS expression were higher in ApoE -/- than in wild type mouse aortic arches. Angiotensin II infusion (3.2 mg/kg/day, for 6 days, via subcutaneous osmotic pump) in ApoE -/- mice enhanced endothelial and adventitial VCAM-1 and iNOS expression, but reduced medial smooth muscle iNOS expression associated with reduced phosphorylation of ERK and RSK-1. These results indicate that angiotensin II can differentially modulate inflammatory gene expression in aortic smooth muscle cells

  19. Effect of gamma rays on electrically evoked contractions of non-vascular smooth muscles (rat vas deferens)

    International Nuclear Information System (INIS)

    Azroony, R.; Ksies, F.; Alya, G.

    2002-10-01

    We have tried, in this experiment, to study the modifications of non-vascular smooth muscles contraction induced via gamma rays. Smooth muscular fibers were isolated from the vas deferens of an adult rat and contractions were electrically evoked. Our results show that irradiation activates the VOC (Voltage Operated Channel) type of ionic channels which causes an increasing in the inward flux of Ca 2+ and then causes an increasing in the inner calcium concentration [Ca 2] i, the matter which means an increasing in the force of muscular contraction. Concerning to the response of vas deferens smooth muscles to the activation of membrane receptors, we have tried to study the effects of gamma rays on activating adrenergic and cholinergic receptors, also, we have tried to show the effects of different doses of gamma rays (1, 3, 5, 7 Gy) on regulating the contractile response of this type of smooth muscles. And results show that: - Irradiation increases contraction force, mediated by adrenergic and cholinergic receptors, in a dose dependent manner, with E m ax 1 Gy m axc 3 Gy m ax 5 Gy m ax 7 Gy. There is an important shift on irradiated rats (3, 5, 7 Gy) where the maximum effect of Acetylcholine (E m ax) can be obtained in lower concentrations of Acetylcholine. These results mean that irradiation activates the inward flux of Ca 2+ through the ROC (Receptors Operated Channels) type of ionic channels, which rely, in their activation, on activating the membrane receptors. By comparing these results with the effects of gamma rays on activating vascular adrenergic and cholinergic receptors, we concluded that: Non-vascular smooth muscles (vas deferens) are less sensitive to irradiation in comparing with vascular smooth muscles (venae portal hepatica), and irradiation increases the sensitivity of cholinergic receptors to acetylcholine in the smooth muscular fibers of vas deferens while; if decreases this sensitivity in the smooth muscular fibers of venae portal hepatica

  20. Guiding the orientation of smooth muscle cells on random and aligned polyurethane/collagen nanofibers.

    Science.gov (United States)

    Jia, Lin; Prabhakaran, Molamma P; Qin, Xiaohong; Ramakrishna, Seeram

    2014-09-01

    Fabricating scaffolds that can simulate the architecture and functionality of native extracellular matrix is a huge challenge in vascular tissue engineering. Various kinds of materials are engineered via nano-technological approaches to meet the current challenges in vascular tissue regeneration. During this study, nanofibers from pure polyurethane and hybrid polyurethane/collagen in two different morphologies (random and aligned) and in three different ratios of polyurethane:collagen (75:25; 50:50; 25:75) are fabricated by electrospinning. The fiber diameters of the nanofibrous scaffolds are in the range of 174-453 nm and 145-419 for random and aligned fibers, respectively, where they closely mimic the nanoscale dimensions of native extracellular matrix. The aligned polyurethane/collagen nanofibers expressed anisotropic wettability with mechanical properties which is suitable for regeneration of the artery. After 12 days of human aortic smooth muscle cells culture on different scaffolds, the proliferation of smooth muscle cells on hybrid polyurethane/collagen (3:1) nanofibers was 173% and 212% higher than on pure polyurethane scaffolds for random and aligned scaffolds, respectively. The results of cell morphology and protein staining showed that the aligned polyurethane/collagen (3:1) scaffold promote smooth muscle cells alignment through contact guidance, while the random polyurethane/collagen (3:1) also guided cell orientation most probably due to the inherent biochemical composition. Our studies demonstrate the potential of aligned and random polyurethane/collagen (3:1) as promising substrates for vascular tissue regeneration. © The Author(s) 2014 Reprints and permissions: sagepub.co.uk/journalsPermissions.nav.

  1. Static pressure accelerates ox-LDL-induced cholesterol accumulation via SREBP-1-mediated caveolin-1 downregulation in cultured vascular smooth muscle cells

    International Nuclear Information System (INIS)

    Luo, Di-xian; Xia, Cheng-lai; Li, Jun-mu; Xiong, Yan; Yuan, Hao-yu; TANG, Zhen-Wang; Zeng, Yixin; Liao, Duan-fang

    2010-01-01

    Research highlights: → Vertical static pressure accelerates ox-LDL-induced cholesterol accumulation in cultured vascular smooth muscle cells. → Static pressure induces SREBP-1 activation. → Static pressure downregulates the expressions of caveolin-1 by activating SREBP-1. → Static pressure also downregulates the transcription of ABCA1 by activating SREBP-1. → Static pressure increases ox-LDL-induced cholesterol accumulation by SREBP-1-mediated caveolin-1 downregulation in vascular smooth muscle cells cultured in vitro. -- Abstract: Objective: To investigate the effect of static pressure on cholesterol accumulation in vascular smooth muscle cells (VSMCs) and its mechanism. Methods: Rat-derived VSMC cell line A10 treated with 50 mg/L ox-LDL and different static pressures (0, 60, 90, 120, 150, 180 mm Hg) in a custom-made pressure incubator for 48 h. Intracellular lipid droplets and lipid levels were assayed by oil red O staining and HPLC; The mRNA levels of caveolin-1 and ABCA1, the protein levels of caveolin-1 SREBP-1 and mature SREBP-1 were respectively detected by RT-PCR or western blot. ALLN, an inhibitor of SREBP metabolism, was used to elevate SREBP-1 protein level in VSMCs treated with static pressure. Results: Static pressures significantly not only increase intracellular lipid droplets in VSMCs, but also elevate cellular lipid content in a pressure-dependent manner. Intracellular free cholesterol (FC), cholesterol ester (CE), total cholesterol (TC) were respectively increased from 60.5 ± 2.8 mg/g, 31.8 ± 0.7 mg/g, 92.3 ± 2.1 mg/g at atmosphere pressure (ATM, 0 mm Hg) to 150.8 ± 9.4 mg/g, 235.9 ± 3.0 mg/g, 386.7 ± 6.4 mg/g at 180 mm Hg. At the same time, static pressures decrease the mRNA and protein levels of caveolin-1, and induce the activation and nuclear translocation of SREBP-1. ALLN increases the protein level of mature SREBP-1 and decreases caveolin-1 expression, so that cellular lipid levels were upregulated. Conclusion: Static

  2. Static pressure accelerates ox-LDL-induced cholesterol accumulation via SREBP-1-mediated caveolin-1 downregulation in cultured vascular smooth muscle cells

    Energy Technology Data Exchange (ETDEWEB)

    Luo, Di-xian, E-mail: luodixian_2@163.com [Department of Pharmacology, School of Pharmaceutics, Central South University, Changsha 410083, Hunan (China); Institute of Pharmacy and Pharmacology, College of Science and Technology, University of South China, Hengyang 421001, Hunan (China); First People' s Hospital of Chenzhou City, Chenzhou 423000, Hunan (China); Xia, Cheng-lai [Institute of Pharmacy and Pharmacology, College of Science and Technology, University of South China, Hengyang 421001, Hunan (China); Department of Pharmacy, Third Affiliated Hospital Medical College of Guangzhou, Guangzhou 510150, Guangdong (China); Li, Jun-mu [Institute of Pharmacy and Pharmacology, College of Science and Technology, University of South China, Hengyang 421001, Hunan (China); Xiong, Yan [Department of Pharmacology, School of Pharmaceutics, Central South University, Changsha 410083, Hunan (China); Yuan, Hao-yu [Institute of Pharmacy and Pharmacology, College of Science and Technology, University of South China, Hengyang 421001, Hunan (China); Lusong Center for Disease Control and Prevention, Zhuzhou 412000, Hunan (China); TANG, Zhen-Wang; Zeng, Yixin [Institute of Pharmacy and Pharmacology, College of Science and Technology, University of South China, Hengyang 421001, Hunan (China); Liao, Duan-fang, E-mail: dfliao66@yahoo.com.cn [Institute of Pharmacy and Pharmacology, College of Science and Technology, University of South China, Hengyang 421001, Hunan (China); Department of Traditional Chinese Diagnostics, School of Pharmacy, Hunan University of Chinese Medicine, Changsha 420108, Hunan (China)

    2010-12-03

    Research highlights: {yields} Vertical static pressure accelerates ox-LDL-induced cholesterol accumulation in cultured vascular smooth muscle cells. {yields} Static pressure induces SREBP-1 activation. {yields} Static pressure downregulates the expressions of caveolin-1 by activating SREBP-1. {yields} Static pressure also downregulates the transcription of ABCA1 by activating SREBP-1. {yields} Static pressure increases ox-LDL-induced cholesterol accumulation by SREBP-1-mediated caveolin-1 downregulation in vascular smooth muscle cells cultured in vitro. -- Abstract: Objective: To investigate the effect of static pressure on cholesterol accumulation in vascular smooth muscle cells (VSMCs) and its mechanism. Methods: Rat-derived VSMC cell line A10 treated with 50 mg/L ox-LDL and different static pressures (0, 60, 90, 120, 150, 180 mm Hg) in a custom-made pressure incubator for 48 h. Intracellular lipid droplets and lipid levels were assayed by oil red O staining and HPLC; The mRNA levels of caveolin-1 and ABCA1, the protein levels of caveolin-1 SREBP-1 and mature SREBP-1 were respectively detected by RT-PCR or western blot. ALLN, an inhibitor of SREBP metabolism, was used to elevate SREBP-1 protein level in VSMCs treated with static pressure. Results: Static pressures significantly not only increase intracellular lipid droplets in VSMCs, but also elevate cellular lipid content in a pressure-dependent manner. Intracellular free cholesterol (FC), cholesterol ester (CE), total cholesterol (TC) were respectively increased from 60.5 {+-} 2.8 mg/g, 31.8 {+-} 0.7 mg/g, 92.3 {+-} 2.1 mg/g at atmosphere pressure (ATM, 0 mm Hg) to 150.8 {+-} 9.4 mg/g, 235.9 {+-} 3.0 mg/g, 386.7 {+-} 6.4 mg/g at 180 mm Hg. At the same time, static pressures decrease the mRNA and protein levels of caveolin-1, and induce the activation and nuclear translocation of SREBP-1. ALLN increases the protein level of mature SREBP-1 and decreases caveolin-1 expression, so that cellular lipid levels were

  3. Intracellular Na+ regulation of Na+ pump sites in cultured vascular smooth muscle cells

    International Nuclear Information System (INIS)

    Allen, J.C.; Navran, S.S.; Seidel, C.L.; Dennison, D.K.; Amann, J.M.; Jemelka, S.K.

    1989-01-01

    Enzymatically dispersed cells from canine saphenous vein and femoral artery were grown in fetal calf serum and studied at day 0 (freshly dispersed) through confluence in primary culture. Intracellular Na levels (Nai), but not intracellular K (Ki), were increased after 24 h in culture and then decreased to a steady state by 4 days. Na+ pump site number [( 3 H] ouabain binding) increased through day 3 and remained elevated. Nai was still elevated at 2 days when the Na+ pump site number began to increase. Total pump turnover (maximum ouabain-inhibited 86 Rb uptake) reflected the increase in Na+ pump site number. These key events precede the observed increases in both protein production and cellular proliferation. If the same cells are maintained in defined medium, without fetal calf serum, Nai, Ki, and the number of [ 3 H]ouabain binding sites do not change with time. These data are consistent with the suggestion that the initial mitogenic response of vascular smooth muscle cells to fetal calf serum involves an increased Na+ influx, and a Nai accumulation, caused by low Na+ pump density. The synthesis of new pump sites effects a decrease in the accumulated Nai, which may be related to cell proliferation

  4. Epigenetic regulation of vascular smooth muscle cell proliferation and neointima formation by histone deacetylase inhibition.

    Science.gov (United States)

    Findeisen, Hannes M; Gizard, Florence; Zhao, Yue; Qing, Hua; Heywood, Elizabeth B; Jones, Karrie L; Cohn, Dianne; Bruemmer, Dennis

    2011-04-01

    Proliferation of smooth muscle cells (SMC) in response to vascular injury is central to neointimal vascular remodeling. There is accumulating evidence that histone acetylation constitutes a major epigenetic modification for the transcriptional control of proliferative gene expression; however, the physiological role of histone acetylation for proliferative vascular disease remains elusive. In the present study, we investigated the role of histone deacetylase (HDAC) inhibition in SMC proliferation and neointimal remodeling. We demonstrate that mitogens induce transcription of HDAC 1, 2, and 3 in SMC. Short interfering RNA-mediated knockdown of either HDAC 1, 2, or 3 and pharmacological inhibition of HDAC prevented mitogen-induced SMC proliferation. The mechanisms underlying this reduction of SMC proliferation by HDAC inhibition involve a growth arrest in the G(1) phase of the cell cycle that is due to an inhibition of retinoblastoma protein phosphorylation. HDAC inhibition resulted in a transcriptional and posttranscriptional regulation of the cyclin-dependent kinase inhibitors p21(Cip1) and p27(Kip). Furthermore, HDAC inhibition repressed mitogen-induced cyclin D1 mRNA expression and cyclin D1 promoter activity. As a result of this differential cell cycle-regulatory gene expression by HDAC inhibition, the retinoblastoma protein retains a transcriptional repression of its downstream target genes required for S phase entry. Finally, we provide evidence that these observations are applicable in vivo by demonstrating that HDAC inhibition decreased neointima formation and expression of cyclin D1 in a murine model of vascular injury. These findings identify HDAC as a critical component of a transcriptional cascade regulating SMC proliferation and suggest that HDAC might play a pivotal role in the development of proliferative vascular diseases, including atherosclerosis and in-stent restenosis.

  5. Specialized mouse embryonic stem cells for studying vascular development.

    Science.gov (United States)

    Glaser, Drew E; Burns, Andrew B; Hatano, Rachel; Medrzycki, Magdalena; Fan, Yuhong; McCloskey, Kara E

    2014-01-01

    Vascular progenitor cells are desirable in a variety of therapeutic strategies; however, the lineage commitment of endothelial and smooth muscle cell from a common progenitor is not well-understood. Here, we report the generation of the first dual reporter mouse embryonic stem cell (mESC) lines designed to facilitate the study of vascular endothelial and smooth muscle development in vitro. These mESC lines express green fluorescent protein (GFP) under the endothelial promoter, Tie-2, and Discomsoma sp. red fluorescent protein (RFP) under the promoter for alpha-smooth muscle actin (α-SMA). The lines were then characterized for morphology, marker expression, and pluripotency. The mESC colonies were found to exhibit dome-shaped morphology, alkaline phosphotase activity, as well as expression of Oct 3/4 and stage-specific embryonic antigen-1. The mESC colonies were also found to display normal karyotypes and are able to generate cells from all three germ layers, verifying pluripotency. Tissue staining confirmed the coexpression of VE (vascular endothelial)-cadherin with the Tie-2 GFP+ expression on endothelial structures and smooth muscle myosin heavy chain with the α-SMA RFP+ smooth muscle cells. Lastly, it was verified that the developing mESC do express Tie-2 GFP+ and α-SMA RFP+ cells during differentiation and that the GFP+ cells colocalize with the vascular-like structures surrounded by α-SMA-RFP cells. These dual reporter vascular-specific mESC permit visualization and cell tracking of individual endothelial and smooth muscle cells over time and in multiple dimensions, a powerful new tool for studying vascular development in real time.

  6. Inhibition of MAPK and PKC pathways by 60Co γ-radiation in cultured vascular smooth muscle cells

    International Nuclear Information System (INIS)

    Jia Guanghong; Ma Yexin; Xiao Jianming

    2002-01-01

    Objective: To investigate the signal transduction pathways inhibited by 60 Co γ-radiation in cultured vascular smooth muscle cells (VSMC). Methods: The cultured VSMC were irradiated with 60 Co γ-radiation of 3.5, 7.0 and 14 Gy respectively. VSMC proliferation was measured by 3 H-TdR incorporation, while PKC, MAPK activities were determined by radioactivity assay. Results: Proliferation of VSMC was inhibited by 7.0, 14 Gy 60 Co γ-irradiation and the activities of PKC, MAPK were decreased significantly. Conclusion: Inhibitory effect of 7.0, 14 Gy 60 Co γ-irradiation on proliferation of VSMC might be resulted from decrease of the activity of PKC, MAPK

  7. Potential role of insulin signaling on vascular smooth muscle cell migration, proliferation, and inflammation pathways.

    Science.gov (United States)

    Cersosimo, Eugenio; Xu, Xiaojing; Musi, Nicolas

    2012-02-15

    To investigate the role of insulin signaling pathways in migration, proliferation, and inflammation of vascular smooth muscle cells (VSMCs), we examined the expression of active components of the phosphatidyl inositol 3 (PI-3) kinase (p-Akt) and mitogen-activated protein kinase (MAPK) (p-Erk) in primary cultures of VSMCs from human coronary arteries. VSMCs were treated in a dose-response manner with insulin (0, 1, 10, and 100 nM) for 20 min, and Akt and Erk phosphorylation were measured by Western blot analysis. In separate experiments, we evaluated the effect of 200 μM palmitate, in the presence and absence of 8 μM pioglitazone, on insulin-stimulated (100 nM for 20 min) Akt and Erk phosphorylation. The phosphorylation of Akt and Erk in VSMCs exhibited a dose dependency with a three- to fourfold increase, respectively, at the highest dose (100 nM). In the presence of palmitate, insulin-induced Akt phosphorylation was completely abolished, and there was a threefold increase in p-Erk. With addition of pioglitazone, the phosphorylation of Akt by insulin remained unchanged, whereas insulin-stimulated Erk phosphorylation was reduced by pioglitazone. These data in VSMCs indicate that high palmitate decreases insulin-stimulated Akt phosphorylation and stimulates MAPK, whereas preexposure peroxisome proliferator-activated receptor-γ agonist pioglitazone preserves Akt phosphorylation and simultaneously attenuates MAPK signaling. Our results suggest that metabolic and mitogenic insulin signals have different sensitivity, are independently regulated, and may play a role in arterial smooth muscle cells migration, proliferation, and inflammation in conditions of acute hyperinsulinemia.

  8. Stimulation of aortic smooth muscle cell mitogenesis by serotonin

    International Nuclear Information System (INIS)

    Nemecek, G.M.; Coughlin, S.R.; Handley, D.A.; Moskowitz, M.A.

    1986-01-01

    Bovine aortic smooth muscle cells in vitro responded to 1 nM to 10 μM serotonin with increased incorporation of [ 3 H]thymidine into DNA. The mitogenic effect of serotonin was half-maximal at 80 nM and maximal above 1 μM. At a concentration of 1 μM, serotonin stimulated smooth muscle cell mitogenesis to the same extent as human platelet-derived growth factor (PDGF) at 12 ng/ml. Tryptamine was ≅ 1/10th as potent as serotonin as a mitogen for smooth muscle cells. Other indoles that are structurally related to serotonin (D- and L-tryptophan, 5-hydroxy-L-tryptophan, N-acetyl-5-hydroxytryptamine, melatonin, 5-hydroxyindoleacetic acid, and 5-hydroxytryptophol) and quipazine were inactive. The stimulatory effect of serotonin on smooth muscle cell DNA synthesis required prolonged (20-24 hr) exposure to the agonist and was attenuated in the presence of serotonin D receptor antagonists. When smooth muscle cells were incubated with submaximal concentrations of serotonin and PDGF, synergistic rather than additive mitogenic responses were observed. These data indicate that serotonin has a significant mitogenic effect on smooth muscle cells in vitro, which appears to be mediated by specific plasma membrane receptors

  9. Effects of One Resistance Exercise Session on Vascular Smooth Muscle of Hypertensive Rats

    International Nuclear Information System (INIS)

    Silva, Tharciano Luiz Teixeira Braga da; Mota, Marcelo Mendonça; Fontes, Milene Tavares; Araújo, João Eliakim dos Santos; Carvalho, Vitor Oliveira; Bonjardim, Leonardo Rigoldi; Santos, Márcio Roberto Viana

    2015-01-01

    Hypertension is a public health problem and increases the incidence of cardiovascular diseases. To evaluate the effects of a resistance exercise session on the contractile and relaxing mechanisms of vascular smooth muscle in mesenteric arteries of N G -nitro L-arginine methyl ester (L-NAME)-induced hypertensive rats. Wistar rats were divided into three groups: control (C), hypertensive (H), and exercised hypertensive (EH). Hypertension was induced by administration of 20 mg/kg of L-NAME for 7 days prior to experimental protocols. The resistance exercise protocol consisted of 10 sets of 10 repetitions and intensity of 40% of one repetition maximum. The reactivity of vascular smooth muscle was evaluated by concentration‑response curves to phenylephrine (PHEN), potassium chloride (KCl) and sodium nitroprusside (SNP). Rats treated with L-NAME showed an increase (p < 0.001) in systolic blood pressure (SBP), diastolic blood pressure (DBP) and mean arterial pressure (MAP) compared to the initial period of induction. No difference in PHEN sensitivity was observed between groups H and EH. Acute resistance exercise reduced (p < 0.001) the contractile response induced by KCl at concentrations of 40 and 60 mM in group EH. Greater (p < 0.01) smooth muscle sensitivity to NPS was observed in group EH as compared to group H. One resistance exercise session reduces the contractile response induced by KCl in addition to increasing the sensitivity of smooth muscle to NO in mesenteric arteries of hypertensive rats

  10. Effects of One Resistance Exercise Session on Vascular Smooth Muscle of Hypertensive Rats

    Energy Technology Data Exchange (ETDEWEB)

    Silva, Tharciano Luiz Teixeira Braga da; Mota, Marcelo Mendonça; Fontes, Milene Tavares; Araújo, João Eliakim dos Santos; Carvalho, Vitor Oliveira; Bonjardim, Leonardo Rigoldi; Santos, Márcio Roberto Viana, E-mail: marciorvsantos@bol.com.br [Universidade Federal de Sergipe, Universidade de São Paulo (Brazil)

    2015-08-15

    Hypertension is a public health problem and increases the incidence of cardiovascular diseases. To evaluate the effects of a resistance exercise session on the contractile and relaxing mechanisms of vascular smooth muscle in mesenteric arteries of N{sup G}-nitro L-arginine methyl ester (L-NAME)-induced hypertensive rats. Wistar rats were divided into three groups: control (C), hypertensive (H), and exercised hypertensive (EH). Hypertension was induced by administration of 20 mg/kg of L-NAME for 7 days prior to experimental protocols. The resistance exercise protocol consisted of 10 sets of 10 repetitions and intensity of 40% of one repetition maximum. The reactivity of vascular smooth muscle was evaluated by concentration‑response curves to phenylephrine (PHEN), potassium chloride (KCl) and sodium nitroprusside (SNP). Rats treated with L-NAME showed an increase (p < 0.001) in systolic blood pressure (SBP), diastolic blood pressure (DBP) and mean arterial pressure (MAP) compared to the initial period of induction. No difference in PHEN sensitivity was observed between groups H and EH. Acute resistance exercise reduced (p < 0.001) the contractile response induced by KCl at concentrations of 40 and 60 mM in group EH. Greater (p < 0.01) smooth muscle sensitivity to NPS was observed in group EH as compared to group H. One resistance exercise session reduces the contractile response induced by KCl in addition to increasing the sensitivity of smooth muscle to NO in mesenteric arteries of hypertensive rats.

  11. Effects of One Resistance Exercise Session on Vascular Smooth Muscle of Hypertensive Rats

    Directory of Open Access Journals (Sweden)

    Tharciano Luiz Teixeira Braga da Silva

    2015-01-01

    Full Text Available Abstract Background: Hypertension is a public health problem and increases the incidence of cardiovascular diseases. Objective: To evaluate the effects of a resistance exercise session on the contractile and relaxing mechanisms of vascular smooth muscle in mesenteric arteries of NG-nitro L-arginine methyl ester (L-NAME-induced hypertensive rats. Methods: Wistar rats were divided into three groups: control (C, hypertensive (H, and exercised hypertensive (EH. Hypertension was induced by administration of 20 mg/kg of L-NAME for 7 days prior to experimental protocols. The resistance exercise protocol consisted of 10 sets of 10 repetitions and intensity of 40% of one repetition maximum. The reactivity of vascular smooth muscle was evaluated by concentration‑response curves to phenylephrine (PHEN, potassium chloride (KCl and sodium nitroprusside (SNP. Results: Rats treated with L-NAME showed an increase (p < 0.001 in systolic blood pressure (SBP, diastolic blood pressure (DBP and mean arterial pressure (MAP compared to the initial period of induction. No difference in PHEN sensitivity was observed between groups H and EH. Acute resistance exercise reduced (p < 0.001 the contractile response induced by KCl at concentrations of 40 and 60 mM in group EH. Greater (p < 0.01 smooth muscle sensitivity to NPS was observed in group EH as compared to group H. Conclusion: One resistance exercise session reduces the contractile response induced by KCl in addition to increasing the sensitivity of smooth muscle to NO in mesenteric arteries of hypertensive rats.

  12. RhoA/ROCK signaling regulates smooth muscle phenotypic modulation and vascular remodeling via the JNK pathway and vimentin cytoskeleton.

    Science.gov (United States)

    Tang, Lian; Dai, Fan; Liu, Yan; Yu, Xiaoqiang; Huang, Chao; Wang, Yuqin; Yao, Wenjuan

    2018-05-20

    The RhoA/ROCK signaling pathway regulates cell morphology, adhesion, proliferation, and migration. In this study, we investigated the regulatory role of RhoA/ROCK signaling on PDGF-BB-mediated smooth muscle phenotypic modulation and vascular remodeling and clarified the molecular mechanisms behind these effects. PDGF-BB treatment induced the activation of RhoA, ROCK, PDGF-Rβ, and the expression of PDGF-Rβ in HA-VSMCs (human aortic vascular smooth muscle cells). PDGF-Rβ inhibition and RhoA suppression blocked PDGF-BB-induced RhoA activation and ROCK induction. In addition, PDGF-BB-mediated cell proliferation and migration were suppressed by PDGF-Rβ inhibition, RhoA suppression, and ROCK inhibition, suggesting that PDGF-BB promotes phenotypic modulation of HA-VSMCs by activating the RhoA/ROCK pathway via the PDGF receptor. Moreover, suppressing both ROCK1 and ROCK2 blocked cell cycle progression from G0/G1 to S phase by decreasing the transcription and protein expression of cyclin D1, CDK2, and CDK4 via JNK/c-Jun pathway, thus reducing cell proliferation in PDGF-BB-treated HA-VSMCs. ROCK1 deletion, rather than ROCK2 suppression, significantly inhibited PDGF-BB-induced migration by reducing the expression of vimentin and preventing the remodeling of vimentin and phospho-vimentin. Furthermore, ROCK1 deletion suppressed vimentin by inhibiting the phosphorylation of Smad2/3 and the nuclear translocation of Smad4. These findings suggested that ROCK1 and ROCK2 might play different roles in PDGF-BB-mediated cell proliferation and migration in HA-VSMCs. In addition, PDGF-BB and its receptor participated in neointima formation and vascular remodeling by promoting cell cycle protein expression via the JNK pathway and enhancing vimentin expression in a rat balloon injury model; effects that were inhibited by treatment with fasudil. Together, the results of this study reveal a novel mechanism through which RhoA/ROCK signaling regulates smooth muscle phenotypic modulation and

  13. Isolation of pulmonary artery smooth muscle cells from neonatal mice.

    Science.gov (United States)

    Lee, Keng Jin; Czech, Lyubov; Waypa, Gregory B; Farrow, Kathryn N

    2013-10-19

    Pulmonary hypertension is a significant cause of morbidity and mortality in infants. Historically, there has been significant study of the signaling pathways involved in vascular smooth muscle contraction in PASMC from fetal sheep. While sheep make an excellent model of term pulmonary hypertension, they are very expensive and lack the advantage of genetic manipulation found in mice. Conversely, the inability to isolate PASMC from mice was a significant limitation of that system. Here we described the isolation of primary cultures of mouse PASMC from P7, P14, and P21 mice using a variation of the previously described technique of Marshall et al. that was previously used to isolate rat PASMC. These murine PASMC represent a novel tool for the study of signaling pathways in the neonatal period. Briefly, a slurry of 0.5% (w/v) agarose + 0.5% iron particles in M199 media is infused into the pulmonary vascular bed via the right ventricle (RV). The iron particles are 0.2 μM in diameter and cannot pass through the pulmonary capillary bed. Thus, the iron lodges in the small pulmonary arteries (PA). The lungs are inflated with agarose, removed and dissociated. The iron-containing vessels are pulled down with a magnet. After collagenase (80 U/ml) treatment and further dissociation, the vessels are put into a tissue culture dish in M199 media containing 20% fetal bovine serum (FBS), and antibiotics (M199 complete media) to allow cell migration onto the culture dish. This initial plate of cells is a 50-50 mixture of fibroblasts and PASMC. Thus, the pull down procedure is repeated multiple times to achieve a more pure PASMC population and remove any residual iron. Smooth muscle cell identity is confirmed by immunostaining for smooth muscle myosin and desmin.

  14. Hsc70 regulates cell surface ASIC2 expression and vascular smooth muscle cell migration.

    Science.gov (United States)

    Grifoni, Samira C; McKey, Susan E; Drummond, Heather A

    2008-05-01

    Recent studies suggest members of the degenerin (DEG)/epithelial Na(+) channel (ENaC)/acid-sensing ion channel (ASIC) protein family play an important role in vascular smooth muscle cell (VSMC) migration. In a previous investigation, we found suppression of a certain DEG/ENaC/ASIC member, ASIC2, increased VSMC chemotactic migration, raising the possibility that ASIC2 may play an inhibitory role. Because ASIC2 protein was retained in the cytoplasm, we reasoned increasing surface expression of ASIC2 might unmask the inhibitory role of ASIC2 in VSMC migration so we could test the hypothesis that ASIC2 inhibits VSMC migration. Therefore, we used the chemical chaperone glycerol to enhance ASIC2 expression. Glycerol 1) increased cytoplasm ASIC2 expression, 2) permitted detection of ASIC2 at the cell surface, and 3) inhibited platelet-derived growth factor (PDGF)-bb mediated VSMC migration. Furthermore, ASIC2 silencing completely abolished the inhibitory effect of glycerol on migration, suggesting upregulation of ASIC2 is responsible for glycerol-induced inhibition of VSMC migration. Because other investigators have shown that glycerol regulates ENaC/ASIC via interactions with a certain heat shock protein, heat shock protein 70 (Hsc70), we wanted to determine the importance of Hsc70 on ASIC2 expression in VSMCs. We found that Hsc70 silencing increases ASIC2 cell surface expression and inhibits VSMC migration, which is abolished by cosilencing ASIC2. These data demonstrate that Hsc70 inhibits ASIC2 expression, and, when the inhibitory effect of Hsc70 is removed, ASIC2 expression increases, resulting in reduced VSMC migration. Because VSMC migration contributes to vasculogenesis and remodeling following vascular injury, our findings raise the possibility that ASIC2-Hsc70 interactions may play a role in these processes.

  15. Resveratrol blocks interleukin-18-EMMPRIN cross-regulation and smooth muscle cell migration

    OpenAIRE

    Venkatesan, Balachandar; Valente, Anthony J.; Reddy, Venkatapuram Seenu; Siwik, Deborah A.; Chandrasekar, Bysani

    2009-01-01

    Vascular smooth muscle cell (SMC) migration is an important mechanism in atherogenesis and postangioplasty arterial remodeling. Previously, we demonstrated that the proinflammatory cytokine interleukin (IL)-18 is a potent inducer of SMC migration. Since extracellular matrix metalloproteinase inducer (EMMPRIN) stimulates ECM degradation and facilitates cell migration, we investigated whether IL-18 and EMMPRIN regulate each other's expression, whether their cross talk induces SMC migration, and...

  16. Selective Expression of an Endogenous Inhibitor of FAK Regulates Proliferation and Migration of Vascular Smooth Muscle Cells

    Science.gov (United States)

    Taylor, Joan M.; Mack, Christopher P.; Nolan, Kate; Regan, Christopher P.; Owens, Gary K.; Parsons, J. Thomas

    2001-01-01

    Extracellular matrix signaling via integrin receptors is important for smooth muscle cell (SMC) differentiation during vasculogenesis and for phenotypic modulation of SMCs during atherosclerosis. We previously reported that the noncatalytic carboxyl-terminal protein binding domain of focal adhesion kinase (FAK) is expressed as a separate protein termed FAK-related nonkinase (FRNK) and that ectopic expression of FRNK can attenuate FAK activity and integrin-dependent signaling (A. Richardson and J. T. Parsons, Nature 380:538–540, 1996). Herein we report that in contrast to FAK, which is expressed ubiquitously, FRNK is expressed selectively in SMCs, with particularly high levels observed in conduit blood vessels. FRNK expression was low during embryonic development, was significantly upregulated in the postnatal period, and returned to low but detectable levels in adult tissues. FRNK expression was also dramatically upregulated following balloon-induced carotid artery injury. In cultured rat aortic smooth muscle cells, overexpression of FRNK attenuated platelet-derived growth factor (PDGF)-BB-induced migration and also dramatically inhibited [3H]thymidine incorporation upon stimulation with PDGF-BB or 10% serum. These effects were concomitant with a reduction in SMC proliferation. Taken together, these data indicate that FRNK acts as an endogenous inhibitor of FAK signaling in SMCs. Furthermore, increased FRNK expression following vascular injury or during development may alter the SMC phenotype by negatively regulating proliferative and migratory signals. PMID:11238893

  17. Cellular function and signaling pathways of vascular smooth muscle cells modulated by sphingosine 1-phosphate

    Directory of Open Access Journals (Sweden)

    Takuji Machida

    2016-12-01

    Full Text Available Sphingosine 1-phosphate (S1P plays important roles in cardiovascular pathophysiology. S1P1 and/or S1P3, rather than S1P2 receptors, seem to be predominantly expressed in vascular endothelial cells, while S1P2 and/or S1P3, rather than S1P1 receptors, seem to be predominantly expressed in vascular smooth muscle cells (VSMCs. S1P has multiple actions, such as proliferation, inhibition or stimulation of migration, and vasoconstriction or release of vasoactive mediators. S1P induces an increase of the intracellular Ca2+ concentration in many cell types, including VSMCs. Activation of S1P3 seems to play an important role in Ca2+ mobilization. S1P induces cyclooxygenase-2 expression in VSMCs via both S1P2 and S1P3 receptors. S1P2 receptor activation in VSMCs inhibits inducible nitric oxide synthase (iNOS expression. At the local site of vascular injury, vasoactive mediators such as prostaglandins and NO produced by VSMCs are considered primarily as a defensive and compensatory mechanism for the lack of endothelial function to prevent further pathology. Therefore, selective S1P2 receptor antagonists may have the potential to be therapeutic agents, in view of their antagonism of iNOS inhibition by S1P. Further progress in studies of the precise mechanisms of S1P may provide useful knowledge for the development of new S1P-related drugs for the treatment of cardiovascular diseases.

  18. New aspects of vascular remodelling: the involvement of all vascular cell types.

    Science.gov (United States)

    McGrath, John C; Deighan, Clare; Briones, Ana M; Shafaroudi, Majid Malekzadeh; McBride, Melissa; Adler, Jeremy; Arribas, Silvia M; Vila, Elisabet; Daly, Craig J

    2005-07-01

    Conventionally, the architecture of arteries is based around the close-packed smooth muscle cells and extracellular matrix. However, the adventitia and endothelium are now viewed as key players in vascular growth and repair. A new dynamic picture has emerged of blood vessels in a constant state of self-maintenance. Recent work raises fundamental questions about the cellular heterogeneity of arteries and the time course and triggering of normal and pathological remodelling. A common denominator emerging in hypertensive remodelling is an early increase in adventitial cell density suggesting that adventitial cells drive remodelling and may initiate subsequent changes such as re-arrangement of smooth muscle cells and extracellular matrix. The organization of vascular smooth muscle cells follows regular arrangements that can be modelled mathematically. In hypertension, new patterns can be quantified in these terms and give insights to how structure affects function. As with smooth muscle, little is known about the organization of the vascular endothelium, or its role in vascular remodelling. Current observations suggest that there may be a close relationship between the helical organization of smooth muscle cells and the underlying pattern of endothelial cells. The function of myoendothelial connections is a topic of great current interest and may relate to the structure of the internal elastic lamina through which the connections must pass. In hypertensive remodelling this must present an organizational challenge. The objective of this paper is to show how the functions of blood vessels depend on their architecture and a continuous interaction of different cell types and extracellular proteins.

  19. PHYSICAL CONTACT BETWEEN HUMAN VASCULAR ENDOTHELIAL AND SMOOTH MUSCLE CELLS MODULATES CYTOSOLIC AND NUCLEAR CALCIUM HOMEOSTASIS.

    Science.gov (United States)

    Hassan, Ghada S; Jacques, Danielle; D'Orleans-Juste, Pedro; Magder, Sheldon; Bkaily, Ghassan

    2018-05-14

    The interaction between vascular endothelial cells (VECs) and vascular smooth muscle cells (VSMCs) plays an important role in the modulation of vascular tone. There is however no information on whether direct physical communication regulates the intracellular calcium levels of human VECs (hVECs) and/or hVSMCs . Thus, the objective of the study is to verify whether co-culture of hVECs and hVSMCs modulates cytosolic ([Ca2+]c) and nuclear calcium ([Ca2+]n) levels via physical contact and/or factors released by both cell types. Quantitative 3D confocal microscopy for [Ca2+]c and [Ca2+]n measurement was performed in cultured hVECs or hVSMCs or in co-culture of hVECs-hVSMCs. Our results show that: 1) physical contact between hVECs-hVECs or hVSMCs-hVSMCs does not affect [Ca2+]c and [Ca2+]n in these two cell types; 2) physical contact between hVECs and hVSMCs induces a significant increase only of [Ca2+]n of hVECs without affecting the level of [Ca2+]c and [Ca2+]n of hVSMCs; and 3) preconditioned culture medium of hVECs or hVSMCs does not affect [Ca2+]c and [Ca2+]n of both types of cells. We concluded that physical contact between hVECs and hVSMCs only modulates [Ca2+]n in hVECs. The increase of [Ca2+]n in hVECs may modulate nuclear functions that are calcium dependent.

  20. Synthetic smooth muscle in the outer blood plexus of the rhinarium skin of Lemur catta L.

    Science.gov (United States)

    Elofsson, Rolf; Kröger, Ronald H H

    2017-01-01

    The skin of the lemur nose tip (rhinarium) has arterioles in the outer vascular plexus that are endowed with an unusual coat of smooth muscle cells. Comparison with the arterioles of the same area in a number of unrelated mammalians shows that the lemur pattern is unique. The vascular smooth muscle cells belong to the synthetic type. The function of synthetic smooth muscles around the terminal vessels in the lemur rhinarium is unclear but may have additional functions beyond regulation of vessel diameter.

  1. Ouabain binding to cultured vascular smooth muscle cells of the spontaneously hypertensive rat

    International Nuclear Information System (INIS)

    Hopp, L.; Khalil, F.; Tamura, H.; Kino, M.; Searle, B.M.; Tokushige, A.; Aviv, A.

    1986-01-01

    The binding of ouabain and K + to the Na + pump were analyzed in serially passed cultured vascular smooth muscle cells (VSMCs) originating from spontaneously hypertensive (SH) Wistar-Kyoto (WKY), and American Wistar (W) rats. The techniques have utilized analyses of displacement of [ 3 H]ouabain by both unlabeled ouabain and K + from specific binding sites on the VSMCs. The authors have found that 1) each of the VSMC preparations from the three rat strains appeared to demonstrate one population of specific ouabain receptors (Na + pumps); 2) the number of Na + pump units of both the SH and WKY rats was significantly lower than the number of Na + pump units of W rat VSMCs; 3) the equilibrium dissociation constant values (μM) for ouabain in VSMCs of SH and WKY rats were similar but were significantly higher than that of VSMCs derived from W rats; and 4) among the VSMCs originating from the three rat strains, the apparent equilibrium dissociation constant value for K + (mM) was the lowest in those of the SH rat compared with VSMCs of the WKY rat and W rat. Previous studies have demonstrated increased passive Na + and K + transport rate constants of SH rat VSMCs compared with either W or WKY rat cells. These findings suggest the possibility of higher permeabilities of the SH cells. They propose that the combined effect of a low number of Na + pump units with higher permeabilities to Na + and K + predisposes VSMCs of the SH rat to disturbances in their cellular ionic regulation. These genetic defects, if they occur in vivo, may lead to an increase in the vascular tone

  2. Effect of hypertensive rat plasma on ion transport of cultured vascular smooth muscle

    International Nuclear Information System (INIS)

    Magargal, W.W.; Overbeck, H.W.

    1986-01-01

    We layered fresh, unprocessed plasma from healthy rats with early (less than or equal to 7 days) or benign, chronic (greater than 3 wk) one-kidney, one-clip hypertension and from paired one-kidney normotensive control rats over confluent primary-cultured rat aortic smooth muscle cells. Plasma from all rats increased cellular ouabain-sensitive 86 Rb + uptake and sodium content and decreased ouabain-insensitive 86 Rb + uptake compared with uptakes and content in the presence of balanced salt solution (P less than 0.01). Cells incubated in the presence of plasma from rats with early (P less than 0.02) or chronic hypertension (P less than 0.01) had significantly reduced ouabain-sensitive 86 Rb + uptake when compared with cells incubated in normotensive plasma, but their intracellular Na+ contents were not lower. We no longer detected this uptake difference when chronic hypertensives drank 0.9% NaCl instead of water. Plasma from hypertensive rats also altered ouabain-insensitive 86 Rb + uptake by the cultured cells. These findings of this new, reproducible, and specific assay system support the hypothesis that plasma factors inhibit the membrane sodium-potassium pump in vascular smooth muscle cells in this form of hypertension. The abnormality occurs in both early and chronic stages, but may not be related to sodium intake. The data also provide evidence for plasma factors in hypertension altering membrane K+ permeability

  3. Differential gene expression profiling of human adipose stem cells differentiating into smooth muscle-like cells by TGFβ1/BMP4

    Energy Technology Data Exchange (ETDEWEB)

    Elçin, Ayşe Eser; Parmaksiz, Mahmut; Dogan, Arin; Seker, Sukran; Durkut, Serap; Dalva, Klara; Elçin, Yaşar Murat, E-mail: elcinmurat@gmail.com

    2017-03-15

    Regenerative repair of the vascular system is challenging from the perspectives of translational medicine and tissue engineering. There are fundamental hurdles in front of creating bioartificial arteries, which involve recaputilation of the three-layered structure under laboratory settings. Obtaining and maintaining smooth muscle characteristics is an important limitation, as the transdifferentiated cells fail to display mature phenotype. This study aims to shed light on the smooth muscle differentiation of human adipose stem cells (hASCs). To this end, we first acquired hASCs from lipoaspirate samples. Upon characterization, the cells were induced to differentiate into smooth muscle (SM)-like cells using a variety of inducer combinations. Among all, TGFβ1/BMP4 combination had the highest differentiation efficiency, based on immunohistochemical analyses. hSM-like cell samples were compared to hASCs and to the positive control, human coronary artery-smooth muscle cells (hCA-SMCs) through gene transcription profiling. Microarray findings revealed the activation of gene groups that function in smooth muscle differentiation, signaling pathways, extracellular modeling and cell proliferation. Our results underline the effectiveness of the growth factors and suggest some potential variables for detecting the SM-like cell characteristics. Evidence in transcriptome level was used to evaluate the TGFβ1/BMP4 combination as a previously unexplored effector for the smooth muscle differentiation of adipose stem cells. - Highlights: • Human adipose stem cells (hASCs) were isolated, characterized and cultured. • Growth factor combinations were evaluated for their effectiveness in differentiation using IHC. • hASCs were differentiated into smooth muscle (SM)-like cells using TGF-β1 and BMP4 combination. • Microarray analysis was performed for hASCs, SM-like cells and coronary artery-SMCs. • Microarray data was used to perform hierarchical clustering and interpretation

  4. Oxygenation decreases elastin secretion from rat ductus arteriosus smooth muscle cells.

    Science.gov (United States)

    Kawakami, Shoji; Minamisawa, Susumu

    2015-08-01

    The ductus arteriosus (DA), a fetal arterial connection between the main pulmonary artery and the descending aorta, normally closes immediately after birth. The oxygen concentration in the blood rises after birth, and in the DA this increase in oxygen concentration causes functional closure, which is induced by smooth muscle contraction. Previous studies have demonstrated that hypoxia and/or oxygenation affect vascular remodeling of various vessels. Therefore, we hypothesized that the rise in oxygen concentration would affect the vascular structure of the DA due to production of proteins secreted from DA smooth muscle cells (SMC). Liquid chromatography-tandem mass spectrometry was used to comprehensively investigate the secreted proteins in the supernatant of rat DA SMC harvested under hypoxic conditions (1% oxygen) or under normoxic conditions (21% oxygen). We found that the rise in oxygen concentration reduced the secretion of elastin from DA SMC. On reverse transcription-polymerase chain reaction, the expression of elastin mRNA was not significantly changed in DA SMC from hypoxic to normoxic conditions. Given that elastin forms internal elastic lamina and elastic fibers in the vascular muscle layers, and that a rise in oxygen concentration reduced the secretion of elastin, this suggests that the rise in blood oxygen concentration after birth reduces the secretion of elastin, and therefore may play a role in DA structural remodeling after birth. © 2015 Japan Pediatric Society.

  5. Inhibition of NF-κB activity in rabbit vascular smooth muscle cells by lovastatin

    International Nuclear Information System (INIS)

    Luan Zhaoxia; Lan Xiaoli

    2003-01-01

    Nuclear factor NF-κB is believed to play an important role in regulating the production of matrix metalloproteinase (MMPs), which induce atherosclerosis, restenosis and plaque rupture. We incubated rabbit vascular smooth muscle cells (RVSMCs) with 5 μmol/L lovastatin in the presence of IL-1-α and PDGF BB (20 μg/L, respectively) to study whether lovastatin inhibited NF-κB binding activity induced by IL-1 and PDGF. The NF-κB activity was detected by electrophoretic mobility shift assay (EMSA); MMP-1 and MMP-3 were measured by western blotting; and MMP-9 was detected by zymography. The result showed that lovastatin strongly reduced NF-κB activity upregulated by IL-1 combined with PDGF, and lovastatin also dose-dependently inhibited the expression of MMP-1, -3 and -9 induced by IL-1 and PDGF. It suggested that the beneficial effects of statins may extend to mechanisms beyond cholesterol reduction

  6. Inhibition of vascular smooth muscle cell proliferation by Gentiana lutea root extracts.

    Directory of Open Access Journals (Sweden)

    Rushendhiran Kesavan

    Full Text Available Gentiana lutea belonging to the Gentianaceae family of flowering plants are routinely used in traditional Serbian medicine for their beneficial gastro-intestinal and anti-inflammatory properties. The aim of the study was to determine whether aqueous root extracts of Gentiana lutea consisting of gentiopicroside, gentisin, bellidifolin-8-O-glucoside, demethylbellidifolin-8-O-glucoside, isovitexin, swertiamarin and amarogentin prevents proliferation of aortic smooth muscle cells in response to PDGF-BB. Cell proliferation and cell cycle analysis were performed based on alamar blue assay and propidium iodide labeling respectively. In primary cultures of rat aortic smooth muscle cells (RASMCs, PDGF-BB (20 ng/ml induced a two-fold increase in cell proliferation which was significantly blocked by the root extract (1 mg/ml. The root extract also prevented the S-phase entry of synchronized cells in response to PDGF. Furthermore, PDGF-BB induced ERK1/2 activation and consequent increase in cellular nitric oxide (NO levels were also blocked by the extract. These effects of extract were due to blockade of PDGF-BB induced expression of iNOS, cyclin D1 and proliferating cell nuclear antigen (PCNA. Docking analysis of the extract components on MEK1, the upstream ERK1/2 activating kinase using AutoDock4, indicated a likely binding of isovitexin to the inhibitor binding site of MEK1. Experiments performed with purified isovitexin demonstrated that it successfully blocks PDGF-induced ERK1/2 activation and proliferation of RASMCs in cell culture. Thus, Gentiana lutea can provide novel candidates for prevention and treatment of atherosclerosis.

  7. Inhibition of vascular smooth muscle cell proliferation by Gentiana lutea root extracts.

    Science.gov (United States)

    Kesavan, Rushendhiran; Potunuru, Uma Rani; Nastasijević, Branislav; T, Avaneesh; Joksić, Gordana; Dixit, Madhulika

    2013-01-01

    Gentiana lutea belonging to the Gentianaceae family of flowering plants are routinely used in traditional Serbian medicine for their beneficial gastro-intestinal and anti-inflammatory properties. The aim of the study was to determine whether aqueous root extracts of Gentiana lutea consisting of gentiopicroside, gentisin, bellidifolin-8-O-glucoside, demethylbellidifolin-8-O-glucoside, isovitexin, swertiamarin and amarogentin prevents proliferation of aortic smooth muscle cells in response to PDGF-BB. Cell proliferation and cell cycle analysis were performed based on alamar blue assay and propidium iodide labeling respectively. In primary cultures of rat aortic smooth muscle cells (RASMCs), PDGF-BB (20 ng/ml) induced a two-fold increase in cell proliferation which was significantly blocked by the root extract (1 mg/ml). The root extract also prevented the S-phase entry of synchronized cells in response to PDGF. Furthermore, PDGF-BB induced ERK1/2 activation and consequent increase in cellular nitric oxide (NO) levels were also blocked by the extract. These effects of extract were due to blockade of PDGF-BB induced expression of iNOS, cyclin D1 and proliferating cell nuclear antigen (PCNA). Docking analysis of the extract components on MEK1, the upstream ERK1/2 activating kinase using AutoDock4, indicated a likely binding of isovitexin to the inhibitor binding site of MEK1. Experiments performed with purified isovitexin demonstrated that it successfully blocks PDGF-induced ERK1/2 activation and proliferation of RASMCs in cell culture. Thus, Gentiana lutea can provide novel candidates for prevention and treatment of atherosclerosis.

  8. Control of Vascular Smooth Muscle Cell Growth by Connexin 43

    Directory of Open Access Journals (Sweden)

    Chintamani eJoshi

    2012-06-01

    Full Text Available Connexin 43 (Cx43, the principal gap junction protein in vascular smooth muscle cells (VSMCs, regulates movement of ions and other signaling molecules through gap junction intercellular communication (GJIC and plays important roles in maintaining normal vessel function; however, many of the signaling mechanisms controlling Cx43 in VSMCs are not clearly described. The goal of this study was to investigate mechanisms of Cx43 regulation with respect to VSMC proliferation. Treatment of rat primary VSMCs with the cAMP analog 8Br-cAMP, the soluble guanylate cyclase (sGC stimulator BAY 41-2272 (BAY, or the Cx inducer diallyl disulfide (DADS significantly reduced proliferation after 72 h compared to vehicle controls. Bromodeoxyuridine uptake revealed reduction (p<.001 in DNA synthesis after 6 h and flow cytometry showed reduced (40% S phase cell numbers after 16 h in DADS-treated cells compared to controls. Cx43 expression significantly increased after 270 min treatment with 8Br-cAMP, 8Br-cGMP, BAY or DADS. Inhibition of PKA, PKG or PKC reversed 8Br-cAMP-stimulated increases in Cx43 expression, whereas only PKG or PKC inhibition reversed 8Br-cGMP- and BAY-stimulated increases in total Cx43. Interestingly, stimulation of Cx43 expression by DADS was not dependent on PKA, PKG or PKC. Using fluorescence recovery after photobleaching, only 8Br-cAMP or DADS increased GJIC with 8Br-cAMP mediated by PKC and DADS mediated by PKG. Further, DADS significantly increased phosphorylation at the MAPK-sensitive serine (Ser255 and Ser279, the cell cycle regulatory kinase-sensitive Ser262 and the PKC-sensitive Ser368 after 30 min while 8Br-cAMP significantly increased phosphorylation only at Ser279 compared to controls. This study demonstrates that 8Br-cAMP- and DADS-enhanced GJIC rather than Cx43 expression and/or phosphorylation plays an important role in regulation of VSMC proliferation and provides new insights into the growth-regulatory capacities of Cx43 in VSMCs.

  9. The Complex Role of Store Operated Calcium Entry Pathways and Related Proteins in the Function of Cardiac, Skeletal and Vascular Smooth Muscle Cells

    Directory of Open Access Journals (Sweden)

    Javier Avila-Medina

    2018-03-01

    Full Text Available Cardiac, skeletal, and smooth muscle cells shared the common feature of contraction in response to different stimuli. Agonist-induced muscle's contraction is triggered by a cytosolic free Ca2+ concentration increase due to a rapid Ca2+ release from intracellular stores and a transmembrane Ca2+ influx, mainly through L-type Ca2+ channels. Compelling evidences have demonstrated that Ca2+ might also enter through other cationic channels such as Store-Operated Ca2+ Channels (SOCCs, involved in several physiological functions and pathological conditions. The opening of SOCCs is regulated by the filling state of the intracellular Ca2+ store, the sarcoplasmic reticulum, which communicates to the plasma membrane channels through the Stromal Interaction Molecule 1/2 (STIM1/2 protein. In muscle cells, SOCCs can be mainly non-selective cation channels formed by Orai1 and other members of the Transient Receptor Potential-Canonical (TRPC channels family, as well as highly selective Ca2+ Release-Activated Ca2+ (CRAC channels, formed exclusively by subunits of Orai proteins likely organized in macromolecular complexes. This review summarizes the current knowledge of the complex role of Store Operated Calcium Entry (SOCE pathways and related proteins in the function of cardiac, skeletal, and vascular smooth muscle cells.

  10. Proliferation and extracellular matrix synthesis of smooth muscle cells cultured from human coronary atherosclerotic and restenotic lesions

    NARCIS (Netherlands)

    D.C. MacLeod (Donald); B.H. Strauss (Bradley); J. Escaned (Javier); V.A.W.M. Umans (Victor); R-J. van Suylen (Robert-Jan); A. Verkerk (Anton); P.J. de Feyter (Pim); P.W.J.C. Serruys (Patrick); M. de Jong (Marcel)

    1994-01-01

    textabstractOBJECTIVES. The purpose of this study was to examine the proliferative capacity and extracellular matrix synthesis of human coronary plaque cells in vitro. BACKGROUND. Common to both primary atherosclerosis and restenosis are vascular smooth muscle cell proliferation and production of

  11. The mechanism of inhibitory effect of γ-ray irradiation on rat vascular smooth muscle cells

    International Nuclear Information System (INIS)

    Zhuang Yongzhi; Wang Junjie; Zhang Zhanchun; Jia Tingzhen

    2001-01-01

    Objective: To investigate the inhibitory effect of γ-ray irradiation on rat vascular smooth muscle cells (VSMCs). Methods: Dose-survival curve of VSMCs was figured by colony formation. The effect of γ-ray irradiation on viability and proliferation of VSMCs was observed by 3 H incorporation. Flow cytometry and DNA Ladder were used to detect the apoptosis effect of γ-ray irradiation on VSMCs. Results: The values of D 0 , D q , D 37 and N for VSMCs were 1.95 Gy, 1.76 Gy, 3.71 Gy and 2.47, respectively. The inhibitory effect of γ-ray irradiation on VSMCs proliferation was dose-dependent, being stronger along with increase of dose. VSMCs did not undergo apoptosis within 48 hours after γ-ray irradiation. Conclusion: γ-ray irradiation could inhibit the proliferation of VSMCs, the main mechanism of which is the killing effect and inhibition of mitosis of VSMCs

  12. Compressive elasticity of three-dimensional nanofiber matrix directs mesenchymal stem cell differentiation to vascular cells with endothelial or smooth muscle cell markers.

    Science.gov (United States)

    Wingate, K; Bonani, W; Tan, Y; Bryant, S J; Tan, W

    2012-04-01

    The importance of mesenchymal stem cells (MSC) in vascular regeneration is becoming increasingly recognized. However, few in vitro studies have been performed to identify the effects of environmental elasticity on the differentiation of MSC into vascular cell types. Electrospinning and photopolymerization techniques were used to fabricate a three-dimensional (3-D) polyethylene glycol dimethacrylate nanofiber hydrogel matrix with tunable elasticity for use as a cellular substrate. Compression testing demonstrated that the elastic modulus of the hydrated 3-D matrices ranged from 2 to 15 kPa, similar to the in vivo elasticity of the intima basement membrane and media layer. MSC seeded on rigid matrices (8-15 kPa) showed an increase in cell area compared with those seeded on soft matrices (2-5 kPa). Furthermore, the matrix elasticity guided the cells to express different vascular-specific phenotypes with high differentiation efficiency. Around 95% of MSC seeded on the 3-D matrices with an elasticity of 3 kPa showed Flk-1 endothelial markers within 24h, while only 20% of MSC seeded on the matrices with elasticity >8 kPa demonstrated Flk-1 marker. In contrast, ∼80% of MSC seeded on 3-D matrices with elasticity >8 kPa demonstrated smooth muscle α-actin marker within 24h, while fewer than 10% of MSC seeded on 3-D matrices with elasticity elasticity of the substrate could be a powerful tool for vascular tissue regeneration. Copyright © 2012 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

  13. Bioenergetic profile of human coronary artery smooth muscle cells and effect of metabolic intervention.

    Directory of Open Access Journals (Sweden)

    Mingming Yang

    Full Text Available Bioenergetics of artery smooth muscle cells is critical in cardiovascular health and disease. An acute rise in metabolic demand causes vasodilation in systemic circulation while a chronic shift in bioenergetic profile may lead to vascular diseases. A decrease in intracellular ATP level may trigger physiological responses while dedifferentiation of contractile smooth muscle cells to a proliferative and migratory phenotype is often observed during pathological processes. Although it is now possible to dissect multiple building blocks of bioenergetic components quantitatively, detailed cellular bioenergetics of artery smooth muscle cells is still largely unknown. Thus, we profiled cellular bioenergetics of human coronary artery smooth muscle cells and effects of metabolic intervention. Mitochondria and glycolysis stress tests utilizing Seahorse technology revealed that mitochondrial oxidative phosphorylation accounted for 54.5% of ATP production at rest with the remaining 45.5% due to glycolysis. Stress tests also showed that oxidative phosphorylation and glycolysis can increase to a maximum of 3.5 fold and 1.25 fold, respectively, indicating that the former has a high reserve capacity. Analysis of bioenergetic profile indicated that aging cells have lower resting oxidative phosphorylation and reduced reserve capacity. Intracellular ATP level of a single cell was estimated to be over 1.1 mM. Application of metabolic modulators caused significant changes in mitochondria membrane potential, intracellular ATP level and ATP:ADP ratio. The detailed breakdown of cellular bioenergetics showed that proliferating human coronary artery smooth muscle cells rely more or less equally on oxidative phosphorylation and glycolysis at rest. These cells have high respiratory reserve capacity and low glycolysis reserve capacity. Metabolic intervention influences both intracellular ATP concentration and ATP:ADP ratio, where subtler changes may be detected by the latter.

  14. Synthesis and Protective Effects of Kaempferol-3'-sulfonate on Hydrogen Peroxide-induced injury in Vascular Smooth Muscle Cells.

    Science.gov (United States)

    Yang, Xinbin; Wang, Qin; Wang, Chunmei; Qin, Xiaolin; Huang, Yu; Zeng, Renquan

    2016-06-01

    A novel water-soluble sulfated derivative, kaempferol-3'-sulfonate acid sodium (KS) with the composition of [C15 H9 O9 SNa]·2.5H2 O, was synthesized and characterized by elemental analysis, IR, (1) H NMR, (13) C NMR, and HRMS. Its protective effects on human vascular smooth muscle cells injured by hydrogen peroxide were evaluated by CCK-8 method, flow cytometry, and Western blotting. The experimental results indicated that the KS can significantly increase cell viability and reduce apoptosis on H2 O2 -injured VSMCs, as well as reverse the effects of H2 O2 on Bcl-2, Bad, and caspase-3 expressions. In addition, LDH leakage, MDA levels, and SOD and GSH activities were also measured with spectrophotometry. The results indicated that the KS acted as antioxidant preventing LDH leakage and MDA production, while increasing intracellular SOD and GSH activities. These findings revealed that KS might potentially serve as an effective antioxidant agent for prevention and treatment of vascular disease caused by H2 O2 -injured VSMCs. © 2015 John Wiley & Sons A/S.

  15. Essential Roles of Raf/Extracellular Signal-regulated Kinase/Mitogen-activated Protein Kinase Pathway, YY1, and Ca2+ Influx in Growth Arrest of Human Vascular Smooth Muscle Cells by Bilirubin*

    Science.gov (United States)

    Stoeckius, Marlon; Erat, Anna; Fujikawa, Tatsuya; Hiromura, Makoto; Koulova, Anna; Otterbein, Leo; Bianchi, Cesario; Tobiasch, Edda; Dagon, Yossi; Sellke, Frank W.; Usheva, Anny

    2012-01-01

    The biological effects of bilirubin, still poorly understood, are concentration-dependent ranging from cell protection to toxicity. Here we present data that at high nontoxic physiological concentrations, bilirubin inhibits growth of proliferating human coronary artery smooth muscle cells by three events. It impairs the activation of Raf/ERK/MAPK pathway and the cellular Raf and cyclin D1 content that results in retinoblastoma protein hypophosphorylation on amino acids S608 and S780. These events impede the release of YY1 to the nuclei and its availability to regulate the expression of genes and to support cellular proliferation. Moreover, altered calcium influx and calpain II protease activation leads to proteolytical degradation of transcription factor YY1. We conclude that in the serum-stimulated human vascular smooth muscle primary cell cultures, bilirubin favors growth arrest, and we propose that this activity is regulated by its interaction with the Raf/ERK/MAPK pathway, effect on cyclin D1 and Raf content, altered retinoblastoma protein profile of hypophosphorylation, calcium influx, and YY1 proteolysis. We propose that these activities together culminate in diminished 5 S and 45 S ribosomal RNA synthesis and cell growth arrest. The observations provide important mechanistic insight into the molecular mechanisms underlying the transition of human vascular smooth muscle cells from proliferative to contractile phenotype and the role of bilirubin in this transition. PMID:22262839

  16. Expression of lectin-like oxidized LDL receptor-1 in smooth muscle cells after vascular injury

    International Nuclear Information System (INIS)

    Eto, Hideyuki; Miyata, Masaaki; Kume, Noriaki; Minami, Manabu; Itabe, Hiroyuki; Orihara, Koji; Hamasaki, Shuichi; Biro, Sadatoshi; Otsuji, Yutaka; Kita, Toru; Tei, Chuwa

    2006-01-01

    Lectin-like oxidized LDL receptor-1 (LOX-1) is an oxidized LDL receptor, and its role in restenosis after angioplasty remains unknown. We used a balloon-injury model of rabbit aorta, and reverse transcription-polymerase chain reaction revealed that LOX-1 mRNA expression was modest in the non-injured aorta, reached a peak level 2 days after injury, and remained elevated until 24 weeks after injury. Immunohistochemistry and in situ hybridization showed that LOX-1 was not detected in the media of non-injured aorta but expressed in both medial and neointimal smooth muscle cells (SMC) at 2 and 24 weeks after injury. Low concentrations of ox-LDL (10 μg/mL) stimulated the cultured SMC proliferation, which was inhibited by antisense oligonucleotides of LOX-1 mRNA. Double immunofluorescense staining showed the colocalization of LOX-1 and proliferating cell nuclear antigen in human restenotic lesion. These results suggest that LOX-1 mediates ox-LDL-induced SMC proliferation and plays a role in neointimal formation after vascular injury

  17. Major vault protein in cardiac and smooth muscle.

    Science.gov (United States)

    Shults, Nataliia V; Das, Dividutta; Suzuki, Yuichiro J

    Major vault protein (MVP) is the major component of the vault particle whose functions are not well understood. One proposed function of the vault is to serve as a mechanism of drug transport, which confers drug resistance in cancer cells. We show that MVP can be found in cardiac and smooth muscle. In human airway smooth muscle cells, knocking down MVP was found to cause cell death, suggesting that MVP serves as a cell survival factor. Further, our laboratory found that MVP is S-glutathionylated in response to ligand/receptor-mediated cell signaling. The S-glutathionylation of MVP appears to regulate protein-protein interactions between MVP and a protein called myosin heavy chain 9 (MYH9). Through MYH9 and Vsp34, MVP may form a complex with Beclin-1 that regulates autophagic cell death. In pulmonary vascular smooth muscle, proteasome inhibition promotes the ubiquitination of MVP, which may function as a mechanism of proteasome inhibition-mediated cell death. Investigating the functions and the regulatory mechanisms of MVP and vault particles is an exciting new area of research in cardiovascular/pulmonary pathophysiology.

  18. Bone Morphogenetic Protein-Induced Msx1 and Msx2 Inhibit Myocardin-Dependent Smooth Muscle Gene Transcription▿

    OpenAIRE

    Hayashi, Ken'ichiro; Nakamura, Seiji; Nishida, Wataru; Sobue, Kenji

    2006-01-01

    During the onset and progression of atherosclerosis, the vascular smooth muscle cell (VSMC) phenotype changes from differentiated to dedifferentiated, and in some cases, this change is accompanied by osteogenic transition, resulting in vascular calcification. One characteristic of dedifferentiated VSMCs is the down-regulation of smooth muscle cell (SMC) marker gene expression. Bone morphogenetic proteins (BMPs), which are involved in the induction of osteogenic gene ex...

  19. Dynamic and diverse changes in the functional properties of vascular smooth muscle cells in pulmonary hypertension.

    Science.gov (United States)

    Stenmark, Kurt R; Frid, Maria G; Graham, Brian B; Tuder, Rubin M

    2018-03-15

    Pulmonary hypertension (PH) is the end result of interaction between pulmonary vascular tone and a complex series of cellular and molecular events termed 'vascular remodelling'. The remodelling process, which can involve the entirety of pulmonary arterial vasculature, almost universally involves medial thickening, driven by increased numbers and hypertrophy of its principal cellular constituent, smooth muscle cells (SMCs). It is noted, however that SMCs comprise heterogeneous populations of cells, which can exhibit markedly different proliferative, inflammatory, and extracellular matrix production changes during remodelling. We further consider that these functional changes in SMCs of different phenotype and their role in PH are dynamic and may undergo significant changes over time (which we will refer to as cellular plasticity); no single property can account for the complexity of the contribution of SMC to pulmonary vascular remodelling. Thus, the approaches used to pharmacologically manipulate PH by targeting the SMC phenotype(s) must take into account processes that underlie dominant phenotypes that drive the disease. We present evidence for time- and location-specific changes in SMC proliferation in various animal models of PH; we highlight the transient nature (rather than continuous) of SMC proliferation, emphasizing that the heterogenic SMC populations that reside in different locations along the pulmonary vascular tree exhibit distinct responses to the stresses associated with the development of PH. We also consider that cells that have often been termed 'SMCs' may arise from many origins, including endothelial cells, fibroblasts and resident or circulating progenitors, and thus may contribute via distinct signalling pathways to the remodelling process. Ultimately, PH is characterized by long-lived, apoptosis-resistant SMC. In line with this key pathogenic characteristic, we address the acquisition of a pro-inflammatory phenotype by SMC that is essential

  20. Protocatechuic aldehyde inhibits migration and proliferation of vascular smooth muscle cells and intravascular thrombosis

    Energy Technology Data Exchange (ETDEWEB)

    Moon, Chang Yoon [The Hotchkiss School, Lakeville, CT (United States); Endocrinology, Brain Korea 21 Project for Medical Science, Institute of Endocrine Research, and Severance Integrative Research Institute for Cerebral and Cardiovascular Disease, Yonsei University College of Medicine, Seoul (Korea, Republic of); Ku, Cheol Ryong [Endocrinology, Brain Korea 21 Project for Medical Science, Institute of Endocrine Research, and Severance Integrative Research Institute for Cerebral and Cardiovascular Disease, Yonsei University College of Medicine, Seoul (Korea, Republic of); Cho, Yoon Hee, E-mail: wooriminji@gmail.com [Endocrinology, Brain Korea 21 Project for Medical Science, Institute of Endocrine Research, and Severance Integrative Research Institute for Cerebral and Cardiovascular Disease, Yonsei University College of Medicine, Seoul (Korea, Republic of); Lee, Eun Jig, E-mail: ejlee423@yuhs.ac [Endocrinology, Brain Korea 21 Project for Medical Science, Institute of Endocrine Research, and Severance Integrative Research Institute for Cerebral and Cardiovascular Disease, Yonsei University College of Medicine, Seoul (Korea, Republic of); Endocrinology, Northwestern University Feinberg School of Medicine, Chicago, IL (United States)

    2012-06-22

    Highlights: Black-Right-Pointing-Pointer Protocatechuic aldehyde (PCA) inhibits ROS production in VSMCs. Black-Right-Pointing-Pointer PCA inhibits proliferation and migration in PDGF-induced VSMCs. Black-Right-Pointing-Pointer PCA has anti-platelet effects in ex vivo rat whole blood. Black-Right-Pointing-Pointer We report the potential therapeutic role of PCA in atherosclerosis. -- Abstract: The migration and proliferation of vascular smooth muscle cells (VSMCs) and formation of intravascular thrombosis play crucial roles in the development of atherosclerotic lesions. This study examined the effects of protocatechuic aldehyde (PCA), a compound isolated from the aqueous extract of the root of Salvia miltiorrhiza, an herb used in traditional Chinese medicine to treat a variety of vascular diseases, on the migration and proliferation of VSMCs and platelets due to platelet-derived growth factor (PDGF). DNA 5-bromo-2 Prime -deoxy-uridine (BrdU) incorporation and wound-healing assays indicated that PCA significantly attenuated PDGF-induced proliferation and migration of VSMCs at a pharmacologically relevant concentration (100 {mu}M). On a molecular level, we observed down-regulation of the phosphatidylinositol 3-kinase (PI3K)/Akt and the mitogen-activated protein kinase (MAPK) pathways, both of which regulate key enzymes associated with migration and proliferation. We also found that PCA induced S-phase arrest of the VSMC cell cycle and suppressed cyclin D2 expression. In addition, PCA inhibited PDGF-BB-stimulated reactive oxygen species production in VSMCs, indicating that PCA's antioxidant properties may contribute to its suppression of PDGF-induced migration and proliferation in VSMCs. Finally, PCA exhibited an anti-thrombotic effect related to its inhibition of platelet aggregation, confirmed with an aggregometer. Together, these findings suggest a potential therapeutic role of PCA in the treatment of atherosclerosis and angioplasty-induced vascular restenosis.

  1. Single Nisoldipine-Sensitive Calcium Channels in Smooth Muscle Cells Isolated from Rabbit Mesenteric Artery

    Science.gov (United States)

    Worley, Jennings F.; Deitmer, Joachim W.; Nelson, Mark T.

    1986-08-01

    Single smooth muscle cells were enzymatically isolated from the rabbit mesenteric artery. At physiological levels of external Ca, these cells were relaxed and contracted on exposure to norepinephrine, caffeine, or high levels of potassium. The patch-clamp technique was used to measure unitary currents through single channels in the isolated cells. Single channels were selective for divalent cations and exhibited two conductance levels, 8 pS and 15 pS. Both types of channels were voltage-dependent, and channel activity occurred at potentials positive to -40 mV. The activity of both channel types was almost completely inhibited by 50 nM nisoldipine. These channels appear to be the pathways for voltage-dependent Ca influx in vascular smooth muscle and may be the targets of the clinically used dihydropyridines.

  2. Myostatin, a profibrotic factor and the main inhibitor of striated muscle mass, is present in the penile and vascular smooth muscle.

    Science.gov (United States)

    Kovanecz, I; Masouminia, M; Gelfand, R; Vernet, D; Rajfer, J; Gonzalez-Cadavid, N F

    2017-09-01

    Myostatin is present in striated myofibers but, except for myometrial cells, has not been reported within smooth muscle cells (SMC). We investigated in the rat whether myostatin is present in SMC within the penis and the vascular wall and, if so, whether it is transcriptionally expressed and associated with the loss of corporal SMC occurring in certain forms of erectile dysfunction (ED). Myostatin protein was detected by immunohistochemistry/fluorescence and western blots in the perineal striated muscles, and also in the SMC of the penile corpora, arteries and veins, and aorta. Myostatin was found in corporal SMC cultures, and its transcriptional expression (and its receptor) was shown there by DNA microarrays. Myostatin protein was measured by western blots in the penile shaft of rats subjected to bilateral cavernosal nerve resection (BCNR), that were left untreated, or treated (45 days) with muscle-derived stem cells (MDSC), or concurrent daily low-dose sildenafil. Myostatin was not increased by BCNR (compared with sham operated animals), but over expressed after treatment with MDSC. This was reduced by concurrent sildenafil. The presence of myostatin in corporal and vascular SMC, and its overexpression in the corpora by MDSC therapy, may have relevance for the stem cell treatment of corporal fibrosis and ED.

  3. Specification and Diversification of Pericytes and Smooth Muscle Cells from Mesenchymoangioblasts

    Directory of Open Access Journals (Sweden)

    Akhilesh Kumar

    2017-05-01

    Full Text Available Elucidating the pathways that lead to vasculogenic cells, and being able to identify their progenitors and lineage-restricted cells, is critical to the establishment of human pluripotent stem cell (hPSC models for vascular diseases and development of vascular therapies. Here, we find that mesoderm-derived pericytes (PCs and smooth muscle cells (SMCs originate from a clonal mesenchymal progenitor mesenchymoangioblast (MB. In clonogenic cultures, MBs differentiate into primitive PDGFRβ+CD271+CD73− mesenchymal progenitors, which give rise to proliferative PCs, SMCs, and mesenchymal stem/stromal cells. MB-derived PCs can be further specified to CD274+ capillary and DLK1+ arteriolar PCs with a proinflammatory and contractile phenotype, respectively. SMC maturation was induced using a MEK inhibitor. Establishing the vasculogenic lineage tree, along with identification of stage- and lineage-specific markers, provides a platform for interrogating the molecular mechanisms that regulate vasculogenic cell specification and diversification and manufacturing well-defined mural cell populations for vascular engineering and cellular therapies from hPSCs.

  4. LPS Promotes Vascular Smooth Muscle Cells Proliferation Through the TLR4/Rac1/Akt Signalling Pathway

    Directory of Open Access Journals (Sweden)

    Qianran Yin

    2017-12-01

    Full Text Available Background/Aims: Lipopolysaccharide (LPS is a potent activator of vascular smooth muscle cells (VSMCs proliferation, but the underlying mechanism remains unknown. In this study, we knocked down Toll-like receptor 4 (TLR4 and Ras-related C3 botulinum toxin substrate 1 (Rac1 expression using small interfering RNA (siRNA in order to investigate the effects and possible mechanisms of LPS-induced VSMCs proliferation. Methods: VSMCs proliferation was monitored by 5-ethynyl-2’-deoxyuridine staining, and Rac1 activity was measured via Glutathione S-transferase pull-down assay. mRNAs encoding proliferating cell nuclear antigen (PCNA, smooth muscle 22α (SM22α, myosin heavy chain (MYH and transient receptor potential channel 1 (TRPC1 were detected by qRT-PCR. The expression of total Akt, p-Akt (308, p-Akt (473, SM22α, MYH and TRPC1 protein was analysed by Western blot. Results: Treatment with TLR4 siRNA (siTLR4 or Rac1 siRNA (siRac1 significantly decreased LPS-induced VSMCs proliferation. Moreover, LPS-induced activation of Rac1 through TLR4 was observed. Western blot analysis revealed that transfection with siTLR4 or siRac1 inhibited LPS-induced Akt phosphorylation. We discovered that LPS stimulated VSMCs proliferation via phenotypic modulation and that this effect was partially inhibited by pre-treatment with siTLR4 or siRac1. Further, TLR4 and Rac1 are involved in LPS-induced activation of TRPC1. Conclusion: This study suggests that LPS exerts an effect on VSMCs proliferation and that the TLR4/Rac1/Akt signalling pathway mediates this effect.

  5. Localization and function of KLF4 in cytoplasm of vascular smooth muscle cell

    International Nuclear Information System (INIS)

    Liu, Yan; Zheng, Bin; Zhang, Xin-hua; Nie, Chan-juan; Li, Yong-hui; Wen, Jin-kun

    2013-01-01

    Highlights: •PDGF-BB prompts the translocation of KLF4 to the cytoplasm. •PDGF-BB promotes interaction between KLF4 and actin in the cytoplasm. •Phosphorylation and SUMOylation of KLF4 participates in regulation of cytoskeletal organization. •KLF4 regulates cytoskeleton by promoting the expression of contraction-associated genes. -- Abstract: The Krüppel-like factor 4 is a DNA-binding transcriptional regulator that regulates a diverse array of cellular processes, including development, differentiation, proliferation, and apoptosis. The previous studies about KLF4 functions mainly focused on its role as a transcription factor, its functions in the cytoplasm are still unknown. In this study, we found that PDGF-BB could prompt the translocation of KLF4 to the cytoplasm through CRM1-mediated nuclear export pathway in vascular smooth muscle cells (VSMCs) and increased the interaction of KLF4 with actin in the cytoplasm. Further study showed that both KLF4 phosphorylation and SUMOylation induced by PDGF-BB participates in regulation of cytoskeletal organization by stabilizing the actin cytoskeleton in VSMCs. In conclusion, these results identify that KLF4 participates in the cytoskeletal organization by stabilizing cytoskeleton in the cytoplasm of VSMCs

  6. Localization and function of KLF4 in cytoplasm of vascular smooth muscle cell

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Yan [Department of Biochemistry and Molecular Biology, The Key Laboratory of Neurobiology and Vascular Biology (China); The Third Hospital of Hebei Medical University, Shijazhuang (China); Zheng, Bin; Zhang, Xin-hua; Nie, Chan-juan; Li, Yong-hui [Department of Biochemistry and Molecular Biology, The Key Laboratory of Neurobiology and Vascular Biology (China); Wen, Jin-kun, E-mail: wjk@hebmu.edu.cn [Department of Biochemistry and Molecular Biology, The Key Laboratory of Neurobiology and Vascular Biology (China)

    2013-06-28

    Highlights: •PDGF-BB prompts the translocation of KLF4 to the cytoplasm. •PDGF-BB promotes interaction between KLF4 and actin in the cytoplasm. •Phosphorylation and SUMOylation of KLF4 participates in regulation of cytoskeletal organization. •KLF4 regulates cytoskeleton by promoting the expression of contraction-associated genes. -- Abstract: The Krüppel-like factor 4 is a DNA-binding transcriptional regulator that regulates a diverse array of cellular processes, including development, differentiation, proliferation, and apoptosis. The previous studies about KLF4 functions mainly focused on its role as a transcription factor, its functions in the cytoplasm are still unknown. In this study, we found that PDGF-BB could prompt the translocation of KLF4 to the cytoplasm through CRM1-mediated nuclear export pathway in vascular smooth muscle cells (VSMCs) and increased the interaction of KLF4 with actin in the cytoplasm. Further study showed that both KLF4 phosphorylation and SUMOylation induced by PDGF-BB participates in regulation of cytoskeletal organization by stabilizing the actin cytoskeleton in VSMCs. In conclusion, these results identify that KLF4 participates in the cytoskeletal organization by stabilizing cytoskeleton in the cytoplasm of VSMCs.

  7. The angiotensin-(1-7/Mas axis counteracts angiotensin II-dependent and –independent pro-inflammatory signaling in human vascular smooth muscle cells

    Directory of Open Access Journals (Sweden)

    Laura A Villalobos

    2016-12-01

    Full Text Available Background and aims: Targeting inflammation is nowadays considered as a challenging pharmacological strategy to prevent or delay the development of vascular diseases. Angiotensin-(1-7 is a member of the renin-angiotensin system (RAS that binds Mas receptors and has gained growing attention in the last years as a regulator of vascular homeostasis. Here, we explored the capacity of Ang-(1-7 to counteract human aortic smooth muscle cell (HASMC inflammation triggered by RAS-dependent and –independent stimuli, such as Ang II or interleukin (IL-1.Methods and Results: In cultured HASMC, the expression of iNOS and the release of nitric oxide were stimulated by both Ang II and IL-1, as determined by Western blot and indirect immunofluorescence or the Griess method, respectively. iNOS induction was inhibited by Ang-(1-7 in a concentration-dependent manner. This effect was equally blocked by two different Mas receptor antagonists, A779 and D-Pro7-Ang-(1-7, suggesting the participation of a unique Mas receptor subtype. Using pharmacological inhibitors, the induction of iNOS was proven to rely on the consecutive upstream activation of NADPH oxidase and NF-B. Indeed, Ang-(1-7 markedly inhibited the activation of the NADPH oxidase and subsequently of NF-B, as determined by lucigenin-derived chemiluminiscence and electromobility shift assay, respectively.Conclusion: Ang-(1-7 can act as a counter-regulator of the inflammation of vascular smooth muscle cells triggered by Ang II, but also by other stimuli beyond the RAS. Activating or mimicking the Ang-(1-7/Mas axis may represent a pharmacological opportunity to attenuate the pro-inflammatory environment that promotes and sustains the development of vascular diseases.

  8. Palmitic Acid Induces Osteoblastic Differentiation in Vascular Smooth Muscle Cells through ACSL3 and NF-κB, Novel Targets of Eicosapentaenoic Acid

    Science.gov (United States)

    Kageyama, Aiko; Matsui, Hiroki; Ohta, Masahiko; Sambuichi, Keisuke; Kawano, Hiroyuki; Notsu, Tatsuto; Imada, Kazunori; Yokoyama, Tomoyuki; Kurabayashi, Masahiko

    2013-01-01

    Free fatty acids (FFAs), elevated in metabolic syndrome and diabetes, play a crucial role in the development of atherosclerotic cardiovascular disease, and eicosapentaenoic acid (EPA) counteracts many aspects of FFA-induced vascular pathology. Although vascular calcification is invariably associated with atherosclerosis, the mechanisms involved are not completely elucidated. In this study, we tested the hypothesis that EPA prevents the osteoblastic differentiation and mineralization of vascular smooth muscle cells (VSMC) induced by palmitic acid (PA), the most abundant long-chain saturated fatty acid in plasma. PA increased and EPA abolished the expression of the genes for bone-related proteins, including bone morphogenetic protein (BMP)-2, Msx2 and osteopontin in human aortic smooth muscle cells (HASMC). Among the long-chain acyl-CoA synthetase (ACSL) subfamily, ACSL3 expression was predominant in HASMC, and PA robustly increased and EPA efficiently inhibited ACSL3 expression. Importantly, PA-induced osteoblastic differentiation was mediated, at least in part, by ACSL3 activation because acyl-CoA synthetase (ACS) inhibitor or siRNA targeted to ACSL3 completely prevented the PA induction of both BMP-2 and Msx2. Conversely, adenovirus-mediated ACSL3 overexpression enhanced PA-induced BMP-2 and Msx2 expression. In addition, EPA, ACSL3 siRNA and ACS inhibitor attenuated calcium deposition and caspase activation induced by PA. Notably, PA induced activation of NF-κB, and NF-κB inhibitor prevented PA-induction of osteoblastic gene expression and calcium deposition. Immunohistochemistry revealed the prominent expression of ACSL3 in VSMC and macrophages in human non-calcifying and calcifying atherosclerotic plaques from the carotid arteries. These results identify ACSL3 and NF-κB as mediators of PA-induced osteoblastic differentiation and calcium deposition in VSMC and suggest that EPA prevents vascular calcification by inhibiting such a new molecular pathway elicited

  9. Ureter smooth muscle cell orientation in rat is predominantly longitudinal.

    Directory of Open Access Journals (Sweden)

    Bart Spronck

    Full Text Available In ureter peristalsis, the orientation of the contracting smooth muscle cells is essential, yet current descriptions of orientation and composition of the smooth muscle layer in human as well as in rat ureter are inconsistent. The present study aims to improve quantification of smooth muscle orientation in rat ureters as a basis for mechanistic understanding of peristalsis. A crucial step in our approach is to use two-photon laser scanning microscopy and image analysis providing objective, quantitative data on smooth muscle cell orientation in intact ureters, avoiding the usual sectioning artifacts. In 36 rat ureter segments, originating from a proximal, middle or distal site and from a left or right ureter, we found close to the adventitia a well-defined longitudinal smooth muscle orientation. Towards the lamina propria, the orientation gradually became slightly more disperse, yet the main orientation remained longitudinal. We conclude that smooth muscle cell orientation in rat ureter is predominantly longitudinal, though the orientation gradually becomes more disperse towards the proprial side. These findings do not support identification of separate layers. The observed longitudinal orientation suggests that smooth muscle contraction would rather cause local shortening of the ureter, than cause luminal constriction. However, the net-like connective tissue of the ureter wall may translate local longitudinal shortening into co-local luminal constriction, facilitating peristalsis. Our quantitative, minimally invasive approach is a crucial step towards more mechanistic insight into ureter peristalsis, and may also be used to study smooth muscle cell orientation in other tube-like structures like gut and blood vessels.

  10. Ureter smooth muscle cell orientation in rat is predominantly longitudinal.

    Science.gov (United States)

    Spronck, Bart; Merken, Jort J; Reesink, Koen D; Kroon, Wilco; Delhaas, Tammo

    2014-01-01

    In ureter peristalsis, the orientation of the contracting smooth muscle cells is essential, yet current descriptions of orientation and composition of the smooth muscle layer in human as well as in rat ureter are inconsistent. The present study aims to improve quantification of smooth muscle orientation in rat ureters as a basis for mechanistic understanding of peristalsis. A crucial step in our approach is to use two-photon laser scanning microscopy and image analysis providing objective, quantitative data on smooth muscle cell orientation in intact ureters, avoiding the usual sectioning artifacts. In 36 rat ureter segments, originating from a proximal, middle or distal site and from a left or right ureter, we found close to the adventitia a well-defined longitudinal smooth muscle orientation. Towards the lamina propria, the orientation gradually became slightly more disperse, yet the main orientation remained longitudinal. We conclude that smooth muscle cell orientation in rat ureter is predominantly longitudinal, though the orientation gradually becomes more disperse towards the proprial side. These findings do not support identification of separate layers. The observed longitudinal orientation suggests that smooth muscle contraction would rather cause local shortening of the ureter, than cause luminal constriction. However, the net-like connective tissue of the ureter wall may translate local longitudinal shortening into co-local luminal constriction, facilitating peristalsis. Our quantitative, minimally invasive approach is a crucial step towards more mechanistic insight into ureter peristalsis, and may also be used to study smooth muscle cell orientation in other tube-like structures like gut and blood vessels.

  11. Intermittent hypoxia induces the proliferation of rat vascular smooth muscle cell with the increases in epidermal growth factor family and erbB2 receptor

    International Nuclear Information System (INIS)

    Kyotani, Yoji; Ota, Hiroyo; Itaya-Hironaka, Asako; Yamauchi, Akiyo; Sakuramoto-Tsuchida, Sumiyo; Zhao, Jing; Ozawa, Kentaro; Nagayama, Kosuke; Ito, Satoyasu; Takasawa, Shin; Kimura, Hiroshi; Uno, Masayuki; Yoshizumi, Masanori

    2013-01-01

    Obstructive sleep apnea is characterized by intermittent hypoxia (IH), and associated with cardiovascular diseases, such as stroke and heart failure. These cardiovascular diseases have a relation to atherosclerosis marked by the proliferation of vascular smooth muscle cells (VSMCs). In this study, we investigated the influence of IH on cultured rat aortic smooth muscle cell (RASMC). The proliferation of RASMC was significantly increased by IH without changing the level of apoptosis. In order to see what induces RASMC proliferation, we investigated the influence of normoxia (N)-, IH- and sustained hypoxia (SH)-treated cell conditioned media on RASMC proliferation. IH-treated cell conditioned medium significantly increased RASMC proliferation compared with N-treated cell conditioned medium, but SH-treated cell conditioned medium did not. We next investigated the epidermal growth factor (EGF) family as autocrine growth factors. Among the EGF family, we found significant increases in mRNAs for epiregulin (ER), amphiregulin (AR) and neuregulin-1 (NRG1) in IH-treated cells and mature ER in IH-treated cell conditioned medium. We next investigated the changes in erbB family receptors that are receptors for ER, AR and NRG1, and found that erbB2 receptor mRNA and protein expressions were increased by IH, but not by SH. Phosphorylation of erbB2 receptor at Tyr-1248 that mediates intracellular signaling for several physiological effects including cell proliferation was increased by IH, but not by SH. In addition, inhibitor for erbB2 receptor suppressed IH-induced cell proliferation. These results provide the first demonstration that IH induces VSMC proliferation, and suggest that EGF family, such as ER, AR and NRG1, and erbB2 receptor could be involved in the IH-induced VSMC proliferation. - Highlights: ●In vitro system for intermittent hypoxia (IH) and sustained hypoxia (SH). ●IH, but not SH, induces the proliferation of rat vascular smooth muscle cell. ●Epiregulin m

  12. Intermittent hypoxia induces the proliferation of rat vascular smooth muscle cell with the increases in epidermal growth factor family and erbB2 receptor

    Energy Technology Data Exchange (ETDEWEB)

    Kyotani, Yoji, E-mail: cd147@naramed-u.ac.jp [Department of Pharmacology, Nara Medical University School of Medicine, Kashihara 634-8521 (Japan); Department of Pharmacy, Nara Medical University Hospital, Kashihara 634-8522 (Japan); Ota, Hiroyo [Second Department of Internal Medicine, Nara Medical University School of Medicine, Kashihara 634-8522 (Japan); Department of Biochemistry, Nara Medical University School of Medicine, Kashihara 634-8521 (Japan); Itaya-Hironaka, Asako; Yamauchi, Akiyo; Sakuramoto-Tsuchida, Sumiyo [Department of Biochemistry, Nara Medical University School of Medicine, Kashihara 634-8521 (Japan); Zhao, Jing; Ozawa, Kentaro; Nagayama, Kosuke; Ito, Satoyasu [Department of Pharmacology, Nara Medical University School of Medicine, Kashihara 634-8521 (Japan); Takasawa, Shin [Department of Biochemistry, Nara Medical University School of Medicine, Kashihara 634-8521 (Japan); Kimura, Hiroshi [Second Department of Internal Medicine, Nara Medical University School of Medicine, Kashihara 634-8522 (Japan); Uno, Masayuki [Department of Pharmacy, Nara Medical University Hospital, Kashihara 634-8522 (Japan); Yoshizumi, Masanori [Department of Pharmacology, Nara Medical University School of Medicine, Kashihara 634-8521 (Japan)

    2013-11-15

    Obstructive sleep apnea is characterized by intermittent hypoxia (IH), and associated with cardiovascular diseases, such as stroke and heart failure. These cardiovascular diseases have a relation to atherosclerosis marked by the proliferation of vascular smooth muscle cells (VSMCs). In this study, we investigated the influence of IH on cultured rat aortic smooth muscle cell (RASMC). The proliferation of RASMC was significantly increased by IH without changing the level of apoptosis. In order to see what induces RASMC proliferation, we investigated the influence of normoxia (N)-, IH- and sustained hypoxia (SH)-treated cell conditioned media on RASMC proliferation. IH-treated cell conditioned medium significantly increased RASMC proliferation compared with N-treated cell conditioned medium, but SH-treated cell conditioned medium did not. We next investigated the epidermal growth factor (EGF) family as autocrine growth factors. Among the EGF family, we found significant increases in mRNAs for epiregulin (ER), amphiregulin (AR) and neuregulin-1 (NRG1) in IH-treated cells and mature ER in IH-treated cell conditioned medium. We next investigated the changes in erbB family receptors that are receptors for ER, AR and NRG1, and found that erbB2 receptor mRNA and protein expressions were increased by IH, but not by SH. Phosphorylation of erbB2 receptor at Tyr-1248 that mediates intracellular signaling for several physiological effects including cell proliferation was increased by IH, but not by SH. In addition, inhibitor for erbB2 receptor suppressed IH-induced cell proliferation. These results provide the first demonstration that IH induces VSMC proliferation, and suggest that EGF family, such as ER, AR and NRG1, and erbB2 receptor could be involved in the IH-induced VSMC proliferation. - Highlights: ●In vitro system for intermittent hypoxia (IH) and sustained hypoxia (SH). ●IH, but not SH, induces the proliferation of rat vascular smooth muscle cell. ●Epiregulin m

  13. Modeling Cerebrovascular Pathophysiology in Amyloid-β Metabolism using Neural-Crest-Derived Smooth Muscle Cells

    Directory of Open Access Journals (Sweden)

    Christine Cheung

    2014-10-01

    Full Text Available Summary: There is growing recognition of cerebrovascular contributions to neurodegenerative diseases. In the walls of cerebral arteries, amyloid-beta (Aβ accumulation is evident in a majority of aged people and patients with cerebral amyloid angiopathy. Here, we leverage human pluripotent stem cells to generate vascular smooth muscle cells (SMCs from neural crest progenitors, recapitulating brain-vasculature-specific attributes of Aβ metabolism. We confirm that the lipoprotein receptor, LRP1, functions in our neural-crest-derived SMCs to mediate Aβ uptake and intracellular lysosomal degradation. Hypoxia significantly compromises the contribution of SMCs to Aβ clearance by suppressing LRP1 expression. This enabled us to develop an assay of Aβ uptake by using the neural crest-derived SMCs with hypoxia as a stress paradigm. We then tested several vascular protective compounds in a high-throughput format, demonstrating the value of stem-cell-based phenotypic screening for novel therapeutics and drug repurposing, aimed at alleviating amyloid burden. : The contribution of blood vessel pathologies to neurodegenerative disorders is relatively neglected, partly due to inadequate human tissues for research. By using human stem cells, Cheung et al. establish a method of generating vascular smooth muscle cells (SMCs from neural crest progenitors, the primary precursors that give rise to brain blood vessels. These stem-cell-derived SMCs display defective amyloid processing under chronic hypoxia, a phenomenon well documented in the cerebral vasculatures of aged people and patients with Alzheimer’s disease.

  14. Technetium-99m labeled antisense probes uptake in vascular smooth muscle cells

    International Nuclear Information System (INIS)

    Zhang, Y.X.; Qin, G.M.; An, R.; Cao, G.X.; Cao, W.; Gao, Z.R.

    2002-01-01

    In the arterial wall, smooth muscle cells (SMC) normally exist in a quiescent, differentiated state, representing the contractile phenotype. During the development of atherosclerosis SMC change towards the synthetic phenotype going along with proliferation, chemotactic response and increased monocyte binding. The Fas/Fas ligand/caspase death-signaling pathway, Bcl-2 protein family/mitochondria, the tumor suppressive gene p53, and the proto-oncogene c-myc may be activated in atherosclerotic lesions, and mediates vascular apoptosis during the development of atherosclerosis. The atherosclerotic plaques contained 3-4 fold more c-myc mRNA than those in the normal aortic arteries, while increased Bax and Bak coupled with lack/paucity of Bcl-2 and Bcl-xL are associated with SMC apoptosis in advanced lesions. Methods: 1 Oligonucleotide Conjugation: A solution of single stranded amine-derivatized DNA (100-1000μg) was prepared at a concentration of 2 mg/ml in 0.25M sodium bicarbonate, 1 M sodium chloride, 1mM EDTA, pH8.5. Cell uptake studies: 99m Tc- MAG 3 -DNA radioactivity incorporation into porcine coronary smooth muscle cells in the log and plateau phases, respectively, was determined after different times of incubation at 37. The influence of extracellular 99m Tc- MAG 3 -DNA concentration on SMC uptake was also analyzed. [Results] Essentially complete conjugation was achieved by reverse-phase Sep-Pak C18 chromatography analysis. The MAG 3 -DNA was labeled with 99m Tc at room temperature and neutral pH, with a mean labeling efficiency of 80.11%(s.d=2.96%,n=4). The labeled antisense DNA still remained the ability to hybridize with its complementary DNA. After labeling, the stability of the DNA in saline or serum was retained as determined by reverse-phase Sep-Pak C18 chromatography analysis, except a shift at 30 min in serum incubation that suggesting a short time serum protein binding. 99m Tc-MAG 3 -c-myc uptake plateaued at 60 min and was directly proportional to the

  15. Effects of cyclic stretch on proliferation of mesenchymal stem cells and their differentiation to smooth muscle cells

    International Nuclear Information System (INIS)

    Ghazanfari, Samane; Tafazzoli-Shadpour, Mohammad; Shokrgozar, Mohammad Ali

    2009-01-01

    Bone marrow mesenchymal stem cells (MSCs) are capable of differentiating into a variety of cell types such as vascular smooth muscle cells (SMCs). In this study, we investigated influence of cyclic stretch on proliferation of hMSCs for different loading conditions, alignment of actin filaments, and consequent differentiation to SMCs. Isolated cells from bone marrow were exposed to cyclic stretch utilizing a customized device. Cell proliferation was examined by MTT assay, alignment of actin fibers by a designed image processing code, and cell differentiation by fluorescence staining. Results indicated promoted proliferation of hMSCs by cyclic strain, enhanced by elevated strain amplitude and number of cycles. Such loading regulated smooth muscle α-actin, and reoriented actin fibers. Cyclic stretch led to differentiation of hMSCs to SMCs without addition of growth factor. It was concluded that applying appropriate loading treatment on hMSCs could enhance proliferation capability, and produce functional SMCs for engineered tissues.

  16. Ubiquitin carboxyl terminal hydrolase L1 negatively regulates TNFα-mediated vascular smooth muscle cell proliferation via suppressing ERK activation

    International Nuclear Information System (INIS)

    Ichikawa, Tomonaga; Li, Jinqing; Dong, Xiaoyu; Potts, Jay D.; Tang, Dong-Qi; Li, Dong-Sheng; Cui, Taixing

    2010-01-01

    Deubiquitinating enzymes (DUBs) appear to be critical regulators of a multitude of processes such as proliferation, apoptosis, differentiation, and inflammation. We have recently demonstrated that a DUB of ubiquitin carboxyl terminal hydrolase L1 (UCH-L1) inhibits vascular lesion formation via suppressing inflammatory responses in vasculature. However, the precise underlying mechanism remains to be defined. Herein, we report that a posttranscriptional up-regulation of UCH-L1 provides a negative feedback to tumor necrosis factor alpha (TNFα)-mediated activation of extracellular signal-regulated kinases (ERK) and proliferation in vascular smooth muscle cells (VSMCs). In rat adult VSMCs, adenoviral over-expression of UCH-L1 inhibited TNFα-induced activation of ERK and DNA synthesis. In contrast, over-expression of UCH-L1 did not affect platelet derived growth factor (PDGF)-induced VSMC proliferation and activation of growth stimulating cascades including ERK. TNFα hardly altered UCH-L1 mRNA expression and stability; however, up-regulated UCH-L1 protein expression via increasing UCH-L1 translation. These results uncover a novel mechanism by which UCH-L1 suppresses vascular inflammation.

  17. Statins meditate anti-atherosclerotic action in smooth muscle cells by peroxisome proliferator-activated receptor-γ activation

    International Nuclear Information System (INIS)

    Fukuda, Kazuki; Matsumura, Takeshi; Senokuchi, Takafumi; Ishii, Norio; Kinoshita, Hiroyuki; Yamada, Sarie; Murakami, Saiko; Nakao, Saya; Motoshima, Hiroyuki; Kondo, Tatsuya; Kukidome, Daisuke; Kawasaki, Shuji; Kawada, Teruo; Nishikawa, Takeshi; Araki, Eiichi

    2015-01-01

    Highlights: • Statins induce PPARγ activation in vascular smooth muscle cells. • Statin-induced PPARγ activation is mediated by COX-2 expression. • Statins suppress cell migration and proliferation in vascular smooth muscle cells. • Statins inhibit LPS-induced inflammatory responses by PPARγ activation. • Fluvastatin suppress the progression of atherosclerosis and induces PPARγ activation in the aorta of apoE-deficient mice. - Abstract: The peroxisome proliferator-activated receptor-γ (PPARγ) is an important regulator of lipid and glucose metabolism, and its activation is reported to suppress the progression of atherosclerosis. We have reported that 3-hydroxy-3-methylglutaryl coenzyme A reductase inhibitors (statins) activate PPARγ in macrophages. However, it is not yet known whether statins activate PPARγ in other vascular cells. In the present study, we investigated whether statins activate PPARγ in smooth muscle cells (SMCs) and endothelial cells (ECs) and thus mediate anti-atherosclerotic effects. Human aortic SMCs (HASMCs) and human umbilical vein ECs (HUVECs) were used in this study. Fluvastatin and pitavastatin activated PPARγ in HASMCs, but not in HUVECs. Statins induced cyclooxygenase-2 (COX-2) expression in HASMCs, but not in HUVECs. Moreover, treatment with COX-2-siRNA abrogated statin-mediated PPARγ activation in HASMCs. Statins suppressed migration and proliferation of HASMCs, and inhibited lipopolysaccharide-induced expression of monocyte chemoattractant protein-1 (MCP-1) and tumor necrosis factor-α (TNF-α) in HASMCs. These effects of statins were abrogated by treatment with PPARγ-siRNA. Treatment with statins suppressed atherosclerotic lesion formation in Apoe −/− mice. In addition, transcriptional activity of PPARγ and CD36 expression were increased, and the expression of MCP-1 and TNF-α was decreased, in the aorta of statin-treated Apoe −/− mice. In conclusion, statins mediate anti-atherogenic effects through PPAR

  18. MicroRNA-125b Affects Vascular Smooth Muscle Cell Function by Targeting Serum Response Factor

    Directory of Open Access Journals (Sweden)

    Zhibo Chen

    2018-04-01

    Full Text Available Background/Aims: Increasing evidence links microRNAs to the pathogenesis of peripheral vascular disease. We recently found microRNA-125b (miR-125b to be one of the most significantly down‑regulated microRNAs in human arteries with arteriosclerosis obliterans (ASO of the lower extremities. However, its function in the process of ASO remains unclear. This study aimed to investigate the expression, regulatory mechanisms, and functions of miR-125b in the process of ASO. Methods: Using the tissue explants adherent method, vascular smooth muscle cells (VSMCs were prepared for this study. A rat carotid artery balloon injury model was constructed to simulate the development of vascular neointima, and a lentiviral transduction system was used to overexpress serum response factor (SRF or miR-125b. Quantitative real‑time PCR (qRT‑PCR was used to detect the expression levels of miR‑125b and SRF mRNA. Western blotting was performed to determine the expression levels of SRF and Ki67. In situ hybridization analysis was used to analyze the location and expression levels of miR-125b. CCK-8 and EdU assays were used to assess cell proliferation, and transwell and wound closure assays were performed to measure cell migration. Flow cytometry was used to evaluate cell apoptosis, and a dual-luciferase reporter assay was conducted to examine the effects of miR‑125b on SRF. Immunohistochemistry and immunofluorescence analyses were performed to analyze the location and expression levels of SRF and Ki67. Results: miR-125b expression was decreased in ASO arteries and platelet-derived growth factor (PDGF-BB-stimulated VSMCs. miR-125b suppressed VSMC proliferation and migration but promoted VSMC apoptosis. SRF was determined to be a direct target of miR-125b. Exogenous miR-125b expression modulated SRF expression and inhibited vascular neointimal formation in balloon-injured rat carotid arteries. Conclusions: These findings demonstrate a specific role of the mi

  19. A new iridoid and effect on the rat aortic vascular smooth muscle cell proliferation of isolated compounds from Buddleja officinalis.

    Science.gov (United States)

    Tai, Bui Huu; Nhiem, Nguyen Xuan; Quang, Tran Hong; Ngan, Nguyen Thi Thanh; Tung, Nguyen Huu; Kim, Yohan; Lee, Jung-Jin; Myung, Chang-Seon; Cuong, Nguyen Manh; Kim, Young Ho

    2011-06-01

    A new iridoid, named methylscutelloside (1) together with 19 known compounds belonging to the iridoids (2-4), monoterpenoids (5), flavonoids (6-8), triterpenoids (9-14), and phenylethanoids (15-20) were isolated from the flowers of Buddleja officinalis. Their chemical structures were elucidated on the basis of physicochemical properties, and by spectroscopic methods including 1D, 2D NMR, and MS. All isolated compounds were tested in vitro for their effects on the proliferation of rat aortic vascular smooth muscle cells (VSMCs). Among them, iridoids were the main active components and showed significant inhibitory effects on PDGF-BB-induced proliferation in rat aortic VSMCs. Copyright © 2011 Elsevier Ltd. All rights reserved.

  20. Increased NBCn1 expression, Na+/HCO3 co-transport and intracellular pH in human vascular smooth muscle cells with a risk allele for hypertension

    DEFF Research Database (Denmark)

    Ng, Fu Liang; Boedtkjer, Ebbe; Witkowska, Katarzyna

    2017-01-01

    cultures of vascular smooth muscle and endothelial cells. In both cell types, we found genotype-dependent differences for rs13082711 in DNA-nuclear protein interactions, where the risk allele is associated with increased SLC4A7 expression level, NBCn1 availability and function as reflected in elevated...

  1. Angiotensin II upregulates the expression of placental growth factor in human vascular endothelial cells and smooth muscle cells

    Directory of Open Access Journals (Sweden)

    Guo Yingqiang

    2010-05-01

    Full Text Available Abstract Background Atherosclerosis is now recognized as a chronic inflammatory disease. Angiotensin II (Ang II is a critical factor in inflammatory responses, which promotes the pathogenesis of atherosclerosis. Placental growth factor (PlGF is a member of the vascular endothelial growth factor (VEGF family cytokines and is associated with inflammatory progress of atherosclerosis. However, the potential link between PlGF and Ang II has not been investigated. In the current study, whether Ang II could regulate PlGF expression, and the effect of PlGF on cell proliferation, was investigated in human vascular endothelial cells (VECs and smooth muscle cells (VSMCs. Results In growth-arrested human VECs and VSMCs, Ang II induced PlGF mRNA expression after 4 hour treatment, and peaked at 24 hours. 10-6 mol/L Ang II increased PlGF protein production after 8 hour treatment, and peaked at 24 hours. Stimulation with Ang II also induced mRNA expression of VEGF receptor-1 and -2(VEGFR-1 and -2 in these cells. The Ang II type I receptor (AT1R antagonist blocked Ang II-induced PlGF gene expression and protein production. Several intracellular signals elicited by Ang II were involved in PlGF synthesis, including activation of protein kinase C, extracellular signal-regulated kinase 1/2 (ERK1/2 and PI3-kinase. A neutralizing antibody against PlGF partially inhibited the Ang II-induced proliferation of VECs and VSMCs. However, this antibody showed little effect on the basal proliferation in these cells, whereas blocking antibody of VEGF could suppress both basal and Ang II-induced proliferation in VECs and VSMCs. Conclusion Our results showed for the first time that Ang II could induce the gene expression and protein production of PlGF in VECs and VSMCs, which might play an important role in the pathogenesis of vascular inflammation and atherosclerosis.

  2. Increased NBCn1 expression, Na+/ HCO 3 ? co-transport and intracellular pH in human vascular smooth muscle cells with a risk allele for hypertension

    OpenAIRE

    Ng, Fu Liang; Boedtkjer, Ebbe; Witkowska, Kate; Ren, Meixia; Zhang, Ruoxin; Tucker, Arthur; Aalkj?r, Christian; Caulfield, Mark J.; Ye, Shu

    2017-01-01

    Abstract Genome-wide association studies have revealed an association between variation at the SLC4A7 locus and blood pressure. SLC4A7 encodes the electroneutral Na+/ HCO 3 ? co-transporter NBCn1 which regulates intracellular pH (pH i ). We conducted a functional study of variants at this locus in primary cultures of vascular smooth muscle and endothelial cells. In both cell types, we found genotype-dependent differences for rs13082711 in DNA-nuclear protein interactions, where the risk allel...

  3. Epigallocatechin Gallate Attenuates Proliferation and Oxidative Stress in Human Vascular Smooth Muscle Cells Induced by Interleukin-1β via Heme Oxygenase-1

    Directory of Open Access Journals (Sweden)

    Po-Len Liu

    2014-01-01

    Full Text Available Proliferation of vascular smooth muscle cells (VSMCs triggered by inflammatory stimuli and oxidative stress contributes importantly to atherogenesis. The association of green tea consumption with cardiovascular protection has been well documented in epidemiological observations, however, the underlying mechanisms remain unclear. This study aimed to elucidate the effects of the most active green tea catechin derivative, (−-epigallocatechin-3-gallate (EGCG, in human aortic smooth muscle cells (HASMCs, focusing particularly on the role of a potent anti-inflammatory and antioxidative enzyme heme oxygenase-1 (HO-1. We found that pretreatment of EGCG dose- and time-dependently induced HO-1 protein levels in HASMCs. EGCG inhibited interleukin- (IL-1β-induced HASMC proliferation and oxidative stress in a dose-dependent manner. The HO-1 inducer CoPPIX decreased IL-1β-induced cell proliferation, whereas the HO-1 enzyme inhibitor ZnPPIX significantly reversed EGCG-caused growth inhibition in IL-1β-treated HASMCs. At the molecular level, EGCG treatment significantly activated nuclear factor erythroid-2-related factor (Nrf2 transcription activities. These results suggest that EGCG might serve as a complementary and alternative medicine in the treatment of these pathologies by inducing HO-1 expression and subsequently decreasing VSMC proliferation.

  4. α-Iso-Cubebene Inhibits PDGF-Induced Vascular Smooth Muscle Cell Proliferation by Suppressing Osteopontin Expression

    Science.gov (United States)

    Jang, Min A.; Lee, Seung Jin; Baek, Seung Eun; Park, So Youn; Choi, Young Whan; Kim, Chi Dae

    2017-01-01

    α-Iso-cubebene (ICB) is a dibenzocyclooctadiene lignin contained in Schisandra chinensis (SC), a well-known medicinal herb that ameliorates cardiovascular symptoms. Thus, we examined the effect of ICB on vascular smooth muscle cell (VSMC) proliferation, a key feature of diverse vascular diseases. When VSMCs primary cultured from rat thoracic aorta were stimulated with PDGF (1–10 ng/ml), cell proliferation and osteopontin (OPN) expression were concomitantly up-regulated, but these effects were attenuated when cells were treated with MPIIIB10, a neutralizing monoclonal antibody for OPN. In aortic tissues exposed to PDGF, sprouting VSMC numbers increased, which was attenuated in tissues from OPN-deficient mice. Furthermore, VSMC proliferation and OPN expression induced by PDGF were attenuated dose-dependently by ICB (10 or 30 μg/ml). Reporter assays conducted using OPN promoter-luciferase constructs showed that the promoter region 538–234 bp of the transcription start site was responsible for transcriptional activity enhancement by PDGF, which was significantly inhibited by ICB. Putative binding sites for AP-1 and C/EBPβ in the indicated promoter region were suggested by TF Search, and increased binding of AP-1 and C/EBPβ in PDGF-treated VSMCs was demonstrated using a ChIP assay. The increased bindings of AP-1 and C/EBPβ into OPN promoter were attenuated by ICB. Moreover, the PDGF-induced expression of OPN was markedly attenuated in VSMCs transfected with siRNA for AP-1 and C/EBPβ. These results indicate that ICB inhibit VSMC proliferation by inhibiting the AP-1 and C/EBPβ signaling pathways and thus downregulating OPN expression. PMID:28114367

  5. Artery Tertiary Lymphoid Organs Control Aorta Immunity and Protect against Atherosclerosis via Vascular Smooth Muscle Cell Lymphotoxin β Receptors

    Science.gov (United States)

    Hu, Desheng; Mohanta, Sarajo K.; Yin, Changjun; Peng, Li; Ma, Zhe; Srikakulapu, Prasad; Grassia, Gianluca; MacRitchie, Neil; Dever, Gary; Gordon, Peter; Burton, Francis L.; Ialenti, Armando; Sabir, Suleman R.; McInnes, Iain B.; Brewer, James M.; Garside, Paul; Weber, Christian; Lehmann, Thomas; Teupser, Daniel; Habenicht, Livia; Beer, Michael; Grabner, Rolf; Maffia, Pasquale; Weih, Falk; Habenicht, Andreas J.R.

    2015-01-01

    Summary Tertiary lymphoid organs (TLOs) emerge during nonresolving peripheral inflammation, but their impact on disease progression remains unknown. We have found in aged Apoe−/− mice that artery TLOs (ATLOs) controlled highly territorialized aorta T cell responses. ATLOs promoted T cell recruitment, primed CD4+ T cells, generated CD4+, CD8+, T regulatory (Treg) effector and central memory cells, converted naive CD4+ T cells into induced Treg cells, and presented antigen by an unusual set of dendritic cells and B cells. Meanwhile, vascular smooth muscle cell lymphotoxin β receptors (VSMC-LTβRs) protected against atherosclerosis by maintaining structure, cellularity, and size of ATLOs though VSMC-LTβRs did not affect secondary lymphoid organs: Atherosclerosis was markedly exacerbated in Apoe−/−Ltbr−/− and to a similar extent in aged Apoe−/−Ltbrfl/flTagln-cre mice. These data support the conclusion that the immune system employs ATLOs to organize aorta T cell homeostasis during aging and that VSMC-LTβRs participate in atherosclerosis protection via ATLOs. PMID:26084025

  6. A pro-inflammatory role of deubiquitinating enzyme cylindromatosis (CYLD) in vascular smooth muscle cells

    International Nuclear Information System (INIS)

    Liu, Shuai; Lv, Jiaju; Han, Liping; Ichikawa, Tomonaga; Wang, Wenjuan; Li, Siying; Wang, Xing Li; Tang, Dongqi; Cui, Taixing

    2012-01-01

    Highlights: ► Cyld deficiency suppresses pro-inflammatory phenotypic switch of VSMCs. ► Cyld deficiency inhibits MAPK rather than NF-kB activity in inflamed VSMCs. ► CYLD is up-regulated in the coronary artery with neointimal hyperplasia. -- Abstract: CYLD, a deubiquitinating enzyme (DUB), is a critical regulator of diverse cellular processes, ranging from proliferation and differentiation to inflammatory responses, via regulating multiple key signaling cascades such as nuclear factor kappa B (NF-κB) pathway. CYLD has been shown to inhibit vascular lesion formation presumably through suppressing NF-κB activity in vascular cells. However, herein we report a novel role of CYLD in mediating pro-inflammatory responses in vascular smooth muscle cells (VSMCs) via a mechanism independent of NF-κB activity. Adenoviral knockdown of Cyld inhibited basal and the tumor necrosis factor alpha (TNFα)-induced mRNA expression of pro-inflammatory cytokines including monocyte chemotactic protein-1 (Mcp-1), intercellular adhesion molecule (Icam-1) and interleukin-6 (Il-6) in rat adult aortic SMCs (RASMCs). The CYLD deficiency led to increases in the basal NF-κB transcriptional activity in RASMCs; however, did not affect the TNFα-induced NF-κB activity. Intriguingly, the TNFα-induced IκB phosphorylation was enhanced in the CYLD deficient RASMCs. While knocking down of Cyld decreased slightly the basal expression levels of IκBα and IκBβ proteins, it did not alter the kinetics of TNFα-induced IκB protein degradation in RASMCs. These results indicate that CYLD suppresses the basal NF-κB activity and TNFα-induced IκB kinase activation without affecting TNFα-induced NF-κB activity in VSMCs. In addition, knocking down of Cyld suppressed TNFα-induced activation of mitogen activated protein kinases (MAPKs) including extracellular signal-activated kinases (ERK), c-Jun N-terminal kinase (JNK), and p38 in RASMCs. TNFα-induced RASMC migration and monocyte adhesion to

  7. Human lung mast cells modulate the functions of airway smooth muscle cells in asthma.

    Science.gov (United States)

    Alkhouri, H; Hollins, F; Moir, L M; Brightling, C E; Armour, C L; Hughes, J M

    2011-09-01

    Activated mast cell densities are increased on the airway smooth muscle in asthma where they may modulate muscle functions and thus contribute to airway inflammation, remodelling and airflow obstruction. To determine the effects of human lung mast cells on the secretory and proliferative functions of airway smooth muscle cells from donors with and without asthma. Freshly isolated human lung mast cells were stimulated with IgE/anti-IgE. Culture supernatants were collected after 2 and 24 h and the mast cells lysed. The supernatants/lysates were added to serum-deprived, subconfluent airway smooth muscle cells for up to 48 h. Released chemokines and extracellular matrix were measured by ELISA, proliferation was quantified by [(3) H]-thymidine incorporation and cell counting, and intracellular signalling by phospho-arrays. Mast cell 2-h supernatants reduced CCL11 and increased CXCL8 and fibronectin production from both asthmatic and nonasthmatic muscle cells. Leupeptin reversed these effects. Mast cell 24-h supernatants and lysates reduced CCL11 release from both muscle cell types but increased CXCL8 release by nonasthmatic cells. The 24-h supernatants also reduced asthmatic, but not nonasthmatic, muscle cell DNA synthesis and asthmatic cell numbers over 5 days through inhibiting extracellular signal-regulated kinase (ERK) and phosphatidylinositol (PI3)-kinase pathways. However, prostaglandins, thromboxanes, IL-4 and IL-13 were not involved in reducing the proliferation. Mast cell proteases and newly synthesized products differentially modulated the secretory and proliferative functions of airway smooth muscle cells from donors with and without asthma. Thus, mast cells may modulate their own recruitment and airway smooth muscle functions locally in asthma. © 2011 John Wiley & Sons A/S.

  8. Low density lipoprotein uptake by an endothelial-smooth muscle cell bilayer

    International Nuclear Information System (INIS)

    Alexander, J.J.; Miguel, R.; Graham, D.

    1991-01-01

    To study the interaction of endothelial and smooth muscle cells, and the means by which such interaction may affect lipid permeability of the arterial wall, cell bilayers were established by use of a transwell culture system. After confluent growth of both cell types had been achieved, iodine 125 bound to low-density lipoprotein (10 ng protein/ml) was added to the media of the upper well. After a 3-hour incubation period, the iodine 125-bound low-density lipoprotein content of the upper and lower media demonstrated an impedance to lipoprotein movement across the endothelial cell monolayer as compared to the bare porous polycarbonate filter of the transwell (p less than 10(-6)). The presence of smooth muscle cells in the bottom well significantly enhanced the permeability of the endothelial cell layer (p less than 10(-60)). This effect remained unchanged over a 9-day time course. Membrane binding and cellular uptake of low-density lipoprotein by endothelial cells was not altered by smooth muscle cells, indicating that this change in permeability could not be easily attributed to changes in receptor-mediated transport or transcytosis. Membrane binding (p less than 0.02) and cellular uptake (p less than 10(-6)) of low-density lipoprotein by smooth muscle cells in the bilayer, when adjusted for counts available in the smooth muscle cell media, were both reduced in the early incubation period as compared to isolated smooth muscle cells. The disproportionate reduction in uptake as compared to binding would suggest that this was not entirely a receptor-dependent process

  9. The PPARα/p16INK4a Pathway inhibits Vascular Smooth Muscle Cell Proliferation by repressing Cell Cycle-dependent Telomerase Activation

    Science.gov (United States)

    Gizard, Florence; Nomiyama, Takashi; Zhao, Yue; Findeisen, Hannes M.; Heywood, Elizabeth B.; Jones, Karrie L.; Staels, Bart; Bruemmer, Dennis

    2009-01-01

    Peroxisome Proliferator-Activated Receptor (PPAR) α, the molecular target for fibrates used to treat dyslipidemia, exerts pleiotropic effects on vascular cells. In vascular smooth muscle cells (VSMCs), we have previously demonstrated that PPARα activation suppresses G1→S cell cycle progression by targeting the cyclin-dependent kinase inhibitor p16INK4a (p16). In the present study, we demonstrate that this inhibition of VSMC proliferation by PPARα is mediated through a p16-dependent suppression of telomerase activity, which has been implicated in key cellular functions including proliferation. PPARα activation inhibited mitogen-induced telomerase activity by repressing the catalytic subunit telomerase reverse transcriptase (TERT) through negative cross-talk with an E2F-1-dependent trans-activation of the TERT promoter. This trans-repression involved the recruitment of the retinoblastoma (RB) family proteins p107 and p130 to the TERT promoter resulting in impaired E2F-1 binding, an effect which was dependent on p16. The inhibition of cell proliferation by PPARα activation was lost in VSMC following TERT overexpression or knock-down, pointing to a key role of telomerase as a target for the antiproliferative effects of PPARα. Finally, we demonstrate that PPARα agonists suppress telomerase activation during the proliferative response following vascular injury indicating that these findings are applicable in vivo. In concert, these results demonstrate that the anti-proliferative effects of PPARα in VSMCs depend on the suppression of telomerase activity by targeting the p16/RB/E2F transcriptional cascade. PMID:18818403

  10. Cyclic strain-induced endothelial MMP-2: role in vascular smooth muscle cell migration

    International Nuclear Information System (INIS)

    Sweeney, Nicholas von Offenberg; Cummins, Philip M.; Birney, Yvonne A.; Redmond, Eileen M.; Cahill, Paul A.

    2004-01-01

    Matrix metalloproteinases (MMPs) play a vital role in vasculature response to hemodynamic stimuli via the degradation of extracellular matrix substrates. In this study, we investigated the putative role of cyclic strain-induced endothelial MMP-2 (and MMP-9) expression and release in modulating bovine aortic smooth muscle cell (BASMC) migration in vitro. Equibiaxial cyclic strain of bovine aortic endothelial cells (BAECs) leads to elevation in cellular MMP-2 (and MMP-9) expression, activity, and secretion into conditioned media, events which were time- and force-dependent. Subsequent incubation of BASMCs with conditioned media from chronically strained BAECs (5%, 24 h) significantly reduces BASMC migration (38 ± 6%), an inhibitory effect which could be completely reversed by targeted siRNA 'knock-down' of MMP-2 (but not MMP-9) expression and activity in BAECs. Moreover, inhibition of strain-mediated MMP-2 expression in BAECs by protein tyrosine kinase (PTK) blockade with genistein (50 μM) was also found to completely reverse this inhibitory effect on BASMC migration. Finally, direct supplementation of recombinant MMP-2 into the BASMC migration assay was found to have no significant effect on migration. However, the effect on BASMC migration of MMP-2 siRNA transfection in BAECs could be reversed by supplementation of recombinant MMP-2 into BAEC media prior to (and for the duration of) strain. These findings reveal a potentially novel role for strain-induced endothelial MMP-2 in regulating vascular SMC migration

  11. Dense-body aggregates as plastic structures supporting tension in smooth muscle cells.

    Science.gov (United States)

    Zhang, Jie; Herrera, Ana M; Paré, Peter D; Seow, Chun Y

    2010-11-01

    The wall of hollow organs of vertebrates is a unique structure able to generate active tension and maintain a nearly constant passive stiffness over a large volume range. These properties are predominantly attributable to the smooth muscle cells that line the organ wall. Although smooth muscle is known to possess plasticity (i.e., the ability to adapt to large changes in cell length through structural remodeling of contractile apparatus and cytoskeleton), the detailed structural basis for the plasticity is largely unknown. Dense bodies, one of the most prominent structures in smooth muscle cells, have been regarded as the anchoring sites for actin filaments, similar to the Z-disks in striated muscle. Here, we show that the dense bodies and intermediate filaments formed cable-like structures inside airway smooth muscle cells and were able to adjust the cable length according to cell length and tension. Stretching the muscle cell bundle in the relaxed state caused the cables to straighten, indicating that these intracellular structures were connected to the extracellular matrix and could support passive tension. These plastic structures may be responsible for the ability of smooth muscle to maintain a nearly constant tensile stiffness over a large length range. The finding suggests that the structural plasticity of hollow organs may originate from the dense-body cables within the smooth muscle cells.

  12. α-Trinositol inhibits FGF-stimulated growth of smooth muscle and breast cancer cells

    International Nuclear Information System (INIS)

    Siren, Matti J.; Vainiomaeki, Maija; Vaeaenaenen, Kalervo; Haerkoenen, Pirkko

    2004-01-01

    α-Trinositol (D-myo-inositol-1,2,6-trisphosphate), an isomer of the intracellular messenger IP 3 , has been studied for its anti-inflammatory and other effects in animal experiments and in human. The mechanisms of action remain unknown. Several human pathologies are associated with uncontrolled production of fibroblast growth factors (FGFs). FGF-2 induces vascular smooth muscle cell proliferation, which contributes to restenosis after coronary balloon angioplasty. The expression of several FGFs is also increased in tumors. We studied the effects of the water- and lipid-soluble derivatives of α-trinositol on the FGF-2- and/or FGF-8-induced proliferation of human pulmonary artery smooth muscle cells (HPASMC) and S115 mouse breast cancer cells. α-Trinositol decreased the FGF-mediated proliferation of HPASMC and S115 cells. Membrane permeability did not seem obligatory since the lipid-soluble form of α-trinositol was less effective than the water-soluble derivative. These results suggest a new biological function for certain phosphoinositides in the modulation of FGF-regulated processes

  13. Generation and Characterization of Vascular Smooth Muscle Cell Lines Derived from a Patient with a Bicuspid Aortic Valve

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    Pamela Lazar-Karsten

    2016-04-01

    Full Text Available Thoracic aortic dilation is the most common malformation of the proximal aorta and is responsible for 1%–2% of all deaths in industrialized countries. In approximately 50% of patients with a bicuspid aortic valve (BAV, dilation of any or all segments of the aorta occurs. BAV patients with aortic dilation show an increased incidence of cultured vascular smooth muscle cell (VSMC loss. In this study, VSMC, isolated from the ascending aorta of BAV, was treated with Simian virus 40 to generate a BAV-originated VSMC cell line. To exclude any genomic DNA or cross-contamination, highly polymorphic short tandem repeats of the cells were profiled. The cells were then characterized using flow cytometry and karyotyping. The WG-59 cell line created is the first reported VSMC cell line isolated from a BAV patient. Using an RT2 Profiler PCR Array, genes within the TGFβ/BMP family that are dependent on losartan treatment were identified. Endoglin was found to be among the regulated genes and was downregulated in WG-59 cells following treatment with different losartan concentrations, when compared to untreated WG-59 cells.

  14. Leptin promotes osteoblast differentiation and mineralization of primary cultures of vascular smooth muscle cells by inhibiting glycogen synthase kinase (GSK)-3{beta}

    Energy Technology Data Exchange (ETDEWEB)

    Zeadin, Melec G.; Butcher, Martin K.; Shaughnessy, Stephen G. [Department of Medicine, McMaster University, Hamilton, ON (Canada); Thrombosis and Atherosclerosis Research Institute, Hamilton, ON (Canada); Werstuck, Geoff H., E-mail: Geoff.Werstuck@taari.ca [Department of Medicine, McMaster University, Hamilton, ON (Canada); Thrombosis and Atherosclerosis Research Institute, Hamilton, ON (Canada)

    2012-09-07

    Highlights: Black-Right-Pointing-Pointer Leptin promotes osteoblast differentiation of primary smooth muscle cells. Black-Right-Pointing-Pointer Leptin regulates the expression of genes involved in osteoblast differentiation. Black-Right-Pointing-Pointer Constitutively active GSK-3{beta} attenuates leptin-induced osteoblast differentiation. Black-Right-Pointing-Pointer This suggests that leptin signals through GSK-3{beta} to promote osteoblast differentiation. -- Abstract: In this study, we begin to investigate the underlying mechanism of leptin-induced vascular calcification. We found that treatment of cultured bovine aortic smooth muscle cells (BASMCs) with leptin (0.5-4 {mu}g/ml) induced osteoblast differentiation in a dose-dependent manner. Furthermore, we found that leptin significantly increased the mRNA expression of osteopontin and bone sialoprotein, while down-regulating matrix gla protein (MGP) expression in BASMCs. Key factors implicated in osteoblast differentiation, including members of the Wnt signaling pathway, were examined. Exposure to leptin enhanced phosphorylation of GSK-3{beta} on serine-9 thereby inhibiting activity and promoting the nuclear accumulation of {beta}-catenin. Transfection of BASMCs with an adenovirus that expressed constitutively active GSK-3{beta} (Ad-GSK-3{beta} S9A) resulted in a >2-fold increase in GSK-3{beta} activity and a significant decrease in leptin-induced alkaline phosphatase (ALP) activity. In addition, qRT-PCR analysis showed that GSK-3{beta} activation resulted in a significant decrease in the expression of osteopontin and bone sialoprotein, but a marked increase in MGP mRNA expression. When taken together, our results suggest a mechanism by which leptin promotes osteoblast differentiation and vascular calcification in vivo.

  15. Molecular basis for interaction of Na+/K+-ATPase with other transporters in membrane microdomains of vascular smooth muscle cells

    DEFF Research Database (Denmark)

    Hansen, Anne Kirstine; Matchkov, Vladimir; Bouzinova, Elena

    2008-01-01

    Ouabain, a specific inhibitor of the Na+/K+-pump, has previously been shown to interfere with intercellular communication. We have recently demonstrated a mechanism of this action of ouabain (1). We have showed that gap junctions between vascular smooth muscle cells (SMCs) are regulated through...... an interaction between the Na+/K+-pump and the Na+/Ca2+-exchanger leading to an increase in the intracellular calcium concentration in discrete areas near the plasma membrane. This regulation suggests a close association of the proteins in microdomains. We have also suggested that this Na...

  16. Transcriptional Control of Vascular Smooth Muscle Cell Proliferation by Peroxisome Proliferator-Activated Receptor-γ: Therapeutic Implications for Cardiovascular Diseases

    Directory of Open Access Journals (Sweden)

    Florence Gizard

    2008-01-01

    Full Text Available Proliferation of vascular smooth muscle cells (SMCs is a critical process for the development of atherosclerosis and complications of procedures used to treat atherosclerotic diseases, including postangioplasty restenosis, vein graft failure, and transplant vasculopathy. Peroxisome proliferator-activated receptor (PPAR γ is a member of the nuclear hormone receptor superfamily and the molecular target for the thiazolidinediones (TZD, used clinically to treat insulin resistance in patients with type 2 diabetes. In addition to their efficacy to improve insulin sensitivity, TZD exert a broad spectrum of pleiotropic beneficial effects on vascular gene expression programs. In SMCs, PPARγ is prominently upregulated during neointima formation and suppresses the proliferative response to injury of the arterial wall. Among the molecular target genes regulated by PPARγ in SMCs are genes encoding proteins involved in the regulation of cell-cycle progression, cellular senescence, and apoptosis. This inhibition of SMC proliferation is likely to contribute to the prevention of atherosclerosis and postangioplasty restenosis observed in animal models and proof-of-concept clinical studies. This review will summarize the transcriptional target genes regulated by PPARγ in SMCs and outline the therapeutic implications of PPARγ activation for the treatment and prevention of atherosclerosis and its complications.

  17. Hypoxia Enhances Differentiation of Adipose Tissue-Derived Stem Cells toward the Smooth Muscle Phenotype

    Directory of Open Access Journals (Sweden)

    Fang Wang

    2018-02-01

    Full Text Available Smooth muscle differentiated adipose tissue-derived stem cells are a valuable resource for regeneration of gastrointestinal tissues, such as the gut and sphincters. Hypoxia has been shown to promote adipose tissue-derived stem cells proliferation and maintenance of pluripotency, but the influence of hypoxia on their smooth myogenic differentiation remains unexplored. This study investigated the phenotype and contractility of adipose-derived stem cells differentiated toward the smooth myogenic lineage under hypoxic conditions. Oxygen concentrations of 2%, 5%, 10%, and 20% were used during differentiation of adipose tissue-derived stem cells. Real time reverse transcription polymerase chain reaction and immunofluorescence staining were used to detect the expression of smooth muscle cells-specific markers, including early marker smooth muscle alpha actin, middle markers calponin, caldesmon, and late marker smooth muscle myosin heavy chain. The specific contractile properties of cells were verified with both a single cell contraction assay and a gel contraction assay. Five percent oxygen concentration significantly increased the expression levels of α-smooth muscle actin, calponin, and myosin heavy chain in adipose-derived stem cell cultures after 2 weeks of induction (p < 0.01. Cells differentiated in 5% oxygen conditions showed greater contraction effect (p < 0.01. Hypoxia influences differentiation of smooth muscle cells from adipose stem cells and 5% oxygen was the optimal condition to generate smooth muscle cells that contract from adipose stem cells.

  18. Statins meditate anti-atherosclerotic action in smooth muscle cells by peroxisome proliferator-activated receptor-γ activation

    Energy Technology Data Exchange (ETDEWEB)

    Fukuda, Kazuki [Department of Metabolic Medicine, Faculty of Life Sciences, Kumamoto University, Kumamoto (Japan); Matsumura, Takeshi, E-mail: takeshim@gpo.kumamoto-u.ac.jp [Department of Metabolic Medicine, Faculty of Life Sciences, Kumamoto University, Kumamoto (Japan); Senokuchi, Takafumi; Ishii, Norio; Kinoshita, Hiroyuki; Yamada, Sarie; Murakami, Saiko [Department of Metabolic Medicine, Faculty of Life Sciences, Kumamoto University, Kumamoto (Japan); Nakao, Saya [Department of Environmental & Symbiotic Sciences, Prefectural University of Kumamoto, Kumamoto (Japan); Motoshima, Hiroyuki; Kondo, Tatsuya; Kukidome, Daisuke; Kawasaki, Shuji [Department of Metabolic Medicine, Faculty of Life Sciences, Kumamoto University, Kumamoto (Japan); Kawada, Teruo [Laboratory of Nutrition Chemistry, Division of Food Science and Biotechnology, Graduate School of Agriculture, Kyoto University, Kyoto (Japan); Nishikawa, Takeshi; Araki, Eiichi [Department of Metabolic Medicine, Faculty of Life Sciences, Kumamoto University, Kumamoto (Japan)

    2015-01-30

    Highlights: • Statins induce PPARγ activation in vascular smooth muscle cells. • Statin-induced PPARγ activation is mediated by COX-2 expression. • Statins suppress cell migration and proliferation in vascular smooth muscle cells. • Statins inhibit LPS-induced inflammatory responses by PPARγ activation. • Fluvastatin suppress the progression of atherosclerosis and induces PPARγ activation in the aorta of apoE-deficient mice. - Abstract: The peroxisome proliferator-activated receptor-γ (PPARγ) is an important regulator of lipid and glucose metabolism, and its activation is reported to suppress the progression of atherosclerosis. We have reported that 3-hydroxy-3-methylglutaryl coenzyme A reductase inhibitors (statins) activate PPARγ in macrophages. However, it is not yet known whether statins activate PPARγ in other vascular cells. In the present study, we investigated whether statins activate PPARγ in smooth muscle cells (SMCs) and endothelial cells (ECs) and thus mediate anti-atherosclerotic effects. Human aortic SMCs (HASMCs) and human umbilical vein ECs (HUVECs) were used in this study. Fluvastatin and pitavastatin activated PPARγ in HASMCs, but not in HUVECs. Statins induced cyclooxygenase-2 (COX-2) expression in HASMCs, but not in HUVECs. Moreover, treatment with COX-2-siRNA abrogated statin-mediated PPARγ activation in HASMCs. Statins suppressed migration and proliferation of HASMCs, and inhibited lipopolysaccharide-induced expression of monocyte chemoattractant protein-1 (MCP-1) and tumor necrosis factor-α (TNF-α) in HASMCs. These effects of statins were abrogated by treatment with PPARγ-siRNA. Treatment with statins suppressed atherosclerotic lesion formation in Apoe{sup −/−} mice. In addition, transcriptional activity of PPARγ and CD36 expression were increased, and the expression of MCP-1 and TNF-α was decreased, in the aorta of statin-treated Apoe{sup −/−} mice. In conclusion, statins mediate anti-atherogenic effects

  19. Mediators on human airway smooth muscle.

    Science.gov (United States)

    Armour, C; Johnson, P; Anticevich, S; Ammit, A; McKay, K; Hughes, M; Black, J

    1997-01-01

    1. Bronchial hyperresponsiveness in asthma may be due to several abnormalities, but must include alterations in the airway smooth muscle responsiveness and/or volume. 2. Increased responsiveness of airway smooth muscle in vitro can be induced by certain inflammatory cell products and by induction of sensitization (atopy). 3. Increased airway smooth muscle growth can also be induced by inflammatory cell products and atopic serum. 4. Mast cell numbers are increased in the airways of asthmatics and, in our studies, in airway smooth muscle that is sensitized and hyperresponsive. 5. We propose that there is a relationship between mast cells and airway smooth muscle cells which, once an allergic process has been initiated, results in the development of critical features in the lungs in asthma.

  20. Essential role of TGF-beta/Smad pathway on statin dependent vascular smooth muscle cell regulation.

    Directory of Open Access Journals (Sweden)

    Juan Rodríguez-Vita

    Full Text Available BACKGROUND: The 3-hydroxy-3-methylglutaryl CoA reductase inhibitors (also called statins exert proven beneficial effects on cardiovascular diseases. Recent data suggest a protective role for Transforming Growth Factor-beta (TGF-beta in atherosclerosis by regulating the balance between inflammation and extracellular matrix accumulation. However, there are no studies about the effect of statins on TGF-beta/Smad pathway in atherosclerosis and vascular cells. METHODOLOGY: In cultured vascular smooth muscle cells (VSMCs statins enhanced Smad pathway activation caused by TGF-beta. In addition, statins upregulated TGF-beta receptor type II (TRII, and increased TGF-beta synthesis and TGF-beta/Smad-dependent actions. In this sense, statins, through Smad activation, render VSMCs more susceptible to TGF-beta induced apoptosis and increased TGF-beta-mediated ECM production. It is well documented that high doses of statins induce apoptosis in cultured VSMC in the presence of serum; however the precise mechanism of this effect remains to be elucidated. We have found that statins-induced apoptosis was mediated by TGF-beta/Smad pathway. Finally, we have described that RhoA inhibition is a common intracellular mechanisms involved in statins effects. The in vivo relevance of these findings was assessed in an experimental model of atherosclerosis in apolipoprotein E deficient mice: Treatment with Atorvastatin increased Smad3 phosphorylation and TRII overexpression, associated to elevated ECM deposition in the VSMCs within atheroma plaques, while apoptosis was not detected. CONCLUSIONS: Statins enhance TGF-beta/Smad pathway, regulating ligand levels, receptor, main signaling pathway and cellular responses of VSMC, including apoptosis and ECM accumulation. Our findings show that TGF-beta/Smad pathway is essential for statins-dependent actions in VSMCs.

  1. Inhaled tolafentrine reverses pulmonary vascular remodeling via inhibition of smooth muscle cell migration

    Directory of Open Access Journals (Sweden)

    Weissmann Norbert

    2005-11-01

    Full Text Available Abstract Background The aim of the study was to assess the chronic effects of combined phosphodiesterase 3/4 inhibitor tolafentrine, administered by inhalation, during monocrotaline-induced pulmonary arterial hypertension (PAH in rats. Methods CD rats were given a single subcutaneous injection of monocrotaline to induce PAH. Four weeks after, rats were subjected to inhalation of tolafentrine or sham nebulization in an unrestrained, whole body aerosol exposure system. In these animals (i the acute pulmonary vasodilatory efficacy of inhaled tolafentrine (ii the anti-remodeling effect of long-term inhalation of tolafentrine (iii the effects of tolafentrine on the expression profile of 96 genes encoding cell adhesion and extracellular matrix regulation were examined. In addition, the inhibitory effect of tolafentrine on ex vivo isolated pulmonary artery SMC cell migration was also investigated. Results Monocrotaline injection provoked severe PAH (right ventricular systolic pressure increased from 25.9 ± 4.0 to 68.9 ± 3.2 after 4 weeks and 74.9 ± 5.1 mmHg after 6 weeks, cardiac output depression and right heart hypertrophy. The media thickness of the pulmonary arteries and the proportion of muscularization of small precapillary resistance vessels increased dramatically, and the migratory response of ex-vivo isolated pulmonary artery smooth muscle cells (PASMC was increased. Micro-arrays and subsequent confirmation with real time PCR demonstrated upregulation of several extracellular matrix regulation and adhesion genes, such as matrixmetalloproteases (MMP 2, 8, 9, 10, 11, 12, 20, Icam, Itgax, Plat and serpinb2. When chronically nebulized from day 28 to 42 (12 daily aerosol maneuvers, after full establishment of severe pulmonary hypertension, tolafentrine reversed about 60% of all hemodynamic abnormalities, right heart hypertrophy and monocrotaline-induced structural lung vascular changes, including the proportion of pulmonary artery

  2. SREBP inhibits VEGF expression in human smooth muscle cells

    Energy Technology Data Exchange (ETDEWEB)

    Motoyama, Koka [Metabolism, Endocrinology and Molecular Medicine, Osaka City University Graduate School of Medicine, Osaka (Japan); Fukumoto, Shinya [Metabolism, Endocrinology and Molecular Medicine, Osaka City University Graduate School of Medicine, Osaka (Japan); Koyama, Hidenori [Metabolism, Endocrinology and Molecular Medicine, Osaka City University Graduate School of Medicine, Osaka (Japan); Emoto, Masanori [Metabolism, Endocrinology and Molecular Medicine, Osaka City University Graduate School of Medicine, Osaka (Japan); Shimano, Hitoshi [Department of Internal Medicine, Institute of Clinical Medicine, University of Tsukuba, Ibaraki (Japan); Maemura, Koji [Department of Cardiovascular Medicine, Graduate School of Medicine, University of Tokyo, Tokyo (Japan); Nishizawa, Yoshiki [Metabolism, Endocrinology and Molecular Medicine, Osaka City University Graduate School of Medicine, Osaka (Japan)

    2006-03-31

    Sterol regulatory element-binding proteins (SREBPs) are transcription factors that regulate expression of genes encoding enzymes for lipid biosynthesis. SREBPs are activated by HMG-CoA reductase inhibitors (statins). Statins have been also reported to suppress vascular endothelial growth factor (VEGF) expression in vascular smooth muscle cells (VSMCs). Therefore, we hypothesized that SREBPs are involved in statin-mediated regulation of VEGF production in VSMCs. SREBP1 was robustly expressed, and was activated by atorvastatin in VSMCs, as demonstrated by increased levels of the mature nuclear form of SREBP1, and increased promoter activities of a reporter containing sterol regulatory elements by atorvastatin. Moreover, overexpression of SREBP1a dose-dependently suppressed VEGF promoter activity. Site-specific mutation or deletion of the proximal Sp1 sites reduced the inhibitory effects of SREBP1a on VEGF promoter activity. These data demonstrated that SREBP1, activated by atorvastatin, suppressed VEGF expression through the indirect interaction with the proximal tandem Sp1 sites in VSMCs.

  3. SREBP inhibits VEGF expression in human smooth muscle cells

    International Nuclear Information System (INIS)

    Motoyama, Koka; Fukumoto, Shinya; Koyama, Hidenori; Emoto, Masanori; Shimano, Hitoshi; Maemura, Koji; Nishizawa, Yoshiki

    2006-01-01

    Sterol regulatory element-binding proteins (SREBPs) are transcription factors that regulate expression of genes encoding enzymes for lipid biosynthesis. SREBPs are activated by HMG-CoA reductase inhibitors (statins). Statins have been also reported to suppress vascular endothelial growth factor (VEGF) expression in vascular smooth muscle cells (VSMCs). Therefore, we hypothesized that SREBPs are involved in statin-mediated regulation of VEGF production in VSMCs. SREBP1 was robustly expressed, and was activated by atorvastatin in VSMCs, as demonstrated by increased levels of the mature nuclear form of SREBP1, and increased promoter activities of a reporter containing sterol regulatory elements by atorvastatin. Moreover, overexpression of SREBP1a dose-dependently suppressed VEGF promoter activity. Site-specific mutation or deletion of the proximal Sp1 sites reduced the inhibitory effects of SREBP1a on VEGF promoter activity. These data demonstrated that SREBP1, activated by atorvastatin, suppressed VEGF expression through the indirect interaction with the proximal tandem Sp1 sites in VSMCs

  4. Smooth Muscle-Like Cells Generated from Human Mesenchymal Stromal Cells Display Marker Gene Expression and Electrophysiological Competence Comparable to Bladder Smooth Muscle Cells.

    Science.gov (United States)

    Brun, Juliane; Lutz, Katrin A; Neumayer, Katharina M H; Klein, Gerd; Seeger, Tanja; Uynuk-Ool, Tatiana; Wörgötter, Katharina; Schmid, Sandra; Kraushaar, Udo; Guenther, Elke; Rolauffs, Bernd; Aicher, Wilhelm K; Hart, Melanie L

    2015-01-01

    The use of mesenchymal stromal cells (MSCs) differentiated toward a smooth muscle cell (SMC) phenotype may provide an alternative for investigators interested in regenerating urinary tract organs such as the bladder where autologous smooth muscle cells cannot be used or are unavailable. In this study we measured the effects of good manufacturing practice (GMP)-compliant expansion followed by myogenic differentiation of human MSCs on the expression of a range of contractile (from early to late) myogenic markers in relation to the electrophysiological parameters to assess the functional role of the differentiated MSCs and found that differentiation of MSCs associated with electrophysiological competence comparable to bladder SMCs. Within 1-2 weeks of myogenic differentiation, differentiating MSCs significantly expressed alpha smooth muscle actin (αSMA; ACTA2), transgelin (TAGLN), calponin (CNN1), and smooth muscle myosin heavy chain (SM-MHC; MYH11) according to qRT-PCR and/or immunofluorescence and Western blot. Voltage-gated Na+ current levels also increased within the same time period following myogenic differentiation. In contrast to undifferentiated MSCs, differentiated MSCs and bladder SMCs exhibited elevated cytosolic Ca2+ transients in response to K+-induced depolarization and contracted in response to K+ indicating functional maturation of differentiated MSCs. Depolarization was suppressed by Cd2+, an inhibitor of voltage-gated Ca2+-channels. The expression of Na+-channels was pharmacologically identified as the Nav1.4 subtype, while the K+ and Ca2+ ion channels were identified by gene expression of KCNMA1, CACNA1C and CACNA1H which encode for the large conductance Ca2+-activated K+ channel BKCa channels, Cav1.2 L-type Ca2+ channels and Cav3.2 T-type Ca2+ channels, respectively. This protocol may be used to differentiate adult MSCs into smooth muscle-like cells with an intermediate-to-late SMC contractile phenotype exhibiting voltage-gated ion channel

  5. Smooth Muscle-Like Cells Generated from Human Mesenchymal Stromal Cells Display Marker Gene Expression and Electrophysiological Competence Comparable to Bladder Smooth Muscle Cells.

    Directory of Open Access Journals (Sweden)

    Juliane Brun

    Full Text Available The use of mesenchymal stromal cells (MSCs differentiated toward a smooth muscle cell (SMC phenotype may provide an alternative for investigators interested in regenerating urinary tract organs such as the bladder where autologous smooth muscle cells cannot be used or are unavailable. In this study we measured the effects of good manufacturing practice (GMP-compliant expansion followed by myogenic differentiation of human MSCs on the expression of a range of contractile (from early to late myogenic markers in relation to the electrophysiological parameters to assess the functional role of the differentiated MSCs and found that differentiation of MSCs associated with electrophysiological competence comparable to bladder SMCs. Within 1-2 weeks of myogenic differentiation, differentiating MSCs significantly expressed alpha smooth muscle actin (αSMA; ACTA2, transgelin (TAGLN, calponin (CNN1, and smooth muscle myosin heavy chain (SM-MHC; MYH11 according to qRT-PCR and/or immunofluorescence and Western blot. Voltage-gated Na+ current levels also increased within the same time period following myogenic differentiation. In contrast to undifferentiated MSCs, differentiated MSCs and bladder SMCs exhibited elevated cytosolic Ca2+ transients in response to K+-induced depolarization and contracted in response to K+ indicating functional maturation of differentiated MSCs. Depolarization was suppressed by Cd2+, an inhibitor of voltage-gated Ca2+-channels. The expression of Na+-channels was pharmacologically identified as the Nav1.4 subtype, while the K+ and Ca2+ ion channels were identified by gene expression of KCNMA1, CACNA1C and CACNA1H which encode for the large conductance Ca2+-activated K+ channel BKCa channels, Cav1.2 L-type Ca2+ channels and Cav3.2 T-type Ca2+ channels, respectively. This protocol may be used to differentiate adult MSCs into smooth muscle-like cells with an intermediate-to-late SMC contractile phenotype exhibiting voltage-gated ion

  6. Vascular smooth muscle cell differentiation to an osteogenic phenotype involves matrix metalloproteinase-2 modulation by homocysteine.

    Science.gov (United States)

    Liu, Tingjiao; Lin, Jinghan; Ju, Ting; Chu, Lei; Zhang, Liming

    2015-08-01

    Arterial calcification is common in vascular diseases and involves conversion of vascular smooth muscle cells (VSMCs) to an osteoblast phenotype. Clinical studies suggest that the development of atherosclerosis can be promoted by homocysteine (HCY), but the mechanisms remain unclear. Here, we determined whether increases in HCY levels lead to an increase in VSMC calcification and differentiation, and examined the role of an extracellular matrix remodeler, matrix metalloproteinase-2 (MMP-2). Rat VSMCs were exposed to calcification medium in the absence or presence of HCY (10, 100 or 200 μmol/L) or an MMP-2 inhibitor (10(-6) or 10(-5) mol/L). MTT assays were performed to determine the cytotoxicity of the MMP-2 inhibitor in calcification medium containing 200 μmol/L HCY. Calcification was assessed by measurements of calcium deposition and alkaline phosphatase (ALP) activity as well as von Kossa staining. Expression of osteocalcin, bone morphogenetic protein (BMP)-2, and osteopontin, and MMP-2 was determined by immunoblotting. Calcification medium induced osteogenic differentiation of VSMCs. HCY promoted calcification, increased osteocalcin and BMP-2 expression, and decreased expression of osteopontin. MMP-2 expression was increased by HCY in a dose-dependent manner in VSMCs exposed to both control and calcification medium. The MMP-2 inhibitor decreased the calcium content and ALP activity, and attenuated the osteoblastic phenotype of VSMCs. Vascular calcification and osteogenic differentiation of VSMCs were positively regulated by HCY through increased/restored MMP-2 expression, increased expression of calcification proteins, and decreased anti-calcification protein levels. In summary, MMP-2 inhibition may be a protective strategy against VSMC calcification.

  7. p115 RhoGEF activates the Rac1 GTPase signaling cascade in MCP1 chemokine-induced vascular smooth muscle cell migration and proliferation.

    Science.gov (United States)

    Singh, Nikhlesh K; Janjanam, Jagadeesh; Rao, Gadiparthi N

    2017-08-25

    Although the involvement of Rho proteins in the pathogenesis of vascular diseases is well studied, little is known about the role of their upstream regulators, the Rho guanine nucleotide exchange factors (RhoGEFs). Here, we sought to identify the RhoGEFs involved in monocyte chemotactic protein 1 (MCP1)-induced vascular wall remodeling. We found that, among the RhoGEFs tested, MCP1 induced tyrosine phosphorylation of p115 RhoGEF but not of PDZ RhoGEF or leukemia-associated RhoGEF in human aortic smooth muscle cells (HASMCs). Moreover, p115 RhoGEF inhibition suppressed MCP1-induced HASMC migration and proliferation. Consistent with these observations, balloon injury (BI) induced p115 RhoGEF tyrosine phosphorylation in rat common carotid arteries, and siRNA-mediated down-regulation of its levels substantially attenuated BI-induced smooth muscle cell migration and proliferation, resulting in reduced neointima formation. Furthermore, depletion of p115 RhoGEF levels also abrogated MCP1- or BI-induced Rac1-NFATc1-cyclin D1-CDK6-PKN1-CDK4-PAK1 signaling, which, as we reported previously, is involved in vascular wall remodeling. Our findings also show that protein kinase N1 (PKN1) downstream of Rac1-cyclin D1/CDK6 and upstream of CDK4-PAK1 in the p115 RhoGEF-Rac1-NFATc1-cyclin D1-CDK6-PKN1-CDK4-PAK1 signaling axis is involved in the modulation of vascular wall remodeling. Of note, we also observed that CCR2-G i/o -Fyn signaling mediates MCP1-induced p115 RhoGEF and Rac1 GTPase activation. These findings suggest that p115 RhoGEF is critical for MCP1-induced HASMC migration and proliferation in vitro and for injury-induced neointima formation in vivo by modulating Rac1-NFATc1-cyclin D1-CDK6-PKN1-CDK4-PAK1 signaling. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  8. RhoA-Rho kinase and platelet-activating factor stimulation of ovine foetal pulmonary vascular smooth muscle cell proliferation.

    Science.gov (United States)

    Renteria, L S; Austin, M; Lazaro, M; Andrews, M A; Lustina, J; Raj, J U; Ibe, B O

    2013-10-01

    Platelet-activating factor (PAF) is produced by pulmonary vascular smooth muscle cells (PVSMC). We studied effects of Rho kinase on PAF stimulation of PVSMC proliferation in an attempt to understand the role of RhoA/Rho kinase on PAF-induced ovine foetal pulmonary vascular remodelling. Our hypothesis is that PAF acts through Rho kinase, as one of its downstream signals, to induce arterial (SMC-PA) and venous (SMC-PV) cell proliferation in the hypoxic lung environment of the foetus, in utero. Rho kinase and MAPK effects on PAF receptor (PAFR)-mediated cell population expansion, and PAFR expression, were studied by DNA synthesis, western blot analysis and immunocytochemistry. Effects of constructs T19N and G14V on PAF-induced cell proliferation were also investigated. Hypoxia increased PVSMC proliferation and Rho kinase inhibitors, Y-27632 and Fasudil (HA-1077) as well as MAPK inhibitors PD 98059 and SB 203580 attenuated PAF stimulation of cell proliferation. RhoA T19N and G14V stimulated cell proliferation, but co-incubation with PAF did not affect proliferative effects of the constructs. PAFR protein expression was significantly downregulated in both cell types by both Y-27632 and HA-1077, with comparable profiles. Also, cells treated with Y-27632 had less PAF receptor fluorescence with significant disruption of cell morphology. Our results show that Rho kinase non-specifically modulated PAFR-mediated responses by a translational modification of PAFR protein, and suggest that, in vivo, activation of Rho kinase by PAF may be a further pathway to sustain PAFR-mediated PVSMC proliferation. © 2013 John Wiley & Sons Ltd.

  9. Smooth muscle-like tissue constructs with circumferentially oriented cells formed by the cell fiber technology.

    Science.gov (United States)

    Hsiao, Amy Y; Okitsu, Teru; Onoe, Hiroaki; Kiyosawa, Mahiro; Teramae, Hiroki; Iwanaga, Shintaroh; Kazama, Tomohiko; Matsumoto, Taro; Takeuchi, Shoji

    2015-01-01

    The proper functioning of many organs and tissues containing smooth muscles greatly depends on the intricate organization of the smooth muscle cells oriented in appropriate directions. Consequently controlling the cellular orientation in three-dimensional (3D) cellular constructs is an important issue in engineering tissues of smooth muscles. However, the ability to precisely control the cellular orientation at the microscale cannot be achieved by various commonly used 3D tissue engineering building blocks such as spheroids. This paper presents the formation of coiled spring-shaped 3D cellular constructs containing circumferentially oriented smooth muscle-like cells differentiated from dedifferentiated fat (DFAT) cells. By using the cell fiber technology, DFAT cells suspended in a mixture of extracellular proteins possessing an optimized stiffness were encapsulated in the core region of alginate shell microfibers and uniformly aligned to the longitudinal direction. Upon differentiation induction to the smooth muscle lineage, DFAT cell fibers self-assembled to coiled spring structures where the cells became circumferentially oriented. By changing the initial core-shell microfiber diameter, we demonstrated that the spring pitch and diameter could be controlled. 21 days after differentiation induction, the cell fibers contained high percentages of ASMA-positive and calponin-positive cells. Our technology to create these smooth muscle-like spring constructs enabled precise control of cellular alignment and orientation in 3D. These constructs can further serve as tissue engineering building blocks for larger organs and cellular implants used in clinical treatments.

  10. A pro-inflammatory role of deubiquitinating enzyme cylindromatosis (CYLD) in vascular smooth muscle cells

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Shuai [Shandong University Qilu Hospital Research Center for Cell Therapy, Key Laboratory of Cardiovascular Remodeling and Function Research, Qilu Hospital of Shandong University, Jinan 250012 (China); Department of Cell Biology and Anatomy, University of South Carolina School of Medicine, Columbia, SC 29208 (United States); Lv, Jiaju [Department of Urology, Shandong Provincial Hospital, Shandong University, Jinan 250021 (China); Han, Liping; Ichikawa, Tomonaga; Wang, Wenjuan; Li, Siying [Department of Cell Biology and Anatomy, University of South Carolina School of Medicine, Columbia, SC 29208 (United States); Wang, Xing Li [Shandong University Qilu Hospital Research Center for Cell Therapy, Key Laboratory of Cardiovascular Remodeling and Function Research, Qilu Hospital of Shandong University, Jinan 250012 (China); Tang, Dongqi, E-mail: tangdq@pathology.ufl.edu [Department of Pathology, Immunology, and Laboratory Medicine, University of Florida College of Medicine, Gainesville, FL 32610-0275 (United States); Cui, Taixing, E-mail: taixing.cui@uscmed.sc.edu [Department of Cell Biology and Anatomy, University of South Carolina School of Medicine, Columbia, SC 29208 (United States)

    2012-03-30

    Highlights: Black-Right-Pointing-Pointer Cyld deficiency suppresses pro-inflammatory phenotypic switch of VSMCs. Black-Right-Pointing-Pointer Cyld deficiency inhibits MAPK rather than NF-kB activity in inflamed VSMCs. Black-Right-Pointing-Pointer CYLD is up-regulated in the coronary artery with neointimal hyperplasia. -- Abstract: CYLD, a deubiquitinating enzyme (DUB), is a critical regulator of diverse cellular processes, ranging from proliferation and differentiation to inflammatory responses, via regulating multiple key signaling cascades such as nuclear factor kappa B (NF-{kappa}B) pathway. CYLD has been shown to inhibit vascular lesion formation presumably through suppressing NF-{kappa}B activity in vascular cells. However, herein we report a novel role of CYLD in mediating pro-inflammatory responses in vascular smooth muscle cells (VSMCs) via a mechanism independent of NF-{kappa}B activity. Adenoviral knockdown of Cyld inhibited basal and the tumor necrosis factor alpha (TNF{alpha})-induced mRNA expression of pro-inflammatory cytokines including monocyte chemotactic protein-1 (Mcp-1), intercellular adhesion molecule (Icam-1) and interleukin-6 (Il-6) in rat adult aortic SMCs (RASMCs). The CYLD deficiency led to increases in the basal NF-{kappa}B transcriptional activity in RASMCs; however, did not affect the TNF{alpha}-induced NF-{kappa}B activity. Intriguingly, the TNF{alpha}-induced I{kappa}B phosphorylation was enhanced in the CYLD deficient RASMCs. While knocking down of Cyld decreased slightly the basal expression levels of I{kappa}B{alpha} and I{kappa}B{beta} proteins, it did not alter the kinetics of TNF{alpha}-induced I{kappa}B protein degradation in RASMCs. These results indicate that CYLD suppresses the basal NF-{kappa}B activity and TNF{alpha}-induced I{kappa}B kinase activation without affecting TNF{alpha}-induced NF-{kappa}B activity in VSMCs. In addition, knocking down of Cyld suppressed TNF{alpha}-induced activation of mitogen activated protein

  11. Nestin upregulation characterizes vascular remodeling secondary to hypertension in the rat.

    Science.gov (United States)

    Tardif, Kim; Hertig, Vanessa; Duquette, Natacha; Villeneuve, Louis; El-Hamamsy, Ismail; Tanguay, Jean-François; Calderone, Angelino

    2015-05-15

    Proliferation and hypertrophy of vascular smooth muscle cells represent hallmark features of vessel remodeling secondary to hypertension. The intermediate filament protein nestin was recently identified in vascular smooth muscle cells and in other cell types directly participated in proliferation. The present study tested the hypothesis that vessel remodeling secondary to hypertension was characterized by nestin upregulation in vascular smooth muscle cells. Two weeks after suprarenal abdominal aorta constriction of adult male Sprague-Dawley rats, elevated mean arterial pressure increased the media area and thickness of the carotid artery and aorta and concomitantly upregulated nestin protein levels. In the normal adult rat carotid artery, nestin immunoreactivity was observed in a subpopulation of vascular smooth muscle cells, and the density significantly increased following suprarenal abdominal aorta constriction. Filamentous nestin was detected in cultured rat carotid artery- and aorta-derived vascular smooth muscle cells and an analogous paradigm observed in human aorta-derived vascular smooth muscle cells. ANG II and EGF treatment of vascular smooth muscle cells stimulated DNA and protein synthesis and increased nestin protein levels. Lentiviral short-hairpin RNA-mediated nestin depletion of carotid artery-derived vascular smooth muscle cells inhibited peptide growth factor-stimulated DNA synthesis, whereas protein synthesis remained intact. These data have demonstrated that vessel remodeling secondary to hypertension was characterized in part by nestin upregulation in vascular smooth muscle cells. The selective role of nestin in peptide growth factor-stimulated DNA synthesis has revealed that the proliferative and hypertrophic responses of vascular smooth muscle cells were mediated by divergent signaling events. Copyright © 2015 the American Physiological Society.

  12. Cytokine-induced differentiation of multipotent adult progenitor cells into functional smooth muscle cells.

    Science.gov (United States)

    Ross, Jeffrey J; Hong, Zhigang; Willenbring, Ben; Zeng, Lepeng; Isenberg, Brett; Lee, Eu Han; Reyes, Morayma; Keirstead, Susan A; Weir, E Kenneth; Tranquillo, Robert T; Verfaillie, Catherine M

    2006-12-01

    Smooth muscle formation and function are critical in development and postnatal life. Hence, studies aimed at better understanding SMC differentiation are of great importance. Here, we report that multipotent adult progenitor cells (MAPCs) isolated from rat, murine, porcine, and human bone marrow demonstrate the potential to differentiate into cells with an SMC-like phenotype and function. TGF-beta1 alone or combined with PDGF-BB in serum-free medium induces a temporally correct expression of transcripts and proteins consistent with smooth muscle development. Furthermore, SMCs derived from MAPCs (MAPC-SMCs) demonstrated functional L-type calcium channels. MAPC-SMCs entrapped in fibrin vascular molds became circumferentially aligned and generated force in response to KCl, the L-type channel opener FPL64176, or the SMC agonists 5-HT and ET-1, and exhibited complete relaxation in response to the Rho-kinase inhibitor Y-27632. Cyclic distention (5% circumferential strain) for 3 weeks increased responses by 2- to 3-fold, consistent with what occurred in neonatal SMCs. These results provide evidence that MAPC-SMCs are phenotypically and functionally similar to neonatal SMCs and that the in vitro MAPC-SMC differentiation system may be an ideal model for the study of SMC development. Moreover, MAPC-SMCs may lend themselves to tissue engineering applications.

  13. Divergent effects of 17-β-estradiol on human vascular smooth muscle and endothelial cell function diminishes TNF-α-induced neointima formation

    International Nuclear Information System (INIS)

    Nintasen, Rungrat; Riches, Kirsten; Mughal, Romana S.; Viriyavejakul, Parnpen; Chaisri, Urai; Maneerat, Yaowapa; Turner, Neil A.; Porter, Karen E.

    2012-01-01

    Highlights: ► TNF-α augments neointimal hyperplasia in human saphenous vein. ► TNF-α induces detrimental effects on endothelial and smooth muscle cell function. ► Estradiol exerts modulatory effects on TNF-induced vascular cell functions. ► The modulatory effects of estradiol are discriminatory and cell-type specific. -- Abstract: Coronary heart disease (CHD) is a condition characterized by increased levels of proinflammatory cytokines, including tumor necrosis factor-α (TNF-α). TNF-α can induce vascular endothelial cell (EC) and smooth muscle cell (SMC) dysfunction, central events in development of neointimal lesions. The reduced incidence of CHD in young women is believed to be due to the protective effects of estradiol (E2). We therefore investigated the effects of TNF-α on human neointima formation and SMC/EC functions and any modulatory effects of E2. Saphenous vein (SV) segments were cultured in the presence of TNF-α (10 ng/ml), E2 (2.5 nM) or both in combination. Neointimal thickening was augmented by incubation with TNF-α, an effect that was abolished by co-culture with E2. TNF-α increased SV–SMC proliferation in a concentration-dependent manner that was optimal at 10 ng/ml (1.5-fold increase), and abolished by E2 at all concentrations studied (1–50 nM). Surprisingly, E2 itself at low concentrations (1 and 5 nM) stimulated SV–SMC proliferation to a level comparable to that of TNF-α alone. SV–EC migration was significantly impaired by TNF-α (42% of control), and co-culture with E2 partially restored the ability of SV–EC to migrate and repair the wound. In contrast, TNF-α increased SV–SMC migration by 1.7-fold, an effect that was completely reversed by co-incubation with E2. Finally, TNF-α potently induced ICAM-1 and VCAM-1 expression in both SV–EC and SV–SMC. However there was no modulation by E2 in either cell-type. In conclusion, TNF-α induced SV neointima formation, increased SMC proliferation and migration, impaired

  14. Birth weight and characteristics of endothelial and smooth muscle cell cultures from human umbilical cord vessels

    Directory of Open Access Journals (Sweden)

    Lurbe Empar

    2009-04-01

    Full Text Available Abstract Background Low birth weight has been related to an increased risk for developing high blood pressure in adult life. The molecular and cellular analysis of umbilical cord artery and vein may provide information about the early vascular characteristics of an individual. We have assessed several phenotype characteristics of the four vascular cell types derived from human umbilical cords of newborns with different birth weight. Further follow-up studies could show the association of those vascular properties with infancy and adulthood blood pressure. Methods Endothelial and smooth muscle cell cultures were obtained from umbilical cords from two groups of newborns of birth weight less than 2.8 kg or higher than 3.5 kg. The expression of specific endothelial cell markers (von Willebrand factor, CD31, and the binding and internalization of acetylated low-density lipoprotein and the smooth muscle cell specific α-actin have been evaluated. Cell culture viability, proliferation kinetic, growth fraction (expression of Ki67 and percentage of senescent cells (detection of β-galactosidase activity at pH 6.0 have been determined. Endothelial cell projection area was determined by morphometric analysis of cell cultures after CD31 immunodetection. Results The highest variation was found in cell density at the confluence of endothelial cell cultures derived from umbilical cord arteries (66,789 ± 5,093 cells/cm2 vs. 45,630 ± 11,927 cells/cm2, p 2, p Conclusion The analysis of umbilical cord artery endothelial cells, which demonstrated differences in cell size related to birth weight, can provide hints about the cellular and molecular links between lower birth weight and increased adult high blood pressure risk.

  15. Role of integrin-linked kinase in vascular smooth muscle cells: Regulation by statins and angiotensin II

    International Nuclear Information System (INIS)

    Friedrich, Erik B.; Clever, Yvonne P.; Wassmann, Sven; Werner, Nikos; Boehm, Michael; Nickenig, Georg

    2006-01-01

    Our goal was to characterize the role of integrin-linked kinase (ILK) in vascular smooth muscle cells (VSMC), which play a crucial role in atherogenesis. Transfection of VSMC with wild-type and dominant-negative ILK cDNA constructs revealed that ILK mediates migration and proliferation of VSMC but has no effect on VSMC survival. The pro-atherogenic mediator angiotensin II increases ILK protein expression and kinase activity while statin treatment down-regulates ILK in VSMC. Functionally, ILK is necessary for angiotensin II-mediated VSMC migration and proliferation. In VSMC transduced with dominant-negative ILK, statins mediate an additive inhibition of VSMC migration and proliferation, while transfection with wild-type ILK is sufficient to overcome the inhibitory effects of statin treatment on VSMC migration and proliferation. In vivo, ILK is expressed in VSMC of aortic sections from wild-type mice where it is down-regulated following statin treatment and up-regulated following induction of atherosclerosis in apoE-/- mice. These data identify ILK as a novel target in VSMC for anti-atherosclerotic therapy

  16. Cultured smooth muscle cells of the human vesical sphincter are more sensitive to histamine than are detrusor smooth muscle cells.

    Science.gov (United States)

    Neuhaus, Jochen; Oberbach, Andreas; Schwalenberg, Thilo; Stolzenburg, Jens-Uwe

    2006-05-01

    To compare histamine receptor expression in cultured smooth muscle cells from the human detrusor and internal sphincter using receptor-specific agonists. Smooth muscle cells from the bladder dome and internal sphincter were cultured from 5 male patients undergoing cystectomy for bladder cancer therapy. Calcium transients in cells stimulated with carbachol, histamine, histamine receptor 1 (H1R)-specific heptanecarboxamide (HTMT), dimaprit (H2R), and R-(alpha)-methylhistamine (H3R) were measured by calcium imaging. Histamine receptor proteins were detected by Western blot analysis and immunocytochemistry. H1R, H2R, and H3R expression was found in tissue and cultured cells. Carbachol stimulated equal numbers of detrusor and sphincter cells (60% and 51%, respectively). Histamine stimulated significantly more cells than carbachol in detrusor (100%) and sphincter (99.34%) cells. Calcium responses to carbachol in detrusor and sphincter cells were comparable and did not differ from those to histamine in detrusor cells. However, histamine and specific agonists stimulated more sphincter cells than did carbachol (P <0.001), and the calcium increase was greater in sphincter cells than in detrusor cells. Single cell analysis revealed comparable H2R responses in detrusor and sphincter cells, but H1R and H3R-mediated calcium reactions were significantly greater in sphincter cells. Histamine very effectively induces calcium release in smooth muscle cells. In sphincter cells, histamine is even more effective than carbachol regarding the number of reacting cells and the intracellular calcium increase. Some of the variability in the outcome of antihistaminic interstitial cystitis therapies might be caused by the ineffectiveness of the chosen antihistaminic or unintentional weakening of sphincteric function.

  17. SIRT1 deacetylates RFX5 and antagonizes repression of collagen type I (COL1A2) transcription in smooth muscle cells

    Energy Technology Data Exchange (ETDEWEB)

    Xia, Jun [Department of Respiratory Medicine, The First Affiliated Hospital of Nanjing Medical University (China); Department of Respiratory Medicine, Jiangsu Provincial Hospital of Chinese Traditional Medicine (China); Wu, Xiaoyan; Yang, Yuyu; Zhao, Yuhao [Atherosclerosis Research Center, Key Laboratory of Cardiovascular Disease and Molecular Intervention, Department of Pathophysiology, Nanjing Medical University (China); Fang, Mingming [Jiangsu Jiankang Vocational Institute (China); Xie, Weiping, E-mail: wpxienjmu@gmail.com [Department of Respiratory Medicine, The First Affiliated Hospital of Nanjing Medical University (China); Wang, Hong, E-mail: hwangnjmu@gmail.com [Department of Respiratory Medicine, The First Affiliated Hospital of Nanjing Medical University (China); Xu, Yong [Atherosclerosis Research Center, Key Laboratory of Cardiovascular Disease and Molecular Intervention, Department of Pathophysiology, Nanjing Medical University (China)

    2012-11-16

    Highlights: Black-Right-Pointing-Pointer SIRT1 interacts with and deacetylates RFX5. Black-Right-Pointing-Pointer SIRT1 activation attenuates whereas SIRT1 inhibition enhances collagen repression by RFX5 in vascular smooth muscle cells. Black-Right-Pointing-Pointer SIRT1 promotes cytoplasmic localization and proteasomal degradation of RFX5 and cripples promoter recruitment of RFX5. Black-Right-Pointing-Pointer IFN-{gamma} represses SIRT1 expression in vascular smooth muscle cells. Black-Right-Pointing-Pointer SIRT1 agonist alleviates collagen repression by IFN-{gamma} in vascular smooth muscle cells. -- Abstract: Decreased expression of collagen by vascular smooth muscle cells (SMCs) within the atherosclerotic plaque contributes to the thinning of the fibrous cap and poses a great threat to plaque rupture. Elucidation of the mechanism underlying repressed collagen type I (COL1A2) gene would potentially provide novel solutions that can prevent rupture-induced complications. We have previously shown that regulatory factor for X-box (RFX5) binds to the COL1A2 transcription start site and represses its transcription. Here we report that SIRT1, an NAD-dependent, class III deacetylase, forms a complex with RFX5. Over-expression of SIRT1 or NAMPT, which synthesizes NAD+ to activate SIRT1, or treatment with the SIRT1 agonist resveratrol decreases RFX5 acetylation and disrupts repression of the COL1A2 promoter activity by RFX5. On the contrary, knockdown of SIRT1 or treatment with SIRT1 inhibitors induces RFX5 acetylation and enhances the repression of collagen transcription. SIRT1 antagonizes RFX5 activity by promoting its nuclear expulsion and proteasomal degradation hence dampening its binding to the COL1A2 promoter. The pro-inflammatory cytokine IFN-{gamma} represses COL1A2 transcription by down-regulating SIRT1 expression in SMCs. Therefore, our data have identified as novel pathway whereby SIRT1 maintains collagen synthesis in SMCs by modulating RFX5 activity.

  18. RhoA–Rho kinase and Platelet Activating Factor Stimulation of Ovine Fetal Pulmonary Vascular Smooth Muscle Cell Proliferation

    Science.gov (United States)

    Renteria, Lissette S.; Austin, Monique; Lazaro, Mariecon; Andrews, Mari Ashley; Lustina, Jennessee; Raj, J. Usha; Ibe, Basil O.

    2013-01-01

    Objectives Platelet Activating Factor (PAF) is produced by pulmonary vascular smooth muscle Cells (PVSMC). We studied effect of Rho kinase on PAF stimulation of PVSMC proliferation in an attempt to understand a role for RhoA/Rho kinase on PAF-induced ovine fetal pulmonary vascular remodeling. Our hypothesis is that PAF acts through Rho kinase, as one of its downstream signaling, to induce arterial (SMC-PA) and venous (SMC-PV) growth in the hypoxic lung environment of the fetus in utero. Materials and methods Rho kinase and MAPK effects on PAF receptor (PAFR)-mediated cell growth and PAFR expression were studied by DNA synthesis, Western and immunocytochemistry. Effects of constructs T19N and G14V on PAF-induced cell proliferation was also studied. Results Hypoxia increased PVSMC proliferation and the Rho kinase inhibitors, Y-27632 and Fasudil (HA-1077) as well as MAPK inhibitors PD 98059 and SB 203580 attenuated PAF stimulation of cell proliferation. RhoA T19N and G14V stimulated cell proliferation, but co-incubation with PAF did not affect proliferative effects of the constructs. PAFR protein expression was significantly down-regulated in both cell types by both Y-27632 and HA-1077 with comparable profiles. Also cells treated with Y-27632 showed less PAF receptor fluorescence with significant disruption of the cell morphology. Conclusions Our results show that Rho kinase nonspecifically modulates PAFR-mediated responses via a translational modification of PAFR protein and suggest that, in vivo, activation of Rho kinase by PAF may be one other pathway to sustain PAFR-mediated PVSMC growth. PMID:24033386

  19. Fetuin-A and albumin alter cytotoxic effects of calcium phosphate nanoparticles on human vascular smooth muscle cells.

    Directory of Open Access Journals (Sweden)

    Yana Dautova

    Full Text Available Calcification is a detrimental process in vascular ageing and in diseases such as atherosclerosis and arthritis. In particular, small calcium phosphate (CaP crystal deposits are associated with inflammation and atherosclerotic plaque de-stabilisation. We previously reported that CaP particles caused human vascular smooth muscle cell (VSMC death and that serum reduced the toxic effects of the particles. Here, we found that the serum proteins fetuin-A and albumin (≥ 1 µM reduced intracellular Ca2+ elevations and cell death in VSMCs in response to CaP particles. In addition, CaP particles functionalised with fetuin-A, but not albumin, were less toxic than naked CaP particles. Electron microscopic studies revealed that CaP particles were internalised in different ways; via macropinocytosis, membrane invagination or plasma membrane damage, which occurred within 10 minutes of exposure to particles. However, cell death did not occur until approximately 30 minutes, suggesting that plasma membrane repair and survival mechanisms were activated. In the presence of fetuin-A, CaP particle-induced damage was inhibited and CaP/plasma membrane interactions and particle uptake were delayed. Fetuin-A also reduced dissolution of CaP particles under acidic conditions, which may contribute to its cytoprotective effects after CaP particle exposure to VSMCs. These studies are particularly relevant to the calcification observed in blood vessels in patients with kidney disease, where circulating levels of fetuin-A and albumin are low, and in pathological situations where CaP crystal formation outweighs calcification-inhibitory mechanisms.

  20. Resveratrol Increases Serum BDNF Concentrations and Reduces Vascular Smooth Muscle Cells Contractility via a NOS-3-Independent Mechanism

    Directory of Open Access Journals (Sweden)

    Michał Wiciński

    2017-01-01

    Full Text Available Resveratrol is a polyphenol that presents both antineuroinflammatory properties and the ability to interact with NOS-3, what contributes to vasorelaxation. Brain-derived neurotrophic factor (BNDF, a molecule associated with neuroprotection in many neurodegenerative disorders, is considered as an important element of maintaining stable cerebral blood flow. Vascular smooth muscle cells (VSMCs are considered to be an important element in the pathogenesis of neurodegeneration and a potential preventative target by agents which reduce the contractility of the vessels. Our main objectives were to define the relationship between serum and long-term oral resveratrol administration in the rat model, as well as to assess the effect of resveratrol on phenylephrine- (PHE- induced contraction of vascular smooth muscle cells (VSMCs. Moreover, we attempt to define the dependence of contraction mechanisms on endothelial NO synthase. Experiments were performed on Wistar rats (n=17 pretreated with resveratrol (4 weeks; 10 mg/kg p.o. or placebo. Serum BDNF levels were quantified after 2 and 4 weeks of treatment with ELISA. Contraction force was measured on isolated and perfused tail arteries as the increase of perfusion pressure with a constant flow. Values of serum BNDF in week 0 were 1.18±0.12 ng/mL (treated and 1.17±0.13 ng/mL (control (p = ns. After 2 weeks of treatment, BDNF in the treatment group was higher than in controls, 1.52±0.23 ng/mL and 1.24±0.13 ng/mL, respectively. (p=0.02 Following 4 weeks of treatment, BDNF values were higher in the resveratrol group compared to control 1.64±0.31 ng/mL and 1.32±0.26 ng/mL, respectively (p=0.031. EC50 values obtained for PHE in resveratrol pretreated arteries were significantly higher than controls (5.33±1.7 × 10−7 M/L versus 4.53±1.2 × 10−8 M/L, p<0.05. These results show a significant increase in BDNF concentration in the resveratrol pretreated group. The reactivity of resistant

  1. Loss of MURC/Cavin-4 induces JNK and MMP-9 activity enhancement in vascular smooth muscle cells and exacerbates abdominal aortic aneurysm.

    Science.gov (United States)

    Miyagawa, Kotaro; Ogata, Takehiro; Ueyama, Tomomi; Kasahara, Takeru; Nakanishi, Naohiko; Naito, Daisuke; Taniguchi, Takuya; Hamaoka, Tetsuro; Maruyama, Naoki; Nishi, Masahiro; Kimura, Taizo; Yamada, Hiroyuki; Aoki, Hiroki; Matoba, Satoaki

    2017-06-03

    Abdominal aortic aneurysm (AAA) is relatively common in elderly patients with atherosclerosis. MURC (muscle-restricted coiled-coil protein)/Cavin-4 modulating the caveolae function of muscle cells is expressed in cardiomyocytes, skeletal muscle cells and smooth muscle cells. Here, we show a novel functional role of MURC/Cavin-4 in vascular smooth muscle cells (VSMCs) and AAA development. Both wild-type (WT) and MURC/Cavin-4 knockout (MURC -/- ) mice subjected to periaortic application of CaCl 2 developed AAAs. Six weeks after CaCl 2 treatment, internal and external aortic diameters were significantly increased in MURC -/- AAAs compared with WT AAAs, which were accompanied by advanced fibrosis in the tunica media of MURC -/- AAAs. The activity of JNK and matrix metalloproteinase (MMP) -2 and -9 were increased in MURC -/- AAAs compared with WT AAAs at 5 days after CaCl 2 treatment. At 6 weeks after CaCl 2 treatment, MURC -/- AAAs exhibited attenuated JNK activity compared with WT AAAs. There was no difference in the activity of MMP-2 or -9 between saline and CaCl 2 treatments. In MURC/Cavin-4-knockdown VSMCs, TNFα-induced activity of JNK and MMP-9 was enhanced compared with control VSMCs. Furthermore, WT, MURC -/- , apolipoprotein E -/- (ApoE -/- ), and MURC/Cavin-4 and ApoE double-knockout (MURC -/- ApoE -/- ) mice were subjected to angiotensin II (Ang II) infusion. In both ApoE -/- and MURC -/- ApoE -/- mice infused for 4 weeks with Ang II, AAAs were promoted. The internal aortic diameter was significantly increased in Ang II-infused MURC -/- ApoE -/- mice compared with Ang II-infused ApoE -/- mice. In MURC/Cavin-4-knockdown VSMCs, Ang II-induced activity of JNK and MMP-9 was enhanced compared with control VSMCs. Our results suggest that MURC/Cavin-4 in VSMCs modulates AAA progression at the early stage via the activation of JNK and MMP-9. MURC/Cavin-4 is a potential therapeutic target against AAA progression. Copyright © 2017 Elsevier Inc. All rights reserved.

  2. Fibroblast growth factor-2 induces osteogenic differentiation through a Runx2 activation in vascular smooth muscle cells

    Energy Technology Data Exchange (ETDEWEB)

    Nakahara, Takehiro; Sato, Hiroko; Shimizu, Takehisa; Tanaka, Toru; Matsui, Hiroki; Kawai-Kowase, Keiko; Sato, Mahito; Iso, Tatsuya; Arai, Masashi [Department of Medicine and Biological Science, Gunma University Graduate School of Medicine, 3-39-15 Showa-machi, Maebashi, Gunma 371-8511 (Japan); Kurabayashi, Masahiko, E-mail: mkuraba@med.gunma-u.ac.jp [Department of Medicine and Biological Science, Gunma University Graduate School of Medicine, 3-39-15 Showa-machi, Maebashi, Gunma 371-8511 (Japan)

    2010-04-02

    Expression of bone-associated proteins and osteoblastic transcription factor Runx2 in arterial cells has been implicated in the development of vascular calcification. However, the signaling upstream of the Runx2-mediated activation of osteoblastic program in vascular smooth muscle cells (VSMC) is poorly understood. We examined the effects of fibroblast growth factor-2 (FGF-2), an important regulator of bone formation, on osteoblastic differentiation of VSMC. Stimulation of cultured rat aortic SMC (RASMC) with FGF-2 induced the expression of the osteoblastic markers osteopontin (OPN) and osteocalcin. Luciferase assays showed that FGF-2 induced osteocyte-specific element (OSE)-dependent transcription. Downregulation of Runx2 by siRNA repressed the basal and FGF-2-stimulated expression of the OPN gene in RASMC. FGF-2 produced hydrogen peroxide in RASMC, as evaluated by fluorescent probe. Induction of OPN expression by FGF-2 was inhibited not only by PD98059 (MEK1 inhibitor) and PP1 (c-Src inhibitor), but also by an antioxidant, N-acetyl cysteine. Nuclear extracts from FGF-2-treated RASMC exhibited increased DNA-binding of Runx2 to its target sequence. Immunohistochemistry of human coronary atherectomy specimens and calcified aortic tissues showed that expression of FGF receptor-1 and Runx2 was colocalized. In conclusion, these results suggest that FGF-2 plays a role in inducing osteoblastic differentiation of VSMC by activating Runx2 through mitogen-activated protein kinase (MAPK)-dependent- and oxidative stress-sensitive-signaling pathways.

  3. Stem cell-derived vascular endothelial cells and their potential application in regenerative medicine

    Science.gov (United States)

    Although a 'vascular stem cell' population has not been identified or generated, vascular endothelial and mural cells (smooth muscle cells and pericytes) can be derived from currently known pluripotent stem cell sources, including human embryonic stem cells and induced pluripotent stem cells. We rev...

  4. Peptides PHI and VIP: comparison between vascular and nonvascular smooth muscle effect in rabbit uterus

    International Nuclear Information System (INIS)

    Bardrum, B.; Ottesen, B.; Fahrenkrug, J.

    1986-01-01

    The distribution and effects of the two neuropeptides, vasoactive intestinal polypeptide (VIP) and peptide histidine isoleucine amide (PHI), on vascular and nonvascular smooth muscle in the urogenital tract of nonpregnant rabbit female, were investigated. Immunoreactive VIP and PHI were present in all regions except the ovary with the highest concentration in the uterin cervix. By using in vitro tension recordings of myometrial specimens, it was demonstrated that both peptides displayed a dose-dependent inhibition of the mechanical activity. The dose-response curves of VIP and PHI were superimposable with and ID 50 of 3 x 10 -8 mol/l, and their combined effect was additive. In addition, the influence of the two peptides on myometrial blood flow (MBF) was investigated by the xenon-133 washout technique. Both peptides were found to increase MBF with the same potency and efficacy. Their combined effect was additive. In conclusion VIP and PHI are present in the rabbit urogenital tract, and the two peptides are equipotent inhibitors of mechanical nonvascular and vascular smooth muscle activity in the uterus

  5. Initiation and Propagation of Vascular Calcification Is Regulated by a Concert of Platelet- and Smooth Muscle Cell-Derived Extracellular Vesicles

    Directory of Open Access Journals (Sweden)

    Leon J. Schurgers

    2018-04-01

    Full Text Available The ageing population continues to suffer from its primary killer, cardiovascular disease (CVD. Despite recent advances in interventional medicinal and surgical therapies towards the end of the 20th century, the epidemic of cardiovascular disease has not been halted. Yet, rather than receding globally, the burden of CVD has risen to become a top cause of morbidity and mortality worldwide. Most CVD arises from thrombotic rupture of an atherosclerotic plaque, the pathologic thickening of coronary and carotid artery segments and subsequent distal ischemia in heart or brain. In fact, one-fifth of deaths are directly attributable to thrombotic rupture of a vulnerable plaque. Atherosclerotic lesion formation is caused by a concert of interactions between circulating leukocytes and platelets, interacting with the endothelial barrier, signalling into the arterial wall by the release of cytokines and extracellular vesicles (EVs. Both platelet- and cell-derived EVs represent a novel mechanism of cellular communication, particularly by the transport and transfer of cargo and by reprogramming of the recipient cell. These interactions result in phenotypic switching of vascular smooth muscle cells (VSMCs causing migration and proliferation, and subsequent secretion of EVs. Loss of VSMCs attracts perivascular Mesenchymal Stem Cells (MSCs from the adventitia, which are a source of VSMCs and contribute to repair after vascular injury. However, continuous stress stimuli eventually switch phenotype of cells into osteochondrogenic VSMCs facilitating vascular calcification. Although Virchow’s triad is over 100 years old, it is a reality that is accurate today. It can be briefly summarised as changes in the composition of blood (platelet EVs, alterations in the vessel wall (VSMC phenotypic switching, MSC infiltration and EV release and disruption of blood flow (atherothrombosis. In this paper, we review the latest relevant advances in the identification of

  6. Differential Cellular and Molecular Effects of Butyrate and Trichostatin A on Vascular Smooth Muscle Cells

    Directory of Open Access Journals (Sweden)

    Kasturi Ranganna

    2012-09-01

    Full Text Available The histone deacetylase (HDAC inhibitors, butyrate and trichostatin A (TSA, are epigenetic histone modifiers and proliferation inhibitors by downregulating cyclin D1, a positive cell cycle regulator, and upregulating p21Cip1 and INK family of proteins, negative cell cycle regulators. Our recent study indicated cyclin D1 upregulation in vascular smooth muscle cells (VSMC that are proliferation-arrested by butyrate. Here we investigate whether cyclin D1 upregulation is a unique response of VSMC to butyrate or a general response to HDAC inhibitors (HDACi by evaluating the effects of butyrate and TSA on VSMC. While butyrate and TSA inhibit VSMC proliferation via cytostatic and cytotoxic effects, respectively, they downregulate cdk4, cdk6, and cdk2, and upregulate cyclin D3, p21Cip1 and p15INK4B, and cause similar effects on key histone H3 posttranslational modifications. Conversely, cyclin D1 is upregulated by butyrate and inhibited by TSA. Assessment of glycogen synthase 3-dependent phosphorylation, subcellular localization and transcription of cyclin D1 indicates that differential effects of butyrate and TSA on cyclin D1 levels are linked to disparity in cyclin D1 gene expression. Disparity in butyrate- and TSA-induced cyclin D1 may influence transcriptional regulation of genes that are associated with changes in cellular morphology/cellular effects that these HDACi confer on VSMC, as a transcriptional modulator.

  7. Increased rhythmicity in hypertensive arterial smooth muscle is linked to transient receptor potential canonical channels

    DEFF Research Database (Denmark)

    Chen, Xiaoping; Yang, Dachun; Ma, Shuangtao

    2010-01-01

    Vasomotion describes oscillations of arterial vascular tone due to synchronized changes of intracellular calcium concentrations. Since increased calcium influx into vascular smooth muscle cells from spontaneously hypertensive rats (SHR) has been associated with variances of transient receptor pot...

  8. Apelin-13 upregulates Egr-1 expression in rat vascular smooth muscle cells through the PI3K/Akt and PKC signaling pathways

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Qi-Feng [Department of Cardiology, The First Affiliated Hospital of Liaoning Medical University, Jinzhou 121001 (China); Yu, Hong-Wei [Department of Cardiology, Jinzhou Central Hospital, Jinzhou 121001 (China); Sun, Li-Li [Department of Ophthalmology, The Third Affiliated Hospital of Liaoning Medical University, Jinzhou 121001 (China); You, Lu; Tao, Gui-Zhou [Department of Cardiology, The First Affiliated Hospital of Liaoning Medical University, Jinzhou 121001 (China); Qu, Bao-Ze, E-mail: qubaoze1971@hotmail.com [Department of Cardiology, The First Affiliated Hospital of Liaoning Medical University, Jinzhou 121001 (China)

    2015-12-25

    Previous studies have shown that Apelin-13 upregulates early growth response factor-1 (Egr-1) via the extracellular signal-regulated protein kinase (ERK) signaling pathway. Apelin-13 induces proliferation and migration of vascular smooth muscle cells (VSMCs) as well as the upregulation of osteopontin (OPN) via the upregulation of Egr-1. This study was designed to further explore the activity of Apelin-13 in VSMCs by investigating members of the mitogen-activated protein kinase (MAPK) family, in particular Jun kinase (JNK) and p38 mitogen-activated protein kinase (P38). We also examined whether the phosphatidylinositol 3 kinase (PI3K)/protein kinase B (Akt) and protein kinase C (PKC) signaling pathways were involved in the regulation of Egr-1 by Apelin-13. We treated rat aortic VSMCs with Apelin-13 and examined the expression of JNK, p-JNK, P38, and p-P38 to investigate whether Apelin-13-mediated increases in Egr-1 occurred through the JNK and P38 signaling pathways. We then pretreated VSMCs with the Gi protein inhibitor pertussis toxin (PTX) and the Gq inhibitor YM254890, added Apelin-13 and looked for changes in Egr-1 expression. Finally, we pretreated with the PI3K inhibitor LY294002 and the PKC inhibitor GF109203X, and treated with Apelin-13. Our results showed that JNK and P38 did not participate in Apelin-13-mediated increase in Egr-1. Instead, Apelin-13 upregulation of Egr-1 was mediated by a PTX-sensitive Gi protein. Apelin-13 did increase ERK phosphorylation through the PI3K/Akt and PKC signaling pathways, resulting in changes in Egr-1 expression. These data provide important targets for future studies to modulate vascular remodeling. - Highlights: • Apelin-13 mediates Egr-1 upregulation in vascular smooth muscle cells via ERK1/2. • The underlying mechanisms are unknown, but exclude Jnk or p38 pathway activation. • Apelin-13 binds to Gi, activating the PI3K/Akt and PKC signaling cascades. • Consequent ERK phosphorylation results in increased Egr-1

  9. Apelin-13 upregulates Egr-1 expression in rat vascular smooth muscle cells through the PI3K/Akt and PKC signaling pathways

    International Nuclear Information System (INIS)

    Liu, Qi-Feng; Yu, Hong-Wei; Sun, Li-Li; You, Lu; Tao, Gui-Zhou; Qu, Bao-Ze

    2015-01-01

    Previous studies have shown that Apelin-13 upregulates early growth response factor-1 (Egr-1) via the extracellular signal-regulated protein kinase (ERK) signaling pathway. Apelin-13 induces proliferation and migration of vascular smooth muscle cells (VSMCs) as well as the upregulation of osteopontin (OPN) via the upregulation of Egr-1. This study was designed to further explore the activity of Apelin-13 in VSMCs by investigating members of the mitogen-activated protein kinase (MAPK) family, in particular Jun kinase (JNK) and p38 mitogen-activated protein kinase (P38). We also examined whether the phosphatidylinositol 3 kinase (PI3K)/protein kinase B (Akt) and protein kinase C (PKC) signaling pathways were involved in the regulation of Egr-1 by Apelin-13. We treated rat aortic VSMCs with Apelin-13 and examined the expression of JNK, p-JNK, P38, and p-P38 to investigate whether Apelin-13-mediated increases in Egr-1 occurred through the JNK and P38 signaling pathways. We then pretreated VSMCs with the Gi protein inhibitor pertussis toxin (PTX) and the Gq inhibitor YM254890, added Apelin-13 and looked for changes in Egr-1 expression. Finally, we pretreated with the PI3K inhibitor LY294002 and the PKC inhibitor GF109203X, and treated with Apelin-13. Our results showed that JNK and P38 did not participate in Apelin-13-mediated increase in Egr-1. Instead, Apelin-13 upregulation of Egr-1 was mediated by a PTX-sensitive Gi protein. Apelin-13 did increase ERK phosphorylation through the PI3K/Akt and PKC signaling pathways, resulting in changes in Egr-1 expression. These data provide important targets for future studies to modulate vascular remodeling. - Highlights: • Apelin-13 mediates Egr-1 upregulation in vascular smooth muscle cells via ERK1/2. • The underlying mechanisms are unknown, but exclude Jnk or p38 pathway activation. • Apelin-13 binds to Gi, activating the PI3K/Akt and PKC signaling cascades. • Consequent ERK phosphorylation results in increased Egr-1

  10. Regulation of collagen biosynthesis in cultured bovine aortic smooth muscle cells

    International Nuclear Information System (INIS)

    Stepp, M.A.

    1986-01-01

    Aortic smooth muscles cells have been implicated in the etiology of lesions which occur in atherosclerosis and hypertension. Both diseases involve proliferation of smooth muscle cells and accumulation of excessive amounts of extracellular matrix proteins, including collagen type I and type III produced by the smooth muscle cells. To better understand the sites of regulation of collagen biosynthesis and to correlate these with the growth rate of the cells, cultured bovine aortic smooth muscle cells were studied as a function of the number of days (3 to 14) in second passage. Cells grew rapidly up to day 6 when confluence was reached. The total incorporation of [ 3 H]-proline into proteins was highest at day 3 and decreased to a constant level after the cultures reached confluence. In contrast, collagen protein production was lowest before confluence and continued to increase over the entire time course of the experiments. cDNA clones for the α1 and α2 chains of type I and the α1 chain of type III collagen were used to quantitate the steady state level of collagen mRNAs. RNA was tested in a cell-free translation system. Changes in the translational activity of collagen mRNAs parallelled the observed increases in collagen protein production. Thus, at later time points, collagen mRNAs are more active in directing synthesis of preprocollagens, even though less collagen mRNA is present. The conclusion is that the site of regulation of the expression of collagen genes is a function of the growth rate of cultured smooth muscle cells

  11. BeWo cells stimulate smooth muscle cell apoptosis and elastin breakdown in a model of spiral artery transformation

    OpenAIRE

    Harris, L. K.; Keogh, R. J.; Wareing, M.; Baker, P. N.; Cartwright, J. E.; Whitley, G. S.; Aplin, J. D.

    2007-01-01

    BACKGROUND: During pregnancy, extravillous trophoblast invades the uterine wall and enters the spiral arteries. Remodelling ensues, with loss of vascular smooth muscle cells (SMCs) to create high flow, low resistance vessels. Pregnancies complicated by pre-eclampsia are characterized by incomplete arterial remodelling. Endovascular trophoblast is not easily accessible for studies to establish the pathogenesis of pre-eclampsia, so we have developed a model appropriate to carry out mechanistic ...

  12. Effect of 103Pd on proliferation and apoptosis of vascular smooth muscle cells

    International Nuclear Information System (INIS)

    Luo Quanyong; Zhu Jun; Lu Hankui; Zhu Ruisen

    2003-01-01

    This study aimed at the effect of γ-emitting radionuclide 103 Pd on the proliferation and apoptosis of vascular SMCs (smooth muscle cells) in vitro. The cavy aortic SMCs were cultured with culture medium M-199. The experiments were carried out in two groups, one for proliferation test and the other for apoptosis test. In each group, 103 Pd solutions with various radioactivities were respectively added to the culture solution to irradiate SMCs for 72 h, while non-radioactive palladium solution was added to the control. 3 H-thymidine incorporation test and liquid scintillator were used to detect the effect of 103 Pd on the proliferation of SMCs. Flow cytometer was used to detect the apoptotic SMCs. The inhibition rate of SMCs proliferation by 1.85 MBq 103 Pd solution was 2.3%, which was not significant, while the inhibition rate increased from 41.6% to 91.3% as the 103 Pd activity increased from 7.40 MBq to 37 MBq. The apoptosis rate of SMCs was extremely low (less than 4.0%) by 103 Pd with activity from 1.85 MBq to 37 MBq. The results suggest that the proliferation of SMCs can be repressed effectively in a dose-dependent fashion by 103 Pd in vitro. The mechanism of its inhibiting over neointima proliferation is likely to inhibit SMCs proliferation rather than to induce its apoptosis by 103 Pd. 103 Pd can be used as a γ-emitting intravascular brachytherapy radionuclide to inhibit SMCs proliferation

  13. Platelet-derived growth factor regulates K-Cl cotransport in vascular smooth muscle cells.

    Science.gov (United States)

    Zhang, Jing; Lauf, Peter K; Adragna, Norma C

    2003-03-01

    Platelet-derived growth factor (PDGF), a potent serum mitogen for vascular smooth muscle cells (VSMCs), plays an important role in membrane transport regulation and in atherosclerosis. K-Cl cotransport (K-Cl COT/KCC), the coupled-movement of K and Cl, is involved in ion homeostasis. VSMCs possess K-Cl COT activity and the KCC1 and KCC3 isoforms. Here, we report on the effect of PDGF on K-Cl COT activity and mRNA expression in primary cultures of rat VSMCs. K-Cl COT was determined as the Cl-dependent Rb influx and mRNA expression by semiquantitative RT-PCR. Twenty four-hour serum deprivation inhibited basal K-Cl COT activity. Addition of PDGF increased total protein content and K-Cl COT activity in a time-dependent manner. PDGF activated K-Cl COT in a dose-dependent manner, both acutely (10 min) and chronically (12 h). AG-1296, a selective inhibitor of the PDGF receptor tyrosine kinase, abolished these effects. Actinomycin D and cycloheximide had no effect on the acute PDGF activation of K-Cl COT, suggesting posttranslational regulation by the drug. Furthermore, PDGF increased KCC1 and decreased KCC3 mRNA expression in a time-dependent manner. These results indicate that chronic activation of K-Cl COT activity by PDGF may involve regulation of the two KCC mRNA isoforms, with KCC1 playing a dominant role in the mechanism of PDGF-mediated activation.

  14. Overexpression of Catalase in Vascular Smooth Muscle Cells Prevents the Formation of Abdominal Aortic Aneurysms

    Science.gov (United States)

    Parastatidis, Ioannis; Weiss, Daiana; Joseph, Giji; Taylor, W Robert

    2013-01-01

    Objective Elevated levels of oxidative stress have been reported in abdominal aortic aneurysms (AAA), but which reactive oxygen species (ROS) promotes the development of AAA remains unclear. Here we investigate the effect of the hydrogen peroxide (H2O2) degrading enzyme catalase on the formation of AAA. Approach and Results AAA were induced with the application of calcium chloride (CaCl2) on mouse infrarenal aortas. The administration of PEG-catalase, but not saline, attenuated the loss of tunica media and protected against AAA formation (0.91±0.1 mm vs. 0.76±0.09 mm). Similarly, in a transgenic mouse model, catalase over-expression in the vascular smooth muscle cells (VSMC) preserved the thickness of tunica media and inhibited aortic dilatation by 50% (0.85±0.14 mm vs. 0.57±0.08 mm). Further studies showed that injury with CaCl2 decreased catalase expression and activity in the aortic wall. Pharmacologic administration or genetic over-expression of catalase restored catalase activity and subsequently decreased matrix metalloproteinase activity. In addition, a profound reduction in inflammatory markers and VSMC apoptosis was evident in aortas of catalase over-expressing mice. Interestingly, as opposed to infusion of PEG-catalase, chronic over-expression of catalase in VSMC did not alter the total aortic H2O2 levels. Conclusions The data suggest that a reduction in aortic wall catalase activity can predispose to AAA formation. Restoration of catalase activity in the vascular wall enhances aortic VSMC survival and prevents AAA formation primarily through modulation of matrix metalloproteinase activity. PMID:23950141

  15. Betulinic Acid Inhibits Growth of Cultured Vascular Smooth Muscle Cells In Vitro by Inducing G1 Arrest and Apoptosis

    Directory of Open Access Journals (Sweden)

    Raja Kumar Vadivelu

    2012-01-01

    Full Text Available Betulinic acid is a widely available plant-derived triterpene which is reported to possess selective cytotoxic activity against cancer cells of neuroectodermal origin and leukemia. However, the potential of betulinic acid as an antiproliferative and cytotoxic agent on vascular smooth muscle (VSMC is still unclear. This study was carried out to demonstrate the antiproliferative and cytotoxic effect of betulinic acid on VSMCs using 3-[4,5-dimethylthizol-2-yl]-2,5-diphenyltetrazolium bromide (MTT assay, flow cytometry cell cycle assay, BrdU proliferation assay, acridine orange/propidium iodide staining, and comet assay. Result from MTT and BrdU assays indicated that betulinic acid was able to inhibit the growth and proliferation of VSMCs in a dose-dependent manner with IC50 of 3.8 μg/mL significantly (P<0.05. Nevertheless, betulinic acid exhibited G1 cell cycle arrest in flow cytometry cell cycle profiling and low level of DNA damage against VSMC in acridine orange/propidium iodide and comet assay after 24 h of treatment. In conclusion, betulinic acid induced G1 cell cycle arrest and dose-dependent DNA damage on VSMC.

  16. Temperature and nucleotide dependence of calcium release by myo-inositol 1,4,5-trisphosphate in cultured vascular smooth muscle cells

    International Nuclear Information System (INIS)

    Smith, J.B.; Smith, L.; Higgins, B.L.

    1985-01-01

    Inositol 1,4,5-trisphosphate (IP3) rapidly increased 45 Ca 2+ efflux from a nonmitochondrial organelle in cultured vascular smooth muscle cells that were permeabilized with saponin. A nucleotide, preferably ATP, was essential for IP3-evoked 45 Ca 2+ release. Two nonhydrolyzable ATP analogues satisfied the nucleotide requirement for IP3-evoked 45 Ca 2+ release. IP3 strongly stimulated 45 Ca 2+ efflux at low temperatures (1 to 15 degrees C). Decreasing the temperature from 37 to 4 degrees C inhibited the rate of IP3-stimulated efflux by only about 33%. The failure of such low temperatures to strongly inhibit IP3-induced 45 Ca 2+ efflux suggests that IP3 activated a Ca 2+ channel, rather than a carrier, by a ligand-binding, rather than a metabolic, reaction

  17. A 310-bp minimal promoter mediates smooth muscle cell-specific expression of telokin.

    Science.gov (United States)

    Smith, A F; Bigsby, R M; Word, R A; Herring, B P

    1998-05-01

    A cell-specific promoter located in an intron of the smooth muscle myosin light chain kinase gene directs transcription of telokin exclusively in smooth muscle cells. Transgenic mice were generated in which a 310-bp rabbit telokin promoter fragment, extending from -163 to +147, was used to drive expression of simian virus 40 large T antigen. Smooth muscle-specific expression of the T-antigen transgene paralleled that of the endogenous telokin gene in all smooth muscle tissues except uterus. The 310-bp promoter fragment resulted in very low levels of transgene expression in uterus; in contrast, a transgene driven by a 2.4-kb fragment (-2250 to +147) resulted in high levels of transgene expression in uterine smooth muscle. Telokin expression levels correlate with the estrogen status of human myometrial tissues, suggesting that deletion of an estrogen response element (ERE) may account for the low levels of transgene expression driven by the 310-bp rabbit telokin promoter in uterine smooth muscle. Experiments in A10 smooth muscle cells directly showed that reporter gene expression driven by the 2.4-kb, but not 310-bp, promoter fragment could be stimulated two- to threefold by estrogen. This stimulation was mediated through an ERE located between -1447 and -1474. Addition of the ERE to the 310-bp fragment restored estrogen responsiveness in A10 cells. These data demonstrate that in addition to a minimal 310-bp proximal promoter at least one distal cis-acting regulatory element is required for telokin expression in uterine smooth muscle. The distal element may include an ERE between -1447 and -1474.

  18. Mitofusin2 decreases intracellular cholesterol of oxidized LDL-induced foam cells from rat vascular smooth muscle cells.

    Science.gov (United States)

    He, Chao; Chen, Ying; Liu, Chun; Cao, Ming; Fan, Yu-jin; Guo, Xiao-mei

    2013-04-01

    Mitofusin2 (Mfn2) plays a pivotal role in the proliferation and apoptosis of vascular smooth muscle cells (VSMCs). The purpose of this study was to investigate the effects of Mfn2 on the trafficking of intracellular cholesterol in the foam cells derived from rat VSMCs (rVSMCs) and also to investigate the effects of Mfn2 on the expression of adenosine triphosphate-binding cassette subfamily A member 1 (ABCA1), adenosine triphosphate-binding cassette subfamily G member 1 (ABCG1) and peroxisome proliferator-activated receptor gamma (PPARγ). The rVSMCs were co-cultured with oxidized low density lipoprotein (LDL, 80 μg/mL) to produce foam cells and cholesterol accumulation in cells. Before oxidized LDL treatment, different titers (20, 40 and 60 pfu/cell) of recombinant adenovirus containing Mfn2 gene (Adv-Mfn2) were added into the culture medium for 24 h to transfect the Mfn2 gene into the rVSMCs. Then the cells were harvested for analyses. The protein expression of Mfn2 was significantly higher in Adv-Mfn2-transfected group than in untransfected group (PLDL treatment, rVSMCs became irregular and their nuclei became larger, and their plasma abounded with red lipid droplets. However, the number of red lipid droplets was significantly decreased in Adv-Mfn2-transfected group as compared with untransfected group. At 48 h after oxidized LDL treatment, the intracellular cholesterol in rVSMCs was significantly increased (P0.05), the phosporylation levels of PPARγ were significantly decreased in Adv-Mfn2-transfected group as compared with untransfected group (Pcholesterol in oxidized LDL-induced rVSMCs possibly by decreasing PPARγ phosporylation and then increasing protein expression levels of ABCA1 and ABCG1, which may be helpful to suppress the formation of foam cells.

  19. MiR-29-mediated elastin down-regulation contributes to inorganic phosphorus-induced osteoblastic differentiation in vascular smooth muscle cells.

    Science.gov (United States)

    Sudo, Ryo; Sato, Fumiaki; Azechi, Takuya; Wachi, Hiroshi

    2015-12-01

    Vascular calcification increases the risk of cardiovascular mortality. We previously reported that expression of elastin decreases with progression of inorganic phosphorus (Pi)-induced vascular smooth muscle cell (VSMC) calcification. However, the regulatory mechanisms of elastin mRNA expression during vascular calcification remain unclear. MicroRNA-29 family members (miR-29a, b and c) are reported to mediate elastin mRNA expression. Therefore, we aimed to determine the effect of miR-29 on elastin expression and Pi-induced vascular calcification. Calcification of human VSMCs was induced by Pi and evaluated measuring calcium deposition. Pi stimulation promoted Ca deposition and suppressed elastin expression in VSMCs. Knockdown of elastin expression by shRNA also promoted Pi-induced VSMC calcification. Elastin pre-mRNA measurements indicated that Pi stimulation suppressed elastin expression without changing transcriptional activity. Conversely, Pi stimulation increased miR-29a and miR-29b expression. Inhibition of miR-29 recovered elastin expression and suppressed calcification in Pi-treated VSMCs. Furthermore, over-expression of miR-29b promoted Pi-induced VSMC calcification. RT-qPCR analysis showed knockdown of elastin expression in VSMCs induced expression of osteoblast-related genes, similar to Pi stimulation, and recovery of elastin expression by miR-29 inhibition reduced their expression. Our study shows that miR-29-mediated suppression of elastin expression in VSMCs plays a pivotal role in osteoblastic differentiation leading to vascular calcification. © 2015 The Molecular Biology Society of Japan and Wiley Publishing Asia Pty Ltd.

  20. Effect of 125I seeds and 103Pd stents on proliferation of vascular smooth muscle cells

    International Nuclear Information System (INIS)

    Zhu Jun; Zhu Ruisen

    2004-01-01

    To establish the theoretical and practical base for implementing radioactive stents aft PTCA in order to prevent restenosis, in vitro observation was taken over the effects of 12 '5I-seeds and 103 Pd-implanted stents on the vascular smooth muscle cell (VSMC) proliferation. In vitro VSMC model from guinea-pig aortic arteries was established using adherent cell culture methods. The effects of 125 I-seeds and 103 Pd-implanted stents on the VSMC proliferation, with or without fetal bovine serum (FCS), were investigated through cell counting methods and 3 H-TDR implementation tests. It was shown that (1) 10% FCS significantly promoted the DNA synthesis of VSMC (P 125 I-seeds and 103 Pd-implanted stents inhibited the VSMC DNA synthesis in dose-dependent manner, regardless of 10% FCS inducement. At lower radioactive doses, neither 125 I-seeds (18.5-74 kBq) nor 103 Pd-implanted stents (1.48-2.96 MBq) exhibited distinctive effects on the VSMC DNA synthesis (P>0.05); and (3) 48 hour exposure from 125 I-seeds at 128 kBq or 10 '3Pd-implanted stents at 7.4 MBq did not result in VSMC morphological alteration, but 125 I-seeds at 370 kBq caused cells' morphological changes. Therefore both 125 I-seeds and 103 Pd-implanted stents inhibit the in vitro VSMC DNA synthesis, and the inhibition effects are significantly related to their exposure duration and doses. (authors)

  1. Retinoid-induced expression and activity of an immediate early tumor suppressor gene in vascular smooth muscle cells.

    Directory of Open Access Journals (Sweden)

    Jeffrey W Streb

    2011-04-01

    Full Text Available Retinoids are used clinically to treat a number of hyper-proliferative disorders and have been shown in experimental animals to attenuate vascular occlusive diseases, presumably through nuclear receptors bound to retinoic acid response elements (RARE located in target genes. Here, we show that natural or synthetic retinoids rapidly induce mRNA and protein expression of a specific isoform of A-Kinase Anchoring Protein 12 (AKAP12β in cultured smooth muscle cells (SMC as well as the intact vessel wall. Expression kinetics and actinomycin D studies indicate Akap12β is a retinoid-induced, immediate-early gene. Akap12β promoter analyses reveal a conserved RARE mildly induced with atRA in a region that exhibits hyper-acetylation. Immunofluorescence microscopy and protein kinase A (PKA regulatory subunit overlay assays in SMC suggest a physical association between AKAP12β and PKA following retinoid treatment. Consistent with its designation as a tumor suppressor, inducible expression of AKAP12β attenuates SMC growth in vitro. Further, immunohistochemistry studies establish marked decreases in AKAP12 expression in experimentally-injured vessels of mice as well as atheromatous lesions in humans. Collectively, these results demonstrate a novel role for retinoids in the induction of an AKAP tumor suppressor that blocks vascular SMC growth thus providing new molecular insight into how retiniods may exert their anti-proliferative effects in the injured vessel wall.

  2. Resveratrol prevents angiotensin II-induced hypertrophy of vascular smooth muscle cells through the transactivation of growth factor receptors.

    Science.gov (United States)

    Hossain, Ekhtear; Anand-Srivastava, Madhu B

    2017-08-01

    We previously showed that augmented levels of endogenous angiotensin II (AngII) contribute to vascular smooth muscle cell (VSMC) hypertrophy through the transactivation of growth factor receptors in spontaneously hypertensive rats. Resveratrol (RV), a polyphenolic component of red wine, has also been shown to attenuate AngII-evoked VSMC hypertrophy; however, the molecular mechanism mediating this response is obscure. The present study was therefore undertaken to examine whether RV could prevent AngII-induced VSMC hypertrophy through the transactivation of growth factor receptor and associated signaling pathways. AngII treatment of VSMC enhanced the protein synthesis that was attenuated towards control levels by RV pretreatment as well as by the inhibitors of NADPH oxidase, c-Src, and growth factor receptors. Furthermore, RV pretreatment also inhibited enhanced levels of superoxide anion, NADPH oxidase activity, increased expression of NADPH oxidase subunits, and phosphorylation of c-Src, EGF-R, PDGE-R, ERK1/2, and AKT1/2. In conclusion, these results indicate that RV attenuates AngII-induced VSMC hypertrophy through the inhibition of enhanced oxidative stress and activation of c-Src, growth factor receptors, and MAPK/AKT signaling. We suggest that RV could be used as a therapeutic agent in the treatment of vascular complications associated with hypertension and hypertrophy.

  3. Electrospun silk fibroin/poly (L-lactide-ε-caplacton) graft with platelet-rich growth factor for inducing smooth muscle cell growth and infiltration.

    Science.gov (United States)

    Yin, Anlin; Bowlin, Gary L; Luo, Rifang; Zhang, Xingdong; Wang, Yunbing; Mo, Xiumei

    2016-12-01

    The construction of a smooth muscle layer for blood vessel through electrospinning method plays a key role in vascular tissue engineering. However, smooth muscle cells (SMCs) penetration into the electrospun graft to form a smooth muscle layer is limited due to the dense packing of fibers and lack of inducing factors. In this paper, silk fibroin/poly (L-lactide-ε-caplacton) (SF/PLLA-CL) vascular graft loaded with platelet-rich growth factor (PRGF) was fabricated by electrospinning. The in vitro results showed that SMCs cultured in the graft grew fast, and the incorporation of PRGF could induce deeper SMCs infiltrating compared to the SF/PLLA-CL graft alone. Mechanical properties measurement showed that PRGF-incorporated graft had proper tensile stress, suture retention strength, burst pressure and compliance which could match the demand of native blood vessel. The success in the fabrication of PRGF-incorporated SF/PLLA-CL graft to induce fast SMCs growth and their strong penetration into graft has important application for tissue-engineered blood vessels.

  4. Effects of gamma rays on rat vascular smooth muscle fibers

    Energy Technology Data Exchange (ETDEWEB)

    Ghassan, A [Radio-Biology and Health Dept. Syrian Atomic Energy Commission, (Syrian Arab Republic)

    1995-10-01

    Modifications of the Vasomotoricity induced by gamma rays have been investigated. Vascular smooth muscle fibres (VSMF) of rat portal vein have been used in this study. Irradiation procedures using a {sup 60} Co source have been carried out as follows: - Whole body irradiation. - Irradiation of isolated portal vein and isolated VSMF. Our results show that : 1-irradiation reduces the functional competition between Mg{sup 2+} and Ca{sup 2+}, thus hyper magnetic Krebs solutions have a negligible effect on irradiated VSMF. 2- irradiation activates Ca{sup 2+} influx into the VSMF. Thus the effect of hypocalcemic solutions on irradiated VSMF is minor compared with control. 3- Hyperpotassic solutions provoke titanic contractions with high amplitude on the irradiated VSMF compared with control. 5 figs.

  5. Long Non-Coding RNA MEG3 Downregulation Triggers Human Pulmonary Artery Smooth Muscle Cell Proliferation and Migration via the p53 Signaling Pathway

    Directory of Open Access Journals (Sweden)

    Zengxian Sun

    2017-08-01

    Full Text Available Background/Aims: Increasing evidence has demonstrated a significant role of long non-coding RNAs (lncRNAs in diverse biological processes, and many of which are likely to have functional roles in vascular remodeling. However, their functions in pulmonary arterial hypertension (PAH remain largely unknown. Pulmonary vascular remodeling is an important pathological feature of PAH, leading to increased vascular resistance and reduced compliance. Pulmonary artery smooth muscle cells (PASMCs dysfunction is involved in vascular remodeling. Long noncoding RNAs are potential regulators of PASMCs function. Herein, we determined whether long noncoding RNA–maternally expressed gene 3 (MEG3 was involved in PAH-related vascular remodeling. Methods: The arterial wall thickness was examined by hematoxylin and eosin (H&E staining in distal pulmonary arteries (PAs isolated from lungs of healthy volunteers and PAH patients. The expression level of MEG3 was analyzed by qPCR. The effects of MEG3 on human PASMCs were assessed by cell counting Kit-8 assay, BrdU incorporation assay, flow cytometry, scratch-wound assay, immunofluorescence, and western blotting in human PASMCs. Results: We revealed that the expression of MEG3 was significantly downregulated in lung and PAs of patients with PAH. MEG3 knockdown affected PASMCs proliferation and migration in vitro. Moreover, inhibition of MEG3 regulated the cell cycle progression and made more smooth muscle cells from the G0/G1 phase to the G2/M+S phase and the process could stimulate the expression of PCNA, Cyclin A and Cyclin E. In addition, we found that the p53 pathway was involved in MEG3–induced smooth muscle cell proliferation. Conclusions: This study identified MEG3 as a critical regulator in PAH and demonstrated the potential of gene therapy and drug development for treating PAH.

  6. Micro- and nanostructured Al{sub 2}O{sub 3} surfaces for controlled vascular endothelial and smooth muscle cell adhesion and proliferation

    Energy Technology Data Exchange (ETDEWEB)

    Aktas, Cenk, E-mail: cenk.aktas@inm-gmbh.de [INM - Leibniz Institute for New Materials, CVD/Biosurfaces Division, 66123 Saarbruecken (Germany); Doerrschuck, Eva; Schuh, Cathrin [Clinic of Paediatric Cardiology, Saarland University, Building 9, 66424 Homburg (Germany); Miro, Marina Martinez; Lee, Juseok [INM - Leibniz Institute for New Materials, CVD/Biosurfaces Division, 66123 Saarbruecken (Germany); Puetz, Norbert; Wennemuth, Gunther [Department of Anatomy and Cell Biology, Saarland University, Building 61, 66424 Homburg (Germany); Metzger, Wolfgang; Oberringer, Martin [Department of Trauma-, Hand- and Reconstructive Surgery, Saarland University, Building 57, 66424 Homburg (Germany); Veith, Michael [INM - Leibniz Institute for New Materials, CVD/Biosurfaces Division, 66123 Saarbruecken (Germany); Department of Inorganic Chemistry, University of Saarland, Building C 4 1, 66123 Saarbruecken (Germany); Abdul-Khaliq, Hashim [Clinic of Paediatric Cardiology, Saarland University, Building 9, 66424 Homburg (Germany)

    2012-07-01

    The effect of the micro- and nanotopography on vascular cell-surface interaction is investigated using nano- and microstructured Al{sub 2}O{sub 3} as model substrate. Two different nanostructured Al{sub 2}O{sub 3} surfaces composed of low density (LD) and high density (HD) nanowires (NWs) were synthesized by chemical vapour deposition (CVD) and commercially available microstructured Al{sub 2}O{sub 3} plates were used for comparison. A clear diverging response of human umbilical vein endothelial cells (HUVEC) and human umbilical vein smooth muscle cells (HUVSMC) was observed on these nano- and microstructured surfaces. LD Al{sub 2}O{sub 3} NWs seem to enhance the proliferation of HUVECs selectively. This selective control of the cell-surface interaction by topography may represent a key issue for the future stent material design. - Highlights: Black-Right-Pointing-Pointer Nanostructured alumina surfaces triggers selective adhesion and proliferation of endothelial cells. Black-Right-Pointing-Pointer Catalyst free synthesis of nanowires. Black-Right-Pointing-Pointer Topography induces selective cell response.

  7. The nanostructure of myoendothelial junctions contributes to signal rectification between endothelial and vascular smooth muscle cells

    DEFF Research Database (Denmark)

    Brasen, Jens Christian; Jacobsen, Jens Christian Brings; von Holstein-Rathlou, Niels-Henrik

    2012-01-01

    Micro-anatomical structures in tissues have potential physiological effects. In arteries and arterioles smooth muscle cells and endothelial cells are separated by the internal elastic lamina, but the two cell layers often make contact through micro protrusions called myoendothelial junctions. Cross...... types and the myoendothelial junction. The model is implemented as a 2D axi-symmetrical model and solved using the finite element method. We have simulated diffusion of Ca(2+) and IP(3) between the two cell types and we show that the micro-anatomical structure of the myoendothelial junction in itself...

  8. Effect of uric acid on inflammatory COX-2 and ROS pathways in vascular smooth muscle cells.

    Science.gov (United States)

    Oğuz, Nurgül; Kırça, Mustafa; Çetin, Arzu; Yeşilkaya, Akın

    2017-10-01

    Hyperuricemia is thought to play a role in cardiovascular diseases (CVD), including hypertension, coronary artery disease and atherosclerosis. However, exactly how uric acid contributes to these pathologies is unknown. An underlying mechanism of inflammatory diseases, such as atherosclerosis, includes enhanced production of cyclooxygenase-2 (COX-2) and superoxide anion. Here, we aimed to examine the effect of uric acid on inflammatory COX-2 and superoxide anion production and to determine the role of losartan. Primarily cultured vascular smooth muscle cells (VSMCs) were time and dose-dependently induced by uric acid and COX-2 and superoxide anion levels were measured. COX-2 levels were determined by ELISA, and superoxide anion was measured by the superoxide dismutase (SOD)-inhibitable reduction of ferricytochrome c method. Uric acid elevated COX-2 levels in a time-dependent manner. Angiotensin-II receptor blocker, losartan, diminished uric-acid-induced COX-2 elevation. Uric acid also increased superoxide anion level in VSMCs. Uric acid plays an important role in CVD pathogenesis by inducing inflammatory COX-2 and ROS pathways. This is the first study demonstrating losartan's ability to reduce uric-acid-induced COX-2 elevation.

  9. Divergent effects of 17-{beta}-estradiol on human vascular smooth muscle and endothelial cell function diminishes TNF-{alpha}-induced neointima formation

    Energy Technology Data Exchange (ETDEWEB)

    Nintasen, Rungrat [Division of Cardiovascular Medicine, Leeds Institute of Genetics, Health and Therapeutics, University of Leeds, Leeds LS2 9JT (United Kingdom); Multidisciplinary Cardiovascular Research Center (MCRC), University of Leeds, Leeds LS2 9JT (United Kingdom); Department of Tropical Pathology, Faculty of Tropical Medicine, Mahidol University (Thailand); Riches, Kirsten; Mughal, Romana S. [Division of Cardiovascular Medicine, Leeds Institute of Genetics, Health and Therapeutics, University of Leeds, Leeds LS2 9JT (United Kingdom); Multidisciplinary Cardiovascular Research Center (MCRC), University of Leeds, Leeds LS2 9JT (United Kingdom); Viriyavejakul, Parnpen; Chaisri, Urai; Maneerat, Yaowapa [Department of Tropical Pathology, Faculty of Tropical Medicine, Mahidol University (Thailand); Turner, Neil A. [Division of Cardiovascular Medicine, Leeds Institute of Genetics, Health and Therapeutics, University of Leeds, Leeds LS2 9JT (United Kingdom); Multidisciplinary Cardiovascular Research Center (MCRC), University of Leeds, Leeds LS2 9JT (United Kingdom); Porter, Karen E., E-mail: medkep@leeds.ac.uk [Division of Cardiovascular Medicine, Leeds Institute of Genetics, Health and Therapeutics, University of Leeds, Leeds LS2 9JT (United Kingdom); Multidisciplinary Cardiovascular Research Center (MCRC), University of Leeds, Leeds LS2 9JT (United Kingdom)

    2012-04-20

    Highlights: Black-Right-Pointing-Pointer TNF-{alpha} augments neointimal hyperplasia in human saphenous vein. Black-Right-Pointing-Pointer TNF-{alpha} induces detrimental effects on endothelial and smooth muscle cell function. Black-Right-Pointing-Pointer Estradiol exerts modulatory effects on TNF-induced vascular cell functions. Black-Right-Pointing-Pointer The modulatory effects of estradiol are discriminatory and cell-type specific. -- Abstract: Coronary heart disease (CHD) is a condition characterized by increased levels of proinflammatory cytokines, including tumor necrosis factor-{alpha} (TNF-{alpha}). TNF-{alpha} can induce vascular endothelial cell (EC) and smooth muscle cell (SMC) dysfunction, central events in development of neointimal lesions. The reduced incidence of CHD in young women is believed to be due to the protective effects of estradiol (E2). We therefore investigated the effects of TNF-{alpha} on human neointima formation and SMC/EC functions and any modulatory effects of E2. Saphenous vein (SV) segments were cultured in the presence of TNF-{alpha} (10 ng/ml), E2 (2.5 nM) or both in combination. Neointimal thickening was augmented by incubation with TNF-{alpha}, an effect that was abolished by co-culture with E2. TNF-{alpha} increased SV-SMC proliferation in a concentration-dependent manner that was optimal at 10 ng/ml (1.5-fold increase), and abolished by E2 at all concentrations studied (1-50 nM). Surprisingly, E2 itself at low concentrations (1 and 5 nM) stimulated SV-SMC proliferation to a level comparable to that of TNF-{alpha} alone. SV-EC migration was significantly impaired by TNF-{alpha} (42% of control), and co-culture with E2 partially restored the ability of SV-EC to migrate and repair the wound. In contrast, TNF-{alpha} increased SV-SMC migration by 1.7-fold, an effect that was completely reversed by co-incubation with E2. Finally, TNF-{alpha} potently induced ICAM-1 and VCAM-1 expression in both SV-EC and SV-SMC. However there

  10. Identification and characterization of [6]-shogaol from ginger as inhibitor of vascular smooth muscle cell proliferation.

    Science.gov (United States)

    Liu, Rongxia; Heiss, Elke H; Sider, Nadine; Schinkovitz, Andreas; Gröblacher, Barbara; Guo, Dean; Bucar, Franz; Bauer, Rudolf; Dirsch, Verena M; Atanasov, Atanas G

    2015-05-01

    Vascular smooth muscle cell (VSMC) proliferation is involved in the pathogenesis of cardiovascular disease, making the identification of new counteracting agents and their mechanisms of action relevant. Ginger and its constituents have been reported to improve cardiovascular health, but no studies exist addressing a potential interference with VSMC proliferation. The dichloromethane extract of ginger inhibited VSMC proliferation when monitored by resazurin metabolic conversion (IC50 = 2.5 μg/mL). The examination of major constituents from ginger yielded [6]-shogaol as the most active compound (IC50 = 2.7 μM). In the tested concentration range [6]-shogaol did not exhibit cytotoxicity toward VSMC and did not interfere with endothelial cell proliferation. [6]-shogaol inhibited DNA synthesis and induced accumulation of the VSMC in the G0 /G1 cell-cycle phase accompanied with activation of the nuclear factor-erythroid 2-related factor 2 (Nrf2)/HO-1 pathway. Since [6]-shogaol lost its antiproliferative activity in the presence of the heme oxygenase-1 (HO-1) inhibitor tin protoporphyrin IX, HO-1 induction appears to contribute to the antiproliferative effect. This study demonstrates for the first time inhibitory potential of ginger constituents on VSMC proliferation. The presented data suggest that [6]-shogaol exerts its antiproliferative effect through accumulation of cells in the G0 /G1 cell-cycle phase associated with activation of the Nrf2/HO-1 pathway. © 2015 The Authors. Molecular Nutrition & Food Research published by Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

  11. Bioreactor-induced mesenchymal progenitor cell differentiation and elastic fiber assembly in engineered vascular tissues.

    Science.gov (United States)

    Lin, Shigang; Mequanint, Kibret

    2017-09-01

    In vitro maturation of engineered vascular tissues (EVT) requires the appropriate incorporation of smooth muscle cells (SMC) and extracellular matrix (ECM) components similar to native arteries. To this end, the aim of the current study was to fabricate 4mm inner diameter vascular tissues using mesenchymal progenitor cells seeded into tubular scaffolds. A dual-pump bioreactor operating either in perfusion or pulsatile perfusion mode was used to generate physiological-like stimuli to promote progenitor cell differentiation, extracellular elastin production, and tissue maturation. Our data demonstrated that pulsatile forces and perfusion of 3D tubular constructs from both the lumenal and ablumenal sides with culture media significantly improved tissue assembly, effectively inducing mesenchymal progenitor cell differentiation to SMCs with contemporaneous elastin production. With bioreactor cultivation, progenitor cells differentiated toward smooth muscle lineage characterized by the expression of smooth muscle (SM)-specific markers smooth muscle alpha actin (SM-α-actin) and smooth muscle myosin heavy chain (SM-MHC). More importantly, pulsatile perfusion bioreactor cultivation enhanced the synthesis of tropoelastin and its extracellular cross-linking into elastic fiber compared with static culture controls. Taken together, the current study demonstrated progenitor cell differentiation and vascular tissue assembly, and provides insights into elastin synthesis and assembly to fibers. Incorporation of elastin into engineered vascular tissues represents a critical design goal for both mechanical and biological functions. In the present study, we seeded porous tubular scaffolds with multipotent mesenchymal progenitor cells and cultured in dual-pump pulsatile perfusion bioreactor. Physiological-like stimuli generated by bioreactor not only induced mesenchymal progenitor cell differentiation to vascular smooth muscle lineage but also actively promoted elastin synthesis and

  12. Airway smooth muscle cells : regulators of airway inflammation

    NARCIS (Netherlands)

    Zuyderduyn, Suzanne

    2007-01-01

    Airways from asthmatic subjects are more responsive to bronchoconstrictive stimuli than airways from healthy subjects. Airway smooth muscle (ASM) cells mediate contraction of the airways by responding to the bronchoconstrictive stimuli, which was thought to be the primary role of ASM cells. In this

  13. Progenitor cells in pulmonary vascular remodeling

    Science.gov (United States)

    Yeager, Michael E.; Frid, Maria G.; Stenmark, Kurt R.

    2011-01-01

    Pulmonary hypertension is characterized by cellular and structural changes in the walls of pulmonary arteries. Intimal thickening and fibrosis, medial hypertrophy and fibroproliferative changes in the adventitia are commonly observed, as is the extension of smooth muscle into the previously non-muscularized vessels. A majority of these changes are associated with the enhanced presence of α-SM-actin+ cells and inflammatory cells. Atypical abundances of functionally distinct endothelial cells, particularly in the intima (plexiform lesions), and also in the perivascular regions, are also described. At present, neither the origin(s) of these cells nor the molecular mechanisms responsible for their accumulation, in any of the three compartments of the vessel wall, have been fully elucidated. The possibility that they arise from either resident vascular progenitors or bone marrow–derived progenitor cells is now well established. Resident vascular progenitor cells have been demonstrated to exist within the vessel wall, and in response to certain stimuli, to expand and express myofibroblastic, endothelial or even hematopoietic markers. Bone marrow–derived or circulating progenitor cells have also been shown to be recruited to sites of vascular injury and to assume both endothelial and SM-like phenotypes. Here, we review the data supporting the contributory role of vascular progenitors (including endothelial progenitor cells, smooth muscle progenitor cells, pericytes, and fibrocytes) in vascular remodeling. A more complete understanding of the processes by which progenitor cells modulate pulmonary vascular remodeling will undoubtedly herald a renaissance of therapies extending beyond the control of vascular tonicity and reduction of pulmonary artery pressure. PMID:22034593

  14. Fluvastatin inhibits AGE-induced cell proliferation and migration via an ERK5-dependent Nrf2 pathway in vascular smooth muscle cells.

    Directory of Open Access Journals (Sweden)

    Ae-Rang Hwang

    Full Text Available Advanced glycation endproduct (AGE-induced vascular smooth muscle cell (VSMC proliferation and reactive oxygen species (ROS production are emerging as important mechanisms of diabetic vasculopathy, but little is known about the molecular mechanism responsible for the antioxidative effects of statins on AGEs. It has been reported that statins exert pleiotropic effects on the cardiovascular system due to decreases in AGE-induced cell proliferation, migration, and vascular inflammation. Thus, in the present study, the authors investigated the molecular mechanism by which statins decrease AGE-induced cell proliferation and VSMC migration. In cultured VSMCs, statins upregulated Nrf2-related antioxidant gene, NQO1 and HO-1, via an ERK5-dependent Nrf2 pathway. Inhibition of ERK5 by siRNA or BIX02189 (a specific ERK5 inhibitor reduced the statin-induced upregulations of Nrf2, NQO1, and HO-1. Furthermore, fluvastatin was found to significantly increase ARE promoter activity through ERK5 signaling, and to inhibit AGE-induced VSMC proliferation and migration as determined by MTT assay, cell counting, FACS analysis, a wound scratch assay, and a migration chamber assay. In addition, AGE-induced proliferation was diminished in the presence of Ad-CA-MEK5α encoding a constitutively active mutant form of MEK5α (an upstream kinase of ERK5, whereas depletion of Nrf2 restored statin-mediated reduction of AGE-induced cell proliferation. Moreover, fluvastatin suppressed the protein expressions of cyclin D1 and Cdk4, but induced p27, and blocked VSMC proliferation by regulating cell cycle. These results suggest statin-induced activation of an ERK5-dependent Nrf2 pathway reduces VSMC proliferation and migration induced by AGEs, and that the ERK5-Nrf2 signal module be viewed as a potential therapeutic target of vasculopathy in patients with diabetes and complications of the disease.

  15. Regulation of capacitative and non-capacitative Ca2+ entry in A7r5 vascular smooth muscle cells

    Directory of Open Access Journals (Sweden)

    COLIN W TAYLOR

    2004-01-01

    Full Text Available A capacitative Ca2+ entry (CCE pathway, activated by depletion of intracellular Ca2+ stores, is thought to mediate much of the Ca2+ entry evoked by receptors that stimulate phospholipase C (PLC. However, in A7r5 vascular smooth muscle cells, vasopressin, which stimulates PLC, empties intracellular Ca2+ stores but simultaneously inhibits their ability to activate CCE. The diacylglycerol produced with the IP3 that empties the stores is metabolized to arachidonic and this leads to activation of nitric oxide (NO synthase, production of NO and cyclic GMP, and consequent activation of protein kinase G. The latter inhibits CCE. In parallel, NO directly activates a non-capacitative Ca2+ entry (NCCE pathway, which is entirely responsible for the Ca2+ entry that occurs in the presence of vasopressin. This reciprocal regulation of two Ca2+ entry pathways ensures that there is sequential activation of first NCCE in the presence of vasopressin, and then a transient activation of CCE when vasopressin is removed. We suggest that the two routes for Ca2+ entry may selectively direct Ca2+ to processes that mediate activation and then recovery of the cell.

  16. Fibronectin promotes differentiation of neural crest progenitors endowed with smooth muscle cell potential

    International Nuclear Information System (INIS)

    Costa-Silva, Bruno; Coelho da Costa, Meline; Melo, Fernanda Rosene; Neves, Cynara Mendes; Alvarez-Silva, Marcio; Calloni, Giordano Wosgrau; Trentin, Andrea Goncalves

    2009-01-01

    The neural crest (NC) is a model system used to investigate multipotency during vertebrate development. Environmental factors control NC cell fate decisions. Despite the well-known influence of extracellular matrix molecules in NC cell migration, the issue of whether they also influence NC cell differentiation has not been addressed at the single cell level. By analyzing mass and clonal cultures of mouse cephalic and quail trunk NC cells, we show for the first time that fibronectin (FN) promotes differentiation into the smooth muscle cell phenotype without affecting differentiation into glia, neurons, and melanocytes. Time course analysis indicated that the FN-induced effect was not related to massive cell death or proliferation of smooth muscle cells. Finally, by comparing clonal cultures of quail trunk NC cells grown on FN and collagen type IV (CLIV), we found that FN strongly increased both NC cell survival and the proportion of unipotent and oligopotent NC progenitors endowed with smooth muscle potential. In contrast, melanocytic progenitors were prominent in clonogenic NC cells grown on CLIV. Taken together, these results show that FN promotes NC cell differentiation along the smooth muscle lineage, and therefore plays an important role in fate decisions of NC progenitor cells

  17. IGF-I Stimulates Cooperative Interaction between the IGF-I Receptor and CSK Homologous Kinase that Regulates SHPS-1 Phosphorylation in Vascular Smooth Muscle Cells

    Science.gov (United States)

    Radhakrishnan, Yashwanth; Shen, Xinchun; Maile, Laura A.; Xi, Gang

    2011-01-01

    IGF-I plays an important role in smooth muscle cell proliferation and migration. In vascular smooth muscle cells cultured in 25 mm glucose, IGF-I stimulated a significant increase in Src homology 2 domain containing protein tyrosine phosphatase substrate-1 (SHPS-1) phosphorylation compared with 5 mm glucose and this increase was required for smooth muscle cell proliferation. A proteome-wide screen revealed that carboxyl-terminal SRC kinase homologous kinase (CTK) bound directly to phosphotyrosines in the SHPS-1 cytoplasmic domain. Because the kinase(s) that phosphorylates these tyrosines in response to IGF-I is unknown, we determined the roles of IGF-I receptor (IGF-IR) and CTK in mediating SHPS-1 phosphorylation. After IGF-I stimulation, CTK was recruited to IGF-IR and subsequently to phospho-SHPS-1. Expression of an IGF-IR mutant that eliminated CTK binding reduced CTK transfer to SHPS-1, SHPS-1 phosphorylation, and cell proliferation. IGF-IR phosphorylated SHPS-1, which provided a binding site for CTK. CTK recruitment to SHPS-1 resulted in a further enhancement of SHPS-1 phosphorylation. CTK knockdown also impaired IGF-I-stimulated SHPS-1 phosphorylation and downstream signaling. Analysis of specific tyrosines showed that mutation of tyrosines 428/452 in SHPS-1 to phenylalanine reduced SHPS-1 phosphorylation but allowed CTK binding. In contrast, the mutation of tyrosines 469/495 inhibited IGF-IR-mediated the phosphorylation of SHPS-1 and CTK binding, suggesting that IGF-IR phosphorylated Y469/495, allowing CTK binding, and that CTK subsequently phosphorylated Y428/452. Based on the above findings, we conclude that after IGF-I stimulation, CTK is recruited to IGF-IR and its recruitment facilitates CTK's subsequent association with phospho-SHPS-1. This results in the enhanced CTK transfer to SHPS-1, and the two kinases then fully phosphorylate SHPS-1, which is necessary for IGF-I stimulated cellular proliferation. PMID:21799000

  18. Adhesion, Growth, and Maturation of Vascular Smooth Muscle Cells on Low-Density Polyethylene Grafted with Bioactive Substances

    Directory of Open Access Journals (Sweden)

    Martin Parizek

    2013-01-01

    Full Text Available The attractiveness of synthetic polymers for cell colonization can be affected by physical, chemical, and biological modification of the polymer surface. In this study, low-density polyethylene (LDPE was treated by an Ar+ plasma discharge and then grafted with biologically active substances, namely, glycine (Gly, polyethylene glycol (PEG, bovine serum albumin (BSA, colloidal carbon particles (C, or BSA+C. All modifications increased the oxygen content, the wettability, and the surface free energy of the materials compared to the pristine LDPE, but these changes were most pronounced in LDPE with Gly or PEG, where all the three values were higher than in the only plasma-treated samples. When seeded with vascular smooth muscle cells (VSMCs, the Gly- or PEG-grafted samples increased mainly the spreading and concentration of focal adhesion proteins talin and vinculin in these cells. LDPE grafted with BSA or BSA+C showed a similar oxygen content and similar wettability, as the samples only treated with plasma, but the nano- and submicron-scale irregularities on their surface were more pronounced and of a different shape. These samples promoted predominantly the growth, the formation of a confluent layer, and phenotypic maturation of VSMC, demonstrated by higher concentrations of contractile proteins alpha-actin and SM1 and SM2 myosins. Thus, the behavior of VSMC on LDPE can be regulated by the type of bioactive substances that are grafted.

  19. Increased atrial natriuretic factor receptor density in cultured vascular smooth muscle cells of the spontaneously hypertensive rat

    International Nuclear Information System (INIS)

    Khalil, F.; Fine, B.; Kuriyama, S.; Hatori, N.; Nakamura, A.; Nakamura, M.; Aviv, A.

    1987-01-01

    To explore the role of the atrial natriuretic factor (ANF) system in the pathophysiology of hypertension we examined the binding kinetics of synthetic ANF to cultured vascular smooth muscle cells (VSMCs) derived from the spontaneously hypertensive rat (SHR) and two normotensive controls-the Wistar Kyoto (WKY) and American Wistar (W). The number of maximal binding sites (Bmax) per cell (mean +/- SEM; X10(3] were: SHR = 278.0 +/- 33.0, WKY = 28.3 +/- 7.1 and W = 26.6 +/- 4.2. The differences between the SHR and normotensive strains were significant at p less than 0.001. The equilibrium dissociation constant (Kd; X 10(-9)M) was higher in SHR VSMCs (0.94 +/- 0.14) than in WKY (0.22 +/- 0.09; p less than 0.01) and W (0.39 +/- 0.14; p less than 0.02) cells. The plasma levels of the immunoreactive ANF were higher in SHR than the normotensive controls. We suggest that the relatively greater ANF receptor density in cultured VSMCs of the SHR represents a response to the in vitro environment which is relatively more deficient in ANF for VSMCs of the SHR as compared with the normotensive rats. Thus, the capacity of the SHR VSMC to regulate ANF receptor density appears to be independent of the blood pressure level

  20. Nitric oxide signaling pathway regulates potassium chloride cotransporter-1 mRNA expression in vascular smooth muscle cells.

    Science.gov (United States)

    Di Fulvio, M; Lauf, P K; Adragna, N C

    2001-11-30

    Rat vascular smooth muscle cells (VSMCs) express at least two mRNAs for K-Cl cotransporters (KCC): KCC1 and KCC3. cGMP-dependent protein kinase I regulates KCC3 mRNA expression in these cells. Here, we show evidence implicating the nitric oxide (NO)/cGMP signaling pathway in the expression of KCC1 mRNA, considered to be the major cell volume regulator. VSMCs, expressing soluble guanylyl cyclase (sGC) and PKG-I isoforms showed a time- and concentration-dependent increase in KCC1 mRNA levels after treatment with sodium nitroprusside as demonstrated by semiquantitative RT-PCR. sGC-dependent regulation of KCC1 mRNA expression was confirmed using YC-1, a NO-independent sGC stimulator. The sGC inhibitor LY83583 blocked the effects of sodium nitroprusside and YC-1. Moreover, 8-Br-cGMP increased KCC1 mRNA expression in a concentration- and time-dependent fashion. The 8-Br-cGMP effect was partially blocked by KT5823 but not by actinomycin D. However, actinomycin D and cycloheximide increased basal KCC1 mRNA in an additive manner, suggesting different mechanisms of action for both drugs. These findings suggest that in VSMCs, the NO/cGMP-signaling pathway participates in KCC1 mRNA regulation at the post-transcriptional level.

  1. Suppressive activities and mechanisms of ugonin J on vascular smooth muscle cells and balloon angioplasty-induced neointimal hyperplasia.

    Science.gov (United States)

    Pan, Chun-Hsu; Li, Pei-Chuan; Chien, Yi-Chung; Yeh, Wan-Ting; Liaw, Chih-Chuang; Sheu, Ming-Jyh; Wu, Chieh-Hsi

    2018-02-01

    Neointimal hyperplasia (or restenosis) is primarily attributed to excessive proliferation and migration of vascular smooth muscle cells (VSMCs). In this study, we investigated the inhibitory effects and mechanisms of ugonin J on VSMC proliferation and migration as well as neointimal formation. Cell viability and the cell-cycle distribution were, respectively, analyzed using an MTT assay and flow cytometry. Cell migration was examined using a wound-healing analysis and a transwell assay. Protein expressions and gelatinase activities were, respectively, measured using Western blot and gelatin zymography. Balloon angioplasty-induced neointimal formation was induced in a rat carotid artery model and then examined using immunohistochemical staining. Ugonin J induced cell-cycle arrest at the G 0 /G 1 phase and apoptosis to inhibit VSMC growth. Ugonin J also exhibited marked suppressive activity on VSMC migration. Ugonin J significantly reduced activations of focal adhesion kinase, phosphoinositide 3-kinase, v-akt murine thymoma viral oncogene homolog 1, and extracellular signal-regulated kinase 1/2 proteins. Moreover, ugonin J obviously reduced expressions and activity levels of matrix metalloproteinase-2 and matrix metalloproteinase-9. In vivo data indicated that ugonin J prevented balloon angioplasty-induced neointimal hyperplasia. Our study suggested that ugonin J has the potential for application in the prevention of balloon injury-induced neointimal formation. Copyright © 2017 John Wiley & Sons, Ltd.

  2. Microparticle Shedding from Neural Progenitor Cells and Vascular Compartment Cells Is Increased in Ischemic Stroke.

    Science.gov (United States)

    Chiva-Blanch, Gemma; Suades, Rosa; Crespo, Javier; Peña, Esther; Padró, Teresa; Jiménez-Xarrié, Elena; Martí-Fàbregas, Joan; Badimon, Lina

    2016-01-01

    Ischemic stroke has shown to induce platelet and endothelial microparticle shedding, but whether stroke induces microparticle shedding from additional blood and vascular compartment cells is unclear. Neural precursor cells have been shown to replace dying neurons at sites of brain injury; however, if neural precursor cell activation is associated to microparticle shedding, and whether this activation is maintained at long term and associates to stroke type and severity remains unknown. We analyzed neural precursor cells and blood and vascular compartment cells microparticle shedding after an acute ischemic stroke. Forty-four patients were included in the study within the first 48h after the onset of stroke. The cerebral lesion size was evaluated at 3-7 days of the stroke. Circulating microparticles from neural precursor cells and blood and vascular compartment cells (platelets, endothelial cells, erythrocytes, leukocytes, lymphocytes, monocytes and smooth muscle cells) were analyzed by flow cytometry at the onset of stroke and at 7 and 90 days. Forty-four age-matched high cardiovascular risk subjects without documented vascular disease were used as controls. Compared to high cardiovascular risk controls, patients showed higher number of neural precursor cell- and all blood and vascular compartment cell-derived microparticles at the onset of stroke, and after 7 and 90 days. At 90 days, neural precursor cell-derived microparticles decreased and smooth muscle cell-derived microparticles increased compared to levels at the onset of stroke, but only in those patients with the highest stroke-induced cerebral lesions. Stroke increases blood and vascular compartment cell and neural precursor cell microparticle shedding, an effect that is chronically maintained up to 90 days after the ischemic event. These results show that stroke induces a generalized blood and vascular cell activation and the initiation of neuronal cell repair process after stroke. Larger cerebral lesions

  3. Microparticle Shedding from Neural Progenitor Cells and Vascular Compartment Cells Is Increased in Ischemic Stroke.

    Directory of Open Access Journals (Sweden)

    Gemma Chiva-Blanch

    Full Text Available Ischemic stroke has shown to induce platelet and endothelial microparticle shedding, but whether stroke induces microparticle shedding from additional blood and vascular compartment cells is unclear. Neural precursor cells have been shown to replace dying neurons at sites of brain injury; however, if neural precursor cell activation is associated to microparticle shedding, and whether this activation is maintained at long term and associates to stroke type and severity remains unknown. We analyzed neural precursor cells and blood and vascular compartment cells microparticle shedding after an acute ischemic stroke.Forty-four patients were included in the study within the first 48h after the onset of stroke. The cerebral lesion size was evaluated at 3-7 days of the stroke. Circulating microparticles from neural precursor cells and blood and vascular compartment cells (platelets, endothelial cells, erythrocytes, leukocytes, lymphocytes, monocytes and smooth muscle cells were analyzed by flow cytometry at the onset of stroke and at 7 and 90 days. Forty-four age-matched high cardiovascular risk subjects without documented vascular disease were used as controls.Compared to high cardiovascular risk controls, patients showed higher number of neural precursor cell- and all blood and vascular compartment cell-derived microparticles at the onset of stroke, and after 7 and 90 days. At 90 days, neural precursor cell-derived microparticles decreased and smooth muscle cell-derived microparticles increased compared to levels at the onset of stroke, but only in those patients with the highest stroke-induced cerebral lesions.Stroke increases blood and vascular compartment cell and neural precursor cell microparticle shedding, an effect that is chronically maintained up to 90 days after the ischemic event. These results show that stroke induces a generalized blood and vascular cell activation and the initiation of neuronal cell repair process after stroke. Larger

  4. Endothelial and Smooth Muscle Cell Interaction via FoxM1 Signaling Mediates Vascular Remodeling and Pulmonary Hypertension.

    Science.gov (United States)

    Dai, Zhiyu; Zhu, Maggie M; Peng, Yi; Jin, Hua; Machireddy, Narsa; Qian, Zhijian; Zhang, Xianming; Zhao, You-Yang

    2018-04-17

    Angioproliferative vasculopathy is a hallmark of pulmonary arterial hypertension (PAH). However, little is known how endothelial cell (EC) and smooth muscle cell (SMC) crosstalk regulates the angioproliferative vascular remodeling. We aimed to investigate the role of EC and SMC interaction and underlying signaling pathways in PH development. SMC-specific Foxm1 or Cxcr4 knockout mice, EC-specific Foxm1 or Egln1 knockout mice, as well as EC-specific Egln1/Cxcl12 double knockout mice were used to assess the role of FoxM1 on SMC proliferation and PH. Lung tissues and cells from PAH patients were employed to validate clinical relevance. FoxM1 inhibitor Thiostrepton was used in Sugen 5416/hypoxia- and monocrotaline-challenged rats. FoxM1 expression was markedly upregulated in lungs and pulmonary arterial SMCs of idiopathic PAH patients and 4 discrete PH rodent models. Mice with SMC- (but not EC-) specific deletion of Foxm1 were protected from hypoxia- or Sugen 5416/hypoxia-induced PH. The upregulation of FoxM1 in SMCs induced by multiple EC-derived factors (PDGF-B, CXCL12, ET-1 and MIF) mediated SMC proliferation. Genetic deletion of endothelial Cxcl12 in Egln1Tie2Cre mice or loss of its cognate receptor Cxcr4 in SMCs in hypoxia-treated mice inhibited FoxM1 expression, SMC proliferation and PH. Accordingly, pharmacological inhibition of FoxM1 inhibited severe PH in both Sugen 5416/hypoxia and monocrotaline-challenged rats. Multiple factors derived from dysfunctional ECs induced FoxM1 expression in SMCs and activated FoxM1-dependent SMC proliferation which contributes to pulmonary vascular remodeling and PH. Thus, targeting FoxM1 signaling represents a novel strategy for treatment of IPAH.

  5. Phytochemical uptake following human consumption of Montmorency tart cherry (L. Prunus cerasus) and influence of phenolic acids on vascular smooth muscle cells in vitro.

    Science.gov (United States)

    Keane, Karen M; Bell, Phillip G; Lodge, John K; Constantinou, Costas L; Jenkinson, Sarah E; Bass, Rosemary; Howatson, Glyn

    2016-06-01

    To investigate the phytochemical uptake following human consumption of Montmorency tart cherry (L. Prunus cerasus) and influence of selected phenolic acids on vascular smooth muscle cells in vitro. In a randomised, double-blinded, crossover design, 12 healthy males consumed either 30 or 60 mL of Montmorency tart cherry concentrate. Following analysis of the juice composition, venous blood samples were taken before and 1, 2, 3, 5 and 8 h post-consumption of the beverage. In addition to examining some aspects of the concentrate contents, plasma concentrations of protocatechuic acid (PCA), vanillic acid (VA) and chlorogenic (CHL) acid were analysed by reversed-phase high-performance liquid chromatography (HPLC) with diode array for quantitation and mass spectrometry detection (LCMS) for qualitative purposes. Vascular smooth muscle cell migration and proliferation were also assessed in vitro. Both the 30 and 60 mL doses of Montmorency cherry concentrate contained high amounts of total phenolics (71.37 ± 0.11; 142.73 ± 0.22 mg/L) and total anthocyanins (62.47 ± 0.31; 31.24 ± 0.16 mg/L), as well as large quantities of CHL (0.205 ± 0.24; 0.410 ± 0.48 mg/L) and VA (0.253 ± 0.84; 0.506 ± 1.68 mg/L). HPLC/LCMS identified two dihydroxybenzoic acids (PCA and VA) in plasma following MC concentrate consumption. Both compounds were most abundant 1-2 h post-initial ingestion with traces detectable at 8 h post-ingestion. Cell migration was significantly influenced by the combination of PCA and VA, but not in isolation. There was no effect of the compounds on cell proliferation. These data show new information that phenolic compounds thought to exert vasoactive properties are bioavailable in vivo following MC consumption and subsequently can influence cell behaviour. These data may be useful for the design and interpretation of intervention studies investigating the health effects of Montmorency cherries.

  6. BAG3 promotes the phenotypic transformation of primary rat vascular smooth muscle cells via TRAIL.

    Science.gov (United States)

    Fu, Yao; Chang, Ye; Chen, Shuang; Li, Yuan; Chen, Yintao; Sun, Guozhe; Yu, Shasha; Ye, Ning; Li, Chao; Sun, Yingxian

    2018-05-01

    Under normal physiological condition, the mature vascular smooth muscle cells (VSMCs) show differentiated phenotype. In response to various environmental stimuluses, VSMCs convert from the differentiated phenotype to dedifferentiated phenotype characterized by the increased ability of proliferation/migration and the reduction of contractile ability. The phenotypic transformation of VSMCs played an important role in atherosclerosis. Both Bcl-2-associated athanogene 3 (BAG3) and tumor necrosis factor-related apopt-osis inducing ligand (TRAIL) involved in apoptosis. The relationship between BAG3 and TRAIL and their effects the proliferation and migration in VSMCs are rarely reported. This study investigated the effects of BAG3 on the phenotypic modulation and the potential underlying mechanisms in primary rat VSMCs. Primary rat VSMCs were extracted and cultured in vitro. Cell proliferation was detected by cell counting, real-time cell analyzer (RTCA) and EdU incorporation. Cell migration was detected by wound healing, Transwell and RTCA. BAG3 and TRAIL were detected using real-time PCR and western blotting and the secreted proteins in the cultured media by dot blot. The expression of BAG3 increased with continued passages in cultured primary VSMCs. BAG3 promoted the proliferation and migration of primary rat VSMC in a time-dependent manner. BAG3 significantly increased the expression of TRAIL while had no effects on its receptors. TRAIL knockdown or blocking by neutralizing antibody inhibited the proliferation of VSMCs induced by BAG3. TRAIL knockdown exerted no obvious influence on the migration of VSMCs. Based on this study, we report for the first time that BAG3 was expressed in cultured primary rat VSMCs and the expression of BAG3 increased with continued passages. Furthermore, BAG3 promoted the proliferation of VSMCs via increasing the expression of TRAIL. In addition, we also demonstrated that BAG3 promoted the migration of VSMCs independent of TRAIL

  7. Vascular smooth muscle cell apoptosis is an early trigger for hypothyroid atherosclerosis.

    Science.gov (United States)

    Wang, Pei; Xu, Tian-Ying; Guan, Yun-Feng; Zhao, Yan; Li, Zhi-Yong; Lan, Xiao-Hong; Wang, Xia; Yang, Peng-Yuan; Kang, Zhi-Min; Vanhoutte, Paul M; Miao, Chao-Yu

    2014-06-01

    Endothelial dysfunction is an initial and vascular smooth muscle cell (VSMC) apoptosis, a later step of atherosclerosis. Hypothyroidism accelerates atherosclerosis. However, the early events responsible for this pro-atherosclerotic effect are unclear. Rats were resistant to induction of atherosclerosis by high cholesterol diet alone, but became susceptible in hypothyroid state achieved by administration of propylthiouracil (PTU) for 6 weeks. VSMC dysfunction and apoptosis were obvious within 1 week after PTU treatment, without signs of endothelial dysfunction. This early VSMC damage was caused by hypothyroidism but not the high cholesterol diet. In ApoE knockout mice, PTU-induced hypothyroidism triggered early VSMC apoptosis, increased oxidative stress, and accelerated atherosclerosis development. Thyroid hormone supplementation (T4, 10, or 50 μg/kg) prevented atherogenic phenotypes in hypothyroid rats and mice. In rats, thyroidectomy caused severe hypothyroidism 5 days after operation, which also led to rapid VSMC dysfunction and apoptosis. In vitro studies did not show a direct toxic effect of PTU on VSMCs. In contrast, thyroid hormone (T3, 0.75 μg/L plus T4, 50 nmol/L) exerted a direct protection against VSMC apoptosis, which was reduced by knockdown of TRα1, rather than TRβ1 and TRβ2 receptors. TRα1-mediated inhibition of apoptotic signalling of JNKs and caspase-3 contributed to the anti-apoptotic action of thyroid hormone. These findings provide an in vivo example for VSMC apoptosis as an early trigger of hypothyroidism-associated atherosclerosis, and reveal activation of TRα1 receptors to prevent VSMC apoptosis as a therapeutic strategy in this disease. Published on behalf of the European Society of Cardiology. All rights reserved. © The Author 2014. For permissions please email: journals.permissions@oup.com.

  8. Hyperplasia of smooth muscle in mild to moderate asthma without changes in cell size or gene expression.

    Science.gov (United States)

    Woodruff, Prescott G; Dolganov, Gregory M; Ferrando, Ronald E; Donnelly, Samantha; Hays, Steven R; Solberg, Owen D; Carter, Roderick; Wong, Hofer H; Cadbury, Peggy S; Fahy, John V

    2004-05-01

    Bronchial hyperresponsiveness in mild to moderate asthma may result from airway smooth muscle cell proliferation or acquisition of a hypercontractile phenotype. Because these cells have not been well characterized in mild to moderate asthma, we examined the morphometric and gene expression characteristics of smooth muscle cells in this subgroup of patients with asthma. Using bronchial biopsies from 14 subjects with mild to moderate asthma and 15 control subjects, we quantified smooth muscle cell morphology by stereology and the expression of a panel of genes related to a hypercontractile phenotype of airway smooth muscle, using laser microdissection and two-step real-time polymerase chain reaction. We found that airway smooth muscle cell size was similar in both groups, but cell number was nearly twofold higher in subjects with asthma (p = 0.03), and the amount of smooth muscle in the submucosa was increased 50-83% (p 0.1). We conclude that airway smooth muscle proliferation is a pathologic characteristic of subjects with mild to moderate asthma. However, smooth muscle cells in mild to moderate asthma do not show hypertrophy or gene expression changes of a hypercontractile phenotype observed in vitro.

  9. BMP-2 Overexpression Augments Vascular Smooth Muscle Cell Motility by Upregulating Myosin Va via Erk Signaling

    Directory of Open Access Journals (Sweden)

    Ming Zhang

    2014-01-01

    Full Text Available Background. The disruption of physiologic vascular smooth muscle cell (VSMC migration initiates atherosclerosis development. The biochemical mechanisms leading to dysfunctional VSMC motility remain unknown. Recently, cytokine BMP-2 has been implicated in various vascular physiologic and pathologic processes. However, whether BMP-2 has any effect upon VSMC motility, or by what manner, has never been investigated. Methods. VSMCs were adenovirally transfected to genetically overexpress BMP-2. VSMC motility was detected by modified Boyden chamber assay, confocal time-lapse video assay, and a colony wounding assay. Gene chip array and RT-PCR were employed to identify genes potentially regulated by BMP-2. Western blot and real-time PCR detected the expression of myosin Va and the phosphorylation of extracellular signal-regulated kinases 1/2 (Erk1/2. Immunofluorescence analysis revealed myosin Va expression locale. Intracellular Ca2+ oscillations were recorded. Results. VSMC migration was augmented in VSMCs overexpressing BMP-2 in a dose-dependent manner. siRNA-mediated knockdown of myosin Va inhibited VSMC motility. Both myosin Va mRNA and protein expression significantly increased after BMP-2 administration and were inhibited by Erk1/2 inhibitor U0126. BMP-2 induced Ca2+ oscillations, generated largely by a “cytosolic oscillator”. Conclusion. BMP-2 significantly increased VSMCs migration and myosin Va expression, via the Erk signaling pathway and intracellular Ca2+ oscillations. We provide additional insight into the pathophysiology of atherosclerosis, and inhibition of BMP-2-induced myosin Va expression may represent a potential therapeutic strategy.

  10. Mechanical stretch augments insulin-induced vascular smooth muscle cell proliferation by insulin-like growth factor-1 receptor

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Gang [Department of Pharmacology, Faculty of Medicine, Kagawa University, Kagawa (Japan); Department of Anesthesiology, First Affiliated Hospital of China Medical University, Shenyang (China); Hitomi, Hirofumi, E-mail: hitomi@kms.ac.jp [Department of Pharmacology, Faculty of Medicine, Kagawa University, Kagawa (Japan); Hosomi, Naohisa [Department of Cardiorenal and Cerebrovascular Medicine, Faculty of Medicine, Kagawa University, Kagawa (Japan); Lei, Bai; Nakano, Daisuke [Department of Pharmacology, Faculty of Medicine, Kagawa University, Kagawa (Japan); Deguchi, Kazushi; Mori, Hirohito; Masaki, Tsutomu [Department of Gastroenterology and Neurology, Faculty of Medicine, Kagawa University, Kagawa (Japan); Ma, Hong [Department of Anesthesiology, First Affiliated Hospital of China Medical University, Shenyang (China); Griendling, Kathy K. [Department of Medicine, Division of Cardiology, Emory University School of Medicine, Atlanta, GA (United States); Nishiyama, Akira [Department of Pharmacology, Faculty of Medicine, Kagawa University, Kagawa (Japan)

    2011-10-15

    Insulin resistance and hypertension have been implicated in the pathogenesis of cardiovascular disease; however, little is known about the roles of insulin and mechanical force in vascular smooth muscle cell (VSMC) remodeling. We investigated the contribution of mechanical stretch to insulin-induced VSMC proliferation. Thymidine incorporation was stimulated by insulin in stretched VSMCs, but not in un-stretched VSMCs. Insulin increased 2-deoxy-glucose incorporation in both stretched and un-stretched VSMCs. Mechanical stretch augmented insulin-induced extracellular signal-regulated kinase (ERK) and Akt phosphorylation. Inhibitors of epidermal growth factor (EGF) receptor tyrosine kinase and Src attenuated insulin-induced ERK and Akt phosphorylation, as well as thymidine incorporation, whereas 2-deoxy-glucose incorporation was not affected by these inhibitors. Moreover, stretch augmented insulin-like growth factor (IGF)-1 receptor expression, although it did not alter the expression of insulin receptor and insulin receptor substrate-1. Insulin-induced ERK and Akt activation, and thymidine incorporation were inhibited by siRNA for the IGF-1 receptor. Mechanical stretch augments insulin-induced VSMC proliferation via upregulation of IGF-1 receptor, and downstream Src/EGF receptor-mediated ERK and Akt activation. Similar to in vitro experiment, IGF-1 receptor expression was also augmented in hypertensive rats. These results provide a basis for clarifying the molecular mechanisms of vascular remodeling in hypertensive patients with hyperinsulinemia. -- Highlights: {yields} Mechanical stretch augments insulin-induced VSMC proliferation via IGF-1 receptor. {yields} Src/EGFR-mediated ERK and Akt phosphorylation are augmented in stretched VSMCs. {yields} Similar to in vitro experiment, IGF-1 receptor is increased in hypertensive rats. {yields} Results provide possible mechanisms of vascular remodeling in hypertension with DM.

  11. Pathologic bladder microenvironment attenuates smooth muscle differentiation of skin derived precursor cells: implications for tissue regeneration.

    Directory of Open Access Journals (Sweden)

    Cornelia Tolg

    Full Text Available Smooth muscle cell containing organs (bladder, heart, blood vessels are damaged by a variety of pathological conditions necessitating surgery or organ replacement. Currently, regeneration of contractile tissues is hampered by lack of functional smooth muscle cells. Multipotent skin derived progenitor cells (SKPs can easily be isolated from adult skin and can be differentiated in vitro into contractile smooth muscle cells by exposure to FBS. Here we demonstrate an inhibitory effect of a pathologic contractile organ microenvironment on smooth muscle cell differentiation of SKPs. In vivo, urinary bladder strain induces microenvironmental changes leading to de-differentiation of fully differentiated bladder smooth muscle cells. Co-culture of SKPs with organoids isolated from ex vivo stretched bladders or exposure of SKPs to diffusible factors released by stretched bladders (e.g. bFGF suppresses expression of smooth muscle markers (alpha SMactin, calponin, myocardin, myosin heavy chain as demonstrated by qPCR and immunofluorescent staining. Rapamycin, an inhibitor of mTOR signalling, previously observed to prevent bladder strain induced de-differentiation of fully differentiated smooth muscle cells in vitro, inhibits FBS-induced smooth muscle cell differentiation of undifferentiated SKPs. These results suggest that intended precursor cell differentiation may be paradoxically suppressed by the disease context for which regeneration may be required. Organ-specific microenvironment contexts, particularly prevailing disease, may play a significant role in modulating or attenuating an intended stem cell phenotypic fate, possibly explaining the variable and inefficient differentiation of stem cell constructs in in vivo settings. These observations must be considered in drafting any regeneration strategies.

  12. Expression profile and protein translation of TMEM16A in murine smooth muscle

    DEFF Research Database (Denmark)

    Davis, Alison J; Forrest, Abigail S; Jepps, Thomas Andrew

    2010-01-01

    Recently, overexpression of the genes TMEM16A and TMEM16B has been shown to produce currents qualitatively similar to native Ca(2+)-activated Cl(-) currents (I(ClCa)) in vascular smooth muscle. However, there is no information about this new gene family in vascular smooth muscle, where Cl(-) chan...

  13. Andrographolide induces vascular smooth muscle cell apoptosis through a SHP-1-PP2A-p38MAPK-p53 cascade.

    Science.gov (United States)

    Chen, Yu-Ying; Hsieh, Cheng-Ying; Jayakumar, Thanasekaran; Lin, Kuan-Hung; Chou, Duen-Suey; Lu, Wan-Jung; Hsu, Ming-Jen; Sheu, Joen-Rong

    2014-07-10

    The abnormal growth of vascular smooth muscle cells (VSMCs) is considered a critical pathogenic process in inflammatory vascular diseases. We have previously demonstrated that protein phosphatase 2 A (PP2A)-mediated NF-κB dephosphorylation contributes to the anti-inflammatory properties of andrographolide, a novel NF-κB inhibitor. In this study, we investigated whether andrographolide causes apoptosis, and characterized its apoptotic mechanisms in rat VSMCs. Andrographolide activated the p38 mitogen-activated protein kinase (p38MAPK), leading to p53 phosphorylation. Phosphorylated p53 subsequently transactivated the expression of Bax, a pro-apoptotic protein. Transfection with pp2a small interfering RNA (siRNA) suppressed andrographolide-induced p38MAPK activation, p53 phosphorylation, and caspase 3 activation. Andrographolide also activated the Src homology 1 domain-containing protein tyrosine phosphatase (SHP-1), and induced PP2A dephosphorylation, both of which were inhibited by the SHP-1 inhibitor sodium stibogluconate (SSG) or shp-1 siRNA. SSG or shp-1 siRNA prevented andrographolide-induced apoptosis. These results suggest that andrographolide activates the PP2A-p38MAPK-p53-Bax cascade, causing mitochondrial dysfunction and VSMC death through an SHP-1-dependent mechanism.

  14. The chemokine and scavenger receptor CXCL16/SR-PSOX is expressed in human vascular smooth muscle cells and is induced by interferon γ

    International Nuclear Information System (INIS)

    Wagsaeter, Dick; Olofsson, Peder S.; Norgren, Lars; Stenberg, Bjoern; Sirsjoe, Allan

    2004-01-01

    Atherosclerosis is an inflammatory disease that is characterised by the involvement of chemokines that are important for the recruitment of leukocytes and scavenger receptors that mediate foam cell formation. Several cytokines are involved in the regulation of chemokines and scavenger receptors in atherosclerosis. CXCL16 is a chemokine and scavenger receptor and found in macrophages in human atherosclerotic lesions. Using double-labelled immunohistochemistry, we identified that smooth muscle cells in human lesions express CXCL16. We then analysed the effects of IFN-γ, TNF-α, IL-12, IL-15, IL-18, and LPS on CXCL16 expression in cultured aortic smooth muscle cells. IFN-γ was the most potent CXCL16 inducer and increased mRNA, soluble form, membrane form, and total cellular levels of CXCL16. The IFN-γ induction of CXCL16 was also associated with increased uptake of oxLDL into these cells. Taken together, smooth muscle cells express CXCL16 in atherosclerotic lesions, which may play a role in the attraction of T cells to atherosclerotic lesions and contribute to the cellular internalisation of modified LDL

  15. Magnesium Inhibits Wnt/β-Catenin Activity and Reverses the Osteogenic Transformation of Vascular Smooth Muscle Cells

    Science.gov (United States)

    Montes de Oca, Addy; Guerrero, Fatima; Martinez-Moreno, Julio M.; Madueño, Juan A.; Herencia, Carmen; Peralta, Alan; Almaden, Yolanda; Lopez, Ignacio; Aguilera-Tejero, Escolastico; Gundlach, Kristina; Büchel, Janine; Peter, Mirjam E.; Passlick-Deetjen, Jutta; Rodriguez, Mariano; Muñoz-Castañeda, Juan R.

    2014-01-01

    Magnesium reduces vascular smooth muscle cell (VSMC) calcification in vitro but the mechanism has not been revealed so far. This work used only slightly increased magnesium levels and aimed at determining: a) whether inhibition of magnesium transport into the cell influences VSMC calcification, b) whether Wnt/β-catenin signaling, a key mediator of osteogenic differentiation, is modified by magnesium and c) whether magnesium can influence already established vascular calcification. Human VSMC incubated with high phosphate (3.3 mM) and moderately elevated magnesium (1.4 mM) significantly reduced VSMC calcification and expression of the osteogenic transcription factors Cbfa-1 and osterix, and up-regulated expression of the natural calcification inhibitors matrix Gla protein (MGP) and osteoprotegerin (OPG). The protective effects of magnesium on calcification and expression of osteogenic markers were no longer observed in VSMC cultured with an inhibitor of cellular magnesium transport (2-aminoethoxy-diphenylborate [2-APB]). High phosphate induced activation of Wnt/β-catenin pathway as demonstrated by the translocation of β-catenin into the nucleus, increased expression of the frizzled-3 gene, and downregulation of Dkk-1 gene, a specific antagonist of the Wnt/β-catenin signaling pathway. The addition of magnesium however inhibited phosphate-induced activation of Wnt/β-catenin signaling pathway. Furthermore, TRPM7 silencing using siRNA resulted in activation of Wnt/β-catenin signaling pathway. Additional experiments were performed to test the ability of magnesium to halt the progression of already established VSMC calcification in vitro. The delayed addition of magnesium decreased calcium content, down-regulated Cbfa-1 and osterix and up-regulated MGP and OPG, when compared with a control group. This effect was not observed when 2-APB was added. In conclusion, magnesium transport through the cell membrane is important to inhibit VSMC calcification in vitro

  16. NONOates regulate KCl cotransporter-1 and -3 mRNA expression in vascular smooth muscle cells.

    Science.gov (United States)

    Di Fulvio, Mauricio; Lauf, Peter K; Shah, Shalin; Adragna, Norma C

    2003-05-01

    Nitric oxide (NO) donors regulate KCl cotransport (KCC) activity and cotransporter-1 and -3 (KCC1 and KCC3) mRNA expression in sheep erythrocytes and in primary cultures of rat vascular smooth muscle cells (VSMCs), respectively. In this study, we used NONOates as rapid and slow NO releasers to provide direct evidence implicating NO as a regulator of KCC3 gene expression at the mRNA level. In addition, we used the expression of KCC3 mRNA to further investigate the mechanism of action of these NO donors at the cellular level. Treatment of VSMCs with rapid NO releasers, like NOC-5 and NOC-9, as well as with the direct NO-independent soluble guanylyl cyclase (sGC) stimulator YC-1, acutely increased KCC3 mRNA expression in a concentration- and time-dependent manner. The slow NO releaser NOC-18 had no effect on KCC3 gene expression. A specific NO scavenger completely prevented the NONOate-induced KCC3 mRNA expression. Inhibition of sGC with LY-83583 blocked the NONOate- and YC-1-induced KCC3 mRNA expression. This study shows that in primary cultures of rat VSMCs, the fast NO releasers NOC-9 and NOC-5, but not the slow NO releaser NOC-18, acutely upregulate KCC3 mRNA expression in a NO/sGC-dependent manner.

  17. Interleukin-6 downregulated vascular smooth muscle cell contractile proteins via ATG4B-mediated autophagy in thoracic aortic dissection.

    Science.gov (United States)

    An, Zhao; Qiao, Fan; Lu, Qijue; Ma, Ye; Liu, Yang; Lu, Fanglin; Xu, Zhiyun

    2017-12-01

    Interleukin-6 (IL-6) overexpression played an important role in the pathogenesis of thoracic aortic dissection (TAD). Our previous study found enhanced autophagy accompanying with contractile proteins α smooth muscle actin (α-SMA) and smooth muscle 22α (SM22α) degradation in TAD aortic vascular smooth muscle cells (VSMCs). Autophagy is an important way for intracellular proteins degradation, while IL-6 has been found as a contributing factor of autophagy in some cancers. These indicated IL-6 might contribute to the occurrence of TAD by promoting autophagy-induced contractile proteins degradation, which has not been investigated. The aim of the present study is to verify this hypothesis and investigate the mechanism of it. We collected 10 TAD and 10 control aortic specimens from patients underwent TAD surgical repair and coronary artery bypass grafting, respectively. Quantitative real-time polymerase chain reaction was used to detect mRNA expression. Protein expression level was assessed by enzyme-linked immunosorbent assay, western blot, and immunohistochemistry. Microtubule-associated protein 1 light chain 3 beta overexpression adenovirus with green and red fluorescent protein tags and transmission electron microscopy were used to detect autophagy level in VSMCs. 3-Methyladenine (3-MA) and chloroquine were used to block autophagy in human VSMCs. Experiment results showed that the expression of IL-6 was significantly increased accompanying with up-regulated autophagy in TAD aortic wall compared with controls. In vitro results showed that IL-6 stimulation decreased the expression of VSMCs contractile proteins α-SMA and SM22α accompanying with up-regulated autophagy. Blocking autophagy with 3-MA or chloroquine inhibited IL-6 induced α-SMA and SM22α degradation. Further investigation showed that autophagy-related 4B cysteine peptidase (ATG4B) was significantly overexpressed in TAD aortic wall and played important role in IL-6 induced autophagy up

  18. Recovery of prostacyclin synthesis in vascular smooth muscle cells following self-inactivation and requirement for growth factors

    International Nuclear Information System (INIS)

    Bailey, J.M.; Hla, T.T.; Pash, J.M.

    1986-01-01

    The cyclooxygenase enzyme system is a prime example of a metabolic pathway that is regulated by self inactivation. This is believed to occur in part via the irreversible reaction of the endoperoxide intermediate species with the cyclooxygenase enzyme. This inactivation and recovery of activity is similar to the inactivation observed with aspirin which irreversibly acetylates the enzyme. Self inactivation was studied in cultured rat and bovine aorta smooth muscle cells. The production of the prostanoid PGI2 was demonstrated by incubation of a monolayer of cells with 12 μM C-14 labeled arachidonic acid. Products were analyzed by thin layer chromatography and identified by their comigration with authentic standards and confirmed by gas chromatography/mass spectrometry. Preincubation of the cells for 10 minutes with arachidonic acid at concentrations as low as 1 μg/mL inactivated the cells to a second challenge with radiolabeled arachidonic acid. Recovery from self inactivation took place over a three hour time period and was similar to the recovery observed with aspirin pretreatment. Recovery was inhibited by addition of 10 μg/mL cycloheximide to the medium indicating that it involves synthesis of cyclooxygenase protein. Epidermal growth factor was identified as a serum factor responsible for the rapid recovery of cyclooxygenase activity in rat and bovine aorta smooth muscle cells

  19. Actin isoform and alpha 1B-adrenoceptor gene expression in aortic and coronary smooth muscle is influenced by cyclical stretch.

    Science.gov (United States)

    Lundberg, M S; Sadhu, D N; Grumman, V E; Chilian, W M; Ramos, K S

    1995-09-01

    The occurrence of vascular domains with specific biological and pharmacological characteristics suggests that smooth muscle cells in different arteries may respond differentially to a wide range of environmental stimuli. To determine if some of these vessel-specific differences may be attributable to mechano-sensitive gene regulation, the influence of cyclical stretch on the expression of actin isoform and alpha 1B-adrenoceptor genes was examined in aortic and coronary smooth muscle cells. Cells were seeded on an elastin substrate and subjected to maximal stretching (24% elongation) and relaxation cycles at a frequency of 120 cycles/min in a Flexercell strain unit for 72 h. Total RNA was extracted and hybridized to radiolabeled cDNA probes to assess gene expression. Stretch caused a greater reduction of actin isoform mRNA levels in aortic smooth muscle cells as compared to cells from the coronary artery. Steady-state mRNA levels of alpha 1B-adrenoceptor were also decreased by cyclical stretch in both cell types but the magnitude of the response was greater in coronary smooth muscle cells. No changes in alpha 1B-adrenoceptor or beta/gamma-actin steady-state mRNA levels were observed in H4IIE cells, a nonvascular, immortalized cell line. The relative gene expression of heat shock protein 70 was not influenced by the cyclic stretch regimen in any of these cell types. These results suggest that stretch may participate in the regulation of gene expression in vascular smooth muscle cells and that this response exhibits some degree of cell-specificity.

  20. ASIC proteins regulate smooth muscle cell migration.

    Science.gov (United States)

    Grifoni, Samira C; Jernigan, Nikki L; Hamilton, Gina; Drummond, Heather A

    2008-03-01

    The purpose of the present study was to investigate Acid Sensing Ion Channel (ASIC) protein expression and importance in cellular migration. We recently demonstrated that Epithelial Na(+)Channel (ENaC) proteins are required for vascular smooth muscle cell (VSMC) migration; however, the role of the closely related ASIC proteins has not been addressed. We used RT-PCR and immunolabeling to determine expression of ASIC1, ASIC2, ASIC3 and ASIC4 in A10 cells. We used small interference RNA to silence individual ASIC expression and determine the importance of ASIC proteins in wound healing and chemotaxis (PDGF-bb)-initiated migration. We found ASIC1, ASIC2, and ASIC3, but not ASIC4, expression in A10 cells. ASIC1, ASIC2, and ASIC3 siRNA molecules significantly suppressed expression of their respective proteins compared to non-targeting siRNA (RISC) transfected controls by 63%, 44%, and 55%, respectively. Wound healing was inhibited by 10, 20, and 26% compared to RISC controls following suppression of ASIC1, ASIC2, and ASIC3, respectively. Chemotactic migration was inhibited by 30% and 45%, respectively, following suppression of ASIC1 and ASIC3. ASIC2 suppression produced a small, but significant, increase in chemotactic migration (4%). Our data indicate that ASIC expression is required for normal migration and may suggest a novel role for ASIC proteins in cellular migration.

  1. Expression pattern and function of tyrosine receptor kinase B isoforms in rat mesenteric arterial smooth muscle cells

    International Nuclear Information System (INIS)

    Otani, Kosuke; Okada, Muneyoshi; Yamawaki, Hideyuki

    2015-01-01

    Tyrosine receptor kinaseB (TrkB) is a high affinity receptor for brain-derived neurotrophic factor (BDNF). TrkB isoforms involve full length TrkB (TrkB FL) and truncated TrkB type1 (TrkB T1) and type 2 (TrkB T2) in rats. The aim of present study was to explore their expression pattern and function in mesenteric arterial smooth muscle cells (MASMCs). The expression of TrkB isoform protein and mRNA was examined by Western blotting, immunofluorescence and quantitative RT-PCR analyses. Cell proliferation was measured by a bromodeoxyuridine (BrdU) incorporation assay. Cell migration was measured by a Boyden chamber assay. Cell morphology was observed with a phase-contrast microscope. Protein and mRNA expression of BDNF and TrkB isoforms was confirmed in MASMCs. Expression level of TrkB FL was less, while that of TrkB T1 was the highest in MASMCs. Although BDNF increased phosphorylation of ERK, it had no influence on migration and proliferation of MASMCs. TrkB T1 gene knockdown by a RNA interference induced morphological changes and reduced expression level of α-smooth muscle actin (α-SMA) in MASMCs. Similar morphological changes and reduced α-SMA expression were induced in MASMCs by a Rho kinase inhibitor, Y-27632. In conclusion, we for the first time demonstrate that TrkB T1 expressed highly in MASMCs contributes to maintain normal cell morphology possibly via regulation of Rho activity. This study firstly defined expression level of TrkB isoforms and partly revealed their functions in peripheral vascular cells. - Highlights: • BDNF-TrkB axis mediates neurogenesis, growth, differentiation and survival. • Expression pattern and function of TrkB in vascular smooth muscle remain unclear. • Expression of TrkB FL is low, while that of TrkB T1 is the highest. • TrkB T1 contributes to maintain normal morphology possibly via activating Rho.

  2. Expression pattern and function of tyrosine receptor kinase B isoforms in rat mesenteric arterial smooth muscle cells

    Energy Technology Data Exchange (ETDEWEB)

    Otani, Kosuke; Okada, Muneyoshi; Yamawaki, Hideyuki, E-mail: yamawaki@vmas.kitasato-u.ac.jp

    2015-11-27

    Tyrosine receptor kinaseB (TrkB) is a high affinity receptor for brain-derived neurotrophic factor (BDNF). TrkB isoforms involve full length TrkB (TrkB FL) and truncated TrkB type1 (TrkB T1) and type 2 (TrkB T2) in rats. The aim of present study was to explore their expression pattern and function in mesenteric arterial smooth muscle cells (MASMCs). The expression of TrkB isoform protein and mRNA was examined by Western blotting, immunofluorescence and quantitative RT-PCR analyses. Cell proliferation was measured by a bromodeoxyuridine (BrdU) incorporation assay. Cell migration was measured by a Boyden chamber assay. Cell morphology was observed with a phase-contrast microscope. Protein and mRNA expression of BDNF and TrkB isoforms was confirmed in MASMCs. Expression level of TrkB FL was less, while that of TrkB T1 was the highest in MASMCs. Although BDNF increased phosphorylation of ERK, it had no influence on migration and proliferation of MASMCs. TrkB T1 gene knockdown by a RNA interference induced morphological changes and reduced expression level of α-smooth muscle actin (α-SMA) in MASMCs. Similar morphological changes and reduced α-SMA expression were induced in MASMCs by a Rho kinase inhibitor, Y-27632. In conclusion, we for the first time demonstrate that TrkB T1 expressed highly in MASMCs contributes to maintain normal cell morphology possibly via regulation of Rho activity. This study firstly defined expression level of TrkB isoforms and partly revealed their functions in peripheral vascular cells. - Highlights: • BDNF-TrkB axis mediates neurogenesis, growth, differentiation and survival. • Expression pattern and function of TrkB in vascular smooth muscle remain unclear. • Expression of TrkB FL is low, while that of TrkB T1 is the highest. • TrkB T1 contributes to maintain normal morphology possibly via activating Rho.

  3. Aortic smooth muscle cell proteoglycan synthesis in relation to atherosclerosis

    International Nuclear Information System (INIS)

    Edwards, I.J.

    1989-01-01

    Proteoglycans (PG) are implicated in atherogenesis by their effects on tissue permeability and cell proliferation and their interaction with plasma low density lipoproteins. Using the pigeon model in which an atherosclerosis-susceptible (WC) and -resistant (SR) breed can be compared, PG synthesis by cultured aortic smooth muscle cells was examined by the use of [ 35 S]-sodium sulfate and [ 3 H]-serine or [ 3 H]-glucosamine as labeling precursors. In both SR and WC cells, the majority of newly synthesized PG were secreted into the media. Chondroitin sulfate (CS) PG and dermatan sulfate (DS) PG were the major PG produced. Total PG production was consistently lower in WC compared to SR cultures due in part to reduce PG synthesis but also to degradation of newly synthesized PG. Since increased DS-PG accompanines atherosclerosis progression, experiments were designed to test the hypothesis that macrophages modulate smooth muscle cell metabolism to cause increase DS-PG production. Cultured WC aortic smooth muscle cells were exposed to the media of cholesteryl ester-loaded pigeon peritoneal macrophages or a macrophage cell line P388D1 and the production of PG examined. Increasing concentration of conditioned media from both types of macrophages caused increased incorporation of 35 S-sulfate into secreted PG, but no change in cell-associated PG. Lipopolysaccharide activation of P388D1 cells enhanced the effect

  4. Vascular smooth muscle cells express the alpha(1A) subunit of a P-/Q-type voltage-dependent Ca(2+)Channel, and It is functionally important in renal afferent arterioles

    DEFF Research Database (Denmark)

    Hansen, Pernille B. Lærkegaard; Jensen, Boye L.; Andreasen, D

    2000-01-01

    In the present study, we tested whether the alpha(1A) subunit, which encodes a neuronal isoform of voltage-dependent Ca(2+) channels (VDCCs) (P-/Q-type), was present and functional in vascular smooth muscle and renal resistance vessels. By reverse transcription-polymerase chain reaction...... preglomerular resistance vessels and aorta, as well as mesangial cells, and that P-type VDCCs contribute to Ca(2+) influx in aortic and renal VSMCs and are involved in depolarization-mediated contraction in renal afferent arterioles....

  5. Influence of Androgen Receptor in Vascular Cells on Reperfusion following Hindlimb Ischaemia.

    Directory of Open Access Journals (Sweden)

    Junxi Wu

    Full Text Available Studies in global androgen receptor knockout (G-ARKO and orchidectomised mice suggest that androgen accelerates reperfusion of the ischaemic hindlimb by stimulating angiogenesis. This investigation used novel, vascular cell-specific ARKO mice to address the hypothesis that the impaired hindlimb reperfusion in G-ARKO mice was due to loss of AR from cells in the vascular wall.Mice with selective deletion of AR (ARKO from vascular smooth muscle cells (SM-ARKO, endothelial cells (VE-ARKO, or both (SM/VE-ARKO were compared with wild type (WT controls. Hindlimb ischaemia was induced in these mice by ligation and removal of the femoral artery. Post-operative reperfusion was reduced in SM-ARKO and SM/VE-ARKO mice. Immunohistochemistry indicated that this was accompanied by a reduced density of smooth muscle actin-positive vessels but no change in the density of isolectin B4-positive vessels in the gastrocnemius muscle. Deletion of AR from the endothelium (VE-ARKO did not alter post-operative reperfusion or vessel density. In an ex vivo (aortic ring culture model of angiogenesis, AR was not detected in vascular outgrowths and angiogenesis was not altered by vascular ARKO or by exposure to dihydrotestosterone (DHT 10-10-10-7M; 6 days.These results suggest that loss of AR from vascular smooth muscle, but not from the endothelium, contributes to impaired reperfusion in the ischaemic hindlimb of G-ARKO. Impaired reperfusion was associated with reduced collateral formation rather than reduced angiogenesis.

  6. β-adrenergic relaxation of smooth muscle: differences between cells and tissues

    International Nuclear Information System (INIS)

    Scheid, C.R.

    1987-01-01

    The present studies were carried out in an attempt to resolve the controversy about the Na + dependence of β-adrenergic relaxation in smooth muscle. Previous studies on isolated smooth muscle cells from the toad stomach had suggested that at least some of the actions of β-adrenergic agents, including a stimulatory effect on 45 Ca efflux, were dependent on the presence of a normal transmembrane Na + gradient. Studies by other investigators using tissues derived from mammalian sources had suggested that the relaxing effect of β-adrenergic agents was Na + independent. Uncertainty remained as to whether these discrepancies reflected differences between cells and tissues or differences between species. Thus, in the present studies, the authors utilized both tissues and cells from the same source, the stomach muscle of the toad Bufo marinus, and assessed the Na + dependence of β-adrenergic relaxation. They found that elimination of a normal Na + gradient abolished β-adrenergic relaxation of isolated cells. In tissues, however, similar manipulations had no effect on relaxation. The reasons for this discrepancy are unclear but do not appear to be attributable to changes in smooth muscle function following enzymatic dispersion. Thus the controversy concerning the mechanisms of β-adrenergic relaxation may reflect inherent differences between tissues and cells

  7. Simultaneous Increases in Proliferation and Apoptosis of Vascular Smooth Muscle Cells Accelerate Diabetic Mouse Venous Atherosclerosis

    Science.gov (United States)

    Liu, Shuying; Zhang, Zhengyu; Wang, Jingjing; Zhou, Yuhuan; Liu, Kefeng; Huang, Jintao; Chen, Dadi; Wang, Junmei; Li, Chaohong

    2015-01-01

    Aims This study was designed to demonstrate simultaneous increases in proliferation and apoptosis of vascular smooth muscle cells (VSMCs) leading to accelerated vein graft remodeling and to explore the underlying mechanisms. Methods Vein grafts were performed in non-diabetic and diabetic mice. The cultured quiescent VSMCs were subjected to mechanical stretch stress (SS) and/or advanced glycosylation end products (AGEs). Harvested vein grafts and treated VSMCs were used to detect cell proliferation, apoptosis, mitogen-activated protein kinases (MAPKs) activation and SM-α-actin expression. Results Significantly thicker vessel walls and greater increases in proliferation and apoptosis were observed in diabetic vein grafts than those in non-diabetic. Both SS and AGEs were found to induce different activation of three members of MAPKs and simultaneous increases in proliferation and apoptosis of VSMCs, and combined treatment with both had a synergistic effect. VSMCs with strong SM-α-actin expression represented more activated JNKs or p38MAPK, and cell apoptosis, while the cells with weak SM-α-actin expression demonstrated preferential activation of ERKs and cell proliferation. In contrast, inhibition of MAPKs signals triggered significant decreases in VSMC proliferation, and apoptosis. Treatment of the cells with RNA interference of receptor of AGEs (RAGE) also resulted in significant decreases in both proliferation and apoptosis. Conclusions Increased pressure-induced SS triggers simultaneous increases in proliferation and apoptosis of VSMCs in the vein grafts leading to vein arterializations, which can be synergistically accelerated by high glucose-induced AGEs resulting in vein graft atherosclerosis. Either SS or AGEs and their combination induce simultaneous increases in proliferation and apoptosis of VSMCs via different activation of three members of MAPKs resulting from different VSMC subtypes classified by SM-α-actin expression levels. PMID:26488175

  8. Poly(ADP-ribose polymerase 1 is indispensable for transforming growth factor-β Induced Smad3 activation in vascular smooth muscle cell.

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    Dan Huang

    Full Text Available BACKGROUND: Transforming growth factor type-β (TGF-β/Smad pathway plays an essential role in vascular fibrosis. Reactive oxygen species (ROS generation also mediates TGF-β signaling-induced vascular fibrosis, suggesting that some sort of interaction exists between Smad and redox pathways. However, the underlying molecular mechanism is largely unknown. This study aims to investigate the influence of poly(ADP-ribose polymerase 1 (PARP1, a downstream effector of ROS, on TGF-β signaling transduction through Smad3 pathway in rat vascular smooth muscle cells (VSMCs. METHODS AND RESULTS: TGF-β1 treatment promoted PARP1 activation through induction of ROS generation in rat VSMCs. TGF-β1-induced phosphorylation and nuclear accumulation of Smad3 was prevented by treatment of cells with PARP inhibitor, 3-aminobenzamide (3AB or N-(6-oxo-5,6-dihydrophenanthridin-2-yl-2-(N,N-dimethylaminoacetami (PJ34, or PARP1 siRNA. TGF-β1 treatment promoted poly(ADP-ribosylation of Smad3 via activation of PARP1 in the nucleus. Poly(ADP-ribosylation enhanced Smad-Smad binding element (SBE complex formation in nuclear extracts and increased DNA binding activity of Smad3. Pretreatment with 3AB, PJ34, or PARP1 siRNA prevented TGF-β1-induced Smad3 transactivation and expression of Smad3 target genes, including collagen Iα1, collagen IIIα1 and tissue inhibitor of metalloproteinase 1, in rat VSMCs. CONCLUSIONS: PARP1 is indispensable for TGF-β1 induced Smad3 activation in rat VSMCs. Targeting PARP1 may be a promising therapeutic approach against vascular diseases induced by dysregulation of TGF-β/Smad3 pathway.

  9. Angiogenesis is induced by airway smooth muscle strain.

    Science.gov (United States)

    Hasaneen, Nadia A; Zucker, Stanley; Lin, Richard Z; Vaday, Gayle G; Panettieri, Reynold A; Foda, Hussein D

    2007-10-01

    Angiogenesis is an important feature of airway remodeling in both chronic asthma and chronic obstructive pulmonary disease (COPD). Airways in those conditions are exposed to excessive mechanical strain during periods of acute exacerbations. We recently reported that mechanical strain of human airway smooth muscle (HASM) led to an increase in their proliferation and migration. Sustained growth in airway smooth muscle in vivo requires an increase in the nutritional supply to these muscles, hence angiogenesis. In this study, we examined the hypothesis that cyclic mechanical strain of HASM produces factors promoting angiogenic events in the surrounding vascular endothelial cells. Our results show: 1) a significant increase in human lung microvascular endothelial cell (HMVEC-L) proliferation, migration, and tube formation following incubation in conditioned media (CM) from HASM cells exposed to mechanical strain; 2) mechanical strain of HASM cells induced VEGF expression and release; 3) VEGF neutralizing antibodies inhibited the proliferation, migration, and tube formations of HMVEC-L induced by the strained airway smooth muscle CM; 4) mechanical strain of HASM induced a significant increase in hypoxia-inducible factor-1alpha (HIF-1alpha) mRNA and protein, a transcription factor required for VEGF gene transcription; and 5) mechanical strain of HASM induced HIF-1alpha/VEGF through dual phosphatidylinositol 3-kinase (PI3K)/Akt/mammalian target of rapamycin (mTOR) and ERK pathways. In conclusion, exposing HASM cells to mechanical strain induces signal transduction pathway through PI3K/Akt/mTOR and ERK pathways that lead to an increase in HIF-1alpha, a transcription factor required for VEGF expression. VEGF release by mechanical strain of HASM may contribute to the angiogenesis seen with repeated exacerbation of asthma and COPD.

  10. Norepinephrine-induced alteration in the coupling of α1-adrenergic receptor occupancy to calcium efflux in rabbit aortic smooth muscle cells

    International Nuclear Information System (INIS)

    Colucci, W.S.; Alexander, R.W.

    1986-01-01

    To determine whether α-adrenergic desensitization of vascular smooth muscle is due to an alteration in α 1 -adrenergic receptor coupling, the authors determined the relationship between receptor occupancy and maximal receptor-coupled Ca 2+ efflux in cultured rabbit aortic smooth muscle cells (i) under basal conditions as defined by receptor inactivation with phenoxybenzamine and (ii) after 48 hr of exposure to several concentrations of 1-norepinephrine (NE). Neither phenoxybenzamine nor NE exposure caused a change in binding affinity for [ 3 H]prazosin or NE. Maximal [ 3 H]prazosin binding capacity and maximal NE-stimulated 45 Ca 2+ efflux decreased progressively with exposure of incubated cells to increasing concentrations of phenoxybenzamine or NE. An approximately 80% decrease in maximal [ 3 H]prazosin binding capacity caused by either phenoxybenzamine or NE resulted in complete loss of NE-stimulated 45 Ca 2+ efflux, indicating that under these conditions approximately 20% of α 1 -adrenergic receptors are not coupled to the Ca 2+ efflux. Under basal conditions, the relationship between maximal [ 3 H]prazosin binding capacity and maximal NE-stimulated 45 Ca 2+ efflux was markedly nonlinear, so that a near maximal response could be elicited by occupancy of only approximately 40% of the receptors. Thus, an alteration in occupancy-response coupling at a step proximal to Ca 2+ mobilization and/or influx, rather than a reduction in receptor number, is of primary importance in the process of agonist-induced α-adrenergic receptor desensitization of vascular smooth muscle cells

  11. Sensitization of vascular smooth muscle cell to TNF-α-mediated death in the presence of palmitate

    International Nuclear Information System (INIS)

    Rho, Mun-Chual; Ah Lee, Kyeong; Mi Kim, Sun; Sik Lee, Chang; Jeong Jang, Min; Kook Kim, Young; Sun Lee, Hyun; Hyun Choi, Yung; Yong Rhim, Byung; Kim, Koanhoi

    2007-01-01

    Saturated free fatty acids (FFAs), including palmitate, can activate the intrinsic death pathway in cells. However, the relationship between FFAs and receptor-mediated death pathway is still unknown. In this study, we have investigated whether FFAs are able to trigger receptor-mediated death. In addition, to clarify the mechanisms responsible for the activation, we examined the biochemical changes in dying vascular smooth muscle cell (VSMC) and the effects of various molecules to the receptor-mediated VSMC death. Tumor necrosis factor (TNF)-α-mediated VSMC death occurred in the presence of sub-cytotoxic concentration of palmitate as determined by assessing viability and DNA degradation, while the cytokine did not influence VSMC viability in the presence of oleate. The VSMC death was inhibited by the gene transfer of a dominant-negative Fas-associated death domain-containing protein and the baculovirus p35, but not by the bcl-xL or the c-Jun N-terminal kinase (JNK) binding domain of JNK-interacting protein-1, in tests utilizing recombinant adenoviruses. The VSMC death was also inhibited by a neutralizing anti-TNF receptor 1 antibody, the caspase inhibitor z-VAD, and the cathepsin B inhibitor CA074, a finding indicative of the role of both caspases and cathepsin B in this process. Consistent with this finding, caspase-3 activation and an increase in cytosolic cathepsin B activity were detected in the dying VSMC. Palmitate inhibited an increase of TNF-α-mediated nuclear factor kappa B (NF-κB) activity, the survival pathway activated by the cytokine, by hindering the translocation of the NF-κB subunit of p65 from the cytosol into the nucleus. The gene transfer of inhibitor of NF-κB predisposed VSMC to palmitate-induced cell death. To the best of our knowledge, this study is the first report to demonstrate the activation of TNF-α-mediated cell death in the presence of palmitate. The current study proposes that FFAs would take part in deleterious vascular

  12. Alpha1a-Adrenoceptor Genetic Variant Triggers Vascular Smooth Muscle Cell Hyperproliferation and Agonist Induced Hypertrophy via EGFR Transactivation Pathway.

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    Irina Gradinaru

    Full Text Available α1a Adrenergic receptors (α1aARs are the predominant AR subtype in human vascular smooth muscle cells (SMCs. α1aARs in resistance vessels are crucial in the control of blood pressure, yet the impact of naturally occurring human α1aAR genetic variants in cardiovascular disorders remains poorly understood. To this end, we present novel findings demonstrating that 3D cultures of vascular SMCs expressing human α1aAR-247R (247R genetic variant demonstrate significantly increased SMC contractility compared with cells expressing the α1aAR-WT (WT receptor. Stable expression of 247R genetic variant also triggers MMP/EGFR-transactivation dependent serum- and agonist-independent (constitutive hyperproliferation and agonist-dependent hypertrophy of SMCs. Agonist stimulation reduces contractility Using pathway-specific inhibitors we determined that the observed hyperproliferation of 247R-expressing cells is triggered via β-arrestin1/Src/MMP-2/EGFR/ERK-dependent mechanism. MMP-2-specific siRNA inhibited 247R-triggered hyperproliferation indicating MMP-2 involvement in 247R-triggered hyperproliferation in SMCs. β-arrestin1-specific shRNA also inhibited 247R-triggered hyperproliferation but did not affect hypertrophy in 247R-expressing SMCs, indicating that agonist-dependent hypertrophy is independent of β-arrestin1. Our data reveal that in different cardiovascular cells the same human receptor genetic variant can activate alternative modulators of the same signaling pathway. Thus, our findings in SMCs demonstrate that depending on the type of cells expressing the same receptor (or receptor variant, different target-specific inhibitors could be used to modulate aberrant hyperproliferative or hypertrophic pathways in order to restore normal phenotype.

  13. Atorvastatin Protects Vascular Smooth Muscle Cells From TGF-β1-Stimulated Calcification by Inducing Autophagy via Suppression of the β-Catenin Pathway

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    Demin Liu

    2014-01-01

    Full Text Available Background: Arterial calcification is a major event in the progression of atherosclerosis. It is reported that statins exhibit various protective effects against vascular smooth muscle cell (VSMC inflammation and proliferation in cardiovascular remodeling. Although statins counteract atherosclerosis, the molecular mechanisms of statins on the calcium release from VSMCs have not been clearly elucidated. Methods: Calcium content of VSMCs was measured using enzyme-linked immunosorbent assay (ELISA. The expression of proteins involved in cellular transdifferentiation was analyzed by western blot. Cell autophagy was measured by fluorescence microscopic analysis for acridine orange staining and transmission electron microscopy analysis. The autophagic inhibitors (3-MA, chloroquine, NH4Cl and bafilomycin A1 and β-catenin inhibitor JW74 were used to assess the effects of atorvastatin on autophagy and the involvement of β-catenin on cell calcification respectively. Furthermore, cell transfection was performed to overexpress β-catenin. Results: In VSMCs, atorvastatin significantly suppressed transforming growth factor-β1 (TGF-β1-stimulated calcification, accompanied by the induction of autophagy. Downregulation of autophagy with autophagic inhibitors significantly suppressed the inhibitory effect of atorvastatin on cell calcification. Moreover, the beneficial effect of atorvastatin on calcification and autophagy was reversed by β-catenin overexpression. Conversely, JW74 supplement enhanced this effect. Conclusion: These data demonstrated that atorvastatin protect VSMC from TGF-β1-stimulated calcification by inducing autophagy through suppression of the β-catenin pathway, identifying autophagy induction might be a therapeutic strategy for use in vascular calcification.

  14. Stimulatory interactions between human coronary smooth muscle cells and dendritic cells.

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    Sara Paccosi

    Full Text Available Despite inflammatory and immune mechanisms participating to atherogenesis and dendritic cells (DCs driving immune and non-immune tissue injury response, the interactions between DCs and vascular smooth muscle cells (VSMCs possibly relevant to vascular pathology including atherogenesis are still unclear. To address this issue, immature DCs (iDCs generated from CD14+ cells isolated from healthy donors were matured either with cytokines (mDCs, or co-cultured (ccDCs with human coronary artery VSMCs (CASMCs using transwell chambers. Co-culture induced DC immunophenotypical and functional maturation similar to cytokines, as demonstrated by flow cytometry and mixed lymphocyte reaction. In turn, factors from mDCs and ccDCs induced CASMC migration. MCP-1 and TNFα, secreted from DCs, and IL-6 and MCP-1, secreted from CASMCs, were primarily involved. mDCs adhesion to CASMCs was enhanced by CASMC pre-treatment with IFNγ and TNFα ICAM-1 and VCAM-1 were involved, since the expression of specific mRNAs for these molecules increased and adhesion was inhibited by neutralizing antibodies to the counter-receptors CD11c and CD18. Adhesion was also inhibited by CASMC pre-treatment with the HMG-CoA-reductase inhibitor atorvastatin and the PPARγ agonist rosiglitazone, which suggests a further mechanism for the anti-inflammatory action of these drugs. Adhesion of DCs to VSMCs was shown also in vivo in rat carotid 7 to 21 days after crush and incision injury. The findings indicate that DCs and VSMCs can interact with reciprocal stimulation, possibly leading to perpetuate inflammation and vascular wall remodelling, and that the interaction is enhanced by a cytokine-rich inflammatory environment and down-regulated by HMGCoA-reductase inhibitors and PPARγ agonists.

  15. Andrographolide, a Novel NF-κB Inhibitor, Induces Vascular Smooth Muscle Cell Apoptosis via a Ceramide-p47phox-ROS Signaling Cascade

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    Yu-Ying Chen

    2013-01-01

    Full Text Available Atherosclerosis is linked with the development of many cardiovascular complications. Abnormal proliferation of vascular smooth muscle cells (VSMCs plays a crucial role in the development of atherosclerosis. Accordingly, the apoptosis of VSMCs, which occurs in the progression of vascular proliferation, may provide a beneficial strategy for managing cardiovascular diseases. Andrographolide, a novel nuclear factor-κB inhibitor, is the most active and critical constituent isolated from the leaves of Andrographis paniculata. Recent studies have indicated that andrographolide is a potential therapeutic agent for treating cancer through the induction of apoptosis. In this study, the apoptosis-inducing activity and mechanisms in andrographolide-treated rat VSMCs were characterized. Andrographolide significantly induced reactive oxygen species (ROS formation, p53 activation, Bax, and active caspase-3 expression, and these phenomena were suppressed by pretreating the cells with N-acetyl-L-cysteine, a ROS scavenger, or diphenylene iodonium, a nicotinamide adenine dinucleotide phosphate (NADPH oxidase (Nox inhibitor. Furthermore, p47phox, a Nox subunit protein, was phosphorylated in andrographolide-treated rat VSMCs. However, pretreatment with 3-O-methyl-sphingomyelin, a neutral sphingomyelinase inhibitor, significantly inhibited andrographolide-induced p47phox phosphorylation as well as Bax and active caspase-3 expression. Our results collectively demonstrate that andrographolide-reduced cell viability can be attributed to apoptosis in VSMCs, and this apoptosis-inducing activity was associated with the ceramide-p47phox-ROS signaling cascade.

  16. Adenovirus-assisted lipofection: efficient in vitro gene transfer of luciferase and cytosine deaminase to human smooth muscle cells.

    Science.gov (United States)

    Kreuzer, J; Denger, S; Reifers, F; Beisel, C; Haack, K; Gebert, J; Kübler, W

    1996-07-01

    Smooth muscle cells (SMC) are a central cell type involved in multiple processes of coronary artery diseases including restenosis and therefore are major target cells for different aspects of gene transfer. Previous attempts to transfect primary arterial cells using different techniques like liposomes, CaPO4 and electroporation resulted in only low transfection efficiency. The development of recombinant adenoviruses dramatically improved the delivery of foreign genes into different cell types including SMC. However, cloning and identification of recombinants remain difficult and time-consuming techniques. The present study demonstrates that a complex consisting of reporter plasmid encoding firefly luciferase (pLUC), polycationic liposomes and replication-deficient adenovirus was able to yield very high in vitro transfection of primary human smooth muscle cells under optimized conditions. The technique of adenovirus-assisted lipofection (AAL) increases transfer and expression of plasmid DNA in human smooth muscle cells in vitro up to 1000-fold compared to lipofection. To verify the applicability of AAL for gene transfer into human smooth muscle cells we studied a gene therapy approach to suppress proliferation of SMC in vitro, using the prokaryotic cytosine deaminase gene (CD) which enables transfected mammalian cells to deaminate 5-fluorocytosine (5-FC) to the highly toxic 5-fluorouracil (5-FU). The effect of a transient CD expression on RNA synthesis was investigated by means of a cotransfection with a RSV-CD expression plasmid and the luciferase reporter plasmid. Western blot analysis demonstrated high expression of CD protein in transfected SMC. Cotransfected SMC demonstrated two-fold less luciferase activity in the presence of 5-FC (5 mmol/l) after 48 h compared to cells transfected with a non-CD coding plasmid. The data demonstrate that a transient expression of CD could be sufficient to reduce the capacity of protein synthesis in human SMC. This simple and

  17. Structural properties of lipid reconstructs and lipid composition of normotensive and hypertensive rat vascular smooth muscle cell membranes

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    T.R. Oliveira

    2009-09-01

    Full Text Available Multiple cell membrane alterations have been reported to be the cause of various forms of hypertension. The present study focuses on the lipid portion of the membranes, characterizing the microviscosity of membranes reconstituted with lipids extracted from the aorta and mesenteric arteries of spontaneously hypertensive (SHR and normotensive control rat strains (WKY and NWR. Membrane-incorporated phospholipid spin labels were used to monitor the bilayer structure at different depths. The packing of lipids extracted from both aorta and mesenteric arteries of normotensive and hypertensive rats was similar. Lipid extract analysis showed similar phospholipid composition for all membranes. However, cholesterol content was lower in SHR arteries than in normotensive animal arteries. These findings contrast with the fact that the SHR aorta is hyporeactive while the SHR mesenteric artery is hyperreactive to vasopressor agents when compared to the vessels of normotensive animal strains. Hence, factors other than microviscosity of bulk lipids contribute to the vascular smooth muscle reactivity and hypertension of SHR. The excess cholesterol in the arteries of normotensive animal strains apparently is not dissolved in bulk lipids and is not directly related to vascular reactivity since it is present in both the aorta and mesenteric arteries. The lower cholesterol concentrations in SHR arteries may in fact result from metabolic differences due to the hypertensive state or to genes that co-segregate with those that determine hypertension during the process of strain selection.

  18. Emerging Role of Angiotensin Type 2 Receptor (AT2R)/Akt/NO Pathway in Vascular Smooth Muscle Cell in the Hyperthyroidism

    Science.gov (United States)

    Carrillo-Sepúlveda, Maria Alícia; Ceravolo, Graziela S.; Furstenau, Cristina R.; Monteiro, Priscilla de Souza; Bruno-Fortes, Zuleica; Carvalho, Maria Helena; Laurindo, Francisco R.; Tostes, Rita C.; Webb, R. Clinton; Barreto-Chaves, Maria Luiza M.

    2013-01-01

    Hyperthyroidism is characterized by increased vascular relaxation and decreased vascular contraction and is associated with augmented levels of triiodothyronine (T3) that contribute to the diminished systemic vascular resistance found in this condition. T3 leads to augmented NO production via PI3K/Akt signaling pathway, which in turn causes vascular smooth muscle cell (VSMC) relaxation; however, the underlying mechanisms involved remain largely unknown. Evidence from human and animal studies demonstrates that the renin-angiotensin system (RAS) plays a crucial role in vascular function and also mediates some of cardiovascular effects found during hyperthyroidism. Thus, in this study, we hypothesized that type 2 angiotensin II receptor (AT2R), a key component of RAS vasodilatory actions, mediates T3 induced-decreased vascular contraction. Marked induction of AT2R expression was observed in aortas from T3-induced hyperthyroid rats (Hyper). These vessels showed decreased protein levels of the contractile apparatus: α-actin, calponin and phosphorylated myosin light chain (p-MLC). Vascular reactivity studies showed that denuded aortic rings from Hyper rats exhibited decreased maximal contractile response to angiotensin II (AngII), which was attenuated in aortic rings pre-incubated with an AT2R blocker. Further study showed that cultured VSMC stimulated with T3 (0.1 µmol/L) for 24 hours had increased AT2R gene and protein expression. Augmented NO levels and decreased p-MLC levels were found in VSMC stimulated with T3, both of which were reversed by a PI3K/Akt inhibitor and AT2R blocker. These findings indicate for the first time that the AT2R/Akt/NO pathway contributes to decreased contractile responses in rat aorta, promoted by T3, and this mechanism is independent from the endothelium. PMID:23637941

  19. Emerging role of angiotensin type 2 receptor (AT2R/Akt/NO pathway in vascular smooth muscle cell in the hyperthyroidism.

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    Maria Alícia Carrillo-Sepúlveda

    Full Text Available Hyperthyroidism is characterized by increased vascular relaxation and decreased vascular contraction and is associated with augmented levels of triiodothyronine (T3 that contribute to the diminished systemic vascular resistance found in this condition. T3 leads to augmented NO production via PI3K/Akt signaling pathway, which in turn causes vascular smooth muscle cell (VSMC relaxation; however, the underlying mechanisms involved remain largely unknown. Evidence from human and animal studies demonstrates that the renin-angiotensin system (RAS plays a crucial role in vascular function and also mediates some of cardiovascular effects found during hyperthyroidism. Thus, in this study, we hypothesized that type 2 angiotensin II receptor (AT2R, a key component of RAS vasodilatory actions, mediates T3 induced-decreased vascular contraction. Marked induction of AT2R expression was observed in aortas from T3-induced hyperthyroid rats (Hyper. These vessels showed decreased protein levels of the contractile apparatus: α-actin, calponin and phosphorylated myosin light chain (p-MLC. Vascular reactivity studies showed that denuded aortic rings from Hyper rats exhibited decreased maximal contractile response to angiotensin II (AngII, which was attenuated in aortic rings pre-incubated with an AT2R blocker. Further study showed that cultured VSMC stimulated with T3 (0.1 µmol/L for 24 hours had increased AT2R gene and protein expression. Augmented NO levels and decreased p-MLC levels were found in VSMC stimulated with T3, both of which were reversed by a PI3K/Akt inhibitor and AT2R blocker. These findings indicate for the first time that the AT2R/Akt/NO pathway contributes to decreased contractile responses in rat aorta, promoted by T3, and this mechanism is independent from the endothelium.

  20. Adropin Contributes to Anti-Atherosclerosis by Suppressing Monocyte-Endothelial Cell Adhesion and Smooth Muscle Cell Proliferation

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    Kengo Sato

    2018-04-01

    Full Text Available Adropin, a peptide hormone expressed in liver and brain, is known to improve insulin resistance and endothelial dysfunction. Serum levels of adropin are negatively associated with the severity of coronary artery disease. However, it remains unknown whether adropin could modulate atherogenesis. We assessed the effects of adropin on inflammatory molecule expression and human THP1 monocyte adhesion in human umbilical vein endothelial cells (HUVECs, foam cell formation in THP1 monocyte-derived macrophages, and the migration and proliferation of human aortic smooth muscle cells (HASMCs in vitro and atherogenesis in Apoe−/− mice in vivo. Adropin was expressed in THP1 monocytes, their derived macrophages, HASMCs, and HUVECs. Adropin suppressed tumor necrosis factor α-induced THP1 monocyte adhesion to HUVECs, which was associated with vascular cell adhesion molecule 1 and intercellular adhesion molecule 1 downregulation in HUVECs. Adropin shifted the phenotype to anti-inflammatory M2 rather than pro-inflammatory M1 via peroxisome proliferator-activated receptor γ upregulation during monocyte differentiation into macrophages. Adropin had no significant effects on oxidized low-density lipoprotein-induced foam cell formation in macrophages. In HASMCs, adropin suppressed the migration and proliferation without inducing apoptosis via ERK1/2 and Bax downregulation and phosphoinositide 3-kinase/Akt/Bcl2 upregulation. Chronic administration of adropin to Apoe−/− mice attenuated the development of atherosclerotic lesions in the aorta, with reduced the intra-plaque monocyte/macrophage infiltration and smooth muscle cell content. Thus, adropin could serve as a novel therapeutic target in atherosclerosis and related diseases.

  1. Microtissues Enhance Smooth Muscle Differentiation and Cell Viability of hADSCs for Three Dimensional Bioprinting

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    Jin Yipeng

    2017-07-01

    Full Text Available Smooth muscle differentiated human adipose derived stem cells (hADSCs provide a crucial stem cell source for urinary tissue engineering, but the induction of hADSCs for smooth muscle differentiation still has several issues to overcome, including a relatively long induction time and equipment dependence, which limits access to abundant stem cells within a short period of time for further application. Three-dimensional (3D bioprinting holds great promise in regenerative medicine due to its controllable construction of a designed 3D structure. When evenly mixed with bioink, stem cells can be spatially distributed within a bioprinted 3D structure, thus avoiding drawbacks such as, stem cell detachment in a conventional cell-scaffold strategy. Notwithstanding the advantages mentioned above, cell viability is often compromised during 3D bioprinting, which is often due to pressure during the bioprinting process. The objective of our study was to improve the efficiency of hADSC smooth muscle differentiation and cell viability of a 3D bioprinted structure. Here, we employed the hanging-drop method to generate hADSC microtissues in a smooth muscle inductive medium containing human transforming growth factor β1 and bioprinted the induced microtissues onto a 3D structure. After 3 days of smooth muscle induction, the expression of α-smooth muscle actin and smoothelin was higher in microtissues than in their counterpart monolayer cultured hADSCs, as confirmed by immunofluorescence and western blotting analysis. The semi-quantitative assay showed that the expression of α-smooth muscle actin (α-SMA was 0.218 ± 0.077 in MTs and 0.082 ± 0.007 in Controls; smoothelin expression was 0.319 ± 0.02 in MTs and 0.178 ± 0.06 in Controls. Induced MTs maintained their phenotype after the bioprinting process. Live/dead and cell count kit 8 assays showed that cell viability and cell proliferation in the 3D structure printed with microtissues were higher at all time

  2. Intimal smooth muscle cells are a source but not a sensor of anti-inflammatory CYP450 derived oxylipins

    Energy Technology Data Exchange (ETDEWEB)

    Thomson, Scott [Comparative Biomedical Sciences, Royal Veterinary College, Royal College Street, London NW1 0TU (United Kingdom); Edin, Matthew L.; Lih, Fred B. [Division of Intramural Research, NIEHS/NIH, Research Triangle Park, NC 27709 (United States); Davies, Michael [Comparative Biomedical Sciences, Royal Veterinary College, Royal College Street, London NW1 0TU (United Kingdom); Yaqoob, Muhammad M. [Barts and the London, Queen Mary University, Charterhouse Square, London EC1M 6BQ (United Kingdom); Hammock, Bruce D. [Department of Entomology and Comprehensive Cancer Center, University of California, Davies, CA 95616-8584 (United States); Gilroy, Derek [University College London, University Street, London (United Kingdom); Zeldin, Darryl C. [Division of Intramural Research, NIEHS/NIH, Research Triangle Park, NC 27709 (United States); Bishop-Bailey, David, E-mail: dbishopbailey@rvc.ac.uk [Comparative Biomedical Sciences, Royal Veterinary College, Royal College Street, London NW1 0TU (United Kingdom)

    2015-08-07

    Vascular pathologies are associated with changes in the presence and expression of morphologically distinct vascular smooth muscle cells. In particular, in complex human vascular lesions and models of disease in pigs and rodents, an intimal smooth muscle cell (iSMC) which exhibits a stable epithelioid or rhomboid phenotype in culture is often found to be present in high numbers, and may represent the reemergence of a distinct developmental vascular smooth muscle cell phenotype. The CYP450-oxylipin - soluble epoxide hydrolase (sEH) pathway is currently of great interest in targeting for cardiovascular disease. sEH inhibitors limit the development of hypertension, diabetes, atherosclerosis and aneurysm formation in animal models. We have investigated the expression of CYP450-oxylipin-sEH pathway enzymes and their metabolites in paired intimal (iSMC) and medial (mSMC) cells isolated from rat aorta. iSMC basally released significantly larger amounts of epoxy-oxylipin CYP450 products from eicosapentaenoic acid > docosahexaenoic acid > arachidonic acid > linoleic acid, and expressed higher levels of CYP2C12, CYP2B1, but not CYP2J mRNA compared to mSMC. When stimulated with the pro-inflammatory TLR4 ligand LPS, epoxy-oxylipin production did not change greatly in iSMC. In contrast, LPS induced epoxy-oxylipin products in mSMC and induced CYP2J4. iSMC and mSMC express sEH which metabolizes primary epoxy-oxylipins to their dihydroxy-counterparts. The sEH inhibitors TPPU or AUDA inhibited LPS-induced NFκB activation and iNOS induction in mSMC, but had no effect on NFκB nuclear localization or inducible nitric oxide synthase in iSMC; effects which were recapitulated in part by addition of authentic epoxy-oxylipins. iSMCs are a rich source but not a sensor of anti-inflammatory epoxy-oxylipins. Complex lesions that contain high levels of iSMCs may be more resistant to the protective effects of sEH inhibitors. - Highlights: • We examined oxylipin production in different

  3. Vascular smooth muscle responsiveness to nitric oxide is reduced in healthy adults with increased adiposity

    OpenAIRE

    Christou, Demetra D.; Pierce, Gary L.; Walker, Ashley E.; Hwang, Moon-Hyon; Yoo, Jeung-Ki; Luttrell, Meredith; Meade, Thomas H.; English, Mark; Seals, Douglas R.

    2012-01-01

    Vascular smooth muscle responsiveness to nitric oxide, as assessed by nitroglycerin-induced dilation (NID), is impaired in clinical cardiovascular disease, but its relation to adiposity is unknown. We determined the relation of NID to total and abdominal adiposity in healthy adults varying widely in adiposity. In 224 men and women [age, 18–79 years; body mass index (BMI), 16.4–42.2 kg/m2], we measured NID (brachial artery dilation to 0.4 mg sublingual nitroglycerin), total body adiposity [BMI...

  4. Differentiation of Human Adipose Derived Stem Cells into Smooth Muscle Cells Is Modulated by CaMKIIγ

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    Kaisaier Aji

    2016-01-01

    Full Text Available The multifunctional Ca2+/calmodulin-dependent protein kinase II (CaMKII is known to participate in maintenance and switches of smooth muscle cell (SMC phenotypes. However, which isoform of CaMKII is involved in differentiation of adult mesenchymal stem cells into contractile SMCs remains unclear. In the present study, we detected γ isoform of CaMKII in differentiation of human adipose derived stem cells (hASCs into SMCs that resulted from treatment with TGF-β1 and BMP4 in combination for 7 days. The results showed that CaMKIIγ increased gradually during differentiation of hASCs as determined by real-time PCR and western blot analysis. The siRNA-mediated knockdown of CaMKIIγ decreased the protein levels and transcriptional levels of smooth muscle contractile markers (a-SMA, SM22a, calponin, and SM-MHC, while CaMKIIγ overexpression increases the transcriptional and protein levels of smooth muscle contractile markers. These results suggested that γ isoform of CaMKII plays a significant role in smooth muscle differentiation of hASCs.

  5. PDGF activates K-Cl cotransport through phosphoinositide 3-kinase and protein phosphatase-1 in primary cultures of vascular smooth muscle cells.

    Science.gov (United States)

    Zhang, Jing; Lauf, Peter K; Adragna, Norma C

    2005-07-15

    K-Cl cotransport (K-Cl COT, KCC) is an electroneutrally coupled movement of K and Cl present in most cells. In this work, we studied the pathways of regulation of K-Cl COT by platelet-derived growth factor (PDGF) in primary cultures of vascular smooth muscle cells (VSMCs). Wortmannin and LY 294002 blocked the PDGF-induced K-Cl COT activation, indicating that the phosphoinositide 3-kinase (PI 3-K) pathway is involved. However, PD 98059 had no effect on K-Cl COT activation by PDGF, suggesting that the mitogen-activated protein kinase pathway is not involved under the experimental conditions tested. Involvement of phosphatases was also examined. Sodium orthovanadate, cyclosporin A and okadaic acid had no effect on PDGF-stimulated K-Cl COT. Calyculin A blocked the PDGF-stimulated K-Cl COT by 60%, suggesting that protein phosphatase-1 (PP-1) is a mediator in the PDGF signaling pathway/s. In conclusion, our results indicate that the PDGF-mediated pathways of K-Cl COT regulation involve the signaling molecules PI 3-K and PP-1.

  6. Smitin, a novel smooth muscle titin-like protein, interacts with myosin filaments in vivo and in vitro.

    Science.gov (United States)

    Kim, Kyoungtae; Keller, Thomas C S

    2002-01-07

    Smooth muscle cells use an actin-myosin II-based contractile apparatus to produce force for a variety of physiological functions, including blood pressure regulation and gut peristalsis. The organization of the smooth muscle contractile apparatus resembles that of striated skeletal and cardiac muscle, but remains much more poorly understood. We have found that avian vascular and visceral smooth muscles contain a novel, megadalton protein, smitin, that is similar to striated muscle titin in molecular morphology, localization in a contractile apparatus, and ability to interact with myosin filaments. Smitin, like titin, is a long fibrous molecule with a globular domain on one end. Specific reactivities of an anti-smitin polyclonal antibody and an anti-titin monoclonal antibody suggest that smitin and titin are distinct proteins rather than differentially spliced isoforms encoded by the same gene. Smitin immunofluorescently colocalizes with myosin in chicken gizzard smooth muscle, and interacts with two configurations of smooth muscle myosin filaments in vitro. In physiological ionic strength conditions, smitin and smooth muscle myosin coassemble into irregular aggregates containing large sidepolar myosin filaments. In low ionic strength conditions, smitin and smooth muscle myosin form highly ordered structures containing linear and polygonal end-to-end and side-by-side arrays of small bipolar myosin filaments. We have used immunogold localization and sucrose density gradient cosedimentation analyses to confirm association of smitin with both the sidepolar and bipolar smooth muscle myosin filaments. These findings suggest that the titin-like protein smitin may play a central role in organizing myosin filaments in the contractile apparatus and perhaps in other structures in smooth muscle cells.

  7. Genipin inhibits TNF-α-induced vascular smooth muscle cell proliferation and migration via induction of HO-1.

    Directory of Open Access Journals (Sweden)

    Fengrong Jiang

    Full Text Available Vascular smooth muscle cell (VSMC proliferation and migration triggered by inflammatory stimuli contributes importantly to the pathogenesis of atherosclerosis and restenosis. On the other hand, genipin, an aglycon of geniposide, exhibits diverse pharmacological functions such as antitumor and anti-inflammatory effects. The protective effects of genipin on the cardiovascular system have also been reported. However, the molecular mechanism involved remains unknown. This study aimed to elucidate the precise function of genipin in VSMCs, focusing particularly on the role of heme oxygenase-1 (HO-1, a potent anti-inflammatory enzyme. We found that pretreatment of genipin induced HO-1 mRNA and protein levels, as well as its activity in VSMCs. Genipin inhibited TNF-α-induced VSMC proliferation and migration in a dose-dependent manner. At the molecular level, genipin prevented ERK/MAPK and Akt phosphorylation while left p38 MAPK and JNK unchanged. Genipin also blocked the increase of ROS generation induced by TNF-α. More importantly, the specific HO-1 siRNA partially abolished the beneficial effects of genipin on VSMCs. These results suggest that genipin may serve as a novel drug in the treatment of these pathologies by inducing HO-1 expression/activity and subsequently decreasing VSMC proliferation and migration.

  8. Erythroxylum pungens elicits vasorelaxation by reducing intracellular calcium concentration in vascular smooth muscle cells of rats

    Directory of Open Access Journals (Sweden)

    Aurylene C. Oliveira

    2012-01-01

    Full Text Available The cardiovascular effects elicited by the ethanolic extract obtained from the roots of Erythroxylum pungens O.E. Schulz, Erythroxylaceae (EEEP and the vasorelaxant effect induced by its main tropane alkaloid (pungencine were investigated. In normotensive rats, administration of EEEP (1, 10, 30 and 60 mg/kg i.v., randomly produced dose-dependent hypotension (-2±1, -7±0.5 -17.6±1, -24±1 Δ mmHg, n=5 followed by tachycardia (3±0.5, 7±2, 7.1±1, 10±5 Δ bpm, n=5. In intact phenylephrine (Phe, 10 µM-pre-contracted rings, EEEP (0.01-500 µg/mL induced concentration-dependent vasorelaxation (EC50 13.7±5.5 µg/mL, Maximal Response= 92±2.6%, and this effect was unchanged after the removal of the vascular endothelium (EC50 27.2±4.7 µg/ml, Maximal Response= 88.3±3.3 %. In KCl (80 mM-pre-contracted-endothelium-denuded rings, EEEP elicited concentration-dependent relaxation (EC50= 128.2±11.2 µg/mL, Maximal Response 76.8±3.4%. Vasorelaxation has also been achieved with tonic contractions evoked by the L-type Ca2+ channel agonist Bay K 8644 (EC50 80.2±9.1 µg/mL, Maximal Response 86.3±8.3%. In addition, in a depolarizing medium, EEEP inhibited CaCl2 (30-500 µg/mL induced contractions and caused a concentration-dependent rightward shift of the relaxation curves. Lastly, the tropane alkaloid pungencine caused vasorelaxation in mesenteric arteries resembling to the EEEP responses. These results suggests that EEEP induces hypotension and vasorelaxation, at least in part, due to the reduction in [Ca2+]i in vascular smooth muscle cells.

  9. Magnolol inhibits migration of vascular smooth muscle cells via cytoskeletal remodeling pathway to attenuate neointima formation

    International Nuclear Information System (INIS)

    Karki, Rajendra; Kim, Seong-Bin; Kim, Dong-Wook

    2013-01-01

    Background: Increased proliferation and migration of vascular smooth muscle cells (VSMCs) contribute importantly to the formation of both atherosclerotic and restenotic lesions. The objective of this study was to investigate the effect of magnolol on VSMC migration. Methods: The proteolytic activity of matrix metalloproteinases (MMPs) in tumor necrosis factor alpha (TNF-α) stimulated VSMCs was performed by gelatin zymography. VSMC migration was assessed by wound healing and Boyden chamber methods. Collagen induced VSMC adhesion was determined by spectrofluorimeter and stress fibers formation was evaluated by fluorescence microscope. The expression of signaling molecules involved in stress fibers formation was determined by western blot. The phosphorylation of myosin light chain (MLC20) was determined by urea-glycerol polyacrylamide gel electrophoresis. Immunohistochemistry was performed to determine the expression of β1-integrin and collagen type I in the injured carotid arteries of rats on day 35 after vascular injury. Results: VSMC migration was strongly inhibited by magnolol without affecting MMPs expression. Also, magnolol inhibited β1-integrin expression, FAK phosphorylation and RhoA and Cdc42 activation to inhibit the collagen induced stress fibers formation. Moreover, magnolol inhibited the phosphorylation of MLC20. Our in vivo results showed that magnolol inhibited β1-integrin expression, collagen type I deposition and FAK phosphorylation in injured carotid arteries without affecting MMP-2 activity. Conclusions: Magnolol inhibited VSMC migration via inhibition of cytoskeletal remodeling pathway to attenuate neointima formation. General significance: This study provides a rationale for further evaluation of magnolol for the management of atherosclerosis and restenosis. - Highlights: • Magnolol strongly inhibited migration of VSMCs. • Magnolol inhibited stress fibers formation. • MLC20 phosphorylation was also inhibited by magnolol. • Anti

  10. Magnolol inhibits migration of vascular smooth muscle cells via cytoskeletal remodeling pathway to attenuate neointima formation

    Energy Technology Data Exchange (ETDEWEB)

    Karki, Rajendra [Division of Pharmacology and Toxicology, School of Pharmacy, University of Missouri-Kansas City (United States); Department of Oriental Medicine Resources, Mokpo National University (Korea, Republic of); Kim, Seong-Bin [Jeollanamdo Development Institute for Korean Traditional Medicine, Jangheung gun, Jeollanamdo (Korea, Republic of); Kim, Dong-Wook, E-mail: dbkim@mokpo.ac.kr [Department of Oriental Medicine Resources, Mokpo National University (Korea, Republic of)

    2013-12-10

    Background: Increased proliferation and migration of vascular smooth muscle cells (VSMCs) contribute importantly to the formation of both atherosclerotic and restenotic lesions. The objective of this study was to investigate the effect of magnolol on VSMC migration. Methods: The proteolytic activity of matrix metalloproteinases (MMPs) in tumor necrosis factor alpha (TNF-α) stimulated VSMCs was performed by gelatin zymography. VSMC migration was assessed by wound healing and Boyden chamber methods. Collagen induced VSMC adhesion was determined by spectrofluorimeter and stress fibers formation was evaluated by fluorescence microscope. The expression of signaling molecules involved in stress fibers formation was determined by western blot. The phosphorylation of myosin light chain (MLC20) was determined by urea-glycerol polyacrylamide gel electrophoresis. Immunohistochemistry was performed to determine the expression of β1-integrin and collagen type I in the injured carotid arteries of rats on day 35 after vascular injury. Results: VSMC migration was strongly inhibited by magnolol without affecting MMPs expression. Also, magnolol inhibited β1-integrin expression, FAK phosphorylation and RhoA and Cdc42 activation to inhibit the collagen induced stress fibers formation. Moreover, magnolol inhibited the phosphorylation of MLC20. Our in vivo results showed that magnolol inhibited β1-integrin expression, collagen type I deposition and FAK phosphorylation in injured carotid arteries without affecting MMP-2 activity. Conclusions: Magnolol inhibited VSMC migration via inhibition of cytoskeletal remodeling pathway to attenuate neointima formation. General significance: This study provides a rationale for further evaluation of magnolol for the management of atherosclerosis and restenosis. - Highlights: • Magnolol strongly inhibited migration of VSMCs. • Magnolol inhibited stress fibers formation. • MLC20 phosphorylation was also inhibited by magnolol. • Anti

  11. Role of Smooth Muscle in Intestinal Inflammation

    Directory of Open Access Journals (Sweden)

    Stephen M Collins

    1996-01-01

    Full Text Available The notion that smooth muscle function is altered in inflammation is prompted by clinical observations of altered motility in patients with inflammatory bowel disease (IBD. While altered motility may reflect inflammation-induced changes in intrinsic or extrinsic nerves to the gut, changes in gut hormone release and changes in muscle function, recent studies have provided in vitro evidence of altered muscle contractility in muscle resected from patients with ulcerative colitis or Crohn’s disease. In addition, the observation that smooth muscle cells are more numerous and prominent in the strictured bowel of IBD patients compared with controls suggests that inflammation may alter the growth of intestinal smooth muscle. Thus, inflammation is associated with changes in smooth muscle growth and contractility that, in turn, contribute to important symptoms of IBD including diarrhea (from altered motility and pain (via either altered motility or stricture formation. The involvement of smooth muscle in this context may be as an innocent bystander, where cells and products of the inflammatory process induce alterations in muscle contractility and growth. However, it is likely that intestinal muscle cells play a more active role in the inflammatory process via the elaboration of mediators and trophic factors, including cytokines, and via the production of collagen. The concept of muscle cells as active participants in the intestinal inflammatory process is a new concept that is under intense study. This report summarizes current knowledge as it relates to these two aspects of altered muscle function (growth and contractility in the inflamed intestine, and will focus on mechanisms underlying these changes, based on data obtained from animal models of intestinal inflammation.

  12. Nodular smooth muscle metaplasia in multiple peritoneal endometriosis.

    Science.gov (United States)

    Kim, Hyun-Soo; Yoon, Gun; Ha, Sang Yun; Song, Sang Yong

    2015-01-01

    We report here an unusual presentation of peritoneal endometriosis with smooth muscle metaplasia as multiple protruding masses on the lateral pelvic wall. Smooth muscle metaplasia is a common finding in rectovaginal endometriosis, whereas in peritoneal endometriosis, smooth muscle metaplasia is uncommon and its nodular presentation on the pelvic wall is even rarer. To the best of our knowledge, this is the first case of nodular smooth muscle metaplasia occurring in peritoneal endometriosis. As observed in this case, when performing laparoscopic surgery in order to excise malignant tumors of intra-abdominal or pelvic organs, it can be difficult for surgeons to distinguish the metastatic tumors from benign nodular pelvic wall lesions, including endometriosis, based on the gross findings only. Therefore, an intraoperative frozen section biopsy of the pelvic wall nodules should be performed to evaluate the peritoneal involvement by malignant tumors. Moreover, this report implies that peritoneal endometriosis, as well as rectovaginal endometriosis, can clinically present as nodular lesions if obvious smooth muscle metaplasia is present. The pathological investigation of smooth muscle cells in peritoneal lesions can contribute not only to the precise diagnosis but also to the structure and function of smooth muscle cells and related cells involved in the histogenesis of peritoneal endometriosis.

  13. Vasostatin-2 inhibits cell proliferation and adhesion in vascular smooth muscle cells, which are associated with the progression of atherosclerosis

    Energy Technology Data Exchange (ETDEWEB)

    Hou, Jianghong, E-mail: jianghonghou@163.com [Department of Cardiovascular, Weinan Center Hospital, The Middle of Victory Avenue, Linwei District, Weinan City 714000 (China); Xue, Xiaolin [Department of Cardiovascular, The First Affiliated Hospital, College of Medicine, Xi' an Jiaotong University, Xi' an 710061 (China); Li, Junnong [Department of Cardiovascular, Weinan Center Hospital, The Middle of Victory Avenue, Linwei District, Weinan City 714000 (China)

    2016-01-22

    Recently, the serum expression level of vasostatin-2 was found to be reduced and is being studied as an important indicator to assess the presence and severity of coronary artery disease; the functional properties of vasostatin-2 and its relationship with the development of atherosclerosis remains unclear. In this study, we attempted to detect the expression of vasostatin-2 and its impact on human vascular smooth muscle cells (VSMCs). Quantitative real-time PCR (qRT-PCR) and western blot were used to assess the expression level of vasostatin-2 in VSMCs between those from atherosclerosis and disease-free donors; we found that vasostatin-2 was significantly down-regulated in atherosclerosis patient tissues and cell lines. In addition, the over-expression of vasostatin-2 apparently inhibits cell proliferation and migration in VSMCs. Gain-of-function in vitro experiments further show that vasostatin-2 over-expression significantly inhibits inflammatory cytokines release in VSMCs. In addition, cell adhesion experimental analysis showed that soluble adhesion molecules (sICAM-1, sVCAM-1) had decreased expression when vasostatin-2 was over-expressed in VSMCs. Therefore, our results indicate that vasostatin-2 is an atherosclerosis-related factor that can inhibit cell proliferation, inflammatory response and cell adhesion in VSMCs. Taken together, our results indicate that vasostatin-2 could serve as a potential diagnostic biomarker and therapeutic option for human atherosclerosis in the near future. - Highlights: • Vasostatin-2 levels were down-regulated in atherosclerosis patient tissues and VSMCs. • Ectopic expression of vasostatin-2 directly affects cell proliferation and migration in vitro. • Ectopic expression of vasostatin-2 protein affects pro-inflammatory cytokines release in VSMCs. • Ectopic expression of vasostatin-2 protein affects cell adhesion in VSMCs.

  14. Vascular Mural Cells Promote Noradrenergic Differentiation of Embryonic Sympathetic Neurons

    Directory of Open Access Journals (Sweden)

    Vitor Fortuna

    2015-06-01

    Full Text Available The sympathetic nervous system controls smooth muscle tone and heart rate in the cardiovascular system. Postganglionic sympathetic neurons (SNs develop in close proximity to the dorsal aorta (DA and innervate visceral smooth muscle targets. Here, we use the zebrafish embryo to ask whether the DA is required for SN development. We show that noradrenergic (NA differentiation of SN precursors temporally coincides with vascular mural cell (VMC recruitment to the DA and vascular maturation. Blocking vascular maturation inhibits VMC recruitment and blocks NA differentiation of SN precursors. Inhibition of platelet-derived growth factor receptor (PDGFR signaling prevents VMC differentiation and also blocks NA differentiation of SN precursors. NA differentiation is normal in cloche mutants that are devoid of endothelial cells but have VMCs. Thus, PDGFR-mediated mural cell recruitment mediates neurovascular interactions between the aorta and sympathetic precursors and promotes their noradrenergic differentiation.

  15. The Hippo pathway mediates inhibition of vascular smooth muscle cell proliferation by cAMP.

    Science.gov (United States)

    Kimura, Tomomi E; Duggirala, Aparna; Smith, Madeleine C; White, Stephen; Sala-Newby, Graciela B; Newby, Andrew C; Bond, Mark

    2016-01-01

    Inhibition of vascular smooth muscle cell (VSMC) proliferation by intracellular cAMP prevents excessive neointima formation and hence angioplasty restenosis and vein-graft failure. These protective effects are mediated via actin-cytoskeleton remodelling and subsequent regulation of gene expression by mechanisms that are incompletely understood. Here we investigated the role of components of the growth-regulatory Hippo pathway, specifically the transcription factor TEAD and its co-factors YAP and TAZ in VSMC. Elevation of cAMP using forskolin, dibutyryl-cAMP or the physiological agonists, Cicaprost or adenosine, significantly increased phosphorylation and nuclear export YAP and TAZ and inhibited TEAD-luciferase report gene activity. Similar effects were obtained by inhibiting RhoA activity with C3-transferase, its downstream kinase, ROCK, with Y27632, or actin-polymerisation with Latrunculin-B. Conversely, expression of constitutively-active RhoA reversed the inhibitory effects of forskolin on TEAD-luciferase. Forskolin significantly inhibited the mRNA expression of the pro-mitogenic genes, CCN1, CTGF, c-MYC and TGFB2 and this was reversed by expression of constitutively-active YAP or TAZ phospho-mutants. Inhibition of YAP and TAZ function with RNAi or Verteporfin significantly reduced VSMC proliferation. Furthermore, the anti-mitogenic effects of forskolin were reversed by overexpression of constitutively-active YAP or TAZ. Taken together, these data demonstrate that cAMP-induced actin-cytoskeleton remodelling inhibits YAP/TAZ-TEAD dependent expression of pro-mitogenic genes in VSMC. This mechanism contributes novel insight into the anti-mitogenic effects of cAMP in VSMC and suggests a new target for intervention. Copyright © 2015 The Authors. Published by Elsevier Ltd.. All rights reserved.

  16. Nonprenylated Xanthones from Gentiana lutea, Frasera caroliniensis, and Centaurium erythraea as Novel Inhibitors of Vascular Smooth Muscle Cell Proliferation.

    Science.gov (United States)

    Waltenberger, Birgit; Liu, Rongxia; Atanasov, Atanas G; Schwaiger, Stefan; Heiss, Elke H; Dirsch, Verena M; Stuppner, Hermann

    2015-11-13

    Aberrant proliferation of vascular smooth muscle cells (VSMC) plays a major role in restenosis, the pathological renarrowing of the blood vessel lumen after surgical treatment of stenosis. Since available anti-proliferative pharmaceuticals produce unfavorable side effects, there is high demand for the identification of novel VSMC proliferation inhibitors. A natural product screening approach using a resazurin conversion assay enabled the identification of gentisin (1) from Gentiana lutea as a novel inhibitor of VSMC proliferation with an IC50 value of 7.84 µM. Aiming to identify further anti-proliferative compounds, 13 additional nonprenylated xanthones, isolated from different plant species, were also tested. While some compounds showed no or moderate activity at 30 µM, 1-hydroxy-2,3,4,5-tetramethoxyxanthone (4), swerchirin (6), and methylswertianin (7) showed IC50 values between 10.2 and 12.5 µM. The anti-proliferative effect of 1, 4, 6, and 7 was confirmed by the quantification of DNA synthesis (BrdU incorporation) in VSMC. Cell death quantification (determined by LDH release in the culture medium) revealed that the compounds are not cytotoxic in the investigated concentration range. In conclusion, nonprenylated xanthones are identified as novel, non-toxic VSMC proliferation inhibitors, which might contribute to the development of new therapeutic applications to combat restenosis.

  17. Nonprenylated Xanthones from Gentiana lutea, Frasera caroliniensis, and Centaurium erythraea as Novel Inhibitors of Vascular Smooth Muscle Cell Proliferation

    Directory of Open Access Journals (Sweden)

    Birgit Waltenberger

    2015-11-01

    Full Text Available Aberrant proliferation of vascular smooth muscle cells (VSMC plays a major role in restenosis, the pathological renarrowing of the blood vessel lumen after surgical treatment of stenosis. Since available anti-proliferative pharmaceuticals produce unfavorable side effects, there is high demand for the identification of novel VSMC proliferation inhibitors. A natural product screening approach using a resazurin conversion assay enabled the identification of gentisin (1 from Gentiana lutea as a novel inhibitor of VSMC proliferation with an IC50 value of 7.84 µM. Aiming to identify further anti-proliferative compounds, 13 additional nonprenylated xanthones, isolated from different plant species, were also tested. While some compounds showed no or moderate activity at 30 µM, 1-hydroxy-2,3,4,5-tetramethoxyxanthone (4, swerchirin (6, and methylswertianin (7 showed IC50 values between 10.2 and 12.5 µM. The anti-proliferative effect of 1, 4, 6, and 7 was confirmed by the quantification of DNA synthesis (BrdU incorporation in VSMC. Cell death quantification (determined by LDH release in the culture medium revealed that the compounds are not cytotoxic in the investigated concentration range. In conclusion, nonprenylated xanthones are identified as novel, non-toxic VSMC proliferation inhibitors, which might contribute to the development of new therapeutic applications to combat restenosis.

  18. Interstitial cells of Cajal in human small intestine. Ultrastructural identification and organization between the main smooth muscle layers

    DEFF Research Database (Denmark)

    Rumessen, Jüri Johannes; Thuneberg, Lars

    1991-01-01

    Anatomy, interstitial cells of Cajal, small intestine, gut motility, pacemaker cells, smooth muscle......Anatomy, interstitial cells of Cajal, small intestine, gut motility, pacemaker cells, smooth muscle...

  19. A cyclic GMP-dependent calcium-activated chloride current in smooth-muscle cells from rat mesenteric resistance arteries

    DEFF Research Database (Denmark)

    Matchkov, Vladimir; Aalkjær, Christian; Nilsson, Holger

    2004-01-01

    We have previously demonstrated the presence of a cyclic GMP (cGMP)-dependent calcium-activated inward current in vascular smooth-muscle cells, and suggested this to be of importance in synchronizing smooth-muscle contraction. Here we demonstrate the characteristics of this current. Using......M) in the pipette solution. The current was found to be a calcium-activated chloride current with an absolute requirement for cyclic GMP (EC50 6.4 microM). The current could be activated by the constitutively active subunit of PKG. Current activation was blocked by the protein kinase G antagonist Rp-8-Br-PET-cGMP...... differed from those of the calcium-activated chloride current in pulmonary myocytes, which was cGMP-independent, exhibited a high sensitivity to inhibition by niflumic acid, was unaffected by zinc ions, and showed outward current rectification as has previously been reported for this current. Under...

  20. Regulation of GPCR-mediated smooth muscle contraction : implications for asthma and pulmonary hypertension

    NARCIS (Netherlands)

    Wright, D B; Tripathi, S; Sikarwar, A; Santosh, K T; Perez-Zoghbi, J; Ojo, O O; Irechukwu, N; Ward, J P T; Schaafsma, D

    Contractile G-protein-coupled receptors (GPCRs) have emerged as key regulators of smooth muscle contraction, both under healthy and diseased conditions. This brief review will discuss some key topics and novel insights regarding GPCR-mediated airway and vascular smooth muscle contraction as

  1. Mechanisms of mechanical strain memory in airway smooth muscle.

    Science.gov (United States)

    Kim, Hak Rim; Hai, Chi-Ming

    2005-10-01

    We evaluated the hypothesis that mechanical deformation of airway smooth muscle induces structural remodeling of airway smooth muscle cells, thereby modulating mechanical performance in subsequent contractions. This hypothesis implied that past experience of mechanical deformation was retained (or "memorized") as structural changes in airway smooth muscle cells, which modulated the cell's subsequent contractile responses. We termed this phenomenon mechanical strain memory. Preshortening has been found to induce attenuation of both force and isotonic shortening velocity in cholinergic receptor-activated airway smooth muscle. Rapid stretching of cholinergic receptor-activated airway smooth muscle from an initial length to a final length resulted in post-stretch force and myosin light chain phosphorylation that correlated significantly with initial length. Thus post-stretch muscle strips appeared to retain memory of the initial length prior to rapid stretch (mechanical strain memory). Cytoskeletal recruitment of actin- and integrin-binding proteins and Erk 1/2 MAPK appeared to be important mechanisms of mechanical strain memory. Sinusoidal length oscillation led to force attenuation during oscillation and in subsequent contractions in intact airway smooth muscle, and p38 MAPK appeared to be an important mechanism. In contrast, application of local mechanical strain to cultured airway smooth muscle cells induced local actin polymerization and cytoskeletal stiffening. It is conceivable that deep inspiration-induced bronchoprotection may be a manifestation of mechanical strain memory such that mechanical deformation from past breathing cycles modulated the mechanical performance of airway smooth muscle in subsequent cycles in a continuous and dynamic manner.

  2. Iptakalim inhibits PDGF-BB-induced human airway smooth muscle cells proliferation and migration

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Wenrui; Kong, Hui; Zeng, Xiaoning; Wang, Jingjing; Wang, Zailiang; Yan, Xiaopei; Wang, Yanli; Xie, Weiping, E-mail: wpxie@njmu.edu.cn; Wang, Hong, E-mail: hongwang@njmu.edu.cn

    2015-08-15

    Chronic airway diseases are characterized by airway remodeling which is attributed partly to the proliferation and migration of airway smooth muscle cells (ASMCs). ATP-sensitive potassium (K{sub ATP}) channels have been identified in ASMCs. Mount evidence has suggested that K{sub ATP} channel openers can reduce airway hyperresponsiveness and alleviate airway remodeling. Opening K{sup +} channels triggers K{sup +} efflux, which leading to membrane hyperpolarization, preventing Ca{sup 2+}entry through closing voltage-operated Ca{sup 2+} channels. Intracellular Ca{sup 2+} is the most important regulator of muscle contraction, cell proliferation and migration. K{sup +} efflux decreases Ca{sup 2+} influx, which consequently influences ASMCs proliferation and migration. As a K{sub ATP} channel opener, iptakalim (Ipt) has been reported to restrain the proliferation of pulmonary arterial smooth muscle cells (PASMCs) involved in vascular remodeling, while little is known about its impact on ASMCs. The present study was designed to investigate the effects of Ipt on human ASMCs and the mechanisms underlying. Results obtained from cell counting kit-8 (CCK-8), flow cytometry and 5-ethynyl-2′-deoxyuridine (EdU) incorporation showed that Ipt significantly inhibited platelet-derived growth factor (PDGF)-BB-induced ASMCs proliferation. ASMCs migration induced by PDGF-BB was also suppressed by Ipt in transwell migration and scratch assay. Besides, the phosphorylation of Ca{sup 2+}/calmodulin-dependent kinase II (CaMKII), extracellular regulated protein kinases 1/2 (ERK1/2), protein kinase B (Akt), and cyclic adenosine monophosphate (cAMP) response element binding protein (CREB) were as well alleviated by Ipt administration. Furthermore, we found that the inhibition of Ipt on the PDGF-BB-induced proliferation and migration in human ASMCs was blocked by glibenclamide (Gli), a selective K{sub ATP} channel antagonist. These findings provide a strong evidence to support that Ipt

  3. Gentiana lutea exerts anti-atherosclerotic effects by preventing endothelial inflammation and smooth muscle cell migration.

    Science.gov (United States)

    Kesavan, R; Chandel, S; Upadhyay, S; Bendre, R; Ganugula, R; Potunuru, U R; Giri, H; Sahu, G; Kumar, P Uday; Reddy, G Bhanuprakash; Joksic, G; Bera, A K; Dixit, Madhulika

    2016-04-01

    Studies suggest that Gentiana lutea (GL), and its component isovitexin, may exhibit anti-atherosclerotic properties. In this study we sought to investigate the protective mechanism of GL aqueous root extract and isovitexin on endothelial inflammation, smooth muscle cell migation, and on the onset and progression of atherosclerosis in streptozotocin (STZ)-induced diabetic rats. Our results show that both GL extract and isovitexin, block leukocyte adhesion and generation of reactive oxygen species in human umbilical vein endothelial cells (HUVECs) and rat aortic smooth muscle cells (RASMCs), following TNF-alpha and platelet derived growth factor-BB (PDGF-BB) challenges respectively. Both the extract and isovitexin blocked TNF-α induced expression of ICAM-1 and VCAM-1 in HUVECs. PDGF-BB induced migration of RASMCs and phospholipase C-γ activation, were also abrogated by GL extract and isovitexin. Fura-2 based ratiometric measurements demonstrated that, both the extact, and isovitexin, inhibit PDGF-BB mediated intracellular calcium rise in RASMCs. Supplementation of regular diet with 2% GL root powder for STZ rats, reduced total cholesterol in blood. Oil Red O staining demonstrated decreased lipid accumulation in aortic wall of diabetic animals upon treatment with GL. Medial thickness and deposition of collagen in the aortic segment of diabetic rats were also reduced upon supplementation. Immunohistochemistry demonstrated reduced expression of vascular cell adhesion molecule-1 (VCAM-1), inducible nitric oxide synthase (iNOS), and vascular endothelial cadherin (VE-cadherin) in aortic segments of diabetic rats following GL treatment. Thus, our results support that GL root extract/powder and isovitexin exhibit anti-atherosclerotic activities. Copyright © 2016 The Italian Society of Diabetology, the Italian Society for the Study of Atherosclerosis, the Italian Society of Human Nutrition, and the Department of Clinical Medicine and Surgery, Federico II University

  4. Microtubule Regulation of Kv7 Channels Orchestrates cAMP-Mediated Vasorelaxations in Rat Arterial Smooth Muscle

    DEFF Research Database (Denmark)

    Lindman, Johanna; Khammy, Makhala M; Lundegaard, Pia R

    2018-01-01

    Microtubules can regulate GPCR (G protein-coupled receptor) signaling in various cell types. In vascular smooth muscle, activation of the β-adrenoceptor leads to production of cAMP to mediate a vasorelaxation. Little is known about the role of microtubules in smooth muscle, and given the importance...... of renal and mesenteric arteries that the microtubule stabilizer, paclitaxel, prevented. Sharp microelectrode experiments showed that colchicine treatment caused increased hyperpolarization of mesenteric artery segments in response to isoprenaline. Application of the Kv7 channel blocker, XE991, attenuated...

  5. Role of blood and vascular smooth muscle in the vasoactivity of nitrite

    Science.gov (United States)

    Liu, Taiming; Schroeder, Hobe J.; Barcelo, Lisa; Bragg, Shannon L.; Terry, Michael H.; Wilson, Sean M.; Power, Gordon G.

    2014-01-01

    Recent evidence from humans and rats indicates that nitrite is a vasodilator under hypoxic conditions by reacting with metal-containing proteins to produce nitric oxide (NO). We tested the hypothesis that near-physiological concentrations of nitrite would produce vasodilation in a hypoxia- and concentration-dependent manner in the hind limb of sheep. Anesthetized sheep were instrumented to measure arterial blood pressure and femoral blood flows continuously in both hind limbs. Nitrite was infused into one femoral artery to raise the nitrite concentration in the femoral vein by 10 to 15-fold while the sheep breathed 50%, 14% or 12% oxygen in inspired air. In contrast to reports in humans and rats, the nitrite infusion had no measurable effect on mean femoral blood flows or vascular conductances, regardless of inspired O2 levels. In vitro experiments showed no significant difference in the release of NO from nitrite in sheep and human red blood cells. Further experiments demonstrated nitrite is converted to NO in rat artery homogenates faster than sheep arteries, and that this source of NO production is attenuated in the presence of a heme oxidizer. Finally, western blots indicate that concentrations of the heme-containing protein cytoglobin, but not myoglobin, are markedly lower in sheep arteries compared with rats. Overall, the results demonstrate that nitrite is not a physiological vasodilator in sheep. This is likely due to a lack of conversion of nitrite to NO within the vascular smooth muscle, perhaps due to deficient amounts of the heme-containing protein cytoglobin. PMID:25108012

  6. A new paradigm for the role of smooth muscle cells in the human cervix.

    Science.gov (United States)

    Vink, Joy Y; Qin, Sisi; Brock, Clifton O; Zork, Noelia M; Feltovich, Helen M; Chen, Xiaowei; Urie, Paul; Myers, Kristin M; Hall, Timothy J; Wapner, Ronald; Kitajewski, Jan K; Shawber, Carrie J; Gallos, George

    2016-10-01

    Premature cervical remodeling resulting in spontaneous preterm birth may begin with premature failure or relaxation at the internal os (termed "funneling"). To date, we do not understand why the internal os fails or why funneling occurs in some cases of premature cervical remodeling. Although the human cervix is thought to be mostly collagen with minimal cellular content, cervical smooth muscle cells are present in the cervix and can cause cervical tissue contractility. To understand why the internal os relaxes or why funneling occurs in some cases of premature cervical remodeling, we sought to evaluate cervical smooth muscle cell content and distribution throughout human cervix and correlate if cervical smooth muscle organization influences regional cervical tissue contractility. Using institutional review board-approved protocols, nonpregnant women cervix, whole cervical slices were obtained from the internal os, midcervix, and external os and immunostained with smooth muscle actin. To correlate tissue structure with function, whole slices from the internal and external os were stimulated to contract with 1 μmol/L of oxytocin in organ baths. In separate samples, we tested if the cervix responds to a common tocolytic, nifedipine. Cervical slices from the internal os were treated with oxytocin alone or oxytocin + increasing doses of nifedipine to generate a dose response and half maximal inhibitory concentration. Student t test was used where appropriate. Cervical tissue was collected from 41 women. Immunohistochemistry showed cervical smooth muscle cells at the internal and external os expressed mature smooth muscle cell markers and contraction-associated proteins. The cervix exhibited a gradient of cervical smooth muscle cells. The area of the internal os contained 50-60% cervical smooth muscle cells that were circumferentially organized in the periphery of the stroma, which may resemble a sphincter-like pattern. The external os contained approximately 10

  7. Multipotent embryonic isl1+ progenitor cells lead to cardiac, smooth muscle, and endothelial cell diversification.

    Science.gov (United States)

    Moretti, Alessandra; Caron, Leslie; Nakano, Atsushi; Lam, Jason T; Bernshausen, Alexandra; Chen, Yinhong; Qyang, Yibing; Bu, Lei; Sasaki, Mika; Martin-Puig, Silvia; Sun, Yunfu; Evans, Sylvia M; Laugwitz, Karl-Ludwig; Chien, Kenneth R

    2006-12-15

    Cardiogenesis requires the generation of endothelial, cardiac, and smooth muscle cells, thought to arise from distinct embryonic precursors. We use genetic fate-mapping studies to document that isl1(+) precursors from the second heart field can generate each of these diverse cardiovascular cell types in vivo. Utilizing embryonic stem (ES) cells, we clonally amplified a cellular hierarchy of isl1(+) cardiovascular progenitors, which resemble the developmental precursors in the embryonic heart. The transcriptional signature of isl1(+)/Nkx2.5(+)/flk1(+) defines a multipotent cardiovascular progenitor, which can give rise to cells of all three lineages. These studies document a developmental paradigm for cardiogenesis, where muscle and endothelial lineage diversification arises from a single cell-level decision of a multipotent isl1(+) cardiovascular progenitor cell (MICP). The discovery of ES cell-derived MICPs suggests a strategy for cardiovascular tissue regeneration via their isolation, renewal, and directed differentiation into specific mature cardiac, pacemaker, smooth muscle, and endothelial cell types.

  8. Vascular Mural Cells Promote Noradrenergic Differentiation of Embryonic Sympathetic Neurons.

    Science.gov (United States)

    Fortuna, Vitor; Pardanaud, Luc; Brunet, Isabelle; Ola, Roxana; Ristori, Emma; Santoro, Massimo M; Nicoli, Stefania; Eichmann, Anne

    2015-06-23

    The sympathetic nervous system controls smooth muscle tone and heart rate in the cardiovascular system. Postganglionic sympathetic neurons (SNs) develop in close proximity to the dorsal aorta (DA) and innervate visceral smooth muscle targets. Here, we use the zebrafish embryo to ask whether the DA is required for SN development. We show that noradrenergic (NA) differentiation of SN precursors temporally coincides with vascular mural cell (VMC) recruitment to the DA and vascular maturation. Blocking vascular maturation inhibits VMC recruitment and blocks NA differentiation of SN precursors. Inhibition of platelet-derived growth factor receptor (PDGFR) signaling prevents VMC differentiation and also blocks NA differentiation of SN precursors. NA differentiation is normal in cloche mutants that are devoid of endothelial cells but have VMCs. Thus, PDGFR-mediated mural cell recruitment mediates neurovascular interactions between the aorta and sympathetic precursors and promotes their noradrenergic differentiation. Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.

  9. Vardenafil inhibiting parasympathetic function of tracheal smooth muscle.

    Science.gov (United States)

    Lee, Fei-Peng; Chao, Pin-Zhir; Wang, Hsing-Won

    2018-07-01

    Levitra, a phosphodiesterase-5 (PDE5) inhibitor, is the trade name of vardenafil. Nowadays, it is applied to treatment of erectile dysfunction. PDE5 inhibitors are employed to induce dilatation of the vascular smooth muscle. The effect of Levitra on impotency is well known; however, its effect on the tracheal smooth muscle has rarely been explored. When administered for sexual symptoms via oral intake or inhalation, Levitra might affect the trachea. This study assessed the effects of Levitra on isolated rat tracheal smooth muscle by examining its effect on resting tension of tracheal smooth muscle, contraction caused by 10 -6  M methacholine as a parasympathetic mimetic, and electrically induced tracheal smooth muscle contractions. The results showed that adding methacholine to the incubation medium caused the trachea to contract in a dose-dependent manner. Addition of Levitra at doses of 10 -5  M or above elicited a significant relaxation response to 10 -6  M methacholine-induced contraction. Levitra could inhibit electrical field stimulation-induced spike contraction. It alone had minimal effect on the basal tension of the trachea as the concentration increased. High concentrations of Levitra could inhibit parasympathetic function of the trachea. Levitra when administered via oral intake might reduce asthma attacks in impotent patients because it might inhibit parasympathetic function and reduce methacholine-induced contraction of the tracheal smooth muscle. Copyright © 2018. Published by Elsevier Taiwan LLC.

  10. Vitamin D modulates tissue factor and protease-activated receptor 2 expression in vascular smooth muscle cells.

    Science.gov (United States)

    Martinez-Moreno, Julio M; Herencia, Carmen; Montes de Oca, Addy; Muñoz-Castañeda, Juan R; Rodríguez-Ortiz, M Encarnación; Díaz-Tocados, Juan M; Peralbo-Santaella, Esther; Camargo, Antonio; Canalejo, Antonio; Rodriguez, Mariano; Velasco-Gimena, Francisco; Almaden, Yolanda

    2016-03-01

    Clinical and epidemiologic studies reveal an association between vitamin D deficiency and increased risk of cardiovascular disease. Because vascular smooth muscle cell (VSMC)-derived tissue factor (TF) is suggested to be critical for arterial thrombosis, we investigated whether the vitamin D molecules calcitriol and paricalcitol could reduce the expression of TF induced by the proinflammatory cytokine TNF-α in human aortic VSMCs. We found that, compared with controls, incubation with TNF-α increased TF expression and procoagulant activity in a NF-κB-dependent manner, as deduced from the increased nuclear translocation of nuclear factor κ-light-chain-enhancer of activated B cells protein 65 (p65-NF-κB) and direct interaction of NF-κB to the TF promoter. This was accompanied by the up-regulation of TF signaling mediator protease-activated receptor 2 (PAR-2) expression and by the down-regulation of vitamin D receptor expression in a miR-346-dependent way. However, addition of calcitriol or paricalcitol blunted the TNF-α-induced TF expression and activity (2.01 ± 0.24 and 1.32 ± 0.14 vs. 3.02 ± 0.39 pmol/mg protein, P < 0.05), which was associated with down-regulation of NF-κB signaling and PAR-2 expression, as well as with restored levels of vitamin D receptor and enhanced expression of TF pathway inhibitor. Our data suggest that inflammation promotes a prothrombotic state through the up-regulation of TF function in VSMCs and that the beneficial cardiovascular effects of vitamin D may be partially due to decreases in TF expression and its activity in VSMCs. © FASEB.

  11. Delayed effects of cold atmospheric plasma on vascular cells

    NARCIS (Netherlands)

    Stoffels, Eva; Roks, Anton J. M.; Deelmm, Leo E.

    2008-01-01

    We investigated the long-term behaviour of vascular cells (endothelial and smooth muscle) after exposure to a cold atmospheric plasma source. The cells were treated through a gas-permeable membrane, in order to simulate intravenous treatment with a gas plasma-filled catheter. Such indirect treatment

  12. α-smooth muscle actin is not a marker of fibrogenic cell activity in skeletal muscle fibrosis.

    Directory of Open Access Journals (Sweden)

    Wanming Zhao

    Full Text Available α-Smooth muscle actin (α-SMA is used as a marker for a subset of activated fibrogenic cells, myofibroblasts, which are regarded as important effector cells of tissue fibrogenesis. We address whether α-SMA-expressing myofibroblasts are detectable in fibrotic muscles of mdx5cv mice, a mouse model for Duchenne muscular dystrophy (DMD, and whether the α-SMA expression correlates with the fibrogenic function of intramuscular fibrogenic cells. α-SMA immunostaining signal was not detected in collagen I (GFP-expressing cells in fibrotic muscles of ColI-GFP/mdx5cv mice, but it was readily detected in smooth muscle cells lining intramuscular blood vessel walls. α-SMA expression was detected by quantitative RT-PCR and Western blot in fibrogenic cells sorted from diaphragm and quadriceps muscles of the ColI-GFP/mdx5cv mice. Consistent with the more severe fibrosis in the ColI-GFP/mdx5cv diaphragm, the fibrogenic cells in the diaphragm exerted a stronger fibrogenic function than the fibrogenic cells in the quadriceps as gauged by their extracellular matrix gene expression. However, both gene and protein expression of α-SMA was lower in the diaphragm fibrogenic cells than in the quadriceps fibrogenic cells in the ColI-GFP/mdx5cv mice. We conclude that myofibroblasts are present in fibrotic skeletal muscles, but their expression of α-SMA is not detectable by immunostaining. The level of α-SMA expression by intramuscular fibrogenic cells does not correlate positively with the level of collagen gene expression or the severity of skeletal muscle fibrosis in the mdx5cv mice. α-SMA is not a functional marker of fibrogenic cells in skeletal muscle fibrosis associated with muscular dystrophy.

  13. Perfusion of veins at arterial pressure increases the expression of KLF5 and cell cycle genes in smooth muscle cells

    International Nuclear Information System (INIS)

    Amirak, Emre; Zakkar, Mustafa; Evans, Paul C.; Kemp, Paul R.

    2010-01-01

    Vascular smooth muscle cell (VSMC) proliferation remains a major cause of veno-arterial graft failure. We hypothesised that exposure of venous SMCs to arterial pressure would increase KLF5 expression and that of cell cycle genes. Porcine jugular veins were perfused at arterial or venous pressure in the absence of growth factors. The KLF5, c-myc, cyclin-D and cyclin-E expression were elevated within 24 h of perfusion at arterial pressure but not at venous pressure. Arterial pressure also reduced the decline in SM-myosin heavy chain expression. These data suggest a role for KLF5 in initiating venous SMCs proliferation in response to arterial pressure.

  14. Experimental study of the effect of 103Pd on the proliferation and apoptosis of vascular smooth muscle cells

    International Nuclear Information System (INIS)

    Luo Quanyong; Zhu Jun; Chen Libo; Lu Hankui; Zhu Ruisen

    2002-01-01

    Objective: To investigate the ability of γ emitting radionuclide 103 Pd to inhibit the vascular smooth muscle cell (SMC) proliferation and to induce its apoptosis in vitro. Methods: 103 Pd solution was added to the culture medium to irradiate SMCs for 72 h and non-radioactive Pd solution was added as control. 3 H-TdR incorporation test was used to detect the effect of 103 Pd on the proliferation of SMCs. Flow cytometer was used to detect the apoptotic SMCs. Results: The results showed that inhibition of SMC proliferation was evident and the effects were dose-dependant. Inhibition rate of SMC proliferation by 1.85 MBq 103 Pd was 2.3% , which was not significant. The inhibition rate increased from 41.6% to 91.2% as the dose of 103 Pd increased from 7.4 to 37.0 MBq, and the proliferation of SMCs was repressed significantly then. The apoptosis rate was extremely low (less than 4.0% ) with the 103 pd dose escalating from 1.85 to 37.0 MBq. Conclusions: This study suggests that proliferation of SMCs can be repressed effectively in vitro by 103 pd. 103 Pd can be used to inhibit the neointimal proliferation. 103 Pd radioactive stent implantation can be employed as a possible novel means to prevent restenosis

  15. Heparin modulates human intestinal smooth muscle (HISM) cell proliferation and matrix production

    International Nuclear Information System (INIS)

    Graham, M.; Perr, H.; Drucker, D.E.; Diegelmann, R.F.

    1986-01-01

    (HISM) cell proliferation and collagen production may play a role in the pathogenesis of intestinal stricture in Crohn's disease. The present studies were performed to evaluate the effects of heparin, a known modulator of vascular smooth muscle cells, on HISM cell proliferation and collagen production. Heparin (100 μg/ml) was added daily to HISM cell cultures for cell proliferation studies and for 24 hours at various time points during culture for collagen synthesis studies. Collagen synthesis was determined by the uptake of 3 H proline into collagenase-sensitive protein. Heparin completely inhibited cell proliferation for 7 days, after which cell numbers increased but at a slower rate than controls. Cells released from heparin inhibition demonstrated catch-up growth to control levels. Collagen production was significantly inhibited by 24 hours exposure to heparin but only at those times during culture when collagen synthesis was maximal (8 to 12 days). Non-collagen protein synthesis was inhibited by heparin at all time points during culture. Heparin through its modulation of HISM cells may play an important role in the control of the extracellular matrix of the intestinal wall

  16. Metformin inhibits inflammatory response via AMPK–PTEN pathway in vascular smooth muscle cells

    International Nuclear Information System (INIS)

    Kim, Sun Ae; Choi, Hyoung Chul

    2012-01-01

    Highlights: ► PTEN was induced by metformin and inhibited by compound C and AMPK siRNA. ► Metformin suppressed TNF-α-induced COX-2 and iNOS mRNA expression. ► Compound C and bpv (pic) increased iNOS and COX-2 protein expression. ► NF-κB activation was restored by inhibiting AMPK and PTEN. ► AMPK and PTEN regulated TNF-α-induced ROS production in VSMCs. -- Abstract: Atherosclerosis is a chronic inflammation of the coronary arteries. Vascular smooth muscle cells (VSMCs) stimulated by cytokines and chemokines accelerate the inflammatory response and migrate to the injured endothelium during the progression of atherosclerosis. Activation of AMP activated protein kinase (AMPK), a key sensor maintaining metabolic homeostasis, suppresses the inflammatory response. However, how AMPK regulates the inflammatory response is poorly understood. To identify the mechanism of this response, we focused on phosphatase and tensin homolog (PTEN), which is a negative regulator of inflammation. We investigated that activation of AMPK-induced PTEN expression and suppression of the inflammatory response through the AMPK–PTEN pathway in VSMCs. We treated with the well-known AMPK activator metformin to induce PTEN expression. PTEN was induced by metformin (2 mM) and inhibited by compound C (10 μM) and AMPK siRNA. Tumor necrosis factor-alpha (TNF-α) was used to induce inflammation. The inflammatory response was confirmed by cyclooxygenase (COX)-2, inducible nitric oxide synthase (iNOS) expression, and activation of nuclear factor (NF)-κB. Metformin suppressed COX-2 and iNOS mRNA and protein expression dose dependently. Treatment with compound C and bpv (pic) in the presence of metformin, iNOS and COX-2 protein expression increased. NF-κB activation decreased in response to metformin and was restored by inhibiting AMPK and PTEN. Inhibiting AMPK and PTEN restored ROS levels stimulated with TNF-α. Taken together, PTEN could be a possible downstream regulator of AMPK, and the

  17. Overexpression of membrane sialic acid-specific sialidase Neu3 inhibits matrix metalloproteinase-9 expression in vascular smooth muscle cells

    International Nuclear Information System (INIS)

    Moon, Sung-Kwon; Cho, Seung-Hak; Kim, Kyung-Woon; Jeon, Jae Heung; Ko, Jeong-Heon; Kim, Bo Yeon; Kim, Cheorl-Ho

    2007-01-01

    The ganglioside-specific sialidase Neu3 has been suggested to participate in cell growth, migration, and differentiation. Recent reports suggest that sialidase may be involved in intimal thickening, an early stage in the development of atherosclerosis. However, the role of the Neu3 gene in vascular smooth muscle cells (VSMC) responses has not yet been elucidated. To determine whether a Neu3 is able to modulate VSMC growth, the effect of overexpression of the Neu3 gene on cell proliferation was examined. However, the results show that the overexpression of this gene has no effect on DNA synthesis and ERK phosphorylation in cultured VSMC in the presence of TNF-α. Because atherogenic effects need not be limited to proliferation, we decided to examine whether Neu3 exerted inhibitory effects on matrix metalloproteinase-9 (MMP-9) activity in TNF-α-induced VSMC. The expression of the Neu3 gene led to the inhibition of TNF-α-induced matrix metalloproteinase-9 (MMP-9) expression in VSMC as determined by zymography and immunoblot. Furthermore, Neu3 gene expression strongly decreased MMP-9 promoter activity in response to TNF-α. This inhibition was characterized by the down-regulation of MMP-9, which was transcriptionally regulated at NF-κB and activation protein-1 (AP-1) sites in the MMP-9 promoter. These findings suggest that the Neu3 gene represents a physiological modulator of VSMC responses that may contribute to plaque instability in atherosclerosis

  18. Effects and underlying mechanisms of curcumin on the proliferation of vascular smooth muscle cells induced by Chol:MβCD

    International Nuclear Information System (INIS)

    Qin Li; Yang Yunbo; Tuo Qinhui; Zhu Bingyang; Chen Linxi; Zhang Liang; Liao Duanfang

    2009-01-01

    Proliferation of vascular smooth muscle cells (VSMCs) contributes to the development of various cardiovascular diseases. Curcumin, extracted from Curcumae longae, has been shown a variety of beneficial effects on human health, including anti-atherosclerosis by mechanisms poorly understood. In the present study, we attempted to investigate whether curcumin has any effect on VSMCs proliferation and the potential mechanisms involved. Our data showed curcumin concentration-dependently abrogated the proliferation of primary rat VSMCs induced by Chol:MβCD. To explore the underlying cellular and molecular mechanisms, we found that curcumin was capable of restoring caveolin-1 expression which was reduced by Chol:MβCD treatment. Moreover, curcumin abrogated the increment of phospho-ERK1/2 and nuclear accumulation of ERK1/2 in primary rat VSMCs induced by Chol:MβCD, which led to a suppression of AP-1 promoter activity stimulated by Chol:MβCD. In addition, curcumin was able to reverse cell cycle progression induced by Chol:MβCD, which was further supported by its down-regulation of cyclinD1 and E2F promoter activities in the presence of Chol:MβCD. Taking together, our data suggest curcumin inhibits Chol:MβCD-induced VSMCs proliferation via restoring caveolin-1 expression that leads to the suppression of over-activated ERK signaling and causes cell cycle arrest at G1/S phase. These novel findings support the beneficial potential of curcumin in cardiovascular disease.

  19. Effects of High Glucose on Vascular Endothelial Growth Factor Synthesis and Secretion in Aortic Vascular Smooth Muscle Cells from Obese and Lean Zucker Rats

    Directory of Open Access Journals (Sweden)

    Mariella Trovati

    2012-07-01

    Full Text Available Type 1 diabetes is characterized by insulin deficiency, type 2 by both insulin deficiency and insulin resistance: in both conditions, hyperglycaemia is accompanied by an increased cardiovascular risk, due to increased atherosclerotic plaque formation/instabilization and impaired collateral vessel formation. An important factor in these phenomena is the Vascular Endothelial Growth Factor (VEGF, a molecule produced also by Vascular Smooth Muscle Cells (VSMC. We aimed at evaluating the role of high glucose on VEGF-A164 synthesis and secretion in VSMC from lean insulin-sensitive and obese insulin-resistant Zucker rats (LZR and OZR. In cultured aortic VSMC from LZR and OZR incubated for 24 h with D-glucose (5.5, 15 and 25 mM or with the osmotic controls L-glucose and mannitol, we measured VEGF-A164 synthesis (western, blotting and secretion (western blotting and ELISA. We observed that: (i D-glucose dose-dependently increases VEGF-A164 synthesis and secretion in VSMC from LZR and OZR (n = 6, ANOVA p = 0.002–0.0001; (ii all the effects of 15 and 25 mM D-glucose are attenuated in VSMC from OZR vs. LZR (p = 0.0001; (iii L-glucose and mannitol reproduce the VEGF-A164 modulation induced by D-glucose in VSMC from both LZR and OZR. Thus, glucose increases via an osmotic mechanism VEGF synthesis and secretion in VSMC, an effect attenuated in the presence of insulin resistance.

  20. Human mesenchymal stem cells cultured on silk hydrogels with variable stiffness and growth factor differentiate into mature smooth muscle cell phenotype.

    Science.gov (United States)

    Floren, Michael; Bonani, Walter; Dharmarajan, Anirudh; Motta, Antonella; Migliaresi, Claudio; Tan, Wei

    2016-02-01

    Cell-matrix and cell-biomolecule interactions play critical roles in a diversity of biological events including cell adhesion, growth, differentiation, and apoptosis. Evidence suggests that a concise crosstalk of these environmental factors may be required to direct stem cell differentiation toward matured cell type and function. However, the culmination of these complex interactions to direct stem cells into highly specific phenotypes in vitro is still widely unknown, particularly in the context of implantable biomaterials. In this study, we utilized tunable hydrogels based on a simple high pressure CO2 method and silk fibroin (SF) the structural protein of Bombyx mori silk fibers. Modification of SF protein starting water solution concentration results in hydrogels of variable stiffness while retaining key structural parameters such as matrix pore size and β-sheet crystallinity. To further resolve the complex crosstalk of chemical signals with matrix properties, we chose to investigate the role of 3D hydrogel stiffness and transforming growth factor (TGF-β1), with the aim of correlating the effects on the vascular commitment of human mesenchymal stem cells. Our data revealed the potential to upregulate matured vascular smooth muscle cell phenotype (myosin heavy chain expression) of hMSCs by employing appropriate matrix stiffness and growth factor (within 72h). Overall, our observations suggest that chemical and physical stimuli within the cellular microenvironment are tightly coupled systems involved in the fate decisions of hMSCs. The production of tunable scaffold materials that are biocompatible and further specialized to mimic tissue-specific niche environments will be of considerable value to future tissue engineering platforms. This article investigates the role of silk fibroin hydrogel stiffness and transforming growth factor (TGF-β1), with the aim of correlating the effects on the vascular commitment of human mesenchymal stem cells. Specifically, we

  1. Effects of infection with recombinant adenovirus on human vascular endothelial and smooth muscle cells

    NARCIS (Netherlands)

    Quax, P.H.A.; Lamfers, M.L.M.; Grimbergen, J.M.; Teeling, J.; Hoeben, R.C.; Nieuw Amerongen, G.P. van; Hinsbergh, V.W.M. van

    1996-01-01

    The plasminogen activation (PA) system is involved in vascular remodelling. Modulating its activity in vascular cells might be a way to interfere in processes such as angiogenesis and restenosis. Adenoviral vectors have become a favourable tool for direct gene transfer into vascular cells. In the

  2. Early Transcriptomic Response to LDL and oxLDL in Human Vascular Smooth Muscle Cells.

    Directory of Open Access Journals (Sweden)

    Salvador Damián-Zamacona

    Full Text Available Although nowadays it is well known that the human transcriptome can importantly vary according to external or environmental condition, the reflection of this concept when studying oxidative stress and its direct relationship with gene expression profiling during the process of atherogenesis has not been thoroughly achieved.The ability to analyze genome-wide gene expression through transcriptomics has shown that the genome responds dynamically to diverse stimuli. Here, we describe the transcriptome of human vascular smooth muscle cells (hVSMC stimulated by native and oxidized low-density lipoprotein (nLDL and oxLDL respectively, with the aim of assessing the early molecular changes that induce a response in this cell type resulting in a transcriptomic transformation. This expression has been demonstrated in atherosclerotic plaques in vivo and in vitro, particularly in the light of the oxidative modification hypothesis of atherosclerosis.Total RNA was isolated with TRIzol reagent (Life Technologies and quality estimated using an Agilent 2100 bioanalyzer. The transcriptome of hVSMC under different experimental conditions (1,5 and 24 hours for nLDL and oxLDL was obtained using the GeneChip Human Gene 1.0 ST (Affymetrix designed to measure gene expression of 28,869 well-annotated genes. A fixed fold-change cut-off corresponding to ± 2 was used to identify genes exhibiting the most significant variation and statistical significance (P< 0.05, and 8 genes validated by qPCR using Taqman probes.10 molecular processes were significantly affected in hVSMC: Apoptosis and cell cycle, extracellular matrix remodeling, DNA repair, cholesterol efflux, cGMP biosynthesis, endocytic mechanisms, calcium homeostasis, redox balance, membrane trafficking and finally, the immune response to inflammation. The evidence we present supporting the hypothesis for the involvement of oxidative modification of several processes and metabolic pathways in atherosclerosis is

  3. Macrophages control vascular stem/progenitor cell plasticity through tumor necrosis factor-α-mediated nuclear factor-κB activation.

    Science.gov (United States)

    Wong, Mei Mei; Chen, Yikuan; Margariti, Andriani; Winkler, Bernhard; Campagnolo, Paola; Potter, Claire; Hu, Yanhua; Xu, Qingbo

    2014-03-01

    Vascular lineage differentiation of stem/progenitor cells can contribute to both tissue repair and exacerbation of vascular diseases such as in vein grafts. The role of macrophages in controlling vascular progenitor differentiation is largely unknown and may play an important role in graft development. This study aims to identify the role of macrophages in vascular stem/progenitor cell differentiation and thereafter elucidate the mechanisms that are involved in the macrophage- mediated process. We provide in vitro evidence that macrophages can induce endothelial cell (EC) differentiation of the stem/progenitor cells while simultaneously inhibiting their smooth muscle cell differentiation. Mechanistically, both effects were mediated by macrophage-derived tumor necrosis factor-α (TNF-α) via TNF-α receptor 1 and canonical nuclear factor-κB activation. Although the overexpression of p65 enhanced EC (or attenuated smooth muscle cell) differentiation, p65 or TNF-α receptor 1 knockdown using lentiviral short hairpin RNA inhibited EC (or rescued smooth muscle cell) differentiation in response to TNF-α. Furthermore, TNF-α-mediated EC differentiation was driven by direct binding of nuclear factor-κB (p65) to specific VE-cadherin promoter sequences. Subsequent experiments using an ex vivo decellularized vessel scaffold confirmed an increase in the number of ECs and reduction in smooth muscle cell marker expression in the presence of TNF-α. The lack of TNF-α in a knockout mouse model of vein graft decreased endothelialization and significantly increased thrombosis formation. Our study highlights the role of macrophages in directing vascular stem/progenitor cell lineage commitment through TNF-α-mediated TNF-α receptor 1 and nuclear factor-κB activation that is likely required for endothelial repair in vascular diseases such as vein graft.

  4. Antiproliferative effect of UTP on human arterial and venous smooth muscle cells.

    Science.gov (United States)

    White, P J; Kumari, R; Porter, K E; London, N J; Ng, L L; Boarder, M R

    2000-12-01

    We have investigated the hypothesis that responses associated with proliferation are regulated by extracellular nucleotides such as ATP and UTP in cultured human vascular smooth muscle cells (VSMC) derived from internal mammary artery (IMA) and saphenous vein (SV). Platelet-derived growth factor (PDGF), ATP, and UTP each generated an increase in cytosolic free Ca(2+) concentration ([Ca(2+)](i)) in both IMA- and SV-derived cells in the absence of detectable inositol 1,4,5-trisphosphate production. ATP alone had no effect on [(3)H]thymidine incorporation into DNA, but with a submaximal concentration of PDGF it raised [(3)H]thymidine incorporation in SV- but not IMA-derived cells. UTP alone also was without effect on [(3)H]thymidine incorporation or cell number. However, in both SV- and IMA-derived cells, UTP reduced the PDGF-stimulated [(3)H]thymidine response and PDGF-stimulated cell proliferation. This cannot be explained by an inhibitory effect on the p42/p44 mitogen-activated protein kinase (MAPK) cascade, since this response to PDGF was not attenuated by UTP. We conclude that, in human VSMC of both arterial and venous origin, UTP acts as an anti-proliferative regulator.

  5. The combination of lanthanum chloride and the calcimimetic calindol delays the progression of vascular smooth muscle cells calcification

    Energy Technology Data Exchange (ETDEWEB)

    Ciceri, Paola; Volpi, Elisa; Brenna, Irene; Elli, Francesca [Renal Division and Laboratory of Experimental Nephrology, Dipartimento di Medicina e Chirurgia, Universita di Milano, Milan (Italy); Borghi, Elisa [Dipartimento di Salute Pubblica, Microbiologia e Virologia, Universita di Milano, Milan (Italy); Brancaccio, Diego [Renal Division and Laboratory of Experimental Nephrology, Dipartimento di Medicina e Chirurgia, Universita di Milano, Milan (Italy); Cozzolino, Mario, E-mail: mario.cozzolino@unimi.it [Renal Division and Laboratory of Experimental Nephrology, Dipartimento di Medicina e Chirurgia, Universita di Milano, Milan (Italy)

    2012-02-24

    Highlights: Black-Right-Pointing-Pointer Lanthanum reduces the progression of high phosphate-induced calcium deposition. Black-Right-Pointing-Pointer Calcium receptor agonists and the calcimimetic calindol reduce calcium deposition. Black-Right-Pointing-Pointer Lanthanum and calindol cooperate on reducing calcium deposition. Black-Right-Pointing-Pointer Lanthanum and calindol may interact with the same receptor. -- Abstract: Phosphate (Pi)-binders are commonly used in dialysis patients to control high Pi levels, that associated with vascular calcification (VC). The aim of this study was to investigate the effects of lanthanum chloride (LaCl{sub 3}) on the progression of high Pi-induced VC, in rat vascular smooth muscle cells (VSMCs). Pi-induced Ca deposition was inhibited by LaCl{sub 3}, with a maximal effect at 100 {mu}M (59.0 {+-} 2.5% inhibition). Furthermore, we studied the effects on VC of calcium sensing receptor (CaSR) agonists. Gadolinium chloride, neomycin, spermine, and the calcimimetic calindol significantly inhibited Pi-induced VC (55.9 {+-} 2.2%, 37.3 {+-} 4.7%, 30.2 {+-} 5.7%, and 63.8 {+-} 5.7%, respectively). To investigate the hypothesis that LaCl{sub 3} reduces the progression of VC by interacting with the CaSR, we performed a concentration-response curve of LaCl{sub 3} in presence of a sub-effective concentration of calindol (10 nM). Interestingly, this curve was shifted to the left (IC{sub 50} 9.6 {+-} 2.6 {mu}M), compared to the curve in the presence of LaCl{sub 3} alone (IC{sub 50} 19.0 {+-} 4.8 {mu}M). In conclusion, we demonstrated that lanthanum chloride effectively reduces the progression of high phosphate-induced vascular calcification. In addition, LaCl{sub 3} cooperates with the calcimimetic calindol in decreasing Ca deposition in this in vitro model. These results suggest the potential role of lanthanum in the treatment of VC induced by high Pi.

  6. Inflammation induced by mast cell deficiency rather than the loss of interstitial cells of Cajal causes smooth muscle dysfunction in W/Wv mice

    Science.gov (United States)

    Winston, John H.; Chen, Jinghong; Shi, Xuan-Zheng; Sarna, Sushil K.

    2014-01-01

    The initial hypothesis suggested that the interstitial cells of Cajal (ICC) played an essential role in mediating enteric neuronal input to smooth muscle cells. Much information for this hypothesis came from studies in W/Wv mice lacking ICC. However, mast cells, which play critical roles in regulating inflammation in their microenvironment, are also absent in W/Wv mice. We tested the hypothesis that the depletion of mast cells in W/Wv mice generates inflammation in fundus muscularis externa (ME) that impairs smooth muscle reactivity to Ach, independent of the depletion of ICC. We performed experiments on the fundus ME from wild type (WT) and W/Wv mice before and after reconstitution of mast cells by bone marrow transplant. We found that mast cell deficiency in W/Wv mice significantly increased COX-2 and iNOS expression and decreased smooth muscle reactivity to Ach. Mast cell reconstitution or concurrent blockade of COX-2 and iNOS restored smooth muscle contractility without affecting the suppression of c-kit in W/Wv mice. The expression of nNOS and ChAT were suppressed in W/Wv mice; mast cell reconstitution did not restore them. We conclude that innate inflammation induced by mast cell deficiency in W/Wv mice impairs smooth muscle contractility independent of ICC deficiency. The impairment of smooth muscle contractility and the suppression of the enzymes regulating the synthesis of Ach and NO in W/Wv mice need to be considered in evaluating the role of ICC in regulating smooth muscle and enteric neuronal function in W/Wv mice. PMID:24550836

  7. A sex-related difference in the hypertrophic versus hyperplastic response of vascular smooth muscle cells to repeated passaging in culture

    Czech Academy of Sciences Publication Activity Database

    Bačáková, Lucie; Pellicciari, C.; Bottone, M. G.; Lisá, Věra; Mareš, Vladislav

    2001-01-01

    Roč. 16, č. 3 (2001), s. 675-684 ISSN 0213-3911 R&D Projects: GA AV ČR IAA7011908 Grant - others:FAR(IT) 1998 Institutional research plan: CEZ:AV0Z5011922 Keywords : rat aortic smooth muscle cells * polyploidization * gender differences Subject RIV: FA - Cardiovascular Diseases incl. Cardiotharic Surgery Impact factor: 1.859, year: 2001

  8. Regulation of CCL5 expression in smooth muscle cells following arterial injury.

    Directory of Open Access Journals (Sweden)

    Huan Liu

    Full Text Available Chemokines play a crucial role in inflammation and in the pathophysiology of atherosclerosis by recruiting inflammatory immune cells to the endothelium. Chemokine CCL5 has been shown to be involved in atherosclerosis progression. However, little is known about how CCL5 is regulated in vascular smooth muscle cells. In this study we report that CCL5 mRNA expression was induced and peaked in aorta at day 7 and then declined after balloon artery injury, whereas IP-10 and MCP-1 mRNA expression were induced and peaked at day 3 and then rapidly declined.The expression of CCL5 receptors (CCR1, 3 & 5 were also rapidly induced and then declined except CCR5 which expression was still relatively high at day 14 after balloon injury. In rat smooth muscle cells (SMCs, similar as in aorta CCL5 mRNA expression was induced and kept increasing after LPS plus IFN-gamma stimulation, whereas IP-10 mRNA expression was rapidly induced and then declined. Our data further indicate that induction of CCL5 expression in SMCs was mediated by IRF-1 via binding to the IRF-1 response element in CCL5 promoter. Moreover, p38 MAPK was involved in suppression of CCL5 and IP-10 expression in SMCs through common upstream molecule MKK3. The downstream molecule MK2 was required for p38-mediated CCL5 but not IP-10 inhibition. Our findings indicate that CCL5 induction in aorta and SMCs is mediated by IRF-1 while activation of p38 MAPK signaling inhibits CCL5 and IP-10 expression. Methods targeting MK2 expression could be used to selectively regulate CCL5 but not IP-10 expression in SMCs.

  9. A density gradient of VAPG peptides on a cell-resisting surface achieves selective adhesion and directional migration of smooth muscle cells over fibroblasts.

    Science.gov (United States)

    Yu, Shan; Zuo, Xingang; Shen, Tao; Duan, Yiyuan; Mao, Zhengwei; Gao, Changyou

    2018-05-01

    Selective adhesion and migration of smooth muscle cells (SMCs) over fibroblasts (FIBs) is required to prevent adventitia fibrosis in vascular regeneration. In this study, a uniform cell-resisting layer of poly(ethylene glycol) (PEG) with a density gradient of azide groups was generated on a substrate by immobilizing two kinds of PEG molecules in a gradient manner. A density gradient of alkynyl-functionalized Val-Ala-Pro-Gly (VAPG) peptides was then prepared on the PEG layer via click chemistry. The VAPG density gradient was characterized by fluorescence imaging, revealing the gradual enhancement of the fluorescent intensity along the substrate direction. The adhesion and mobility of SMCs were selectively enhanced on the VAPG density gradient, leading to directional migration toward the higher peptide density (up to 84%). In contrast, the adhesion and mobility of FIBs were significantly weakened. The net displacement of SMCs also significantly increased compared with that on tissue culture polystyrene (TCPS) and that of FIBs on the gradient. The mitogen-activated protein kinase (MAPK) signaling pathways related to cell migration were studied, showing higher expressions of functional proteins from SMCs on the VAPG-modified surface in a density-dependent manner. For the first time the selective adhesion and directional migration of SMCs over FIBs was achieved by an elaborative design of a gradient surface, leading to a new insight in design of novel vascular regenerative materials. Selective cell adhesion and migration guided by regenerative biomaterials are extremely important for the regeneration of targeted tissues, which can avoid the drawbacks of incorrect and uncontrolled responses of tissue cells to implants. For example, selectivity of smooth muscle cells (SMCs) over fibroblasts (FIBs) is required to prevent adventitia fibrosis in vascular regeneration. Herein we prepare a uniform cell-repelling layer, on which SMCs-selective Val-Ala-Pro-Gly (VAPG) peptides

  10. Andrographolide inhibits nuclear factor-κB activation through JNK-Akt-p65 signaling cascade in tumor necrosis factor-α-stimulated vascular smooth muscle cells.

    Science.gov (United States)

    Chen, Yu-Ying; Hsu, Ming-Jen; Hsieh, Cheng-Ying; Lee, Lin-Wen; Chen, Zhih-Cherng; Sheu, Joen-Rong

    2014-01-01

    Critical vascular inflammation leads to vascular dysfunction and cardiovascular diseases, including abdominal aortic aneurysms, hypertension, and atherosclerosis. Andrographolide is the most active and critical constituent isolated from the leaves of Andrographis paniculata, a herbal medicine widely used for treating anti-inflammation in Asia. In this study, we investigated the mechanisms of the inhibitory effects of andrographolide in vascular smooth muscle cells (VSMCs) exposed to a proinflammatory stimulus, tumor necrosis factor-α (TNF-α). Treating TNF-α-stimulated VSMCs with andrographolide suppressed the expression of inducible nitric oxide synthase in a concentration-dependent manner. A reduction in TNF-α-induced c-Jun N-terminal kinase (JNK), Akt, and p65 phosphorylation was observed in andrographolide-treated VSMCs. However, andrographolide affected neither IκBα degradation nor p38 mitogen-activated protein kinase or extracellular signal-regulated kinase 1/2 phosphorylation under these conditions. Both treatment with LY294002, a phosphatidylinositol 3-kinase/Akt inhibitor, and treatment with SP600125, a JNK inhibitor, markedly reversed the andrographolide-mediated inhibition of p65 phosphorylation. In addition, LY294002 and SP600125 both diminished Akt phosphorylation, whereas LY294002 had no effects on JNK phosphorylation. These results collectively suggest that therapeutic interventions using andrographolide can benefit the treatment of vascular inflammatory diseases, and andrographolide-mediated inhibition of NF-κB activity in TNF-α-stimulated VSMCs occurs through the JNK-Akt-p65 signaling cascade, an IκBα-independent mechanism.

  11. Andrographolide Inhibits Nuclear Factor-κB Activation through JNK-Akt-p65 Signaling Cascade in Tumor Necrosis Factor-α-Stimulated Vascular Smooth Muscle Cells

    Directory of Open Access Journals (Sweden)

    Yu-Ying Chen

    2014-01-01

    Full Text Available Critical vascular inflammation leads to vascular dysfunction and cardiovascular diseases, including abdominal aortic aneurysms, hypertension, and atherosclerosis. Andrographolide is the most active and critical constituent isolated from the leaves of Andrographis paniculata, a herbal medicine widely used for treating anti-inflammation in Asia. In this study, we investigated the mechanisms of the inhibitory effects of andrographolide in vascular smooth muscle cells (VSMCs exposed to a proinflammatory stimulus, tumor necrosis factor-α (TNF-α. Treating TNF-α-stimulated VSMCs with andrographolide suppressed the expression of inducible nitric oxide synthase in a concentration-dependent manner. A reduction in TNF-α-induced c-Jun N-terminal kinase (JNK, Akt, and p65 phosphorylation was observed in andrographolide-treated VSMCs. However, andrographolide affected neither IκBα degradation nor p38 mitogen-activated protein kinase or extracellular signal-regulated kinase 1/2 phosphorylation under these conditions. Both treatment with LY294002, a phosphatidylinositol 3-kinase/Akt inhibitor, and treatment with SP600125, a JNK inhibitor, markedly reversed the andrographolide-mediated inhibition of p65 phosphorylation. In addition, LY294002 and SP600125 both diminished Akt phosphorylation, whereas LY294002 had no effects on JNK phosphorylation. These results collectively suggest that therapeutic interventions using andrographolide can benefit the treatment of vascular inflammatory diseases, and andrographolide-mediated inhibition of NF-κB activity in TNF-α-stimulated VSMCs occurs through the JNK-Akt-p65 signaling cascade, an IκBα-independent mechanism.

  12. Mechanism by which nuclear factor-kappa beta (NF-kB regulates ovine fetal pulmonary vascular smooth muscle cell proliferation

    Directory of Open Access Journals (Sweden)

    Uchenna D. Ogbozor

    2015-09-01

    Full Text Available Platelet activating factor (PAF modulates ovine fetal pulmonary hemodynamic. PAF acts through its receptors (PAFR in pulmonary vascular smooth muscle cells (PVSMC to phosphorylate and induce nuclear translocation of NF-kB p65 leading to PVSMC proliferation. However, the interaction of NF-kB p65 and PAF in the nuclear domain to effect PVSMC cell growth is not clearly defined. We used siRNA-dependent translation initiation arrest to study a mechanism by which NF-kB p65 regulates PAF stimulation of PVSMC proliferation. Our hypotheses are: (a PAF induces NF-kB p65 DNA binding and (b NF-kB p65 siRNA attenuates PAF stimulation of PVSMC proliferation. For DNA binding, cells were fed 10 nM PAF with and without PAFR antagonists WEB 2170, CV 3988 or BN 52021 and incubated for 12 h. DNA binding was measured by specific ELISA. For NF-kB p65 siRNA effect, starved cells transfected with the siRNA were incubated for 24 h with and without 10 nM PAF. Cell proliferation was measured by DNA synthesis while expression of NF-kB p65 and PAFR protein was measured by Western blotting. In both studies, the effect of 10% FBS alone was used as the positive control. In general, PAF stimulated DNA binding which was inhibited by PAFR antagonists. siRNAs to NF-kB p65 and PAFR significantly attenuated cell proliferation compared to 10% FBS and PAF effect. Inclusion of PAF in siRNA-treated cells did not reverse inhibitory effect of NF-kB p65 siRNA on DNA synthesis. PAFR expression was inhibited in siRNA-treated cells. These data show that PAF-stimulation of PVSMC proliferation occurs via a PAFR-NF-kB p65 linked pathway.

  13. Mechanism by which nuclear factor-kappa beta (NF-kB) regulates ovine fetal pulmonary vascular smooth muscle cell proliferation.

    Science.gov (United States)

    Ogbozor, Uchenna D; Opene, Michael; Renteria, Lissette S; McBride, Shaemion; Ibe, Basil O

    2015-09-01

    Platelet activating factor (PAF) modulates ovine fetal pulmonary hemodynamic. PAF acts through its receptors (PAFR) in pulmonary vascular smooth muscle cells (PVSMC) to phosphorylate and induce nuclear translocation of NF-kB p65 leading to PVSMC proliferation. However, the interaction of NF-kB p65 and PAF in the nuclear domain to effect PVSMC cell growth is not clearly defined. We used siRNA-dependent translation initiation arrest to study a mechanism by which NF-kB p65 regulates PAF stimulation of PVSMC proliferation. Our hypotheses are: (a) PAF induces NF-kB p65 DNA binding and (b) NF-kB p65 siRNA attenuates PAF stimulation of PVSMC proliferation. For DNA binding, cells were fed 10 nM PAF with and without PAFR antagonists WEB 2170, CV 3988 or BN 52021 and incubated for 12 h. DNA binding was measured by specific ELISA. For NF-kB p65 siRNA effect, starved cells transfected with the siRNA were incubated for 24 h with and without 10 nM PAF. Cell proliferation was measured by DNA synthesis while expression of NF-kB p65 and PAFR protein was measured by Western blotting. In both studies, the effect of 10% FBS alone was used as the positive control. In general, PAF stimulated DNA binding which was inhibited by PAFR antagonists. siRNAs to NF-kB p65 and PAFR significantly attenuated cell proliferation compared to 10% FBS and PAF effect. Inclusion of PAF in siRNA-treated cells did not reverse inhibitory effect of NF-kB p65 siRNA on DNA synthesis. PAFR expression was inhibited in siRNA-treated cells. These data show that PAF-stimulation of PVSMC proliferation occurs via a PAFR-NF-kB p65 linked pathway.

  14. Vitamin K2 inhibits rat vascular smooth muscle cell calcification by restoring the Gas6/Axl/Akt anti-apoptotic pathway.

    Science.gov (United States)

    Qiu, Cuiting; Zheng, Haijun; Tao, Huiren; Yu, Wenjun; Jiang, Xiaoyu; Li, Aiqin; Jin, Hui; Lv, Anlin; Li, Huan

    2017-09-01

    Vascular calcification is associated with cardiovascular disease as a complication of hypertension, hyperlipidemia, diabetes mellitus, and chronic kidney disease. Vitamin K2 (VK2) delays vascular calcification by an unclear mechanism. Moreover, apoptosis modulates vascular smooth muscle cell (VSMC) calcification. This paper aimed to study VK2-modified VSMC calcification and survival cell signaling mediated by growth arrest-specific gene 6 (Gas6) and its tyrosine kinase receptor Axl. Primary-cultured VSMCs were dose-dependently treated with VK2 in the presence of calcification medium for 8 days, or pre-treated for 1 h with/without the Axl inhibitor R428 (2 μmol/L) or the caspase inhibitor Z-VAD-fmk (20 μmol/L) followed by treatment with VK2 (10 μmol/L) or rmGas6 (200 nmol/L) in calcification medium for 8 days. Calcium deposition was determined by the o-cresolphthalein complexone assay and Alizarin Red S staining. Apoptosis was determined by TUNEL and flow cytometry using Annexin V-FITC and propidium iodide staining. Western blotting detected the expressions of Axl, Gas6, p-Akt, Akt, and Bcl2. VK2 significantly inhibited CaCl 2 - and β-sodium glycerophosphate (β-GP)-induced VSMC calcification and apoptosis, which was dependent on restored Gas6 expression and activated downstream signaling by Axl, p-Akt, and Bcl2. Z-VAD-fmk significantly inhibited CaCl 2 - and β-GP-induced VSMC calcification and apoptosis. Augmented recombinant mouse Gas6 protein (rmGas6) expression significantly reduced VSMC calcification and apoptosis. Furthermore, the Gas6/Axl interaction was inhibited by R428, which abolished the preventive effect of VK2 on CaCl 2 - and β-GP-induced apoptosis and calcification. These results suggest that Gas6 is critical in VK2-mediated functions that attenuate CaCl 2 - and β-GP-induced VSMC calcification by blocking apoptosis.

  15. PDZK1 prevents neointima formation via suppression of breakpoint cluster region kinase in vascular smooth muscle.

    Directory of Open Access Journals (Sweden)

    Wan Ru Lee

    Full Text Available Scavenger receptor class B, type I (SR-BI and its adaptor protein PDZK1 mediate responses to HDL cholesterol in endothelium. Whether the receptor-adaptor protein tandem serves functions in other vascular cell types is unknown. The current work determined the roles of SR-BI and PDZK1 in vascular smooth muscle (VSM. To evaluate possible VSM functions of SR-BI and PDZK1 in vivo, neointima formation was assessed 21 days post-ligation in the carotid arteries of wild-type, SR-BI-/- or PDZK1-/- mice. Whereas neointima development was negligible in wild-type and SR-BI-/-, there was marked neointima formation in PDZK1-/- mice. PDZK1 expression was demonstrated in primary mouse VSM cells, and compared to wild-type cells, PDZK1-/- VSM displayed exaggerated proliferation and migration in response to platelet derived growth factor (PDGF. Tandem affinity purification-mass spectrometry revealed that PDZK1 interacts with breakpoint cluster region kinase (Bcr, which contains a C-terminal PDZ binding sequence and is known to enhance responses to PDGF in VSM. PDZK1 interaction with Bcr in VSM was demonstrated by pull-down and by coimmunoprecipitation, and the augmented proliferative response to PDGF in PDZK1-/- VSM was abrogated by Bcr depletion. Furthermore, compared with wild-type Bcr overexpression, the introduction of a Bcr mutant incapable of PDZK1 binding into VSM cells yielded an exaggerated proliferative response to PDGF. Thus, PDZK1 has novel SR-BI-independent function in VSM that affords protection from neointima formation, and this involves PDZK1 suppression of VSM cell proliferation via an inhibitory interaction with Bcr.

  16. Transforming growth factor β-activated kinase 1 negatively regulates interleukin-1α-induced stromal-derived factor-1 expression in vascular smooth muscle cells

    Energy Technology Data Exchange (ETDEWEB)

    Yang, Bin [Department of Hepatobiliary Surgery, Union Hospital, Tongji Medical College, Huangzhong University of Science and Technology, Wuhan 430022 (China); Li, Wei [Department of Gerontology, Union Hospital, Tongji Medical College, Huangzhong University of Science and Technology, Wuhan 430022 (China); Zheng, Qichang [Department of Hepatobiliary Surgery, Union Hospital, Tongji Medical College, Huangzhong University of Science and Technology, Wuhan 430022 (China); Qin, Tao [Department of Hepatobiliary Pancreatic Surgery, People' s Hospital of Zhengzhou University, School of Medicine, Zhengzhou University, Zhengzhou 450003 (China); Wang, Kun; Li, Jinjin; Guo, Bing; Yu, Qihong; Wu, Yuzhe; Gao, Yang; Cheng, Xiang; Hu, Shaobo; Kumar, Stanley Naveen [Department of Hepatobiliary Surgery, Union Hospital, Tongji Medical College, Huangzhong University of Science and Technology, Wuhan 430022 (China); Liu, Sanguang, E-mail: sanguang1998@sina.com [Department of Hepatobiliary Surgery, The Second Hospital, Hebei Medical University, Shijiazhuang 050000 (China); Song, Zifang, E-mail: zsong@hust.edu.cn [Department of Hepatobiliary Surgery, Union Hospital, Tongji Medical College, Huangzhong University of Science and Technology, Wuhan 430022 (China)

    2015-07-17

    Stromal-derived Factor-1 (SDF-1) derived from vascular smooth muscle cells (VSMCs) contributes to vascular repair and remodeling in various vascular diseases. In this study, the mechanism underlying regulation of SDF-1 expression by interleukin-1α (IL-1α) was investigated in primary rat VSMCs. We found IL-1α promotes SDF-1 expression by up-regulating CCAAT-enhancer-binding protein β (C/EBPβ) in an IκB kinase β (IKKβ) signaling-dependent manner. Moreover, IL-1α-induced expression of C/EBPβ and SDF-1 was significantly potentiated by knockdown of transforming growth factor β-activated kinase 1 (TAK1), an upstream activator of IKKβ signaling. In addition, we also demonstrated that TAK1/p38 mitogen-activated protein kinase (p38 MAPK) signaling exerted negative effect on IL-1α-induced expression of C/EBPβ and SDF-1 through counteracting ROS-dependent up-regulation of nuclear factor erythroid 2-related factor 2 (NRF2). In conclusion, TAK1 acts as an important regulator of IL-1α-induced SDF-1 expression in VSMCs, and modulating activity of TAK1 may serve as a potential strategy for modulating vascular repair and remodeling. - Highlights: • IL-1α induces IKKβ signaling-dependent SDF-1 expression by up-regulating C/EBPβ. • Activation of TAK1 by IL-1α negatively regulates C/EBPβ-dependent SDF-1 expression. • IL-1α-induced TAK1/p38 MAPK signaling counteracts ROS-dependent SDF-1 expression. • TAK1 counteracts IL-1α-induced SDF-1 expression by attenuating NRF2 up-regulation.

  17. Transforming growth factor β-activated kinase 1 negatively regulates interleukin-1α-induced stromal-derived factor-1 expression in vascular smooth muscle cells

    International Nuclear Information System (INIS)

    Yang, Bin; Li, Wei; Zheng, Qichang; Qin, Tao; Wang, Kun; Li, Jinjin; Guo, Bing; Yu, Qihong; Wu, Yuzhe; Gao, Yang; Cheng, Xiang; Hu, Shaobo; Kumar, Stanley Naveen; Liu, Sanguang; Song, Zifang

    2015-01-01

    Stromal-derived Factor-1 (SDF-1) derived from vascular smooth muscle cells (VSMCs) contributes to vascular repair and remodeling in various vascular diseases. In this study, the mechanism underlying regulation of SDF-1 expression by interleukin-1α (IL-1α) was investigated in primary rat VSMCs. We found IL-1α promotes SDF-1 expression by up-regulating CCAAT-enhancer-binding protein β (C/EBPβ) in an IκB kinase β (IKKβ) signaling-dependent manner. Moreover, IL-1α-induced expression of C/EBPβ and SDF-1 was significantly potentiated by knockdown of transforming growth factor β-activated kinase 1 (TAK1), an upstream activator of IKKβ signaling. In addition, we also demonstrated that TAK1/p38 mitogen-activated protein kinase (p38 MAPK) signaling exerted negative effect on IL-1α-induced expression of C/EBPβ and SDF-1 through counteracting ROS-dependent up-regulation of nuclear factor erythroid 2-related factor 2 (NRF2). In conclusion, TAK1 acts as an important regulator of IL-1α-induced SDF-1 expression in VSMCs, and modulating activity of TAK1 may serve as a potential strategy for modulating vascular repair and remodeling. - Highlights: • IL-1α induces IKKβ signaling-dependent SDF-1 expression by up-regulating C/EBPβ. • Activation of TAK1 by IL-1α negatively regulates C/EBPβ-dependent SDF-1 expression. • IL-1α-induced TAK1/p38 MAPK signaling counteracts ROS-dependent SDF-1 expression. • TAK1 counteracts IL-1α-induced SDF-1 expression by attenuating NRF2 up-regulation

  18. Opportunities to Target Specific Contractile Abnormalities with Smooth Muscle Protein Kinase Inhibitors

    Directory of Open Access Journals (Sweden)

    Annegret Ulke-Lemée

    2010-05-01

    Full Text Available Smooth muscle is a major component of most hollow organ systems (e.g., airways, vasculature, bladder and gut/gastrointestine; therefore, the coordinated regulation of contraction is a key property of smooth muscle. When smooth muscle functions normally, it contributes to general health and wellness, but its dysfunction is associated with morbidity and mortality. Rho-associated protein kinase (ROCK is central to calcium-independent, actomyosin-mediated contractile force generation in the vasculature, thereby playing a role in smooth muscle contraction, cell motility and adhesion. Recent evidence supports an important role for ROCK in the increased vasoconstriction and remodeling observed in various models of hypertension. This review will provide a commentary on the development of specific ROCK inhibitors and their clinical application. Fasudil will be discussed as an example of bench-to-bedside development of a clinical therapeutic that is used to treat conditions of vascular hypercontractility. Due to the wide spectrum of biological processes regulated by ROCK, many additional clinical indications might also benefit from ROCK inhibition. Apart from the importance of ROCK in smooth muscle contraction, a variety of other protein kinases are known to play similar roles in regulating contractile force. The zipper-interacting protein kinase (ZIPK and integrin-linked kinase (ILK are two well-described regulators of contraction. The relative contribution of each kinase to contraction depends on the muscle bed as well as hormonal and neuronal stimulation. Unfortunately, specific inhibitors for ZIPK and ILK are still in the development phase, but the success of fasudil suggests that inhibitors for these other kinases may also have valuable clinical applications. Notably, the directed inhibition of ZIPK with a pseudosubstrate molecule shows unexpected effects on the contractility of gastrointestinal smooth muscle.

  19. Bradykinin B2 receptor-mediated phosphoinositide hydrolysis in bovine cultured tracheal smooth muscle cells.

    OpenAIRE

    Marsh, K. A.; Hill, S. J.

    1992-01-01

    1. Bovine tracheal smooth muscle cells were established in culture to study agonist-induced phosphoinositide (PI) hydrolysis in this tissue. 2. Bradykinin (0.1 nM-10 microM) evoked a concentration-dependent increase (log EC50 (M) = -9.4 +/- 0.2; n = 8) in the accumulation of total [3H]-inositol phosphates in cultured tracheal smooth muscle cells whereas the selective B1 receptor agonist des-Arg9-bradykinin (10 microM) was significantly less effective (16% of bradykinin maximal response; relat...

  20. DHEA attenuates PDGF-induced phenotypic proliferation of vascular smooth muscle A7r5 cells through redox regulation

    Energy Technology Data Exchange (ETDEWEB)

    Urata, Yoshishige; Goto, Shinji; Kawakatsu, Miho [Department of Biochemistry and Molecular Biology in Disease, Atomic Bomb Disease Institute, Nagasaki University Graduate School of Medical Sciences, 1-12-4 Sakamoto, Nagasaki 852-8523 (Japan); Yodoi, Junji [Department of Biological Responses, Institute for Viral Research, Graduate School of Medicine, Kyoto University, 53 Shogain, Kawahara-cho, Sakyo-ku, Kyoto 606-8397 (Japan); Eto, Masato [Department of Geriatric Medicine, Graduate School of Medicine, University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-8655 (Japan); Akishita, Masahiro, E-mail: akishita-tky@umin.ac.jp [Department of Geriatric Medicine, Graduate School of Medicine, University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-8655 (Japan); Kondo, Takahito [Department of Biochemistry and Molecular Biology in Disease, Atomic Bomb Disease Institute, Nagasaki University Graduate School of Medical Sciences, 1-12-4 Sakamoto, Nagasaki 852-8523 (Japan)

    2010-05-28

    It is known that dehydroepiandrosterone (DHEA) inhibits a phenotypic switch in vascular smooth muscle cells (VSMC) induced by platelet-derived growth factor (PDGF)-BB. However, the mechanism behind the effect of DHEA on VSMC is not clear. Previously we reported that low molecular weight-protein tyrosine phosphatase (LMW-PTP) dephosphorylates PDGF receptor (PDGFR)-{beta} via a redox-dependent mechanism involving glutathione (GSH)/glutaredoxin (GRX)1. Here we demonstrate that the redox regulation of PDGFR-{beta} is involved in the effect of DHEA on VSMC. DHEA suppressed the PDGF-BB-dependent phosphorylation of PDGFR-{beta}. As expected, DHEA increased the levels of GSH and GRX1, and the GSH/GRX1 system maintained the redox state of LMW-PTP. Down-regulation of the expression of LMW-PTP using siRNA restored the suppression of PDGFR-{beta}-phosphorylation by DHEA. A promoter analysis of GRX1 and {gamma}-glutamylcysteine synthetase ({gamma}-GCS), a rate-limiting enzyme of GSH synthesis, showed that DHEA up-regulated the transcriptional activity at the peroxisome proliferator-activated receptor (PPAR) response element, suggesting PPAR{alpha} plays a role in the induction of GRX1 and {gamma}-GCS expression by DHEA. In conclusion, the redox regulation of PDGFR-{beta} is involved in the suppressive effect of DHEA on VSMC proliferation through the up-regulation of GSH/GRX system.

  1. Leptin Inhibits the Proliferation of Vascular Smooth Muscle Cells Induced by Angiotensin II through Nitric Oxide-Dependent Mechanisms

    Directory of Open Access Journals (Sweden)

    Amaia Rodríguez

    2010-01-01

    Full Text Available Objective. This study was designed to investigate whether leptin modifies angiotensin (Ang II-induced proliferation of aortic vascular smooth muscle cells (VSMCs from 10-week-old male Wistar and spontaneously hypertensive rats (SHR, and the possible role of nitric oxide (NO. Methods. NO and NO synthase (NOS activity were assessed by the Griess and 3H-arginine/citrulline conversion assays, respectively. Inducible NOS (iNOS and NADPH oxidase subutnit Nox2 expression was determined by Western-blot. The proliferative responses to Ang II were evaluated through enzymatic methods. Results. Leptin inhibited the Ang II-induced proliferative response of VSMCs from control rats. This inhibitory effect of leptin was abolished by NOS inhibitor, NMMA, and iNOS selective inhibitor, L-NIL, and was not observed in leptin receptor-deficient fa/fa rats. SHR showed increased serum leptin concentrations and lipid peroxidation. Despite a similar leptin-induced iNOS up-regulation, VSMCs from SHR showed an impaired NOS activity and NO production induced by leptin, and an increased basal Nox2 expression. The inhibitory effect of leptin on Ang II-induced VSMC proliferation was attenuated. Conclusion. Leptin blocks the proliferative response to Ang II through NO-dependent mechanisms. The attenuation of this inhibitory effect of leptin in spontaneous hypertension appears to be due to a reduced NO bioavailability in VSMCs.

  2. Pro-inflammatory effects of interleukin-17A on vascular smooth muscle cells involve NAD(P)H- oxidase derived reactive oxygen species.

    Science.gov (United States)

    Pietrowski, Eweline; Bender, Bianca; Huppert, Jula; White, Robin; Luhmann, Heiko J; Kuhlmann, Christoph R W

    2011-01-01

    T cells are known for their contribution to the inflammatory element of atherosclerosis. Recently, it has been demonstrated that the Th17 derived cytokine IL-17 is involved in the pro-inflammatory response of vascular smooth muscle cells (VSMC). The aim of the present study was to examine whether reactive oxygen species (ROS) might be involved in this context. The effect of IL-17A on ROS generation was examined using the fluorescent dye 2'7'-dichlorodihydrofluorescein (H(2)DCF) in primary murine VSMC. IL-17A induced an increase in H(2)DCF fluorescence in VSMC, and this effect was blocked by the NAD(P)H-oxidase inhibitor apocynin and siRNA targeting Nox2. The p38-MAPK inhibitors SB203580 and SB202190 dose-dependently reduced the IL-17A induced ROS production. The IL-17A induced release of the pro-inflammatory cytokines IL-6, G-CSF, GM-CSF and MCP-1 from VSMC, as detected by the Luminex technology, was completely abolished by NAD(P)H-oxidase inhibition. Taken together, our data indicate that IL-17A causes the NAD(P)H-oxidase dependent generation of ROS leading to a pro-inflammatory activation of VSMC. Copyright © 2010 S. Karger AG, Basel.

  3. Terminalia chebula Fructus Inhibits Migration and Proliferation of Vascular Smooth Muscle Cells and Production of Inflammatory Mediators in RAW 264.7

    Directory of Open Access Journals (Sweden)

    Hyun-Ho Lee

    2015-01-01

    Full Text Available Pathogenesis of atherosclerosis and neointima formation after angioplasty involves vascular smooth muscle cells (VSMCs migration and proliferation followed by inflammatory responses mediated by recruited macrophages in the neointima. Terminalia chebula is widely used traditional medicine in Asia for its beneficial effects against cancer, diabetes, and bacterial infection. The study was designed to determine whether Terminalia chebula fructus water extract (TFW suppresses VSMC migration and proliferation and inflammatory mediators production in macrophage (RAW 264.7. Our results showed that TFW possessed strong antioxidative effects in 1,1-diphenyl-2-picryl hydrazyl (DPPH scavenging and lipid peroxidation assays. In addition, TFW reduced nitric oxide (NO production, inducible nitric oxide synthase (iNOS, and cyclooxygenase-2 (COX-2 expression in RAW 264.7 cells. Also, TFW inhibited platelet-derived growth factor (PDGF-BB induced VSMC migration as determined by wound healing and Boyden chamber assays. The antimigratory effect of TFW was due to its inhibitory effect on metalloproteinase-9 (MMP-9 expression, focal adhesion kinase (FAK activation, and Rho-family of small GTPases (Cdc42 and RhoA expression in VSMCs. Furthermore, TFW suppressed PDGF-BB induced VSMC proliferation by downregulation of mitogen activated protein kinases (MAPKs signaling molecules. These results suggest that TFW could be a beneficial resource in the prevention of atherosclerosis.

  4. Inhibition of human arterial smooth muscle (HASM) cell proliferation and collagen synthesis by protamine

    International Nuclear Information System (INIS)

    Drucker, D.E.; Graham, M.F.; Diegelmann, R.F.; Greenfield, L.J.

    1986-01-01

    Atherosclerotic plaques result from vascular smooth muscle cell proliferation and collagen deposition. The authors have been studying factors which modulate HASM cell proliferation and collagen synthesis. HASM cells were isolated from the media of normal human thoracic and infrarenal aortas and grown in vitro. Cell numbers were determined by direct counting and collagen synthesis was measured by incorporation of 3 H-proline into collagenase-digestible protein. In this study, protamine (200 μg/ml) was tested and found to cause a 55% reduction of HASM cell proliferation which was reversible when the cells were returned to control medium or when heparin (100 μg/ml) was added with protamine. Protamine caused a constant 33% decrease in non-collagen protein (NCP) synthesis per cell. In contrast, collagen synthesis was inhibited in dose dependent fashion (88% reduction at 200 μg/ml). Protamine blocks HASM cell proliferation and specifically inhibits collagen production. The exact mechanism of this inhibition is unclear but may be related to a transcriptional event since protamine has a high affinity for DNA

  5. Triptolide inhibits TGF-β1-induced cell proliferation in rat airway smooth muscle cells by suppressing Smad signaling

    Energy Technology Data Exchange (ETDEWEB)

    Chen, Ming; Lv, Zhiqiang; Huang, Linjie [Department of Respiratory Medicine, Sun Yat-Sen Memorial Hospital, Institute for Respiratory disease of Sun Yat-sen University, Sun Yat-sen University, Guangzhou, Guangdong Province 510120 (China); Zhang, Wei [Department of Geratology, the Second People' s Hospital of Shenzhen, Shenzhen 518000 (China); Lin, Xiaoling; Shi, Jianting; Zhang, Wei; Liang, Ruiyun [Department of Respiratory Medicine, Sun Yat-Sen Memorial Hospital, Institute for Respiratory disease of Sun Yat-sen University, Sun Yat-sen University, Guangzhou, Guangdong Province 510120 (China); Jiang, Shanping, E-mail: shanpingjiang@126.com [Department of Respiratory Medicine, Sun Yat-Sen Memorial Hospital, Institute for Respiratory disease of Sun Yat-sen University, Sun Yat-sen University, Guangzhou, Guangdong Province 510120 (China)

    2015-02-15

    Background: We have reported that triptolide can inhibit airway remodeling in a murine model of asthma via TGF-β1/Smad signaling. In the present study, we aimed to investigate the effect of triptolide on airway smooth muscle cells (ASMCs) proliferation and the possible mechanism. Methods: Rat airway smooth muscle cells were cultured and made synchronized, then pretreated with different concentration of triptolide before stimulated by TGF-β1. Cell proliferation was evaluated by MTT assay. Flow cytometry was used to study the influence of triptolide on cell cycle and apoptosis. Signal proteins (Smad2, Smad3 and Smad7) were detected by western blotting analysis. Results: Triptolide significantly inhibited TGF-β1-induced ASMC proliferation (P<0.05). The cell cycle was blocked at G1/S-interphase by triptolide dose dependently. No pro-apoptotic effects were detected under the concentration of triptolide we used. Western blotting analysis showed TGF-β1 induced Smad2 and Smad3 phosphorylation was inhibited by triptolide pretreatment, and the level of Smad7 was increased by triptolide pretreatment. Conclusions: Triptolide may function as an inhibitor of asthma airway remodeling by suppressing ASMCs proliferation via negative regulation of Smad signaling pathway. - Highlights: • In this study, rat airway smooth muscle cells were cultured and made synchronized. • Triptolide inhibited TGF-β1-induced airway smooth muscle cells proliferation. • Triptolide inhibited ASMCs proliferation via negative regulation of Smad signaling pathway.

  6. MicroRNA-31 controls phenotypic modulation of human vascular smooth muscle cells by regulating its target gene cellular repressor of E1A-stimulated genes

    International Nuclear Information System (INIS)

    Wang, Jie; Yan, Cheng-Hui; Li, Yang; Xu, Kai; Tian, Xiao-Xiang; Peng, Cheng-Fei; Tao, Jie; Sun, Ming-Yu; Han, Ya-Ling

    2013-01-01

    Phenotypic modulation of vascular smooth muscle cells (VSMCs) plays a critical role in the pathogenesis of a variety of proliferative vascular diseases. The cellular repressor of E1A-stimulated genes (CREG) has been shown to play an important role in phenotypic modulation of VSMCs. However, the mechanism regulating CREG upstream signaling remains unclear. MicroRNAs (miRNAs) have recently been found to play a critical role in cell differentiation via target-gene regulation. This study aimed to identify a miRNA that binds directly to CREG, and may thus be involved in CREG-mediated VSMC phenotypic modulation. Computational analysis indicated that miR-31 bound to the CREG mRNA 3′ untranslated region (3′-UTR). miR-31 was upregulated in quiescent differentiated VSMCs and downregulated in proliferative cells stimulated by platelet-derived growth factor and serum starvation, demonstrating a negative relationship with the VSMC differentiation marker genes, smooth muscle α-actin, calponin and CREG. Using gain-of-function and loss-of-function approaches, CREG and VSMC differentiation marker gene expression levels were shown to be suppressed by a miR-31 mimic, but increased by a miR-31 inhibitor at both protein and mRNA levels. Notably, miR-31 overexpression or inhibition affected luciferase expression driven by the CREG 3′-UTR containing the miR-31 binding site. Furthermore, miR-31-mediated VSMC phenotypic modulation was inhibited in CREG-knockdown human VSMCs. We also determined miR-31 levels in the serum of patients with coronary artery disease (CAD), with or without in stent restenosis and in healthy controls. miR-31 levels were higher in the serum of CAD patients with restenosis compared to CAD patients without restenosis and in healthy controls. In summary, these data demonstrate that miR-31 not only directly binds to its target gene CREG and modulates the VSMC phenotype through this interaction, but also can be an important biomarker in diseases involving VSMC

  7. Microparticle Shedding by Erythrocytes, Monocytes and Vascular Smooth Muscular Cells Is Reduced by Aspirin in Diabetic Patients.

    Science.gov (United States)

    Chiva-Blanch, Gemma; Suades, Rosa; Padró, Teresa; Vilahur, Gemma; Peña, Esther; Ybarra, Juan; Pou, Jose M; Badimon, Lina

    2016-07-01

    Diabetes mellitus is associated with an enhanced risk for cardiovascular disease and its prevalence is increasing. Diabetes induces metabolic stress on blood and vascular cells, promoting platelet activation and vascular dysfunction. The level of vascular cell activation can be measured by the number and phenotype of microparticles found in the circulation. The aim of this study was to investigate the effect of a platelet-inhibitory dose of aspirin on the number and type of microparticles shed to the circulation. Forty-three diabetic patients were enrolled in the study and received a daily dose of 100mg of aspirin for 10 days to cover the average platelet life-span in the circulation. Before and after the intervention period, circulating microparticles were characterized and quantified by flow cytometry. Type 1 diabetic patients had about twice the number of tissue factor-positive circulating microparticles (derived both from platelets and monocytes) and endothelial-derived E-selectin positive microparticles than type 2 diabetic patients. Aspirin therapy significantly inhibited platelets since cyclooxygenase 1 derived thromboxane generation levels were reduced by 99%. Microparticles derived from erythrocytes, activated monocytes, and smooth muscle cells were significantly reduced after 10 days of aspirin administration. These results indicate that: a) vascular and blood cells in type 1 diabetic patients are exposed to more sustained stress shown by their specific microparticle origin and levels; b) aspirin therapy inhibits vascular wall cell activation and microparticle shedding, and c) the effects of aspirin are similar in type 1 and 2 diabetes. Copyright © 2016 Sociedad Española de Cardiología. Published by Elsevier España, S.L.U. All rights reserved.

  8. [3H]QNB binding and contraction of rabbit colonic smooth muscle cells

    International Nuclear Information System (INIS)

    Ringer, M.J.; Hyman, P.E.; Kao, H.W.; Hsu, C.T.; Tomomasa, T.; Snape, W.J. Jr.

    1987-01-01

    The authors used radioligand binding and studies of cell contraction to characterize muscarinic receptors on dispersed smooth muscle cells from rabbit proximal and distal colon. Cells obtained after serial incubations in collagenase were used to measure binding of tritiated quinuclidinyl benzilate ([ 3 H]QNB). At 37 degree C, specific [ 3 H]QNB binding was saturable and linearly related to cell number. Nonlinear regression analysis was used to determine the affinity of [ 3 H]QNB for its receptor. The IC 50 for the muscarinic agonists bethanechol and oxotremorine were 80 and 0.57 μM, respectively. Hill coefficients were 0.67 for both, suggesting more complex interaction involving receptors of different affinities. In studies of cell contraction, bethanechol stimulated a dose-dependent decrease in cell length with half the maximal contraction occurring at 100 pM. These results suggest that (1) contraction is mediated by binding of bethanechol to M 2 -muscarinic receptors and that (2) there are a large number of spare receptors in colonic smooth muscle

  9. Advanced glycation end products promote the proliferation and migration of primary rat vascular smooth muscle cells via the upregulation of BAG3.

    Science.gov (United States)

    Li, Cunshu; Chang, Ye; Li, Yuan; Chen, Shuang; Chen, Yintao; Ye, Ning; Dai, Dongxue; Sun, Yingxian

    2017-05-01

    The present study was aimed to investigate the role of reactive oxygen species (ROS) on advanced glycation end product (AGE)-induced proliferation and migration of vascular smooth muscle cells (VSMCs) and whether Bcl-2‑associated athanogene 3 (BAG3) is involved in the process. Primary rat VSMCs were extracted and cultured in vitro. Cell viability was detected by MTT assay and cell proliferation was detected by EdU incorporation assay. Cell migration was detected by wound healing and Transwell assays. BAG3 was detected using qPCR and western blot analysis. Transcriptional and translational inhibitors (actinomycin D and cycloheximide, respectively) were used to study the effect of AGEs on the expression of BAG3 in VSMCs. Lentiviral plasmids containing short hairpin RNA (shRNA) against rat BAG3 or control shRNA were transduced into VSMCs. Cellular ROS were detected by 2',7'-dichlorofluorescein diacetate (DCFH-DA) staining. Mitochondrial membrane potential was detected by tetramethylrhodamine methyl ester (TMRE) staining. AGEs significantly increased the expression of BAG3 in a dose-and time-dependent manner. Furthermore, AGEs mainly increased the expression of BAG3 mRNA by increasing the RNA synthesis rather than inhibiting the RNA translation. BAG3 knockdown reduced the proliferation and migration of VSMCs induced by AGEs. BAG3 knockdown reduced the generation of ROS and sustained the mitochondrial membrane potential of VSMCs. Reduction of ROS production by N-acetylcysteine (NAC), a potent antioxidant, also reduced the proliferation and migration of VSMCs. On the whole, the present study demonstrated for the first time that AGEs could increase ROS production and promote the proliferation and migration of VSMCs by upregulating BAG3 expression. This study indicated that BAG3 should be considered as a potential target for the prevention and/or treatment of vascular complications of diabetes.

  10. Metformin inhibits inflammatory response via AMPK-PTEN pathway in vascular smooth muscle cells

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Sun Ae [Department of Pharmacology, Aging-Associated Vascular Disease Research Center, College of Medicine, Yeungnam University, Daegu 705-717 (Korea, Republic of); Choi, Hyoung Chul, E-mail: hcchoi@med.yu.ac.kr [Department of Pharmacology, Aging-Associated Vascular Disease Research Center, College of Medicine, Yeungnam University, Daegu 705-717 (Korea, Republic of)

    2012-09-07

    Highlights: Black-Right-Pointing-Pointer PTEN was induced by metformin and inhibited by compound C and AMPK siRNA. Black-Right-Pointing-Pointer Metformin suppressed TNF-{alpha}-induced COX-2 and iNOS mRNA expression. Black-Right-Pointing-Pointer Compound C and bpv (pic) increased iNOS and COX-2 protein expression. Black-Right-Pointing-Pointer NF-{kappa}B activation was restored by inhibiting AMPK and PTEN. Black-Right-Pointing-Pointer AMPK and PTEN regulated TNF-{alpha}-induced ROS production in VSMCs. -- Abstract: Atherosclerosis is a chronic inflammation of the coronary arteries. Vascular smooth muscle cells (VSMCs) stimulated by cytokines and chemokines accelerate the inflammatory response and migrate to the injured endothelium during the progression of atherosclerosis. Activation of AMP activated protein kinase (AMPK), a key sensor maintaining metabolic homeostasis, suppresses the inflammatory response. However, how AMPK regulates the inflammatory response is poorly understood. To identify the mechanism of this response, we focused on phosphatase and tensin homolog (PTEN), which is a negative regulator of inflammation. We investigated that activation of AMPK-induced PTEN expression and suppression of the inflammatory response through the AMPK-PTEN pathway in VSMCs. We treated with the well-known AMPK activator metformin to induce PTEN expression. PTEN was induced by metformin (2 mM) and inhibited by compound C (10 {mu}M) and AMPK siRNA. Tumor necrosis factor-alpha (TNF-{alpha}) was used to induce inflammation. The inflammatory response was confirmed by cyclooxygenase (COX)-2, inducible nitric oxide synthase (iNOS) expression, and activation of nuclear factor (NF)-{kappa}B. Metformin suppressed COX-2 and iNOS mRNA and protein expression dose dependently. Treatment with compound C and bpv (pic) in the presence of metformin, iNOS and COX-2 protein expression increased. NF-{kappa}B activation decreased in response to metformin and was restored by inhibiting AMPK

  11. Proliferation studies of the endothelial and smooth muscle cells of the mouse mesentery after irradiation

    International Nuclear Information System (INIS)

    Hirst, D.G.; Denekamp, J.; Hobson, B.

    1980-01-01

    A continuous tritium labelling technique was employed to study the effects of external β-radiation on the proliferation of endothelial cells and smooth muscle cells in the mesenteric arterioles of mice. Calculations showed very long turnover times for the two cell populations in control animals (> 2 years for endothelium and > 3 years for smooth muscle). After single doses of 20 and 45 Gy, no significant increase in endothelial proliferation was seen except at 3 weeks. No significant increase in labelling was observed in smooth muscle up to 48 weeks after irradiation. These labelling data have been compared with the pattern of cell depletion of the irradiated endothelium. It was concluded that the depletion was much earlier than expected for a slowly proliferating tissue, if all the cells were cycling very slowly. Such an early depletion is, however, consistent with cell death resulting from a small proportion of the cells having a short cell cycle. The recovery of the endothelial cell numbers between 9 and 12 months was not accompanied by a rise in the fraction of labelled cells. It is suggested that repopulation may occur from outside the treated area. (author)

  12. Danshensu prevents hypoxic pulmonary hypertension in rats by inhibiting the proliferation of pulmonary artery smooth muscle cells via TGF-β-smad3-associated pathway.

    Science.gov (United States)

    Zhang, Ning; Dong, Mingqing; Luo, Ying; Zhao, Feng; Li, Yongjun

    2018-02-05

    Hypoxic pulmonary hypertension is characterized by the remodeling of pulmonary artery. Previously we showed that tanshinone IIA, one lipid-soluble component from the Chinese herb Danshen, ameliorated hypoxic pulmonary hypertension by inhibiting pulmonary artery remodeling. Here we explored the effects of danshensu, one water-soluble component of Danshen, on hypoxic pulmonary hypertension and its mechanism. Rats were exposed to hypobaric hypoxia for 4 weeks to develop hypoxic pulmonary hypertension along with administration of danshensu. Hemodynamics and pulmonary arterial remodeling index were measured. The effects of danshensu on the proliferation of primary pulmonary artery smooth muscle cells and transforming growth factor-β-smad3 pathway were assessed in vitro. Danshensu significantly decreased the right ventricle systolic pressure, the right ventricle hypertrophy and pulmonary vascular remodeling index in hypoxic pulmonary hypertension rats. Danshensu also reduced the increased expression of transforming growth factor-β and phosphorylation of smad3 in pulmonary arteries in hypoxic pulmonary hypertension rats. In vitro, danshensu inhibited the hypoxia- or transforming growth factor-β-induced proliferation of primary pulmonary artery smooth muscle cells. Moreover, danshensu decreased the hypoxia-induced expression and secretion of transforming growth factor in primary pulmonary adventitial fibroblasts and NR8383 cell line, inhibited the hypoxia or transforming growth factor-β-induced phosphorylation of smad3 in rat primary pulmonary artery smooth muscle cells. These results demonstrate that danshensu ameliorates hypoxic pulmonary hypertension in rats by inhibiting the hypoxia-induced proliferation of pulmonary artery smooth muscle cells, and the inhibition effects is associated with transforming growth factor-β-smad3 pathway. Therefore danshensu may be a potential treatment for hypoxic pulmonary hypertension. Copyright © 2017 Elsevier B.V. All rights

  13. From depolarization-dependent contractions in gastrointestinal smooth muscle to aortic pulse-synchronized contractions

    Directory of Open Access Journals (Sweden)

    Marion SB

    2014-03-01

    Full Text Available Sarah B Marion, Allen W MangelRTI Health Solutions, Research Triangle Park, NC, USAAbstract: For decades, it was believed that the diameter of gastrointestinal smooth muscle cells is sufficiently narrow, and that the diffusion of calcium across the plasma membrane is sufficient, to support contractile activity. Thus, depolarization-triggered release of intracellular calcium was not believed to be operative in gastrointestinal smooth muscle. However, after the incubation of muscle segments in solutions devoid of calcium and containing the calcium chelator ethylene glycol tetraacetic acid, an alternative electrical event occurred that was distinct from normal slow waves and spikes. Subsequently, it was demonstrated in gastrointestinal smooth muscle segments that membrane depolarization associated with this alternative electrical event triggered rhythmic contractions by release of intracellular calcium. Although this concept of depolarization-triggered calcium release was iconoclastic, it has now been demonstrated in multiple gastrointestinal smooth muscle preparations. On the basis of these observations, we investigated whether a rhythmic electrical and mechanical event would occur in aortic smooth muscle under the same calcium-free conditions. The incubation of aortic segments in a solution with no added calcium plus ethylene glycol tetraacetic acid induced a fast electrical event without corresponding tension changes. On the basis of the frequency of these fast electrical events, we pursued, contrary to what has been established dogma for more than three centuries, the question of whether the smooth muscle wall of the aorta undergoes rhythmic activation during the cardiac cycle. As with depolarization-triggered contractile activity in gastrointestinal smooth muscle, it was “well known” that rhythmic activation of the aorta does not occur in synchrony with the heartbeat. In a series of experiments, however, it was demonstrated that rhythmic

  14. The Smooth Muscle of the Artery

    Science.gov (United States)

    1975-01-01

    of vascular smooth muscle are contrac- tion, thereby mediating vaso constriction, and the synthesis of the extracellular proteins and polysaccharides ...of the monosaccharides turned out to be different for instance from cornea to aorta (229, 283). In the conditions yed (4 hours incubation at 37 degrees... polysaccharides only. This glyco- protein is not very rich in sugar components (- 5Z) (228, 284), but is a very acidic protein (286). Fig.66 shows

  15. Voltage-dependent inward currents in smooth muscle cells of skeletal muscle arterioles

    Science.gov (United States)

    Shirokov, Roman E.

    2018-01-01

    Voltage-dependent inward currents responsible for the depolarizing phase of action potentials were characterized in smooth muscle cells of 4th order arterioles in mouse skeletal muscle. Currents through L-type Ca2+ channels were expected to be dominant; however, action potentials were not eliminated in nominally Ca2+-free bathing solution or by addition of L-type Ca2+ channel blocker nifedipine (10 μM). Instead, Na+ channel blocker tetrodotoxin (TTX, 1 μM) reduced the maximal velocity of the upstroke at low, but not at normal (2 mM), Ca2+ in the bath. The magnitude of TTX-sensitive currents recorded with 140 mM Na+ was about 20 pA/pF. TTX-sensitive currents decreased five-fold when Ca2+ increased from 2 to 10 mM. The currents reduced three-fold in the presence of 10 mM caffeine, but remained unaltered by 1 mM of isobutylmethylxanthine (IBMX). In addition to L-type Ca2+ currents (15 pA/pF in 20 mM Ca2+), we also found Ca2+ currents that are resistant to 10 μM nifedipine (5 pA/pF in 20 mM Ca2+). Based on their biophysical properties, these Ca2+ currents are likely to be through voltage-gated T-type Ca2+ channels. Our results suggest that Na+ and at least two types (T- and L-) of Ca2+ voltage-gated channels contribute to depolarization of smooth muscle cells in skeletal muscle arterioles. Voltage-gated Na+ channels appear to be under a tight control by Ca2+ signaling. PMID:29694371

  16. Cardiac, skeletal, and smooth muscle mitochondrial respiration: are all mitochondria created equal?

    Science.gov (United States)

    Park, Song-Young; Gifford, Jayson R; Andtbacka, Robert H I; Trinity, Joel D; Hyngstrom, John R; Garten, Ryan S; Diakos, Nikolaos A; Ives, Stephen J; Dela, Flemming; Larsen, Steen; Drakos, Stavros; Richardson, Russell S

    2014-08-01

    Unlike cardiac and skeletal muscle, little is known about vascular smooth muscle mitochondrial respiration. Therefore, the present study examined mitochondrial respiratory rates in smooth muscle of healthy human feed arteries and compared with that of healthy cardiac and skeletal muscles. Cardiac, skeletal, and smooth muscles were harvested from a total of 22 subjects (53 ± 6 yr), and mitochondrial respiration was assessed in permeabilized fibers. Complex I + II, state 3 respiration, an index of oxidative phosphorylation capacity, fell progressively from cardiac to skeletal to smooth muscles (54 ± 1, 39 ± 4, and 15 ± 1 pmol·s(-1)·mg(-1), P respiration rates were normalized by CS (respiration per mitochondrial content), oxidative phosphorylation capacity was no longer different between the three muscle types. Interestingly, complex I state 2 normalized for CS activity, an index of nonphosphorylating respiration per mitochondrial content, increased progressively from cardiac to skeletal to smooth muscles, such that the respiratory control ratio, state 3/state 2 respiration, fell progressively from cardiac to skeletal to smooth muscles (5.3 ± 0.7, 3.2 ± 0.4, and 1.6 ± 0.3 pmol·s(-1)·mg(-1), P respiration highlight the existence of intrinsic functional differences between these muscle mitochondria. This likely influences the efficiency of oxidative phosphorylation and could potentially alter ROS production.

  17. Acute insulin resistance stimulates and insulin sensitization attenuates vascular smooth muscle cell migration and proliferation.

    Science.gov (United States)

    Cersosimo, Eugenio; Xu, Xiaojing; Upala, Sikarin; Triplitt, Curtis; Musi, Nicolas

    2014-08-01

    Differential activation/deactivation of insulin signaling, PI-3K and MAP-K pathways by high glucose and palmitate, with/out the insulin sensitizer pioglitazone (PIO), have been previously shown in vascular smooth muscle cells (VSMCs). To determine the biological impact of these molecular changes, we examined VSMC migration and proliferation ("M"&"P") patterns in similar conditions. VSMCs from healthy human coronary arteries were incubated in growth medium and "M"&"P" were analyzed after exposure to high glucose (25 mmol/L) ± palmitate (200 μmol/L) and ± PIO (8 μmol/L) for 5 h. "M"&"P" were assessed by: (1) polycarbonate membrane barrier with chemo-attractants and extended cell protrusions quantified by optical density (OD595 nm); (2) % change in radius area (2D Assay) using inverted microscopy images; and (3) cell viability assay expressed as cell absorbance (ABS) in media. "M" in 25 mmol/L glucose media increased by ~25% from baseline and % change in radius area rose from ~20% to ~30%. The addition of PIO was accompanied by a significant decrease in "M" from 0.25 ± 0.02 to 0.19 ± 0.02; a comparable decline from 0.25 ± 0.02 to 0.18 ± 0.02 was also seen with 25 mmol/L of glucose +200 μmol/L of palmitate. When PIO was coincubated with high glucose plus palmitate there was a 50% reduction in % change in radius. A ~10% increase in ABS, reflecting augmented "P" in media with 25 mmol/L glucose versus control was documented. The addition of PIO reduced ABS from 0.208 ± 0.03 to 0.183 ± 0.06. Both high glucose and palmitate showed ABS of ~0.140 ± 0.02, which decreased with PIO to ~0.120 ± 0.02, indicating "P" was reduced. These results confirm that high glucose and palmitate stimulate VSMCs migration and proliferation in vitro, which is attenuated by coincubation with the insulin sensitizer PIO. Although, we cannot ascertain whether these functional changes are coincident with the activation/deactivation of signal molecules, our findings are consistent with the

  18. Mast cell numbers in airway smooth muscle and PC(20)AMP in asthma and COPD

    NARCIS (Netherlands)

    Liesker, J. J. W.; ten Hacken, N. H. T.; Rutgers, S. R.; Zeinstra-Smith, M.; Postma, D. S.; Timens, W.

    Introduction: Most patients with asthma and many patients with COPD show bronchial hyperresponsiveness to adenosine (BHRAMP). BHRAMP may be caused by release of mast cell histamine, which induces smooth muscle contraction. Aim of the study: To evaluate whether mast cell numbers in airway smooth

  19. Endometrial stem cell differentiation into smooth muscle cell: a novel approach for bladder tissue engineering in women.

    Science.gov (United States)

    Shoae-Hassani, Alireza; Sharif, Shiva; Seifalian, Alexander M; Mortazavi-Tabatabaei, Seyed Abdolreza; Rezaie, Sassan; Verdi, Javad

    2013-10-01

    To investigate manufacturing smooth muscle cells (SMCs) for regenerative bladder reconstruction from differentiation of endometrial stem cells (EnSCs), as the recent discovery of EnSCs from the lining of women's uteri, opens up the possibility of using these cells for tissue engineering applications, such as building up natural tissue to repair prolapsed pelvic floors as well as building urinary bladder wall. Human EnSCs that were positive for cluster of differentiation 146 (CD146), CD105 and CD90 were isolated and cultured in Dulbecco's modified Eagle/F12 medium supplemented with myogenic growth factors. The myogenic factors included: transforming growth factor β, platelet-derived growth factor, hepatocyte growth factor and vascular endothelial growth factor. Differentiated SMCs on bioabsorbable polyethylene-glycol and collagen hydrogels were checked for SMC markers by real-time reverse-transcriptase polymerase chain reaction (RT-PCR), western blot (WB) and immunocytochemistry (ICC) analyses. Histology confirmed the growth of SMCs in the hydrogel matrices. The myogenic growth factors decreased the proliferation rate of EnSCs, but they differentiated the human EnSCs into SMCs more efficiently on hydrogel matrices and expressed specific SMC markers including α-smooth muscle actin, desmin, vinculin and calponin in RT-PCR, WB and ICC experiments. The survival rate of cultures on the hydrogel-coated matrices was significantly higher than uncoated cultures. Human EnSCs were successfully differentiated into SMCs, using hydrogels as scaffold. EnSCs may be used for autologous bladder wall regeneration without any immunological complications in women. Currently work is in progress using bioabsorbable nanocomposite materials as EnSC scaffolds for developing urinary bladder wall tissue. © 2013 The Authors. BJU International © 2013 BJU International.

  20. Role of cyclic GMP in cells with the properties of smooth muscle cultured from the rat myometrium

    International Nuclear Information System (INIS)

    Krall, J.F.; Morin, A.

    1986-01-01

    Cells growing in culture with previously described properties of rat uterine smooth muscle accumulated 45 Ca 2+ from the medium. Ca 2+ uptake by these cells was stimulated by the addition to the medium of 8-bromo-cGMP but not by 8-bromo-cAMP. Ca 2+ uptake was also stimulated by carbachol and by the nitro-vasodilator nitroprusside. Although cholinergic agonists have been shown previously to stimulate contraction but not cGMP synthesis in the rat myometrium, both carbachol and nitroprusside stimulated cGMP production by the cultured cells. These results suggested the cells had cholinergic receptor-medicated functions that reflected some neurotransmitter-sensitive properties of uterine smooth muscle in situ. When determined by a specific radioligand binding assay, subcellular fractions of the cultured cells bound muscarinic cholinergic agonists and antagonists with affinities expected of the muscarinic receptor. The cells were also sensitive to the β-adrenergic catecholamine agonist isoproterenol, which stimulated cAMP production but not Ca 2+ uptake. Carbachol failed to inhibit isoproterenol-dependent cAMP production, which is an important property of the cholinergic receptor in uterine smooth muscle in situ. These results suggest some but not all acetylcholine-sensitive properties of uterine smooth muscle may be retained in cell culture

  1. Epothilones Suppress Neointimal Thickening in the Rat Carotid Balloon-Injury Model by Inducing Vascular Smooth Muscle Cell Apoptosis through p53-Dependent Signaling Pathway.

    Science.gov (United States)

    Son, Dong Ju; Jung, Jae Chul; Hong, Jin Tae

    2016-01-01

    Microtubule stabilizing agents (MTSA) are known to inhibit vascular smooth muscle cell (VSMC) proliferation and migration, and effectively reduce neointimal hyperplasia and restenosis. Epothilones (EPOs), non-taxane MTSA, have been found to be effective in the inhibition of VSMC proliferation and neointimal formation by cell cycle arrest. However, effect of EPOs on apoptosis in hyper-proliferated VSMCs as a possible way to reduce neointimal formation and its action mechanism related to VSMC viability has not been suited yet. Thus, the purposes of the present study was to investigate whether EPOs are able to inhibit neointimal formation by inducing apoptosis within the region of neointimal hyperplasia in balloon-injured rat carotid artery, as well as underlying action mechanism. Treatment of EPO-B and EPO-D significantly induced apoptotic cell death and mitotic catastrophe in hyper-proliferated VSMCs, resulting in cell growth inhibition. Further, EPOs significantly suppressed VSMC proliferation and induced apoptosis by activation of p53-dependent apoptotic signaling pathway, Bax/cytochrome c/caspase-3. We further demonstrated that the local treatment of carotid arteries with EPOs potently inhibited neointimal lesion formation by induction of apoptosis in rat carotid injury model. Our findings demonstrate a potent anti-neointimal hyperplasia property of EPOs by inducing p53-depedent apoptosis in hyper-proliferated VSMCs.

  2. Attenuation of chondrogenic transformation in vascular smooth muscle by dietary quercetin in the MGP-deficient mouse model.

    Directory of Open Access Journals (Sweden)

    Kelly E Beazley

    Full Text Available Cartilaginous metaplasia of vascular smooth muscle (VSM is characteristic for arterial calcification in diabetes and uremia and in the background of genetic alterations in matrix Gla protein (MGP. A better understanding of the molecular details of this process is critical for the development of novel therapeutic approaches to VSM transformation and arterial calcification.This study aimed to identify the effects of bioflavonoid quercetin on chondrogenic transformation and calcification of VSM in the MGP-null mouse model and upon TGF-β3 stimulation in vitro, and to characterize the associated alterations in cell signaling.Molecular analysis revealed activation of β-catenin signaling in cartilaginous metaplasia in Mgp-/- aortae in vivo and during chondrogenic transformation of VSMCs in vitro. Quercetin intercepted chondrogenic transformation of VSM and blocked activation of β-catenin both in vivo and in vitro. Although dietary quercetin drastically attenuated calcifying cartilaginous metaplasia in Mgp-/- animals, approximately one-half of total vascular calcium mineral remained as depositions along elastic lamellae.Quercetin is potent in preventing VSM chondrogenic transformation caused by diverse stimuli. Combined with the demonstrated efficiency of dietary quercetin in preventing ectopic chondrogenesis in the MGP-null vasculature, these findings indicate a potentially broad therapeutic applicability of this safe for human consumption bioflavonoid in the therapy of cardiovascular conditions linked to cartilaginous metaplasia of VSM. Elastocalcinosis is a major component of MGP-null vascular disease and is controlled by a mechanism different from chondrogenic transformation of VSM and not sensitive to quercetin.

  3. Macrophage secretory products selectively stimulate dermatan sulfate proteoglycan production in cultured arterial smooth muscle cells

    International Nuclear Information System (INIS)

    Edwards, I.J.; Wagner, W.D.; Owens, R.T.

    1990-01-01

    Arterial dermatan sulfate proteoglycan has been shown to increase with atherosclerosis progression, but factors responsible for this increase are unknown. To test the hypothesis that smooth muscle cell proteoglycan synthesis may be modified by macrophage products, pigeon arterial smooth muscle cells were exposed to the media of either cholesteryl ester-loaded pigeon peritoneal macrophages or a macrophage cell line P388D1. Proteoglycans radiolabeled with [35S]sulfate and [3H]serine were isolated from culture media and smooth muscle cells and purified following precipitation with 1-hexadecylpyridinium chloride and chromatography. Increasing concentrations of macrophage-conditioned media were associated with a dose-response increase in [35S]sulfate incorporation into secreted proteoglycans, but there was no change in cell-associated proteoglycans. Incorporation of [3H]serine into total proteoglycan core proteins was not significantly different (5.2 X 10(5) dpm and 5.5 X 10(5) disintegrations per minute (dpm) in control and conditioned media-treated cultures, respectively), but selective effects were observed on individual proteoglycan types. Twofold increases in dermatan sulfate proteoglycan and limited degradation of chondroitin sulfate proteoglycan were apparent based on core proteins separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Immunoinhibition studies indicated that interleukin-1 was involved in the modulation of proteoglycan synthesis by macrophage-conditioned media. These data provide support for the role of macrophages in alteration of the matrix proteoglycans synthesized by smooth muscle cells and provide a mechanism to account for the reported increased dermatan sulfate/chondroitin sulfate ratios in the developing atherosclerotic lesion

  4. MicroRNA-222 Promotes the Proliferation of Pulmonary Arterial Smooth Muscle Cells by Targeting P27 and TIMP3

    Directory of Open Access Journals (Sweden)

    Ying Xu

    2017-08-01

    Full Text Available Background/Aims: Aberrant vascular smooth muscle cell (VSMC proliferation plays an important role in the development of pulmonary artery hypertension (PAH. Dysregulated microRNAs (miRNAs, miRs have been implicated in the progression of PAH. miR-222 has a pro-proliferation effect on VSMCs while it has an anti-proliferation effect on vascular endothelial cells (ECs. As the biological function of a single miRNA could be cell-type specific, the role of miR-222 in pulmonary artery smooth muscle cell (PASMC proliferation is not clear and deserves to be explored. Methods: PASMCs were transfected with miR-222 mimic or inhibitor and PASMC proliferation was determined by Western blot for PCNA, Ki-67 and EdU staining, and cell number counting. The target genes of miR-222 including P27 and TIMP3 were determined by luciferase assay and Western blot. In addition, the functional rescue experiments were performed based on miR-222 inhibitor and siRNAs to target genes. Results: miR-222 mimic promoted PASMC proliferation while miR-222 inhibitor decreased that. TIMP3 was identified to be a direct target gene of miR-222 based on luciferase assay. Meanwhile, P27 and TIMP3 were up-regulated by miR-222 inhibitor and down-regulated by miR-222 mimic. Moreover, P27 siRNA and TIMP3 siRNA could both attenuate the anti-proliferation effect of miR-222 inhibitor in PASMCs, supporting that P27 and TIMP3 are at least partially responsible for the regulatory effect of miR-222 in PASMCs. Conclusion: miR-222 promotes PASMC proliferation at least partially through targeting P27 and TIMP3.

  5. Aspirin-induced AMP-activated protein kinase activation regulates the proliferation of vascular smooth muscle cells from spontaneously hypertensive rats

    International Nuclear Information System (INIS)

    Sung, Jin Young; Choi, Hyoung Chul

    2011-01-01

    Highlights: → Aspirin-induced AMPK phosphorylation was greater in VSMC from SHR than WKY. → Aspirin-induced AMPK phosphorylation inhibited proliferation of VSMC from SHR. → Low basal AMPK phosphorylation in SHR elicits increased VSMC proliferation. → Inhibition of AMPK restored decreased VSMC proliferation by aspirin in SHR. → Aspirin exerts anti-proliferative effect through AMPK activation in VSMC from SHR. -- Abstract: Acetylsalicylic acid (aspirin), used to reduce risk of cardiovascular disease, plays an important role in the regulation of cellular proliferation. However, mechanisms responsible for aspirin-induced growth inhibition are not fully understood. Here, we investigated whether aspirin may exert therapeutic effects via AMP-activated protein kinase (AMPK) activation in vascular smooth muscle cells (VSMC) from wistar kyoto rats (WKY) and spontaneously hypertensive rats (SHR). Aspirin increased AMPK and acetyl-CoA carboxylase phosphorylation in a time- and dose-dependent manner in VSMCs from WKY and SHR, but with greater efficacy in SHR. In SHR, a low basal phosphorylation status of AMPK resulted in increased VSMC proliferation and aspirin-induced AMPK phosphorylation inhibited proliferation of VSMCs. Compound C, an AMPK inhibitor, and AMPK siRNA reduced the aspirin-mediated inhibition of VSMC proliferation, this effect was more pronounced in SHR than in WKY. In VSMCs from SHR, aspirin increased p53 and p21 expression and inhibited the expression of cell cycle associated proteins, such as p-Rb, cyclin D, and cyclin E. These results indicate that in SHR VSMCs aspirin exerts anti-proliferative effects through the induction of AMPK phosphorylation.

  6. Aspirin-induced AMP-activated protein kinase activation regulates the proliferation of vascular smooth muscle cells from spontaneously hypertensive rats

    Energy Technology Data Exchange (ETDEWEB)

    Sung, Jin Young [Department of Pharmacology, College of Medicine, Yeungnam University, Daegu 705-717 (Korea, Republic of); Choi, Hyoung Chul, E-mail: hcchoi@med.yu.ac.kr [Department of Pharmacology, College of Medicine, Yeungnam University, Daegu 705-717 (Korea, Republic of)

    2011-05-06

    Highlights: {yields} Aspirin-induced AMPK phosphorylation was greater in VSMC from SHR than WKY. {yields} Aspirin-induced AMPK phosphorylation inhibited proliferation of VSMC from SHR. {yields} Low basal AMPK phosphorylation in SHR elicits increased VSMC proliferation. {yields} Inhibition of AMPK restored decreased VSMC proliferation by aspirin in SHR. {yields} Aspirin exerts anti-proliferative effect through AMPK activation in VSMC from SHR. -- Abstract: Acetylsalicylic acid (aspirin), used to reduce risk of cardiovascular disease, plays an important role in the regulation of cellular proliferation. However, mechanisms responsible for aspirin-induced growth inhibition are not fully understood. Here, we investigated whether aspirin may exert therapeutic effects via AMP-activated protein kinase (AMPK) activation in vascular smooth muscle cells (VSMC) from wistar kyoto rats (WKY) and spontaneously hypertensive rats (SHR). Aspirin increased AMPK and acetyl-CoA carboxylase phosphorylation in a time- and dose-dependent manner in VSMCs from WKY and SHR, but with greater efficacy in SHR. In SHR, a low basal phosphorylation status of AMPK resulted in increased VSMC proliferation and aspirin-induced AMPK phosphorylation inhibited proliferation of VSMCs. Compound C, an AMPK inhibitor, and AMPK siRNA reduced the aspirin-mediated inhibition of VSMC proliferation, this effect was more pronounced in SHR than in WKY. In VSMCs from SHR, aspirin increased p53 and p21 expression and inhibited the expression of cell cycle associated proteins, such as p-Rb, cyclin D, and cyclin E. These results indicate that in SHR VSMCs aspirin exerts anti-proliferative effects through the induction of AMPK phosphorylation.

  7. MicroRNAs dynamically remodel gastrointestinal smooth muscle cells.

    Directory of Open Access Journals (Sweden)

    Chanjae Park

    2011-04-01

    Full Text Available Smooth muscle cells (SMCs express a unique set of microRNAs (miRNAs which regulate and maintain the differentiation state of SMCs. The goal of this study was to investigate the role of miRNAs during the development of gastrointestinal (GI SMCs in a transgenic animal model. We generated SMC-specific Dicer null animals that express the reporter, green fluorescence protein, in a SMC-specific manner. SMC-specific knockout of Dicer prevented SMC miRNA biogenesis, causing dramatic changes in phenotype, function, and global gene expression in SMCs: the mutant mice developed severe dilation of the intestinal tract associated with the thinning and destruction of the smooth muscle (SM layers; contractile motility in the mutant intestine was dramatically decreased; and SM contractile genes and transcriptional regulators were extensively down-regulated in the mutant SMCs. Profiling and bioinformatic analyses showed that SMC phenotype is regulated by a complex network of positive and negative feedback by SMC miRNAs, serum response factor (SRF, and other transcriptional factors. Taken together, our data suggest that SMC miRNAs are required for the development and survival of SMCs in the GI tract.

  8. EVALUATION OF THE FUNCTIONAL PROPERTIES OF HUMAN ENDOTHELIAL AND SMOOTH MUSCLE CELLS AFTER SEEDING ON THE SURFACE OF NATURAL AND SYNTHETIC MATERIALS

    Directory of Open Access Journals (Sweden)

    Sh. B. Saaya

    2016-01-01

    Full Text Available At present, vascular surgery using small diameter synthetic grafts is associated with a higher incidence of complications (thrombosis, restenosis, intimal hyperplasia than in operations using autologous vessels. However, the occurrence of concomitant pathology, reoperations and multifocal vascular disease limit the use of autologous vein and arteries. The important factor providing a long-term patency is the presence of vascular cells, which produce biologically active substance and provide mechanical properties. Aim. Selection of the optimal scaffold for creating cell-seeded tissue-engineering vessels. Materials and methods. Endothelial (EC and smooth muscle cells (SMC derived from human myocardium were seeded on different surfaces: decellularized homoarteriа, хenopericardium, polytetrafl uoroethylene (PTFE, polyethylene terephthalate (PET, polycaprolactone (PCL and polylactide-co-glycolide (PLGA. Results. Synthetic biodegradable materials polycaprolactone and polylactide-co-glycolide provide cell adhesion. The cells cultured on the polycaprolactone and polylactide-coglycolide scaffolds retain their functional properties: viability and proliferative properties, maintain specifi c endothelial antigens and synthesis of extracellular matrix. Conclusion. Synthetic biodegradable polycaprolactone and polylactide-co-glycolide electrospun scaffolds can be used for creation of cell-fi lled vascular prostheses. 

  9. Oxygen mediates vascular smooth muscle relaxation in hypoxia.

    Directory of Open Access Journals (Sweden)

    Jessica Dada

    Full Text Available The activation of soluble guanylate cyclase (sGC by nitric oxide (NO and other ligands has been extensively investigated for many years. In the present study we considered the effect of molecular oxygen (O2 on sGC both as a direct ligand and its affect on other ligands by measuring cyclic guanosine monophosphate (cGMP production, as an index of activity, as well as investigating smooth muscle relaxation under hypoxic conditions. Our isolated enzyme studies confirm the function of sGC is impaired under hypoxic conditions and produces cGMP in the presence of O2, importantly in the absence of NO. We also show that while O2 could partially affect the magnitude of sGC stimulation by NO when the latter was present in excess, activation by the NO independent, haem-dependent sGC stimulator 3-(5'-hydroxymethyl-2'-furyl-1-benzylindazole (YC-1 was unaffected. Our in vitro investigation of smooth muscle relaxation confirmed that O2 alone in the form of a buffer bolus (equilibrated at 95% O2/5% CO2 had the ability to dilate vessels under hypoxic conditions and that this was dependent upon sGC and independent of eNOS. Our studies confirm that O2 can be a direct and important mediator of vasodilation through an increase in cGMP production. In the wider context, these observations are key to understanding the relative roles of O2 versus NO-induced sGC activation.

  10. Vascular smooth muscle modulates endothelial control of vasoreactivity via reactive oxygen species production through myoendothelial communications.

    Directory of Open Access Journals (Sweden)

    Marie Billaud

    Full Text Available BACKGROUND: Endothelial control of vascular smooth muscle plays a major role in the resulting vasoreactivity implicated in physiological or pathological circulatory processes. However, a comprehensive understanding of endothelial (EC/smooth muscle cells (SMC crosstalk is far from complete. Here, we have examined the role of gap junctions and reactive oxygen species (ROS in this crosstalk and we demonstrate an active contribution of SMC to endothelial control of vasomotor tone. METHODOLOGY/PRINCIPAL FINDINGS: In small intrapulmonary arteries, quantitative RT-PCR, Western Blot analyses and immunofluorescent labeling evidenced connexin (Cx 37, 40 and 43 in EC and/or SMC. Functional experiments showed that the Cx-mimetic peptide targeted against Cx 37 and Cx 43 ((37,43Gap27 (1 reduced contractile and calcium responses to serotonin (5-HT simultaneously recorded in pulmonary arteries and (2 abolished the diffusion in SMC of carboxyfluorescein-AM loaded in EC. Similarly, contractile and calcium responses to 5-HT were decreased by superoxide dismutase and catalase which, catabolise superoxide anion and H(2O(2, respectively. Both Cx- and ROS-mediated effects on the responses to 5-HT were reversed by L-NAME, a NO synthase inhibitor or endothelium removal. Electronic paramagnetic resonance directly demonstrated that 5-HT-induced superoxide anion production originated from the SMC. Finally, whereas 5-HT increased NO production, it also decreased cyclic GMP content in isolated intact arteries. CONCLUSIONS/SIGNIFICANCE: These data demonstrate that agonist-induced ROS production in SMC targeting EC via myoendothelial gap junctions reduces endothelial NO-dependent control of pulmonary vasoreactivity. Such SMC modulation of endothelial control may represent a signaling pathway controlling vasoreactivity under not only physiological but also pathological conditions that often implicate excessive ROS production.

  11. Adhesion and growth of rat aortic smooth muscle cells on lactide-based polymers

    Czech Academy of Sciences Publication Activity Database

    Bačáková, Lucie; Lapčíková, Monika; Kubies, Dana; Rypáček, František

    2003-01-01

    Roč. 534, - (2003), s. 179-189 ISSN 0065-2598 R&D Projects: GA AV ČR IAA4050202; GA MŠk LN00A065 Institutional research plan: CEZ:AV0Z4050913; CEZ:AV0Z5011922 Keywords : endothelial cells(EC) * vascular smooth cells (VSMC) Subject RIV: EI - Biotechnology ; Bionics

  12. Smooth muscle cell phenotypic switching in stroke.

    Science.gov (United States)

    Poittevin, Marine; Lozeron, Pierre; Hilal, Rose; Levy, Bernard I; Merkulova-Rainon, Tatiana; Kubis, Nathalie

    2014-06-01

    Disruption of cerebral blood flow after stroke induces cerebral tissue injury through multiple mechanisms that are not yet fully understood. Smooth muscle cells (SMCs) in blood vessel walls play a key role in cerebral blood flow control. Cerebral ischemia triggers these cells to switch to a phenotype that will be either detrimental or beneficial to brain repair. Moreover, SMC can be primarily affected genetically or by toxic metabolic molecules. After stroke, this pathological phenotype has an impact on the incidence, pattern, severity, and outcome of the cerebral ischemic disease. Although little research has been conducted on the pathological role and molecular mechanisms of SMC in cerebrovascular ischemic diseases, some therapeutic targets have already been identified and could be considered for further pharmacological development. We examine these different aspects in this review.

  13. Nonparenchymal cells cultivated from explants of fibrotic liver resemble endothelial and smooth muscle cells from blood vessel walls

    International Nuclear Information System (INIS)

    Voss, B.; Rauterberg, J.; Pott, G.; Brehmer, U.; Allam, S.; Lehmann, R.; von Bassewitz, D.B.

    1982-01-01

    Tissue specimens from human fibrotic liver obtained by needle biopsy were cultured. Two cell types emerged from the tissue explants. From their morphology and biosynthetic products they resembled smooth muscle cells and endothelial cells from blood vessel walls. In the endothelial cells, factor VIII-associated protein was demonstrated by indirect immunofluorescence. Synthesis of collagen types I and III, basement membrane collagen types IV and V, and fibronectin by both cell types was observed by immunofluorescence microscopy. Homogeneous cultures of smooth muscle cells were observed in subcultures. After incubation with [ 14 C]glycine, collagen was isolated and characterized by CM cellulose chromatography, and consisted mainly of types I and III. These data suggest involvement of mesenchymal cells in hepatic fibrosis; they presumably originate from blood vessel or sinusoidal walls

  14. TNF-α activates death pathway in human aorta smooth muscle cell in the presence of 7-ketocholesterol

    International Nuclear Information System (INIS)

    Lee, Hyun Sun; Chang, Jong Sun; Baek, Jin Ah; Chung, Mi Yeon; Lee, Han Cheol; Rhim, Byung Yong; Sok, Dai Eun; Rho, Mun-Chual; Kim, Young Kook; Kim, Koanhoi

    2005-01-01

    This study was undertaken to investigate whether a physiologically compatible concentration of 7-ketocholesterol had any effect on human vascular smooth muscle cells (HVSMCs). We found that 7-ketocholesterol changed the viability of human aorta smooth muscle cells (HAoSMC) not by cytotoxicity but by activation of tumor necrosis factor-α receptor (TNFR)-mediated death. Whereas TNF-α did not affect the viability in the presence of 7α-hydroxycholesterol or cholesterol, the cytokine induced HAoSMC death in the presence of 7-ketocholesterol as detected by morphology, viability, and fragmentation of chromosomal DNA. The HAoSMC death was inhibited by a neutralizing anti-TNF receptor 1 (TNFR1) antibody and by the caspase inhibitors of z-VAD and z-DEVD. Activations of caspase-8 and -3 were detected from dying HAoSMCs. 7-Ketocholesterol inhibited translocation of the nuclear factor κB (NF-κB) subunits of p65 and p50 from the cytosol into the nucleus, increase of NF-κB activity, and expression of caspase-8 homolog Fas ligand interleukin-1-converting enzyme inhibitory protein by TNF-α. We also found that X-chromosome-linked inhibitor of apoptosis protein was degraded in dying HAoSMC. The present study proposes that 7-ketocholesterol would contribute to the disappearance of HVSMC in the atherosclerotic lesions by enhancing receptor-mediated death. This is the first report demonstrating induction of TNF-α-mediated death by oxysterol in cells

  15. Reversal of sodium pump inhibitor induced vascular smooth muscle contraction with digibind. Stoichiometry and its implications.

    Science.gov (United States)

    Krep, H H; Graves, S W; Price, D A; Lazarus, M; Ensign, A; Soszynski, P A; Hollenberg, N K

    1996-01-01

    The possibility that a circulating sodium pump inhibitor contributes to the pathogenesis of volume-dependent hypertension via an action on vascular smooth muscle (VSM) is supported by multiple lines of investigation, but remains controversial. We had two goals in this study. The first was to compare the pattern of contractile response of rabbit aorta induced by two candidates, ouabain and a labile sodium pump inhibitor that we have identified in the peritoneal dialysate of volume-expanded hypertensive patients with chronic renal failure. Our second goal was to examine the ability of Digibind, a Fab fragment of antisera directed against digoxin, to reverse VSM contraction induced by both agents. Ouabain induced a concentration-dependent contraction, which was delayed in onset, was gradual, and reached a stable plateau after many hours. The labile sodium pump inhibitor induced a qualitatively similar series of responses. Digibind rapidly reversed the contractile responses to both sodium pump inhibitors, with a rate of relaxation that matched that induced by physical removal of the pump inhibitor from the bath. For ouabain, the Digibind:ouabain stoichiometry was highly predictable. When Digibind was present in a molar concentration equivalent to that of ouabain, or less, it had no effect. When the Digibind concentration was twice that of ouabain, complete relaxation occurred. Although the concentration:VSM response relationship for ouabain was steep, the concentration:effect interaction with Digibind was even more steep. The molar concentration of Digibind required to reverse the effects of the labile endogenous inhibitor from peritoneal dialysate was consistently lower than that for ouabain, which is compatible with either greater potency of the labile factor in VSM or greater affinity for Digibind. These findings are compatible with a role for one or more endogenous sodium pump inhibitors as the determinant of vascular smooth muscle tone in the volume

  16. Lipid-soluble smoke particles upregulate vascular smooth muscle ETB receptors via activation of mitogen-activating protein kinases and NF-kappaB pathways

    DEFF Research Database (Denmark)

    Xu, C.B.; Zheng, J.P.; Zhang, W.

    2008-01-01

    Cigarette smoke is a strong risk factor for cardiovascular disease. However, the underlying molecular mechanisms that lead to cigarette smoke-associated cardiovascular disease remain elusive. With functional and molecular methods, we demonstrate for the first time that lipid-soluble cigarette smoke...... particles (dimethylsulfoxide-soluble cigarette smoke particles; DSP) increased the expression of endothelin type B (ET(B)) receptors in arterial smooth muscle cells. The increased ET(B) receptors in arterial smooth muscle cells was documented as enhanced contractility (sensitive myograph technique...

  17. Calcium signaling in smooth muscle.

    Science.gov (United States)

    Hill-Eubanks, David C; Werner, Matthias E; Heppner, Thomas J; Nelson, Mark T

    2011-09-01

    Changes in intracellular Ca(2+) are central to the function of smooth muscle, which lines the walls of all hollow organs. These changes take a variety of forms, from sustained, cell-wide increases to temporally varying, localized changes. The nature of the Ca(2+) signal is a reflection of the source of Ca(2+) (extracellular or intracellular) and the molecular entity responsible for generating it. Depending on the specific channel involved and the detection technology employed, extracellular Ca(2+) entry may be detected optically as graded elevations in intracellular Ca(2+), junctional Ca(2+) transients, Ca(2+) flashes, or Ca(2+) sparklets, whereas release of Ca(2+) from intracellular stores may manifest as Ca(2+) sparks, Ca(2+) puffs, or Ca(2+) waves. These diverse Ca(2+) signals collectively regulate a variety of functions. Some functions, such as contractility, are unique to smooth muscle; others are common to other excitable cells (e.g., modulation of membrane potential) and nonexcitable cells (e.g., regulation of gene expression).

  18. Anti-smooth muscle antibody

    Science.gov (United States)

    ... gov/ency/article/003531.htm Anti-smooth muscle antibody To use the sharing features on this page, please enable JavaScript. Anti-smooth muscle antibody is a blood test that detects the presence ...

  19. Overexpression of functional TrkA receptors after internalisation in human airway smooth muscle cells.

    Science.gov (United States)

    Freund-Michel, Véronique; Frossard, Nelly

    2008-10-01

    Trafficking of the TrkA receptor after stimulation by NGF is of emerging importance in structural cells in the context of airway inflammatory diseases. We have recently reported the expression of functional TrkA receptors in human airway smooth muscle cells (HASMC). We have here studied the TrkA trafficking mechanisms in these cells. TrkA disappearance from the cell membrane was induced within 5 min of NGF (3pM) stimulation. Co-immunoprecipitation of clathrin-TrkA was revealed, and TrkA internalisation inhibited either by clathrin inhibitors or by siRNA inducing downregulation of endogenous clathrin. TrkA internalised receptors were totally degraded in lysosomes, with no recycling phenomenon. Newly synthesized TrkA receptors were thereafter re-expressed at the cell membrane within 10 h. TrkA re-synthesis was inhibited by blockade of clathrin-dependent internalisation, but not of TrkA receptors lysosomal degradation. Finally, we observed that NGF multiple stimulations progressively increased TrkA expression in HASMC, which was associated with an increase in NGF/TrkA-dependent proliferation. In conclusion, we show here the occurrence of clathrin-dependent TrkA internalisation and lysosomal degradation in the airway smooth muscle, followed by upregulated re-synthesis of functional TrkA receptors and increased proliferative effect in the human airway smooth muscle. This may have pathophysiological consequences in airway inflammatory diseases.

  20. Immunohistochemical characterization of endometriosis-associated smooth muscle cells in human peritoneal endometriotic lesions.

    Science.gov (United States)

    Barcena de Arellano, Maria L; Gericke, Jessica; Reichelt, Uta; Okuducu, Ali Fuat; Ebert, Andreas D; Chiantera, Vito; Schneider, Achim; Mechsner, Sylvia

    2011-10-01

    Smooth muscle cells (SMC) are common components of endometriotic lesions. SMC have been characterized previously in peritoneal, ovarian and deep infiltrating endometriotic lesions and adenomyosis. The aim of this retrospective study was to investigate the extent of differentiation in endometriosis-associated SMC (EMaSMC) in peritoneal endometriotic lesions. We obtained biopsies from peritoneal endometriotic lesions (n = 60) and peritoneal sites distant from the endometriotic lesion (n = 60), as well as healthy peritoneum from patients without endometriosis (control tissue, n = 10). These controls were hysterectomy specimens from patients without endometriosis or adenomyosis. Histopathological examination of peritoneal specimens using antibodies against oxytocin receptor (OTR), vasopressin receptor (VPR), smooth muscle myosin heavy chain (SM-MHC), estrogen receptor (ER) or progesterone receptor (PR) was performed. To identify SMC and their level of differentiation, antibodies for smooth muscle actin desmin and caldesmon were used. SMC were detected in all endometriotic lesions. SMC were more abundant in unaffected peritoneum of women with endometriosis (38%) compared with women without endometriosis (6%; P endometriosis.

  1. HSP20 phosphorylation and airway smooth muscle relaxation

    Directory of Open Access Journals (Sweden)

    Mariam Ba

    2009-06-01

    Full Text Available Mariam Ba1, Cherie A Singer1, Manoj Tyagi2, Colleen Brophy3, Josh E Baker4, Christine Cremo4, Andrew Halayko5, William T Gerthoffer21Department of Pharmacology, University of Nevada School of Medicine, Reno, NV, USA; 2Department of Biochemistry and Molecular Biology, University of South Alabama, Mobile, AL, USA; 3Harrington Department of Biochemistry, Arizona State University, Tempe, AZ, USA; 4Department of Biochemistry and Molecular Biology, University of Nevada, Reno, NV, USA; 5Departments of Physiology and Internal Medicine, University of Manitoba, Winnipeg, MB, CanadaAbstract: HSP20 (HSPB6 is a small heat shock protein expressed in smooth muscles that is hypothesized to inhibit contraction when phosphorylated by cAMP-dependent protein kinase. To investigate this hypothesis in airway smooth muscle (ASM we showed that HSP20 was constitutively expressed as well as being inducible in cultured hASM cells by treatment with 1 µM isoproterenol or 10 µM salmeterol. In contrast, a mixture of proinflammatory mediators (interleukin-1β, tumor necrosis factor α, and interferon γ inhibited expression of HSP20 by about 50% in 48 hours. To determine whether phosphorylation of HSP20 is sufficient to induce relaxation, canine tracheal smooth muscle was treated with a cell permeant phosphopeptide that mimics the phosphorylation of HSP20. The HSP20 phosphopeptide antagonized carbacholinduced contraction by 60% with no change in myosin light chain phosphorylation. Recombinant full length HSP20 inhibited skeletal actin binding to smooth muscle myosin subfragment 1 (S1, and recombinant cell permeant TAT-HSP20 S16D mutant reduced F-actin filaments in cultured hASM cells. Carbachol stimulation of canine tracheal smooth muscle tissue caused redistribution of HSP20 from large macromolecular complexes (200–500 kDa to smaller complexes (<60 kDa. The results are consistent with HSP20 expression and macromolecular structure being dynamically regulated in airway

  2. Bone morphogenetic protein-induced MSX1 and MSX2 inhibit myocardin-dependent smooth muscle gene transcription.

    Science.gov (United States)

    Hayashi, Ken'ichiro; Nakamura, Seiji; Nishida, Wataru; Sobue, Kenji

    2006-12-01

    During the onset and progression of atherosclerosis, the vascular smooth muscle cell (VSMC) phenotype changes from differentiated to dedifferentiated, and in some cases, this change is accompanied by osteogenic transition, resulting in vascular calcification. One characteristic of dedifferentiated VSMCs is the down-regulation of smooth muscle cell (SMC) marker gene expression. Bone morphogenetic proteins (BMPs), which are involved in the induction of osteogenic gene expression, are detected in calcified vasculature. In this study, we found that the BMP2-, BMP4-, and BMP6-induced expression of Msx transcription factors (Msx1 and Msx2) preceded the down-regulation of SMC marker expression in cultured differentiated VSMCs. Either Msx1 or Msx2 markedly reduced the myocardin-dependent promoter activities of SMC marker genes (SM22alpha and caldesmon). We further investigated interactions between Msx1 and myocardin/serum response factor (SRF)/CArG-box motif (cis element for SRF) using coimmunoprecipitation, gel-shift, and chromatin immunoprecipitation assays. Our results showed that Msx1 or Msx2 formed a ternary complex with SRF and myocardin and inhibited the binding of SRF or SRF/myocardin to the CArG-box motif, resulting in inhibition of their transcription.

  3. Bone Morphogenetic Protein-Induced Msx1 and Msx2 Inhibit Myocardin-Dependent Smooth Muscle Gene Transcription▿

    Science.gov (United States)

    Hayashi, Ken'ichiro; Nakamura, Seiji; Nishida, Wataru; Sobue, Kenji

    2006-01-01

    During the onset and progression of atherosclerosis, the vascular smooth muscle cell (VSMC) phenotype changes from differentiated to dedifferentiated, and in some cases, this change is accompanied by osteogenic transition, resulting in vascular calcification. One characteristic of dedifferentiated VSMCs is the down-regulation of smooth muscle cell (SMC) marker gene expression. Bone morphogenetic proteins (BMPs), which are involved in the induction of osteogenic gene expression, are detected in calcified vasculature. In this study, we found that the BMP2-, BMP4-, and BMP6-induced expression of Msx transcription factors (Msx1 and Msx2) preceded the down-regulation of SMC marker expression in cultured differentiated VSMCs. Either Msx1 or Msx2 markedly reduced the myocardin-dependent promoter activities of SMC marker genes (SM22α and caldesmon). We further investigated interactions between Msx1 and myocardin/serum response factor (SRF)/CArG-box motif (cis element for SRF) using coimmunoprecipitation, gel-shift, and chromatin immunoprecipitation assays. Our results showed that Msx1 or Msx2 formed a ternary complex with SRF and myocardin and inhibited the binding of SRF or SRF/myocardin to the CArG-box motif, resulting in inhibition of their transcription. PMID:17030628

  4. Shear Stress Induces Phenotypic Modulation of Vascular Smooth Muscle Cells via AMPK/mTOR/ULK1-Mediated Autophagy.

    Science.gov (United States)

    Sun, Liqian; Zhao, Manman; Liu, Aihua; Lv, Ming; Zhang, Jingbo; Li, Youxiang; Yang, Xinjian; Wu, Zhongxue

    2018-03-01

    Phenotypic modulation of vascular smooth muscle cells (VSMCs) is involved in the pathophysiological processes of the intracranial aneurysms (IAs). Although shear stress has been implicated in the proliferation, migration, and phenotypic conversion of VSMCs, the molecular mechanisms underlying these events are currently unknown. In this study, we investigated whether shear stress(SS)-induced VSMC phenotypic modulation was mediated by autophagy involved in adenosine monophosphate-activated protein kinase (AMPK)/mammalian target of rapamycin (mTOR)/Unc-51-like kinase 1 (ULK1) pathway. The results show that shear stress could inhibit the expression of key VSMC contractile genes and induce pro-inflammatory/matrix-remodeling genes levels, contributing to VSMCs phenotypic switching from a contractile to a synthetic phenotype. More importantly, Shear stress also markedly increased the levels of the autophagy marker microtubule-associated protein light chain 3-II (LC3II), Beclin-1, and p62 degradation. The autophagy inhibitor 3-methyladenine (3-MA) significantly blocked shear-induced phenotypic modulation of VSMCs. To further explore the molecular mechanism involved in shear-induced autophagy, we found that shear stress could activate AMPK/mTOR/ULK1 signaling pathway in VSMCs. Compound C, a pharmacological inhibitor of AMPK, significantly reduced the levels of p-AMPK and p-ULK, enhanced p-mTOR level, and finally decreased LC3II and Beclin-1 level, which suggested that activated AMPK/mTOR/ULK1 signaling was related to shear-mediated autophagy. These results indicate that shear stress promotes VSMC phenotypic modulation through the induction of autophagy involved in activating the AMPK/mTOR/ULK1 pathway.

  5. Vasopressin activates Akt/mTOR pathway in smooth muscle cells cultured in high glucose concentration

    Energy Technology Data Exchange (ETDEWEB)

    Montes, Daniela K.; Brenet, Marianne; Muñoz, Vanessa C.; Burgos, Patricia V.; Villanueva, Carolina I. [Department of Physiology, Universidad Austral de Chile, Valdivia 509-9200 (Chile); Figueroa, Carlos D. [Department of Anatomy, Histology and Pathology, Universidad Austral de Chile, Valdivia 509-9200 (Chile); González, Carlos B., E-mail: cbgonzal@uach.cl [Department of Physiology, Universidad Austral de Chile, Valdivia 509-9200 (Chile); Department of Neuroscience and Cell Biology, University of Texas Medical Branch, Galveston, TX 77555 (United States)

    2013-11-29

    Highlights: •AVP induces mTOR phosphorylation in A-10 cells cultured in high glucose concentration. •The mTOR phosphorylation is mediated by the PI3K/Akt pathway activation. •The AVP-induced mTOR phosphorylation inhibited autophagy and stimulated cell proliferation. -- Abstract: Mammalian target of rapamycin (mTOR) complex is a key regulator of autophagy, cell growth and proliferation. Here, we studied the effects of arginine vasopressin (AVP) on mTOR activation in vascular smooth muscle cells cultured in high glucose concentration. AVP induced the mTOR phosphorylation in A-10 cells grown in high glucose, in contrast to cells cultured in normal glucose; wherein, only basal phosphorylation was observed. The AVP-induced mTOR phosphorylation was inhibited by a PI3K inhibitor. Moreover, the AVP-induced mTOR activation inhibited autophagy and increased thymidine incorporation in cells grown in high glucose. This increase was abolished by rapamycin which inhibits the mTORC1 complex formation. Our results suggest that AVP stimulates mTOR phosphorylation by activating the PI3K/Akt signaling pathway and, subsequently, inhibits autophagy and raises cell proliferation in A-10 cells maintained in high glucose concentration.

  6. Cyclic Mechanical Stretch Up-regulates Hepatoma-Derived Growth Factor Expression in Cultured Rat Aortic Smooth Muscle Cells.

    Science.gov (United States)

    Kao, Ying-Hsien; Chen, Po-Han; Sun, Cheuk-Kwan; Chang, Yo-Chen; Lin, Yu-Chun; Tsai, Ming-Shian; Lee, Po-Huang; Cheng, Cheng-I

    2018-02-21

    Hepatoma-derived growth factor (HDGF) is a potent mitogen for vascular smooth muscle cells (SMCs) during embryogenesis and injury repair of vessel walls. Whether mechanical stimuli modulate HDGF expression remains unknown. This study aimed at investigating whether cyclic mechanical stretch plays a regulatory role in HDGF expression and regenerative cytokine production in aortic SMCs. A SMC cell line was grown on a silicone-based elastomer chamber with extracellular matrix coatings (either type I collagen or fibronectin) and received cyclic and uni-axial mechanical stretches with 10% deformation at frequency 1 Hz. Morphological observation showed that fibronectin coating provided better cell adhesion and spreading and that consecutive 6 hours of cyclic mechanical stretch remarkably induced reorientation and realignment of SMCs. Western blotting detection demonstrated that continuous mechanical stimuli elicited up-regulation of HDGF and PCNA, a cell proliferative marker. Signal kinetic profiling study indicated that cyclic mechanical stretch induced signaling activity in RhoA/ROCK and PI3K/Akt cascades. Kinase inhibition study further showed that blockade of PI3K activity suppressed the stretch-induced TNF-a, whereas RhoA/ROCK inhibition significantly blunted the IL-6 production and HDGF over-expression. Moreover, siRNA-mediated HDGF gene silencing significantly suppressed constitutive expression of IL-6, but not TNF-α, in SMCs. These findings support the role of HDGF in maintaining vascular expression of IL-6, which has been regarded a crucial regenerative factor for acute vascular injury. In conclusion, cyclic mechanical stretch may maintain constitutive expression of HDGF in vascular walls and be regarded an important biophysical regulator in vascular regeneration. ©2018 The Author(s).

  7. Shh mediates PDGF-induced contractile-to-synthetic phenotypic modulation in vascular smooth muscle cells through regulation of KLF4

    Energy Technology Data Exchange (ETDEWEB)

    Zeng, Qiu [Department of Vascular Surgery, 1st Affiliated Hospital of Chongqing Medical University, Chongqing 400016 (China); Wei, Bin [Department of Dermatology, 1st Affiliated Hospital of Chongqing Medical University, Chongqing 400016 (China); Zhao, Yu; Wang, Xuehu; Fu, Qining; Liu, Hong [Department of Vascular Surgery, 1st Affiliated Hospital of Chongqing Medical University, Chongqing 400016 (China); Li, Fenghe, E-mail: lfh_cmu@126.com [Department of Vascular Surgery, 1st Affiliated Hospital of Chongqing Medical University, Chongqing 400016 (China)

    2016-07-01

    Platelet-derived growth factor (PDGF) is known to induce phenotypic switching of vascular smooth muscle cells (VSMCs) from contractile to a pathological synthetic state, which played an essential role in proliferation of VSMCs. Sonic hedgehog (Shh) contributes to the proliferation of VSMCs when induced by PDGF. Here, we investigated the probable role of Shh in PDGF-induced VSMC dedifferentiation and its underlying mechanisms. We found that PDGF stimulated Shh expression in VSMCs, which was mediated by activation of PDGFRβ/ERK1/2 cell signaling pathway. Further, we found PDGF-induced VSMC phenotypic modulation was accompanied by up-regulation of Shh/Gli family zinc finger 2 (Gli2) signaling and Krüppel-like factor 4 (KLF4). When inhibited Shh in the presence of PDGF, the expressions of KLF4 and VSMC dedifferentiation markers were down-regulated and the effect of PDGF in inducing VSMC dedifferentiation was blocked. In the absence of PDGF, Shh signaling activation increased the expression of KLF4 and promoted VSMC dedifferentiation. The results indicate Shh participated in the regulation of PDGF-induced VSMC dedifferentiation. Finally, we found that KLF4 was closely involved in this process. On inhibition of KLF4, PDGF induced VSMC dedifferentiation was abrogated, even in the presence of Shh. Taken together, the results provide critical insights into the newly discovered role of Shh in phenotypic modulation of VSMCs which depends on KLF4. - Highlights: • Shh as a downstream effector of PDGF participates in PDGF-induced VSMC phenotypic modulation. • Shh can promote VSMC phenotypic switching from contractile to synthetic state. • Shh mediates VSMC phenotypic modulation through regulation of KLF4.

  8. Shh mediates PDGF-induced contractile-to-synthetic phenotypic modulation in vascular smooth muscle cells through regulation of KLF4

    International Nuclear Information System (INIS)

    Zeng, Qiu; Wei, Bin; Zhao, Yu; Wang, Xuehu; Fu, Qining; Liu, Hong; Li, Fenghe

    2016-01-01

    Platelet-derived growth factor (PDGF) is known to induce phenotypic switching of vascular smooth muscle cells (VSMCs) from contractile to a pathological synthetic state, which played an essential role in proliferation of VSMCs. Sonic hedgehog (Shh) contributes to the proliferation of VSMCs when induced by PDGF. Here, we investigated the probable role of Shh in PDGF-induced VSMC dedifferentiation and its underlying mechanisms. We found that PDGF stimulated Shh expression in VSMCs, which was mediated by activation of PDGFRβ/ERK1/2 cell signaling pathway. Further, we found PDGF-induced VSMC phenotypic modulation was accompanied by up-regulation of Shh/Gli family zinc finger 2 (Gli2) signaling and Krüppel-like factor 4 (KLF4). When inhibited Shh in the presence of PDGF, the expressions of KLF4 and VSMC dedifferentiation markers were down-regulated and the effect of PDGF in inducing VSMC dedifferentiation was blocked. In the absence of PDGF, Shh signaling activation increased the expression of KLF4 and promoted VSMC dedifferentiation. The results indicate Shh participated in the regulation of PDGF-induced VSMC dedifferentiation. Finally, we found that KLF4 was closely involved in this process. On inhibition of KLF4, PDGF induced VSMC dedifferentiation was abrogated, even in the presence of Shh. Taken together, the results provide critical insights into the newly discovered role of Shh in phenotypic modulation of VSMCs which depends on KLF4. - Highlights: • Shh as a downstream effector of PDGF participates in PDGF-induced VSMC phenotypic modulation. • Shh can promote VSMC phenotypic switching from contractile to synthetic state. • Shh mediates VSMC phenotypic modulation through regulation of KLF4.

  9. Phenotype commitment in vascular smooth muscle cells derived from coronary atherosclerotic plaques: differential gene expression of endothelial Nitric Oxide Synthase

    Directory of Open Access Journals (Sweden)

    ML Rossi

    2009-06-01

    Full Text Available Unstable angina and myocardial infarction are the clinical manifestations of the abrupt thrombotic occlusion of an epicardial coronary artery as a result of spontaneous atherosclerotic plaque rupture or fissuring, and the exposure of highly thrombogenic material to blood. It has been demonstrated that the proliferation of vascular smooth muscle cells (VSMCs and impaired bioavailabilty of nitric oxide (NO are among the most important mechanisms involved in the progression of atherosclerosis. It has also been suggested that a NO imbalance in coronary arteries may be involved in myocardial ischemia as a result of vasomotor dysfunction triggering plaque rupture and the thrombotic response. We used 5’ nuclease assays (TaqMan™ PCRs to study gene expression in coronary plaques collected by means of therapeutic directional coronary atherectomy from 15 patients with stable angina (SA and 15 with acute coronary syndromes (ACS without ST elevation. Total RNA was extracted from the 30 plaques and the cDNA was amplified in order to determine endothelial nitric oxide synthase (eNOS gene expression. Analysis of the results showed that the expression of eNOS was significantly higher (p<0.001 in the plaques from the ACS patients. Furthermore, isolated VSMCs from ACS and SA plaques confirmed the above pattern even after 25 plating passages. In situ RT-PCR was also carried out to co-localize the eNOS messengers and the VSMC phenotype.

  10. Transcriptional up-regulation of antioxidant genes by PPARδ inhibits angiotensin II-induced premature senescence in vascular smooth muscle cells

    International Nuclear Information System (INIS)

    Kim, Hyo Jung; Ham, Sun Ah; Paek, Kyung Shin; Hwang, Jung Seok; Jung, Si Young; Kim, Min Young; Jin, Hanna; Kang, Eun Sil; Woo, Im Sun; Kim, Hye Jung; Lee, Jae Heun; Chang, Ki Churl; Han, Chang Woo; Seo, Han Geuk

    2011-01-01

    Research highlights: → Activation of PPARδ by GW501516 significantly inhibited Ang II-induced premature senescence in hVSMCs. → Agonist-activated PPARδ suppressed generation of Ang II-triggered ROS with a concomitant reduction in DNA damage. → GW501516 up-regulated expression of antioxidant genes, such as GPx1, Trx1, Mn-SOD and HO-1. → Knock-down of these antioxidant genes abolished the effects of GW501516 on ROS production and premature senescence. -- Abstract: This study evaluated peroxisome proliferator-activated receptor (PPAR) δ as a potential target for therapeutic intervention in Ang II-induced senescence in human vascular smooth muscle cells (hVSMCs). Activation of PPARδ by GW501516, a specific agonist of PPARδ, significantly inhibited the Ang II-induced premature senescence of hVSMCs. Agonist-activated PPARδ suppressed the generation of Ang II-triggered reactive oxygen species (ROS) with a concomitant reduction in DNA damage. Notably, GW501516 up-regulated the expression of antioxidant genes, such as glutathione peroxidase 1, thioredoxin 1, manganese superoxide dismutase and heme oxygenase 1. siRNA-mediated down-regulation of these antioxidant genes almost completely abolished the effects of GW501516 on ROS production and premature senescence in hVSMCs treated with Ang II. Taken together, the enhanced transcription of antioxidant genes is responsible for the PPARδ-mediated inhibition of premature senescence through sequestration of ROS in hVSMCs treated with Ang II.

  11. Vitisin B, a resveratrol tetramer, inhibits migration through inhibition of PDGF signaling and enhancement of cell adhesiveness in cultured vascular smooth muscle cells

    International Nuclear Information System (INIS)

    Ong, Eng-Thaim; Hwang, Tsong-Long; Huang, Yu-Ling; Lin, Chwan-Fwu; Wu, Wen-Bin

    2011-01-01

    Vascular smooth muscle cells (VSMCs) play an important role in normal vessel formation and in the development and progression of cardiovascular diseases. Grape plants contain resveratrol monomer and oligomers and drinking of wine made from grape has been linked to 'French Paradox'. In this study we evaluated the effect of vitisin B, a resveratrol tetramer, on VSMC behaviors. Vitisin B inhibited basal and PDGF-induced VSMC migration. Strikingly, it did not inhibit VSMC proliferation but inversely enhanced cell cycle progression and proliferation. Among the tested resveratrol oligomers, vitisin B showed an excellent inhibitory activity and selectivity on PDGF signaling. The anti-migratory effect by vitisin B was due to direct inhibition on PDGF signaling but was independent of interference with PDGF binding to VSMCs. Moreover, the enhanced VSMC adhesiveness to matrix contributed to the anti-migratory effect by vitisin B. Fluorescence microscopy revealed an enhanced reorganization of actin cytoskeleton and redistribution of activated focal adhesion proteins from cytosol to the peripheral edge of the cell membrane. This was confirmed by the observation that enhanced adhesiveness was repressed by the Src inhibitor. Finally, among the effects elicited by vitisin B, only the inhibitory effect toward basal migration was partially through estrogen receptor activation. We have demonstrated here that a resveratrol tetramer exhibited dual but opposite actions on VSMCs, one is to inhibit VSMC migration and the other is to promote VSMC proliferation. The anti-migratory effect was through a potent inhibition on PDGF signaling and novel enhancement on cell adhesion. - Highlights: → Several resveratrol oligomers from grape plants are examined on VSMC behaviors. → Tetraoligomer vitisin B shows excellent inhibitory activity and selectivity. → It exerts dual but opposing actions: anti-migratory and pro-proliferative effects. → The anti-migratory effect results from anti

  12. Roxithromycin inhibits VEGF-induced human airway smooth muscle cell proliferation: Opportunities for the treatment of asthma

    International Nuclear Information System (INIS)

    Pei, Qing-Mei; Jiang, Ping; Yang, Min; Qian, Xue-Jiao; Liu, Jiang-Bo; Kim, Sung-Ho

    2016-01-01

    Asthma is a chronic respiratory disease characterized by reversible airway obstruction with persistent airway inflammation and airway remodelling, which is associated with increased airway smooth muscle (ASM) mass. Roxithromycin (RXM) has been widely used in asthma treatment; however, its mechanism of action is poorly understood. Vascular endothelial growth factor (VEGF) has been implicated in inflammatory and airway blood vessel remodelling in patients with asthma, and shown to promote ASM cell proliferation. Here, we investigated the effect of RXM on VEGF-induced ASM cell proliferation and attempted to elucidate the underlying mechanisms of action. We tested the effect of RXM on proliferation and cell cycle progression, as well as on the expression of phospho-VEGF receptor 2 (VEGFR2), phospho-extracellular signal-regulated kinase 1/2 (ERK1/2), phospho-Akt, and caveolin-1 in VEGF-stimulated ASM cells. RXM inhibited VEGF-induced ASM cell proliferation and induced cell cycle arrest. Additionally, VEGF-induced ASM cell proliferation was suppressed by inhibiting the activity of ERK1/2, but not that of Akt. Furthermore, RXM treatment inhibits VEGF-induced activation of VEGFR2 and ERK and downregulation of caveolin-1 in a dose-dependent manner. RXM also inhibited TGF-β-induced VEGF secretion by ASM cells and BEAS-2B cells. Collectively, our findings suggest that RXM inhibits VEGF-induced ASM cell proliferation by suppression of VEGFR2 and ERK1/2 activation and caveolin-1 down-regulation, which may be involved in airway remodelling. Further elucidation of the mechanisms underlying these observations should enable the development of treatments for smooth muscle hyperplasia-associated diseases of the airway such as asthma. - Highlights: • RXM inhibited VEGF-induced ASM cell proliferation and induced cell cycle arrest. • VEGF-induced cell proliferation was suppressed by inhibiting the activity of ERK1/2. • RXM inhibits activation of VEGFR2 and ERK and downregulation

  13. Roxithromycin inhibits VEGF-induced human airway smooth muscle cell proliferation: Opportunities for the treatment of asthma

    Energy Technology Data Exchange (ETDEWEB)

    Pei, Qing-Mei, E-mail: 34713316@qq.com [Department of Radiology, Tianjin Hospital of Integrated Traditional Chinese and Western Medicine, Tianjin (China); Jiang, Ping, E-mail: jiangping@163.com [Department of Respiration, Tianjin First Central Hospital, Tianjin (China); Yang, Min, E-mail: YangMin@163.com [Department of Respiration, Tianjin First Central Hospital, Tianjin (China); Qian, Xue-Jiao, E-mail: qianxuejiao@163.com [Department of Respiration, Tianjin First Central Hospital, Tianjin (China); Liu, Jiang-Bo, E-mail: LJB1984@163.com [Department of Respiration, Tianjin First Central Hospital, Tianjin (China); Kim, Sung-Ho, E-mail: chenghao0726@hotmail.com [Department of Respiration, Tianjin First Central Hospital, Tianjin (China)

    2016-10-01

    Asthma is a chronic respiratory disease characterized by reversible airway obstruction with persistent airway inflammation and airway remodelling, which is associated with increased airway smooth muscle (ASM) mass. Roxithromycin (RXM) has been widely used in asthma treatment; however, its mechanism of action is poorly understood. Vascular endothelial growth factor (VEGF) has been implicated in inflammatory and airway blood vessel remodelling in patients with asthma, and shown to promote ASM cell proliferation. Here, we investigated the effect of RXM on VEGF-induced ASM cell proliferation and attempted to elucidate the underlying mechanisms of action. We tested the effect of RXM on proliferation and cell cycle progression, as well as on the expression of phospho-VEGF receptor 2 (VEGFR2), phospho-extracellular signal-regulated kinase 1/2 (ERK1/2), phospho-Akt, and caveolin-1 in VEGF-stimulated ASM cells. RXM inhibited VEGF-induced ASM cell proliferation and induced cell cycle arrest. Additionally, VEGF-induced ASM cell proliferation was suppressed by inhibiting the activity of ERK1/2, but not that of Akt. Furthermore, RXM treatment inhibits VEGF-induced activation of VEGFR2 and ERK and downregulation of caveolin-1 in a dose-dependent manner. RXM also inhibited TGF-β-induced VEGF secretion by ASM cells and BEAS-2B cells. Collectively, our findings suggest that RXM inhibits VEGF-induced ASM cell proliferation by suppression of VEGFR2 and ERK1/2 activation and caveolin-1 down-regulation, which may be involved in airway remodelling. Further elucidation of the mechanisms underlying these observations should enable the development of treatments for smooth muscle hyperplasia-associated diseases of the airway such as asthma. - Highlights: • RXM inhibited VEGF-induced ASM cell proliferation and induced cell cycle arrest. • VEGF-induced cell proliferation was suppressed by inhibiting the activity of ERK1/2. • RXM inhibits activation of VEGFR2 and ERK and downregulation

  14. Natriuretic peptide receptor-C activation attenuates angiotensin II-induced enhanced oxidative stress and hyperproliferation of aortic vascular smooth muscle cells.

    Science.gov (United States)

    Madiraju, Padma; Hossain, Ekhtear; Anand-Srivastava, Madhu B

    2018-02-07

    We showed previously that natriuretic peptide receptor-C (NPR-C) agonist, C-ANP 4-23 , attenuated the enhanced expression of Giα proteins in vascular smooth muscle cells (VSMC) from spontaneously hypertensive rats (SHR) through the inhibition of enhanced oxidative stress. Since the enhanced levels of endogenous angiotensin II (Ang II) contribute to the overexpression of Giα proteins and augmented oxidative stress in VSMC from SHR, the present study was undertaken to investigate if C-ANP 4-23 could also attenuate angiotensin II (Ang II)-induced oxidative stress and associated signaling. Ang II treatment of aortic VSMC augmented the levels of superoxide anion (O 2 - ), NADPH oxidase activity, and the expression of NADPH oxidase subunits and C-ANP 4-23 treatment attenuated all these to control levels. In addition, Ang II-induced enhanced levels of thiobarbituric acid-reactive substances (TBARS) and protein carbonyl content were also attenuated toward control levels by C-ANP 4-23 treatment. On the other hand, Ang II inhibited the levels of nitric oxide (NO) and augmented the levels of peroxynitrite (OONO - ) in VSMC which were restored to control levels by C-ANP 4-23 treatment. Furthermore, C-ANP 4-23 treatment attenuated Ang II-induced enhanced expression of Giα proteins, phosphorylation of p38, JNK, and ERK 1,2 as well as hyperproliferation of VSMC as determined by DNA synthesis, and metabolic activity. These results indicate that C-ANP 4-23 , via the activation of NPR-C, attenuates Ang II-induced enhanced nitroxidative stress, overexpression of Giα proteins, increased activation of the p38/JNK/ERK 1,2 signaling pathways, and hyperproliferation of VSMC. It may be suggested that C-ANP 4-23 could be used as a therapeutic agent in the treatment of vascular remodeling associated with hypertension and atherosclerosis.

  15. Vascular cell responses to ECM produced by smooth muscle cells on TiO{sub 2} nanotubes

    Energy Technology Data Exchange (ETDEWEB)

    Shen, Fangyu [School of Life Science and Engineering, Southwest Jiaotong University, Chengdu (China); Key Lab of Advanced Technology of Materials of Education Ministry, Southwest Jiaotong University, Chengdu 610031 (China); Zhu, Ying [School of Life Science and Engineering, Southwest Jiaotong University, Chengdu (China); Key Lab of Advanced Technology of Materials of Education Ministry, Southwest Jiaotong University, Chengdu 610031 (China); Wuhan Dragonbio Orthopedic Products CO., LTD, 18, Qinglnghe Road, Hongshan District, Wuhan 430065 (China); Li, Xin [School of Life Science and Engineering, Southwest Jiaotong University, Chengdu (China); Key Lab of Advanced Technology of Materials of Education Ministry, Southwest Jiaotong University, Chengdu 610031 (China); Luo, Rifang, E-mail: lrifang@126.com [School of Life Science and Engineering, Southwest Jiaotong University, Chengdu (China); Key Lab of Advanced Technology of Materials of Education Ministry, Southwest Jiaotong University, Chengdu 610031 (China); Tu, Qiufen [School of Life Science and Engineering, Southwest Jiaotong University, Chengdu (China); Key Lab of Advanced Technology of Materials of Education Ministry, Southwest Jiaotong University, Chengdu 610031 (China); Laboratory of Biosensing and Micro Mechatronics, Southwest Jiaotong University, Chengdu 610031 (China); Wang, Jin, E-mail: jinxxwang@263.net [School of Life Science and Engineering, Southwest Jiaotong University, Chengdu (China); Key Lab of Advanced Technology of Materials of Education Ministry, Southwest Jiaotong University, Chengdu 610031 (China); Huang, Nan [School of Life Science and Engineering, Southwest Jiaotong University, Chengdu (China); Key Lab of Advanced Technology of Materials of Education Ministry, Southwest Jiaotong University, Chengdu 610031 (China)

    2015-09-15

    Graphical abstract: - Highlights: • TiO{sub 2} nanotubes with the tube diameter of 30 nm via anodic oxidation was prepared. • SMCs on TiO{sub 2} nanotubes presented enhanced extracellular matrix secreting. • ECM prepared via decellularization retained the components: Fn, Ln and collagen. • ECM-covered TiO{sub 2} nanotubes significantly improved the proliferation of ECs. - Abstract: There is an increasing interest in developing new methods to promote biocompatibility of biomedical materials. The TiO{sub 2} nanotubes with the tube diameter of 30 nm were prepared by anodization. The response behavior of the human umbilical vein endothelial cell (HUVEC) and human umbilical artery smooth muscle cell (HUASMC) to these different nanotube sizes was investigated. Compared to the flat Ti, the growth and viability of HUVEC are prohibited, but there was no significant difference of HUASMC on 30 nm TiO{sub 2} nanotubes. In this study, extracellular matrix (ECM) as a complex cellular environment which provides structural support to cells and regulates the cells functions was further used to modify the biological properties of TiO{sub 2} nanotubes. The ECM secreted from HUASMC was successfully deposited onto the 30 nm TiO{sub 2} nanotubes. Moreover, immunofluorescence staining of common ECM components, such as fibronectin, laminin and type IV collagen, also indicated the successful ECM-covering on nanotube surfaces. Interestingly, the surface of ECM-covered TiO{sub 2} nanotubes significantly improved the proliferation of HUVECs in vitro. This suggested that the ECM secreted from HUASMCs on the TiO{sub 2} nanotubular surface could further improve the HUVECs adhesion and proliferation.

  16. Increased proinflammatory responses from asthmatic human airway smooth muscle cells in response to rhinovirus infection

    Directory of Open Access Journals (Sweden)

    King Nicholas JC

    2006-05-01

    Full Text Available Abstract Background Exacerbations of asthma are associated with viral respiratory tract infections, of which rhinoviruses (RV are the predominant virus type. Airway smooth muscle is important in asthma pathogenesis, however little is known about the potential interaction of RV and human airway smooth muscle cells (HASM. We hypothesised that rhinovirus induction of inflammatory cytokine release from airway smooth muscle is augmented and differentially regulated in asthmatic compared to normal HASM cells. Methods HASM cells, isolated from either asthmatic or non-asthmatic subjects, were infected with rhinovirus. Cytokine production was assayed by ELISA, ICAM-1 cell surface expression was assessed by FACS, and the transcription regulation of IL-6 was measured by luciferase activity. Results RV-induced IL-6 release was significantly greater in HASM cells derived from asthmatic subjects compared to non-asthmatic subjects. This response was RV specific, as 5% serum- induced IL-6 release was not different in the two cell types. Whilst serum stimulated IL-8 production in cells from both subject groups, RV induced IL-8 production in only asthmatic derived HASM cells. The transcriptional induction of IL-6 was differentially regulated via C/EBP in the asthmatic and NF-κB + AP-1 in the non-asthmatic HASM cells. Conclusion This study demonstrates augmentation and differential transcriptional regulation of RV specific innate immune response in HASM cells derived from asthmatic and non-asthmatics, and may give valuable insight into the mechanisms of RV-induced asthma exacerbations.

  17. Mir-22-3p Inhibits Arterial Smooth Muscle Cell Proliferation and Migration and Neointimal Hyperplasia by Targeting HMGB1 in Arteriosclerosis Obliterans

    Directory of Open Access Journals (Sweden)

    Shui-chuan Huang

    2017-08-01

    Full Text Available Background: Aberrant vascular smooth muscle cell (VSMC proliferation and migration contribute to the development of vascular pathologies, such as atherosclerosis and post-angioplasty restenosis. The aim of this study was to determine whether miR-22-3p plays a role in regulating human artery vascular smooth muscle cell (HASMC function and neointima formation. Methods: Quantitative real-time PCR (qRT-PCR and fluorescence in situ hybridization (FISH were used to detect miR-22-3p expression in human arteries. Cell Counting Kit-8 (CCK-8 and EdU assays were performed to assess cell proliferation, and transwell and wound closure assays were performed to assess cell migration. Moreover, luciferase reporter assays were performed to identify the target genes of miR-22-3p. Finally, a rat carotid artery balloon-injury model was used to determine the role of miR-22-3p in neointima formation. Results: MiR-22-3p expression was downregulated in arteriosclerosis obliterans (ASO arteries compared with normal arteries, as well as in platelet-derived growth factor-BB (PDGF-BB-stimulated HASMCs compared with control cells. MiR-22-3p overexpression had anti-proliferative and anti-migratory effects and dual-luciferase assay showed that high mobility group box-1 (HMGB1 is a direct target of miR-22-3p in HASMCs. Furthermore, miR-22-3p expression was negatively correlated with HMGB1 expression in ASO tissue specimens. Finally, LV-miR-22-3p-mediated miR-22-3p upregulation significantly suppressed neointimal hyperplasia specifically by reducing HMGB1 expression in vivo. Conclusions: Our results indicate that miR-22-3p is a key molecule in regulating HASMC proliferation and migration by targeting HMGB1 and that miR-22-3p and HMGB1 may be therapeutic targets in the treatment of human ASO.

  18. RGD peptide-displaying M13 bacteriophage/PLGA nanofibers as cell-adhesive matrices for smooth muscle cells

    Energy Technology Data Exchange (ETDEWEB)

    Shin, Yong Cheol; Lee, Jong Ho; Jin, Oh Seong; Lee, Eun Ji; Jin, Lin Hua; Kim, Chang Seok; Hong, Suck Won; Han, Dong Wook; Kim, Chun Tae; Oh, Jin Woo [Pusan National University, Busan (Korea, Republic of)

    2015-01-15

    Extracellular matrices (ECMs) are network structures that play an essential role in regulating cellular growth and differentiation. In this study, novel nanofibrous matrices were fabricated by electrospinning M13 bacteriophage and poly(lactic-co-glycolic acid) (PLGA) and were shown to be structurally and functionally similar to natural ECMs. A genetically-engineered M13 bacteriophage was constructed to display Arg-Gly-Asp (RGD) peptides on its surface. The physicochemical properties of RGD peptide-displaying M13 bacteriophage (RGD-M13 phage)/PLGA nanofibers were characterized by using scanning electron microscopy and Fourier-transform infrared spectroscopy. We used immunofluorescence staining to confirm that M13 bacteriophages were homogenously distributed in RGD-M13 phage/PLGA matrices. Furthermore, RGD-M13 phage/PLGA nanofibrous matrices, having excellent biocompatibility, can enhance the behaviors of vascular smooth muscle cells. This result suggests that RGD-M13 phage/PLGA nanofibrous matrices have potentials to serve as tissue engineering scaffolds.

  19. A multiscale active structural model of the arterial wall accounting for smooth muscle dynamics.

    Science.gov (United States)

    Coccarelli, Alberto; Edwards, David Hughes; Aggarwal, Ankush; Nithiarasu, Perumal; Parthimos, Dimitris

    2018-02-01

    Arterial wall dynamics arise from the synergy of passive mechano-elastic properties of the vascular tissue and the active contractile behaviour of smooth muscle cells (SMCs) that form the media layer of vessels. We have developed a computational framework that incorporates both these components to account for vascular responses to mechanical and pharmacological stimuli. To validate the proposed framework and demonstrate its potential for testing hypotheses on the pathogenesis of vascular disease, we have employed a number of pharmacological probes that modulate the arterial wall contractile machinery by selectively inhibiting a range of intracellular signalling pathways. Experimental probes used on ring segments from the rabbit central ear artery are: phenylephrine, a selective α 1-adrenergic receptor agonist that induces vasoconstriction; cyclopiazonic acid (CPA), a specific inhibitor of sarcoplasmic/endoplasmic reticulum Ca 2+ -ATPase; and ryanodine, a diterpenoid that modulates Ca 2+ release from the sarcoplasmic reticulum. These interventions were able to delineate the role of membrane versus intracellular signalling, previously identified as main factors in smooth muscle contraction and the generation of vessel tone. Each SMC was modelled by a system of nonlinear differential equations that account for intracellular ionic signalling, and in particular Ca 2+ dynamics. Cytosolic Ca 2+ concentrations formed the catalytic input to a cross-bridge kinetics model. Contractile output from these cellular components forms the input to the finite-element model of the arterial rings under isometric conditions that reproduces the experimental conditions. The model does not account for the role of the endothelium, as the nitric oxide production was suppressed by the action of L-NAME, and also due to the absence of shear stress on the arterial ring, as the experimental set-up did not involve flow. Simulations generated by the integrated model closely matched experimental

  20. MiR-137 inhibited cell proliferation and migration of vascular smooth muscle cells via targeting IGFBP-5 and modulating the mTOR/STAT3 signaling.

    Directory of Open Access Journals (Sweden)

    Jin Pan

    Full Text Available Abnormal proliferation of vascular smooth muscle cells (VSMCs contributes to the development of cardiovascular diseases. Studies have shown the great impact of microRNAs (miRNAs on the cell proliferation of VSMCs. This study examined the effects of miR-137 on the cell proliferation and migration of VSMCs and also explored the underlying molecular mechanisms. The mRNA and protein expression levels were determined by qRT-PCR and western blot assays, respectively. The CCK-8 assay, wound healing assay and transwell migration assay were performed to measure cell proliferation and migration of VSMCs. The miR-137-targeted 3'untranslated region of insulin-like growth factor-binding protein-5 (IGFBP-5 was confirmed by luciferase reporter assay. Platelet-derived growth factor-bb (PDGF-bb treatment enhanced cell proliferation and suppressed the expression of miR-137 in VSMCs. The gain-of-function and loss-of-function assays showed that overexpression of miR-137 suppressed the cell proliferation and migration, and also inhibited the expression of matrix genes of VSMCs; down-regulation of miR-137 had the opposite effects on VSMCs. Bioinformatics analysis and luciferase report assay results showed that IGFBP-5 was a direct target of miR-137, and miR-137 overexpression suppressed the IGFBP-5 expression and down-regulation of miR-137 increased the IGFBP-5 expression in VSMCs. PDGF-bb treatment also increased the IGFBP-5 mRNA expression. In addition, enforced expression of IGFBP-5 reversed the inhibitory effects of miR-137 on cell proliferation and migration of VSMCs. More importantly, overexpression of miR-137 also suppressed the activity of mTOR/STAT3 signaling in VSMCs. Taken together, the results suggest that miR-137 may suppress cell proliferation and migration of VSMCs via targeting IGFBP-5 and modulating mTOR/STAT3 signaling pathway.

  1. Extracellular matrix of smooth muscle cells: interaction of collagen type V with heparan sulfate proteoglycan

    International Nuclear Information System (INIS)

    Gay, S.; Hoeoek, M.; Gay, R.E.; Magargal, W.W.; Reynertson, R.H.

    1986-01-01

    Alteration in the extracellular matrix produced by smooth muscle cells may play a role in the development of atherosclerotic lesions. Consequently the authors have initiated studies on the structural organization of the extracellular matrix produced by cultured smooth muscle cells. Immunohisotological examination of this matrix using well-characterized mono- and polyclonal antibodies showed a partial codistribution of heparan sulfate (HS) proteoglycans with a number of different matrix components including collagen types I, III, IV, V and VI, laminin and fibronectin. Subsequent binding studies between isolated matrix proteins and HS showed that the polysaccharide interacts strongly with type V collagen and to a lesser extent with fibronectin as well as collagen types III and VI. The interaction between type V and HS was readily inhibited by heparin and highly sulfated HS but not be dermatan sulfate, chondroitin sulfate or HS with a low sulfate content. Furthermore, [ 35 S]-HS proteoglycans isolated from cultured smooth muscle cells could be adsorbed on a column of sepharose conjugated with native type V collagen and eluted in a salt gradient. Hence, the interaction between type V and HS may play a major part in stabilizing the extracellular matrix of the vessel wall

  2. Mechanical stretch modulates microRNA 21 expression, participating in proliferation and apoptosis in cultured human aortic smooth muscle cells.

    Directory of Open Access Journals (Sweden)

    Jian tao Song

    Full Text Available OBJECTIVES: Stretch affects vascular smooth muscle cell proliferation and apoptosis, and several responsible genes have been proposed. We tested whether the expression of microRNA 21 (miR-21 is modulated by stretch and is involved in stretch-induced proliferation and apoptosis of human aortic smooth muscle cells (HASMCs. METHODS AND RESULTS: RT-PCR revealed that elevated stretch (16% elongation, 1 Hz increased miR-21 expression in cultured HASMCs, and moderate stretch (10% elongation, 1 Hz decreased the expression. BrdU incorporation assay and cell counting showed miR-21 involved in the proliferation of HASMCs mediated by stretch, likely by regulating the expression of p27 and phosphorylated retinoblastoma protein (p-Rb. FACS analysis revealed that the complex of miR-21 and programmed cell death protein 4 (PDCD4 participated in regulating apoptosis with stretch. Stretch increased the expression of primary miR-21 and pre-miR-21 in HASMCs. Electrophoretic mobility shift assay (EMSA demonstrated that stretch increased NF-κB and AP-1 activities in HASMCs, and blockade of AP-1 activity by c-jun siRNA significantly suppressed stretch-induced miR-21 expression. CONCLUSIONS: Cyclic stretch modulates miR-21 expression in cultured HASMCs, and miR-21 plays important roles in regulating proliferation and apoptosis mediated by stretch. Stretch upregulates miR-21 expression at least in part at the transcription level and AP-1 is essential for stretch-induced miR-21 expression.

  3. Iduronic Acid in chondroitin/dermatan sulfate affects directional migration of aortic smooth muscle cells

    NARCIS (Netherlands)

    Bartolini, B.; Thelin, M.A.; Svensson, L.; Ghiselli, G.; Kuppevelt, T.H. van; Malmstrom, A.; Maccarana, M.

    2013-01-01

    Aortic smooth muscle cells produce chondroitin/dermatan sulfate (CS/DS) proteoglycans that regulate extracellular matrix organization and cell behavior in normal and pathological conditions. A unique feature of CS/DS proteoglycans is the presence of iduronic acid (IdoA), catalyzed by two DS

  4. Implication of molecular vascular smooth muscle cell heterogeneity among arterial beds in arterial calcification.

    Directory of Open Access Journals (Sweden)

    Olivier Espitia

    Full Text Available Vascular calcification is a strong and independent predictive factor for cardiovascular complications and mortality. Our previous work identified important discrepancies in plaque composition and calcification types between carotid and femoral arteries. The objective of this study is to further characterize and understand the heterogeneity in vascular calcification among vascular beds, and to identify molecular mechanisms underlying this process. We established ECLAGEN biocollection that encompasses human atherosclerotic lesions and healthy arteries from different locations (abdominal, thoracic aorta, carotid, femoral, and infrapopliteal arteries for histological, cell isolation, and transcriptomic analysis. Our results show that lesion composition differs between these locations. Femoral arteries are the most calcified arteries overall. They develop denser calcifications (sheet-like, nodule, and are highly susceptible to osteoid metaplasia. These discrepancies may derive from intrinsic differences between SMCs originating from these locations, as microarray analysis showed specific transcriptomic profiles between primary SMCs isolated from each arterial bed. These molecular differences translated into functional disparities. SMC from femoral arteries showed the highest propensity to mineralize due to an increase in basal TGFβ signaling. Our results suggest that biological heterogeneity of resident vascular cells between arterial beds, reflected by our transcriptomic analysis, is critical in understanding plaque biology and calcification, and may have strong implications in vascular therapeutic approaches.

  5. NADPH oxidase 1 deficiency alters caveolin phosphorylation and angiotensin II-receptor localization in vascular smooth muscle.

    Science.gov (United States)

    Basset, Olivier; Deffert, Christine; Foti, Michelangelo; Bedard, Karen; Jaquet, Vincent; Ogier-Denis, Eric; Krause, Karl-Heinz

    2009-10-01

    The superoxide-generating NADPH oxidase NOX1 is thought to be involved in signaling by the angiotensin II-receptor AT1R. However, underlying signaling steps are poorly understood. In this study, we investigated the effect of AngII on aortic smooth muscle from wild-type and NOX1-deficient mice. NOX1-deficient cells showed decreased basal ROS generation and did not produce ROS in response to AngII. Unexpectedly, AngII-dependent Ca(2+) signaling was markedly decreased in NOX1-deficient cells. Immunostaining demonstrated that AT1R was localized on the plasma membrane in wild-type, but intracellularly in NOX1-deficient cells. Immunohistochemistry and immunoblotting showed a decreased expression of AT1R in the aorta of NOX1-deficient mice. To investigate the basis of the abnormal AT1R targeting, we studied caveolin expression and phosphorylation. The amounts of total caveolin and of caveolae were not different in NOX1-deficient mice, but a marked decrease occurred in the phosphorylated form of caveolin. Exogenous H(2)O(2) or transfection of a NOX1 plasmid restored AngII responses in NOX1-deficient cells. Based on these findings, we propose that NOX1-derived reactive oxygen species regulate cell-surface expression of AT1R through mechanisms including caveolin phosphorylation. The lack cell-surface AT1R expression in smooth muscle could be involved in the decreased blood pressure in NOX1-deficient mice.

  6. Functional Modeling of the Shift in Cellular Calcium Dynamics at the Onset of Synchronization in Smooth Muscle Cells

    DEFF Research Database (Denmark)

    Postnov, D E; Brings Jacobsen, J C; von Holstein-Rathlou, Niels-Henrik

    2011-01-01

    In the present paper we address the nature of synchronization properties found in populations of mesenteric artery smooth muscle cells. We present a minimal model of the onset of synchronization in the individual smooth muscle cell that is manifested as a transition from calcium waves to whole......-cell calcium oscillations. We discuss how different types of ion currents may influence both amplitude and frequency in the regime of whole-cell oscillations. The model may also explain the occurrence of mixed-mode oscillations and chaotic oscillations frequently observed in the experimental system....

  7. Transforming growth factor β inhibits platelet derived growth factor-induced vascular smooth muscle cell proliferation via Akt-independent, Smad-mediated cyclin D1 downregulation.

    Directory of Open Access Journals (Sweden)

    Abel Martin-Garrido

    Full Text Available In adult tissue, vascular smooth muscle cells (VSMCs exist in a differentiated phenotype, which is defined by the expression of contractile proteins and lack of proliferation. After vascular injury, VSMC adopt a synthetic phenotype associated with proliferation, migration and matrix secretion. The transition between phenotypes is a consequence of the extracellular environment, and in particular, is regulated by agonists such as the pro-differentiating cytokine transforming growth factor β (TGFβ and the pro-proliferative cytokine platelet derived growth factor (PDGF. In this study, we investigated the interplay between TGFβ and PDGF with respect to their ability to regulate VSMC proliferation. Stimulation of human aortic VSMC with TGFβ completely blocked proliferation induced by all isoforms of PDGF, as measured by DNA synthesis and total cell number. Mechanistically, PDGF-induced Cyclin D1 mRNA and protein expression was inhibited by TGFβ. TGFβ had no effect on PDGF activation of its receptor and ERK1/2, but inhibited Akt activation. However, constitutively active Akt did not reverse the inhibitory effect of TGFβ on Cyclin D1 expression even though inhibition of the proteasome blocked the effect of TGFβ. siRNA against Smad4 completely reversed the inhibitory effect of TGFβ on PDGF-induced Cyclin D1 expression and restored proliferation in response to PDGF. Moreover, siRNA against KLF5 prevented Cyclin D1 upregulation by PDGF and overexpression of KLF5 partially reversed TGFβ-induced inhibition of Cyclin D1 expression. Taken together, our results demonstrate that KLF5 is required for PDGF-induced Cyclin D1 expression, which is inhibited by TGFβ via a Smad dependent mechanism, resulting in arrest of VSMCs in the G1 phase of the cell cycle.

  8. Transforming growth factor β inhibits platelet derived growth factor-induced vascular smooth muscle cell proliferation via Akt-independent, Smad-mediated cyclin D1 downregulation.

    Science.gov (United States)

    Martin-Garrido, Abel; Williams, Holly C; Lee, Minyoung; Seidel-Rogol, Bonnie; Ci, Xinpei; Dong, Jin-Tang; Lassègue, Bernard; Martín, Alejandra San; Griendling, Kathy K

    2013-01-01

    In adult tissue, vascular smooth muscle cells (VSMCs) exist in a differentiated phenotype, which is defined by the expression of contractile proteins and lack of proliferation. After vascular injury, VSMC adopt a synthetic phenotype associated with proliferation, migration and matrix secretion. The transition between phenotypes is a consequence of the extracellular environment, and in particular, is regulated by agonists such as the pro-differentiating cytokine transforming growth factor β (TGFβ) and the pro-proliferative cytokine platelet derived growth factor (PDGF). In this study, we investigated the interplay between TGFβ and PDGF with respect to their ability to regulate VSMC proliferation. Stimulation of human aortic VSMC with TGFβ completely blocked proliferation induced by all isoforms of PDGF, as measured by DNA synthesis and total cell number. Mechanistically, PDGF-induced Cyclin D1 mRNA and protein expression was inhibited by TGFβ. TGFβ had no effect on PDGF activation of its receptor and ERK1/2, but inhibited Akt activation. However, constitutively active Akt did not reverse the inhibitory effect of TGFβ on Cyclin D1 expression even though inhibition of the proteasome blocked the effect of TGFβ. siRNA against Smad4 completely reversed the inhibitory effect of TGFβ on PDGF-induced Cyclin D1 expression and restored proliferation in response to PDGF. Moreover, siRNA against KLF5 prevented Cyclin D1 upregulation by PDGF and overexpression of KLF5 partially reversed TGFβ-induced inhibition of Cyclin D1 expression. Taken together, our results demonstrate that KLF5 is required for PDGF-induced Cyclin D1 expression, which is inhibited by TGFβ via a Smad dependent mechanism, resulting in arrest of VSMCs in the G1 phase of the cell cycle.

  9. Sodium pump activity and calcium relaxation in vascular smooth muscle of deoxycorticosterone acetate-salt rats

    International Nuclear Information System (INIS)

    Soltis, E.E.; Field, F.P.

    1986-01-01

    The Na + -K + pump activity was determined in femoral arterial smooth muscle from deoxycorticosterone acetate (DOCA)-salt hypertensive rats using potassium relaxation and ouabain-sensitive 86 Rb uptake as indices. The membrane-stabilizing effect of calcium and its relation to Na + -K + pump activity also were examined. Femoral arteries from DOCA-salt rats exhibited a greater relaxation in response to potassium addition after contraction with norepinephrine in a low potassium (0.6 mM) Krebs solution. The concentration of potassium required to produce a 50% relaxation was significantly less in DOCA-salt rats. Ouabain-sensitive 86 Rb uptake was significantly greater at 3, 10, and 20 minutes of 86 Rb incubation in femoral arteries from DOCA-salt rats. Linear regression analysis revealed a significant correlation between the uptake of 86 Rb and time of incubation in both control and DOCA-salt rats. A significant difference in the slopes of the regression lines showed that the rate of uptake was greater in DOCA-salt rats. No difference was observed in ouabain-insensitive 86 Rb uptake. A dose-dependent relaxation in response to increasing concentrations of calcium following contraction to norepinephrine was observed in femoral arteries from control and DOCA-salt rats. The relaxation was directly dependent on the level of extracellular potassium and was blocked by ouabain. Femoral arteries from DOCA-salt rats relaxed to a significantly greater extent in response to calcium at each level of potassium when compared with controls. These results provide further evidence for an increase in Na + -K + pump activity in vascular smooth muscle from DOCA-salt hypertensive rats

  10. Smooth muscle cells of penis in the rat: noninvasive quantification with shear wave elastography.

    Science.gov (United States)

    Zhang, Jia-Jie; Qiao, Xiao-Hui; Gao, Feng; Bai, Ming; Li, Fan; Du, Lian-Fang; Xing, Jin-Fang

    2015-01-01

    Smooth muscle cells (SMCs) of cavernosum play an important role in erection. It is of great significance to quantitatively analyze the level of SMCs in penis. In this study, we investigated the feasibility of shear wave elastography (SWE) on evaluating the level of SMCs in penis quantitatively. Twenty healthy male rats were selected. The SWE imaging of penis was carried out and then immunohistochemistry analysis of penis was performed to analyze the expression of alpha smooth muscle actin in penis. The measurement index of SWE examination was tissue stiffness (TS). The measurement index of immunohistochemistry analysis was positive area percentage of alpha smooth muscle actin (AP). Sixty sets of data of TS and AP were obtained. The results showed that TS was significantly correlated with AP and the correlation coefficient was -0.618 (p penis was successfully quantified in vivo with SWE. SWE can be used clinically for evaluating the level of SMCs in penis quantitatively.

  11. Hematopoietic stem cell capture and directional differentiation into vascular endothelial cells for metal stent-coated chitosan/hyaluronic acid loading CD133 antibody.

    Science.gov (United States)

    Zhang, Shixuan; Zhang, Fan; Feng, Bo; Fan, Qingyu; Yang, Feng; Shang, Debin; Sui, Jinghan; Zhao, Hong

    2015-03-01

    A series of metal stents coated with chitosan/hyaluronic acid (CS/HA) loading antibodies by electrostatic self-assembled method were prepared, and the types of cells captured by antibodies and their differentiation in vascular endothelial cells (ECs) evaluated by molecular biology and scanning electron microscope. The results showed that CD133 stent can selectively capture hematopoietic stem cells (HSC),which directionally differentiate into vascular ECs in peripheral blood by (CS/HA) induction, and simultaneously inhibit migration and proliferation of immune cells and vascular smooth muscle cells (MCs). CD34 stent can capture HSC, hematopoietic progenitor cells that differentiate into vascular ECs and immune cells, promoting smooth MCs growth, leading to thrombosis, inflammation, and rejection. CD133 stent can be implanted into miniature pig heart coronary and can repair vascular damage by capturing own HSC, thus contributing to the rapid natural vascular repair, avoiding inflammation and rejection, thrombosis and restenosis. These studies demonstrated that CD133 stent of HSC capture will be an ideal coated metal stent providing a new therapeutic approach for cardiovascular and cerebrovascular disease.

  12. [Mg2+, ATP-dependent plasma membrane calcium pump of smooth muscle cells. I. Structural organization and properties].

    Science.gov (United States)

    Veklich, T O; Mazur, Iu Iu; Kosterin, S O

    2015-01-01

    Tight control of cytoplasm Ca2+ concentration is essential in cell functioning. Changing of Ca2+ concentration is thorough in smooth muscle cells, because it determines relaxation/constraint process. One of key proteins which control Ca2+ concentration in cytoplasm is Mg2+, ATP-dependent plasma membrane calcium pump. Thus, it is important to find compoumds which allowed one to change Mg2+, ATP-dependent plasma membrane calcium pump activity, as long as this topic is of current interest in biochemical research which regards energy and pharmacomechanical coupling mechanism of muscle excitation and contraction. In this article we generalized literatute and own data about properties of smooth muscle cell plasma membrane Ca(2+)-pump. Stuctural oganization, kinetical properties and molecular biology are considered.

  13. Glycogen synthase kinase 3β regulation of nuclear factor of activated T-cells isoform c1 in the vascular smooth muscle cell response to injury

    International Nuclear Information System (INIS)

    Chow Winsion; Hou Guangpei; Bendeck, Michelle P.

    2008-01-01

    The migration and proliferation of vascular smooth muscle cells (vSMCs) are critical events in neointima formation during atherosclerosis and restenosis. The transcription factor nuclear factor of activated T-cells-isoform c1 (NFATc1) is regulated by atherogenic cytokines, and has been implicated in the migratory and proliferative responses of vSMCs through the regulation of gene expression. In T-cells, calcineurin de-phosphorylates NFATc1, leading to its nuclear import, while glycogen synthase kinase 3 β (GSK3β) phosphorylates NFATc1 and promotes its nuclear export. However, the relationship between NFATc1 and GSK3β has not been studied during SMC migration and proliferation. We investigated this by scrape wounding vSMCs in vitro, and studying wound repair. NFATc1 protein was transiently increased, reaching a peak at 8 h after wounding. Cell fractionation and immunocytochemistry revealed that NFATc1 accumulation in the nucleus was maximal at 4 h after injury, and this was coincident with a significant 9 fold increase in transcriptional activity. Silencing NFATc1 expression with siRNA or inhibition of NFAT with cyclosporin A (CsA) attenuated wound closure by vSMCs. Phospho-GSK3β (inactive) increased to a peak at 30 min after injury, preceding the nuclear accumulation of NFATc1. Overexpression of a constitutively active mutant of GSK3β delayed the nuclear accumulation of NFATc1, caused a 50% decrease in NFAT transcriptional activity, and attenuated vSMC wound repair. We conclude that NFATc1 promotes the vSMC response to injury, and that inhibition of GSK3β is required for the activation of NFAT during wound repair

  14. Elastase effect on the extracellular matrix of rat aortic smooth muscle cells in culture

    International Nuclear Information System (INIS)

    Kispert, J.; Mogayzel, P.J. Jr.; Pratt, C.A.; Toselli, P.; Wolfe, B.L.; Faris, B.; Franzblau, C.

    1986-01-01

    The effect of porcine pancreatic elastase on the extracellular matrix (ECM) of neonatal rat aortic smooth muscle cell cultures was monitored both chemically and ultrastructurally. Initially, the elastin appeared as non-coalesced material closely associated with filaments, presumably microfibrils. The insoluble elastin accumulated in the ECM of cells in culture for 6 weeks accounted for 40-45% of the total protein. After exposure to elastase for 30-60 minutes, the elastin content was reduced to 14-20%. The reduction in the total protein content of the cultures after elastase treatment was due primarily to the loss of elastin. Although the amino acid compositions of the elastin isolated from cultures both before and after elastase treatment were similar, there were striking ultrastructural differences in the amorphous elastin. The elastin assumed a mottled appearance after elastase exposure, similar to that seen in in vivo emphysema models. Pulse experiments with 3 H-valine demonstrated an increase in protein synthesis by the cells 20 hours after elastase exposure, suggesting the potential for elastin repair. The use of this culture system will aid in clarifying the role of elastolysis in pulmonary and vascular injuries

  15. High fructose-mediated attenuation of insulin receptor signaling does not affect PDGF-induced proliferative signaling in vascular smooth muscle cells.

    Science.gov (United States)

    Osman, Islam; Poulose, Ninu; Ganapathy, Vadivel; Segar, Lakshman

    2016-11-15

    Insulin resistance is associated with accelerated atherosclerosis. Although high fructose is known to induce insulin resistance, it remains unclear as to how fructose regulates insulin receptor signaling and proliferative phenotype in vascular smooth muscle cells (VSMCs), which play a major role in atherosclerosis. Using human aortic VSMCs, we investigated the effects of high fructose treatment on insulin receptor substrate-1 (IRS-1) serine phosphorylation, insulin versus platelet-derived growth factor (PDGF)-induced phosphorylation of Akt, S6 ribosomal protein, and extracellular signal-regulated kinase (ERK), and cell cycle proteins. In comparison with PDGF (a potent mitogen), neither fructose nor insulin enhanced VSMC proliferation and cyclin D1 expression. d-[ 14 C(U)]fructose uptake studies revealed a progressive increase in fructose uptake in a time-dependent manner. Concentration-dependent studies with high fructose (5-25mM) showed marked increases in IRS-1 serine phosphorylation, a key adapter protein in insulin receptor signaling. Accordingly, high fructose treatment led to significant diminutions in insulin-induced phosphorylation of downstream signaling components including Akt and S6. In addition, high fructose significantly diminished insulin-induced ERK phosphorylation. Nevertheless, high fructose did not affect PDGF-induced key proliferative signaling events including phosphorylation of Akt, S6, and ERK and expression of cyclin D1 protein. Together, high fructose dysregulates IRS-1 phosphorylation state and proximal insulin receptor signaling in VSMCs, but does not affect PDGF-induced proliferative signaling. These findings suggest that systemic insulin resistance rather than VSMC-specific dysregulation of insulin receptor signaling by high fructose may play a major role in enhancing atherosclerosis and neointimal hyperplasia. Copyright © 2016 Elsevier B.V. All rights reserved.

  16. Estrogen-Induced Maldevelopment of the Penis Involves Down-Regulation of Myosin Heavy Chain 11 (MYH11) Expression, a Biomarker for Smooth Muscle Cell Differentiation1

    Science.gov (United States)

    Okumu, L.A.; Bruinton, Sequoia; Braden, Tim D.; Simon, Liz; Goyal, Hari O.

    2012-01-01

    ABSTRACT Cavernous smooth muscle cells are essential components in penile erection. In this study, we investigated effects of estrogen exposure on biomarkers for smooth muscle cell differentiation in the penis. Neonatal rats received diethylstilbestrol (DES), with or without the estrogen receptor (ESR) antagonist ICI 182,780 (ICI) or the androgen receptor (AR) agonist dihydrotestosterone (DHT), from Postnatal Days 1 to 6. Tissues were collected at 7, 10, or 21 days of age. The smooth muscle cell biomarker MYH11 was studied in depth because microarray data showed it was significantly down-regulated, along with other biomarkers, in DES treatment. Quantitative real time-PCR and Western blot analyses showed 50%–80% reduction (P ≤ 0.05) in Myh11 expression in DES-treated rats compared to that in controls; and ICI and DHT coadministration mitigated the decrease. Temporally, from 7 to 21 days of age, Myh11 expression was onefold increased (P ≥ 0.05) in DES-treated rats versus threefold increased (P ≤ 0.001) in controls, implying the long-lasting inhibitory effect of DES on smooth muscle cell differentiation. Immunohistochemical localization of smooth muscle alpha actin, another biomarker for smooth muscle cell differentiation, showed fewer cavernous smooth muscle cells in DES-treated animals than in controls. Additionally, DES treatment significantly up-regulated Esr1 mRNA expression and suppressed the neonatal testosterone surge by 90%, which was mitigated by ICI coadministration but not by DHT coadministration. Collectively, results provided evidence that DES treatment in neonatal rats inhibited cavernous smooth muscle cell differentiation, as shown by down-regulation of MYH11 expression at the mRNA and protein levels and by reduced immunohistochemical staining of smooth muscle alpha actin. Both the ESR and the AR pathways probably mediate this effect. PMID:22976277

  17. Mast cell chymase induces smooth muscle cell apoptosis by disrupting NF-κB-mediated survival signaling

    International Nuclear Information System (INIS)

    Leskinen, Markus J.; Heikkilae, Hanna M.; Speer, Mei Y.; Hakala, Jukka K.; Laine, Mika; Kovanen, Petri T.; Lindstedt, Ken A.

    2006-01-01

    Chymase released from activated mast cells induces apoptosis of vascular smooth muscle cells (SMCs) in vitro by degrading the pericellular matrix component fibronectin, so causing disruption of focal adhesion complexes and Akt dephosphorylation, which are necessary for cell adhesion and survival. However, the molecular mechanisms of chymase-mediated apoptosis downstream of Akt have remained elusive. Here, we show by means of RT-PCR, Western blotting, EMSA, immunocytochemistry and confocal microscopy, that chymase induces SMC apoptosis by disrupting NF-κB-mediated survival signaling. Following chymase treatment, the translocation of active NF-κB/p65 to the nucleus was partly abolished and the amount of nuclear p65 was reduced. Pretreatment of SMCs with chymase also inhibited LPS- and IL-1β-induced nuclear translocation of p65. The chymase-induced degradation of p65 was mediated by active caspases. Loss of NF-κB-mediated transactivation resulted in downregulation of bcl-2 mRNA and protein expression, leading to mitochondrial swelling and release of cytochrome c. The apoptotic process involved activation of both caspase 9 and caspase 8. The results reveal that, by disrupting the NF-κB-mediated survival-signaling pathway, activated chymase-secreting mast cells can mediate apoptosis of cultured arterial SMCs. Since activated mast cells colocalize with apoptotic SMCs in vulnerable areas of human atherosclerotic plaques, they may participate in the weakening and rupture of atherosclerotic plaques

  18. Corynoxeine isolated from the hook of Uncaria rhynchophylla inhibits rat aortic vascular smooth muscle cell proliferation through the blocking of extracellular signal regulated kinase 1/2 phosphorylation.

    Science.gov (United States)

    Kim, Tack-Joong; Lee, Ju-Hyun; Lee, Jung-Jin; Yu, Ji-Yeon; Hwang, Bang-Yeon; Ye, Sang-Kyu; Shujuan, Li; Gao, Li; Pyo, Myoung-Yun; Yun, Yeo-Pyo

    2008-11-01

    The proliferation of vascular smooth muscle cells (VSMCs) induced by injury to the intima of arteries is an important etiologic factor in vascular proliferative disorders such as atherosclerosis and restenosis. Uncaria rhynchophylla is traditional Chinese herb that has been applied to the treatment of convulsive disorders, such as epilepsy, in China. In the present study, we examined whether corynoxeine exerts inhibitory effects on platelet-derived growth factor (PDGF)-BB-induced rat aortic VSMC proliferation and the possible mechanism of such effects. Pre-treatment of VSMCs with corynoxeine (5-50 microM) for 24 h resulted in significant decreases in cell number without any cytotoxicity; the inhibition percentages were 25.0+/-12.5, 63.0+/-27.5 and 88.0+/-12.5% at 5, 20 and 50 microM, respectively. Also, corynoxeine significantly inhibited the 50 ng/ml PDGF-BB-induced DNA synthesis of VSMCs in a concentration-dependent manner without any cytotoxicity; the inhibitions were 32.8+/-11.0, 51.8+/-8.0 and 76.9+/-7.4% at concentrations of 5, 20 and 50 microM, respectively. Pre-incubation of VSMCs with corynoxeine significantly inhibited PDGF-BB-induced extracellular signal-regulated kinase 1/2 (ERK1/2) activation, whereas corynoxeine had no effects on mitogen-activated protein kinase (MAPK/ERK)-activating kinase 1 and 2 (MEK1/2), Akt, or phospholipase C (PLC)gamma1 activation or on PDGF receptor beta (PDGF-Rbeta) phosphorylation. These results suggest that corynoxeine is a potent ERK1/2 inhibitor of key PDGF-BB-induced VSMC proliferation and may be useful in the prevention and treatment of vascular diseases and restenosis after angioplasty.

  19. Hydrogen sulfide potentiates interleukin-1β-induced nitric oxide production via enhancement of extracellular signal-regulated kinase activation in rat vascular smooth muscle cells

    International Nuclear Information System (INIS)

    Jeong, Sun-Oh; Pae, Hyun-Ock; Oh, Gi-Su; Jeong, Gil-Saeng; Lee, Bok-Soo; Lee, Seoul; Kim, Du Yong; Rhew, Hyun Yul; Lee, Kang-Min; Chung, Hun-Taeg

    2006-01-01

    Hydrogen sulfide (H 2 S) and nitric oxide (NO) are endogenously synthesized from L-cysteine and L-arginine, respectively. They might constitute a cooperative network to regulate their effects. In this study, we investigated whether H 2 S could affect NO production in rat vascular smooth muscle cells (VSMCs) stimulated with interleukin-1β (IL-1β). Although H 2 S by itself showed no effect on NO production, it augmented IL-β-induced NO production and this effect was associated with increased expression of inducible NO synthase (iNOS) and activation of nuclear factor (NF)-κB. IL-1β activated the extracellular signal-regulated kinase 1/2 (ERK1/2), and this activation was also enhanced by H 2 S. Inhibition of ERK1/2 activation by the selective inhibitor U0126 inhibited IL-1β-induced NF-κB activation, iNOS expression, and NO production either in the absence or presence of H 2 S. Our findings suggest that H 2 S enhances NO production and iNOS expression by potentiating IL-1β-induced NF-κB activation through a mechanism involving ERK1/2 signaling cascade in rat VSMCs

  20. Isolation and individual electrical stimulation of single smooth-muscle cells from the urinary bladder of the pig

    NARCIS (Netherlands)

    J.J. Glerum (Jacobus); R. van Mastrigt (Ron); J.C. Romijn (Johannes); D.J. Griffiths (Derek)

    1987-01-01

    textabstractIn contrast to striated muscle, measurements on strips of smooth muscle cannot be uniquely interpreted in terms of an array of contractile units. Therefore scaling down to the single-cell level is necessary to gain detailed understanding of the contractile process in this type of muscle.

  1. Iduronic acid in chondroitin/dermatan sulfate affects directional migration of aortic smooth muscle cells.

    Directory of Open Access Journals (Sweden)

    Barbara Bartolini

    Full Text Available Aortic smooth muscle cells produce chondroitin/dermatan sulfate (CS/DS proteoglycans that regulate extracellular matrix organization and cell behavior in normal and pathological conditions. A unique feature of CS/DS proteoglycans is the presence of iduronic acid (IdoA, catalyzed by two DS epimerases. Functional ablation of DS-epi1, the main epimerase in these cells, resulted in a major reduction of IdoA both on cell surface and in secreted CS/DS proteoglycans. Downregulation of IdoA led to delayed ability to re-populate wounded areas due to loss of directional persistence of migration. DS-epi1-/- aortic smooth muscle cells, however, had not lost the general property of migration showing even increased speed of movement compared to wild type cells. Where the cell membrane adheres to the substratum, stress fibers were denser whereas focal adhesion sites were fewer. Total cellular expression of focal adhesion kinase (FAK and phospho-FAK (pFAK was decreased in mutant cells compared to control cells. As many pathological conditions are dependent on migration, modulation of IdoA content may point to therapeutic strategies for diseases such as cancer and atherosclerosis.

  2. Modeling the dispersion effects of contractile fibers in smooth muscles

    Science.gov (United States)

    Murtada, Sae-Il; Kroon, Martin; Holzapfel, Gerhard A.

    2010-12-01

    Micro-structurally based models for smooth muscle contraction are crucial for a better understanding of pathological conditions such as atherosclerosis, incontinence and asthma. It is meaningful that models consider the underlying mechanical structure and the biochemical activation. Hence, a simple mechanochemical model is proposed that includes the dispersion of the orientation of smooth muscle myofilaments and that is capable to capture available experimental data on smooth muscle contraction. This allows a refined study of the effects of myofilament dispersion on the smooth muscle contraction. A classical biochemical model is used to describe the cross-bridge interactions with the thin filament in smooth muscles in which calcium-dependent myosin phosphorylation is the only regulatory mechanism. A novel mechanical model considers the dispersion of the contractile fiber orientations in smooth muscle cells by means of a strain-energy function in terms of one dispersion parameter. All model parameters have a biophysical meaning and may be estimated through comparisons with experimental data. The contraction of the middle layer of a carotid artery is studied numerically. Using a tube the relationships between the internal pressure and the stretches are investigated as functions of the dispersion parameter, which implies a strong influence of the orientation of smooth muscle myofilaments on the contraction response. It is straightforward to implement this model in a finite element code to better analyze more complex boundary-value problems.

  3. miR-125b targets DNMT3b and mediates p53 DNA methylation involving in the vascular smooth muscle cells proliferation induced by homocysteine

    Energy Technology Data Exchange (ETDEWEB)

    Cao, ChengJian [Key Laboratory of Basic Research in Cardio-Cerebral Vascular Diseases, Ningxia Medical University, Yinchuan (China); Zhang, HuiPing [Department of Prenatal Diagnosis Center, General Hospital of Ningxia Medical University, Yinchuan (China); Zhao, Li [Department of Medical Laboratory, Ningxia Medical University, Yinchuan (China); Zhou, Longxia [Department of Basic Medicine, Ningxia Medical University, Yinchuan (China); Zhang, Minghao; Xu, Hua [Key Laboratory of Basic Research in Cardio-Cerebral Vascular Diseases, Ningxia Medical University, Yinchuan (China); Department of Basic Medicine, Ningxia Medical University, Yinchuan (China); Han, Xuebo [Department of Medical Laboratory, Ningxia Medical University, Yinchuan (China); Li, Guizhong; Yang, Xiaoling [Key Laboratory of Basic Research in Cardio-Cerebral Vascular Diseases, Ningxia Medical University, Yinchuan (China); Department of Basic Medicine, Ningxia Medical University, Yinchuan (China); Jiang, YiDeng, E-mail: jyjcyxy@yeah.net [Key Laboratory of Basic Research in Cardio-Cerebral Vascular Diseases, Ningxia Medical University, Yinchuan (China); Department of Basic Medicine, Ningxia Medical University, Yinchuan (China)

    2016-09-10

    MicroRNAs (miRNAs) are short non-coding RNA and play crucial roles in a wide array of biological processes, including cell proliferation, differentiation and apoptosis. Our previous studies found that homocysteine(Hcy) can stimulate the proliferation of vascular smooth muscle cells (VSMCs), however, the underlying mechanisms were not fully elucidated. Here, we found proliferation of VSMCs induced by Hcy was of correspondence to the miR-125b expression reduced both in vitro and in the ApoE knockout mice, the hypermethylation of p53, its decreased expression, and DNA (cytosine-5)-methyltransferase 3b (DNMT3b) up-regulated. And, we found DNMT3b is a target of miR-125b, which was verified by the Dual-Luciferase reporter assay and western blotting. Besides, the siRNA interference for DNMT3b significantly decreased the methylation level of p53, which unveiled the causative role of DNMT3b in p53 hypermethylation. miR-125b transfection further confirmed its regulative roles on p53 gene methylation status and the VSMCs proliferation. Our data suggested that a miR-125b-DNMT3b-p53 signal pathway may exist in the VSMCs proliferation induced by Hcy.

  4. miR-125b targets DNMT3b and mediates p53 DNA methylation involving in the vascular smooth muscle cells proliferation induced by homocysteine

    International Nuclear Information System (INIS)

    Cao, ChengJian; Zhang, HuiPing; Zhao, Li; Zhou, Longxia; Zhang, Minghao; Xu, Hua; Han, Xuebo; Li, Guizhong; Yang, Xiaoling; Jiang, YiDeng

    2016-01-01

    MicroRNAs (miRNAs) are short non-coding RNA and play crucial roles in a wide array of biological processes, including cell proliferation, differentiation and apoptosis. Our previous studies found that homocysteine(Hcy) can stimulate the proliferation of vascular smooth muscle cells (VSMCs), however, the underlying mechanisms were not fully elucidated. Here, we found proliferation of VSMCs induced by Hcy was of correspondence to the miR-125b expression reduced both in vitro and in the ApoE knockout mice, the hypermethylation of p53, its decreased expression, and DNA (cytosine-5)-methyltransferase 3b (DNMT3b) up-regulated. And, we found DNMT3b is a target of miR-125b, which was verified by the Dual-Luciferase reporter assay and western blotting. Besides, the siRNA interference for DNMT3b significantly decreased the methylation level of p53, which unveiled the causative role of DNMT3b in p53 hypermethylation. miR-125b transfection further confirmed its regulative roles on p53 gene methylation status and the VSMCs proliferation. Our data suggested that a miR-125b-DNMT3b-p53 signal pathway may exist in the VSMCs proliferation induced by Hcy.

  5. Mg(2+,ATP-dependent plasma membrane calcium pump of smooth muscle cells. ІІ. Regulation of activity

    Directory of Open Access Journals (Sweden)

    T. О. Veklich

    2015-04-01

    Full Text Available Plasma membrane Ca2+-pump is one of key proteins, which takes part in Ca2+ exchange in smooth muscle cells. It has a lot of diverse functions from control of basal cytoplasmal Ca2+ concentration to regulation of proteins involved in Ca2+-dependent signal pathway. Ca2+ pump function is often depen­dent on the isoform or even form of alternative splicing. Allowing for a variety of Ca2+-pump functions and properties, which were reviewed in detail in the first part of our review article cycle (Ukr. Biochem. J., 2015; 87(1, the precise control of the mentioned pump activity is very important for cell functioning­. The other part of this article is dedicated to different regulation factors of smooth muscle plasma membrane Ca2+-pump activity: endogenous and exo­genous, biotic and abiotic factors. Special attention is given to literature data and own results about design and the search of selective plasma membrane Ca2+-pump inhibitor which would allow examining its functioning in smooth muscle cells more meticulously.

  6. Slow and sustained nitric oxide releasing compounds inhibit multipotent vascular stem cell proliferation and differentiation without causing cell death

    International Nuclear Information System (INIS)

    Curtis, Brandon M.; Leix, Kyle Alexander; Ji, Yajing; Glaves, Richard Samuel Elliot; Ash, David E.; Mohanty, Dillip K.

    2014-01-01

    Highlights: • Multipotent vascular stem cells (MVSCs) proliferate and differentiate. • Nitric oxide inhibits proliferation of MVSCs. • Nitric oxide inhibits MVSC differentiation to mesenchymal-like stem cells (MSCs). • Smooth muscle cells (SMCs) neither de-differentiate nor proliferate. - Abstract: Atherosclerosis is the leading cause of cerebral and myocardial infarction. It is believed that neointimal growth common in the later stages of atherosclerosis is a result of vascular smooth muscle cell (SMC) de-differentiation in response to endothelial injury. However, the claims of the SMC de-differentiation theory have not been substantiated by monitoring the fate of mature SMCs in response to such injuries. A recent study suggests that atherosclerosis is a consequence of multipotent vascular stem cell (MVSC) differentiation. Nitric oxide (NO) is a well-known mediator against atherosclerosis, in part because of its inhibitory effect on SMC proliferation. Using three different NO-donors, we have investigated the effects of NO on MVSC proliferation. Results indicate that NO inhibits MVSC proliferation in a concentration dependent manner. A slow and sustained delivery of NO proved to inhibit proliferation without causing cell death. On the other hand, larger, single-burst NO concentrations, inhibits proliferation, with concurrent significant cell death. Furthermore, our results indicate that endogenously produced NO inhibits MVSC differentiation to mesenchymal-like stem cells (MSCs) and subsequently to SMC as well

  7. Slow and sustained nitric oxide releasing compounds inhibit multipotent vascular stem cell proliferation and differentiation without causing cell death

    Energy Technology Data Exchange (ETDEWEB)

    Curtis, Brandon M.; Leix, Kyle Alexander [Department of Chemistry, Central Michigan University, Mount Pleasant, MI 48859 (United States); Ji, Yajing [Department of Biomedical Science and Medicine, Michigan State University, East Lansing, MI 48824 (United States); Glaves, Richard Samuel Elliot [Department of Biology, Central Michigan University, Mount Pleasant, MI 48859 (United States); Ash, David E. [Department of Chemistry, Central Michigan University, Mount Pleasant, MI 48859 (United States); Mohanty, Dillip K., E-mail: Mohan1dk@cmich.edu [Department of Chemistry, Central Michigan University, Mount Pleasant, MI 48859 (United States)

    2014-07-18

    Highlights: • Multipotent vascular stem cells (MVSCs) proliferate and differentiate. • Nitric oxide inhibits proliferation of MVSCs. • Nitric oxide inhibits MVSC differentiation to mesenchymal-like stem cells (MSCs). • Smooth muscle cells (SMCs) neither de-differentiate nor proliferate. - Abstract: Atherosclerosis is the leading cause of cerebral and myocardial infarction. It is believed that neointimal growth common in the later stages of atherosclerosis is a result of vascular smooth muscle cell (SMC) de-differentiation in response to endothelial injury. However, the claims of the SMC de-differentiation theory have not been substantiated by monitoring the fate of mature SMCs in response to such injuries. A recent study suggests that atherosclerosis is a consequence of multipotent vascular stem cell (MVSC) differentiation. Nitric oxide (NO) is a well-known mediator against atherosclerosis, in part because of its inhibitory effect on SMC proliferation. Using three different NO-donors, we have investigated the effects of NO on MVSC proliferation. Results indicate that NO inhibits MVSC proliferation in a concentration dependent manner. A slow and sustained delivery of NO proved to inhibit proliferation without causing cell death. On the other hand, larger, single-burst NO concentrations, inhibits proliferation, with concurrent significant cell death. Furthermore, our results indicate that endogenously produced NO inhibits MVSC differentiation to mesenchymal-like stem cells (MSCs) and subsequently to SMC as well.

  8. Chronic Prenatal Hypoxia Down-Regulated BK Channel Β1 Subunits in Mesenteric Artery Smooth Muscle Cells of the Offspring

    Directory of Open Access Journals (Sweden)

    Bailin Liu

    2018-02-01

    Full Text Available Background/Aims: Chronic hypoxia in utero could impair vascular functions in the offspring, underlying mechanisms are unclear. This study investigated functional alteration in large-conductance Ca2+-activated K+ (BK channels in offspring mesenteric arteries following prenatal hypoxia. Methods: Pregnant rats were exposed to normoxic control (21% O2, Con or hypoxic (10.5% O2, Hy conditions from gestational day 5 to 21, their 7-month-old adult male offspring were tested for blood pressure, vascular BK channel functions and expression using patch clamp and wire myograh technique, western blotting, and qRT-PCR. Results: Prenatal hypoxia increased pressor responses and vasoconstrictions to phenylephrine in the offspring. Whole-cell currents density of BK channels and amplitude of spontaneous transient outward currents (STOCs, not the frequency, were significantly reduced in Hy vascular myocytes. The sensitivity of BK channels to voltage, Ca2+, and tamoxifen were reduced in Hy myocytes, whereas the number of channels per patch and the single-channel conductance were unchanged. Prenatal hypoxia impaired NS1102- and tamoxifen-mediated relaxation in mesenteric arteries precontracted with phenylephrine in the presence of Nω-nitro-L-arginine methyl ester. The mRNA and protein expression of BK channel β1, not the α-subunit, was decreased in Hy mesenteric arteries. Conclusions: Impaired BK channel β1-subunits in vascular smooth muscle cells contributed to vascular dysfunction in the offspring exposed to prenatal hypoxia.

  9. Increased IGF-IEc expression and mechano-growth factor production in intestinal muscle of fibrostenotic Crohn's disease and smooth muscle hypertrophy.

    Science.gov (United States)

    Li, Chao; Vu, Kent; Hazelgrove, Krystina; Kuemmerle, John F

    2015-12-01

    The igf1 gene is alternatively spliced as IGF-IEa and IGF-IEc variants in humans. In fibrostenotic Crohn's disease, the fibrogenic cytokine TGF-β1 induces IGF-IEa expression and IGF-I production in intestinal smooth muscle and results in muscle hyperplasia and collagen I production that contribute to stricture formation. Mechano-growth factor (MGF) derived from IGF-IEc induces skeletal and cardiac muscle hypertrophy following stress. We hypothesized that increased IGF-IEc expression and MGF production mediated smooth muscle hypertrophy also characteristic of fibrostenotic Crohn's disease. IGF-IEc transcripts and MGF protein were increased in muscle cells isolated from fibrostenotic intestine under regulation by endogenous TGF-β1. Erk5 and MEF2C were phosphorylated in vivo in fibrostenotic muscle; both were phosphorylated and colocalized to nucleus in response to synthetic MGF in vitro. Smooth muscle-specific protein expression of α-smooth muscle actin, γ-smooth muscle actin, and smoothelin was increased in affected intestine. Erk5 inhibition or MEF2C siRNA blocked smooth muscle-specific gene expression and hypertrophy induced by synthetic MGF. Conditioned media of cultured fibrostenotic muscle induced muscle hypertrophy that was inhibited by immunoneutralization of endogenous MGF or pro-IGF-IEc. The results indicate that TGF-β1-dependent IGF-IEc expression and MGF production in patients with fibrostenotic Crohn's disease regulates smooth muscle cell hypertrophy a critical factor that contributes to intestinal stricture formation. Copyright © 2015 the American Physiological Society.

  10. Eye features in three Danish patients with multisystemic smooth muscle dysfunction syndrome

    DEFF Research Database (Denmark)

    Moller, Hans Ulrik; Fledelius, Hans C; Milewicz, Dianna M

    2012-01-01

    A de novo mutation of the ACTA2 gene encoding the smooth muscle cell α-actin has been established in patients with multisystemic smooth muscle dysfunction syndrome associated with patent ductus arteriosus and mydriasis present at birth....

  11. Terbinafine: effects on platelet-derived growth factor-stimulated smooth muscle cells in vitro and myointimal proliferation in vivo

    International Nuclear Information System (INIS)

    McCarthy, L.; Van Halen, R.G.; St Denny, I.H.; Glinka, K.G.; Handley, D.A.; Stuetz, A.; Nemecek, G.M.

    1987-01-01

    Terbinafine (T; (E)-N-(6,6-dimethyl-2-hepten-4-ynyl)-N-methyl-1-naphthalenemethanamine), an antimycotic agent with antimitogenic activity in fibroblasts, was examined for its effects on platelet-derived growth factor (PDGF)-stimulated aortic smooth muscle cell DNA synthesis in vitro and myointimal proliferation in vivo. Exposure of smooth muscle cells to 1-25 μM T resulted in a concentration-dependent inhibition of PDGF-induced mitogenesis as determined by [ 3 H]thymidine incorporation or cell number. The IC 50 for T was approximately 5 μM. The inhibitory effect of terbinafine persisted in the presence of 0.4-8.0 μg/ml cholesterol or 130 μg/ml mevalonate. Administration of T to rats for 2 d before and 14 d after balloon catheter carotid injury resulted in a 40% decrease in lesion area. These observations indicate that T is both a potent in vitro antagonist of the smooth muscle cell mitogenic response to PDGF and an effective, well-tolerated, orally active inhibitor of myointimal proliferation in vivo

  12. Terbinafine: effects on platelet-derived growth factor-stimulated smooth muscle cells in vitro and myointimal proliferation in vivo

    Energy Technology Data Exchange (ETDEWEB)

    McCarthy, L.; Van Halen, R.G.; St. Denny, I.H.; Glinka, K.G.; Handley, D.A.; Stuetz, A.; Nemecek, G.M.

    1987-05-01

    Terbinafine (T; (E)-N-(6,6-dimethyl-2-hepten-4-ynyl)-N-methyl-1-naphthalenemethanamine), an antimycotic agent with antimitogenic activity in fibroblasts, was examined for its effects on platelet-derived growth factor (PDGF)-stimulated aortic smooth muscle cell DNA synthesis in vitro and myointimal proliferation in vivo. Exposure of smooth muscle cells to 1-25 ..mu..M T resulted in a concentration-dependent inhibition of PDGF-induced mitogenesis as determined by (/sup 3/H)thymidine incorporation or cell number. The IC/sub 50/ for T was approximately 5 ..mu..M. The inhibitory effect of terbinafine persisted in the presence of 0.4-8.0 ..mu..g/ml cholesterol or 130 ..mu..g/ml mevalonate. Administration of T to rats for 2 d before and 14 d after balloon catheter carotid injury resulted in a 40% decrease in lesion area. These observations indicate that T is both a potent in vitro antagonist of the smooth muscle cell mitogenic response to PDGF and an effective, well-tolerated, orally active inhibitor of myointimal proliferation in vivo.

  13. From genetics to response to injury: vascular smooth muscle cells in aneurysms and dissections of the ascending aorta.

    Science.gov (United States)

    Michel, Jean-Baptiste; Jondeau, Guillaume; Milewicz, Dianna M

    2018-03-15

    Vascular smooth muscle cells (vSMCs) play a crucial role in both the pathogenesis of Aneurysms and Dissections of the ascending thoracic aorta (TAAD) in humans and in the associated adaptive compensatory responses, since thrombosis and inflammatory processes are absent in the majority of cases. Aneurysms and dissections share numerous characteristics, including aetiologies and histopathological alterations: vSMC disappearance, medial areas of mucoid degeneration, and extracellular matrix (ECM) breakdown. Three aetiologies predominate in TAAD in humans: (i) genetic causes in heritable familial forms, (ii) an association with bicuspid aortic valves, and (iii) a sporadic degenerative form linked to the aortic aging process. Genetic forms include mutations in vSMC genes encoding for molecules of the ECM or the TGF-β pathways, or participating in vSMC tone. On the other hand, aneurysms and dissections, whatever their aetiologies, are characterized by an increase in wall permeability leading to transmural advection of plasma proteins which could interact with vSMCs and ECM components. In this context, blood-borne plasminogen appears to play an important role, because its outward convection through the wall is increased in TAAD, and it could be converted to active plasmin at the vSMC membrane. Active plasmin can induce vSMC disappearance, proteolysis of adhesive proteins, activation of MMPs and release of TGF-β from its ECM storage sites. Conversely, vSMCs could respond to aneurysmal biomechanical and proteolytic injury by an epigenetic phenotypic switch, including constitutional overexpression and nuclear translocation of Smad2 and an increase in antiprotease and ECM protein synthesis. In contrast, such an epigenetic phenomenon is not observed in dissections. In this context, dysfunction of proteins involved in vSMC tone are interesting to study, particularly in interaction with plasma protein transport through the wall and TGF-β activation, to establish the

  14. A fibroblast-associated antigen: Characterization in fibroblasts and immunoreactivity in smooth muscle differentiated stromal cells

    DEFF Research Database (Denmark)

    Rønnov-Jessen, Lone; Celis, Julio E.; van Deurs, Bo

    1992-01-01

    major brands migrating at apparent Mr of 38,000, 45,000, and 80,000, in addition to many minor bands between Mr 45,000 and 97,000, including Mr 52,000. The Mr 45,000 and 38,000 were associated with the cell membrane and Mr 52,000 as well as Mr 38,000 were associated with the lysosomes. The 1B10......Fibroblasts with smooth muscle differentiation are frequently derived from human breast tissue. Immunofluorescence cytochemistry of a fibroblast-associated antigen recognized by a monoclonal antibody (MAb), 1B10, was analyzed with a view to discriminating smooth muscle differentiated fibroblasts...

  15. Bestrophin-3 (vitelliform macular dystrophy 2-like 3 protein) is essential for the cGMP-dependent calcium-activated chloride conductance in vascular smooth muscle cells

    DEFF Research Database (Denmark)

    Matchkov, Vladimir; Larsen, Per; Bouzinova, Elena V.

    2008-01-01

    have recently characterized a cGMP-dependent Ca(2+)-activated Cl(-) current with unique characteristics in smooth muscle cells. This novel current has been shown to coexist with a "classic" (cGMP-independent) Ca(2+)-activated Cl(-) current and to have characteristics distinct from those previously...... known for Ca(2+)-activated Cl(-) currents. Here, we suggest that a bestrophin, a product of the Best gene family, is responsible for the cGMP-dependent Ca(2+)-activated Cl(-) current based on similarities between the membrane currents produced by heterologous expressions of bestrophins and the cGMP......-dependent Ca(2+)-activated Cl(-) current. This is supported by similarities in the distribution pattern of the cGMP-dependent Ca(2+)-activated Cl(-) current and bestrophin-3 (the product of Best-3 gene) expression in different smooth muscle. Furthermore, downregulation of Best-3 gene expression with small...

  16. Human eosinophil–airway smooth muscle cell interactions

    Directory of Open Access Journals (Sweden)

    J. Margaret Hughes

    2000-01-01

    Full Text Available Eosinophils are present throughout the airway wall of asthmatics. The nature of the interaction between human airway smooth muscle cells (ASMC and eosinophils was investigated in this study. We demonstrated, using light microscopy, that freshly isolated eosinophils from healthy donors rapidly attach to ASMC in vitro. Numbers of attached eosinophils were highest at 2 h, falling to 50% of maximum by 20 h. Eosinophil attachment at 2 h was reduced to 72% of control by anti-VCAM-1, and to 74% at 20 h by anti-ICAM-1. Pre-treatment of ASMC for 24 h with TNF-α, 10 nM, significantly increased eosinophil adhesion to 149 and 157% of control after 2 and 20 h. These results provide evidence that eosinophil interactions with ASMC involve VCAM-1 and ICAM-1 and are modulated by TNF-α.

  17. Regulation and Roles of Urocortins in the Vascular System

    Directory of Open Access Journals (Sweden)

    Kazunori Kageyama

    2012-01-01

    Full Text Available Urocortins (Ucns are members of the corticotropin-releasing factor (CRF family of peptides. Ucns would have potent effects on the cardiovascular system via the CRF receptor type 2 (CRF2 receptor. Regulation and roles of each Ucn have been determined in the vascular system. Ucns have more potent vasodilatory effects than CRF. Human umbilical vein endothelial cells (HUVECs express Ucns1-3 mRNAs, and the receptor, CRF2a receptor mRNA. Ucns1-3 mRNA levels are differentially regulated in HUVECs. Differential regulation of Ucns may suggest differential roles of those in HUVECs. Ucn1 and Ucn2 have strong effects on interleukin (IL-6 gene expression and secretion in rat aortic smooth muscle A7r5 cells. The increase that we observed in IL-6 levels following Ucn treatment of A7r5 cells suggests that smooth muscle cells may be a source of IL-6 secretion under physiological stress conditions. Ucns are important and unique modulators of vascular smooth muscle cells and act directly or indirectly as autocrine and paracrine factors in the vascular system.

  18. Androgen action via testicular arteriole smooth muscle cells is important for Leydig cell function, vasomotion and testicular fluid dynamics.

    Directory of Open Access Journals (Sweden)

    Michelle Welsh

    2010-10-01

    Full Text Available Regulation of blood flow through the testicular microvasculature by vasomotion is thought to be important for normal testis function as it regulates interstitial fluid (IF dynamics which is an important intra-testicular transport medium. Androgens control vasomotion, but how they exert these effects remains unclear. One possibility is by signalling via androgen receptors (AR expressed in testicular arteriole smooth muscle cells. To investigate this and determine the overall importance of this mechanism in testis function, we generated a blood vessel smooth muscle cell-specific AR knockout mouse (SMARKO. Gross reproductive development was normal in SMARKO mice but testis weight was reduced in adulthood compared to control littermates; this reduction was not due to any changes in germ cell volume or to deficits in testosterone, LH or FSH concentrations and did not cause infertility. However, seminiferous tubule lumen volume was reduced in adult SMARKO males while interstitial volume was increased, perhaps indicating altered fluid dynamics; this was associated with compensated Leydig cell failure. Vasomotion was impaired in adult SMARKO males, though overall testis blood flow was normal and there was an increase in the overall blood vessel volume per testis in adult SMARKOs. In conclusion, these results indicate that ablating arteriole smooth muscle AR does not grossly alter spermatogenesis or affect male fertility but does subtly impair Leydig cell function and testicular fluid exchange, possibly by locally regulating microvascular blood flow within the testis.

  19. Angiotensin II increases phosphodiesterase 5A expression in vascular smooth muscle cells: A mechanism by which angiotensin II antagonizes cGMP signaling

    Science.gov (United States)

    Kim, Dongsoo; Aizawa, Toru; Wei, Heng; Pi, Xinchun; Rybalkin, Sergei D.; Berk, Bradford C.; Yan, Chen

    2014-01-01

    Angiotensin II (Ang II) and nitric oxide (NO)/natriuretic peptide (NP) signaling pathways mutually regulate each other. Imbalance of Ang II and NO/NP has been implicated in the pathophysiology of many vascular diseases. cGMP functions as a key mediator in the interaction between Ang II and NO/NP. Cyclic nucleotide phosphodiesterase 5A (PDE5A) is important in modulating cGMP signaling by hydrolyzing cGMP in vascular smooth muscle cells (VSMC). Therefore, we examined whether Ang II negatively modulates intracellular cGMP signaling in VSMC by regulating PDE5A. Ang II rapidly and transiently increased PDE5A mRNA levels in rat aortic VSMC. Upregulation of PDE5A mRNA was associated with a time-dependent increase of both PDE5 protein expression and activity. Increased PDE5A mRNA level was transcription-dependent and mediated by the Ang II type 1 receptor. Ang II-mediated activation of extracellular signal-regulated kinases 1/2 (ERK1/2) was essential for Ang II-induced PDE5A upregulation. Pretreatment of VSMC with Ang II inhibited C-type NP (CNP) stimulated cGMP signaling, such as cGMP dependent protein kinase (PKG)-mediated phosphorylation of vasodilator-stimulated-phosphoprotein (VASP). Ang II-mediated inhibition of PKG was blocked when PDE5 activity was decreased by selective PDE5 inhibitors, suggesting that upregulation of PDE5A expression is an important mechanism for Ang II to attenuate cGMP signaling. PDE5A may also play a critical role in the growth promoting effects of Ang II because inhibition of PDE5A activity significantly decreased Ang II-stimulated VSMC growth. These observations establish a new mechanism by which Ang II antagonizes cGMP signaling and stimulates VSMC growth. PMID:15623434

  20. TLR4 induces CREB-mediated IL-6 production via upregulation of F-spondin to promote vascular smooth muscle cell migration

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Guan-Lin [Institute of Cellular and System Medicine, National Health Research Institutes, Zhunan, Taiwan (China); Graduate Institutes of Life Sciences, National Defense Medical Center, Taipei, Taiwan (China); Wu, Jing-Yiing [Institute of Cellular and System Medicine, National Health Research Institutes, Zhunan, Taiwan (China); Yeh, Chang-Ching [Institute of Cellular and System Medicine, National Health Research Institutes, Zhunan, Taiwan (China); Graduate Institutes of Life Sciences, National Defense Medical Center, Taipei, Taiwan (China); Kuo, Cheng-Chin, E-mail: kuocc@nhri.org.tw [Institute of Cellular and System Medicine, National Health Research Institutes, Zhunan, Taiwan (China); Graduate Institutes of Life Sciences, National Defense Medical Center, Taipei, Taiwan (China); Graduate Institute of Basic Medical Science, China Medical University, Taichung, Taiwan (China)

    2016-05-13

    Toll-like receptor 4 (TLR4) is important in promoting inflammation and vascular smooth muscle cell (VSMC) migration, both of which contribute to atherosclerosis development and progression. But the mechanism underlying the regulation of TLR4 in VSMC migration remains unclear. Stimulation of VSMCs with LPS increased the cellular level of F-spondin which is associated with the regulation of proinflammatory cytokine production. The LPS-induced F-spondin expression depended on TLR4-mediated PI3K/Akt pathway. Suppression of F-spondin level by siRNA inhibited not only F-spondin expression but also LPS-induced phosphorylation of cAMP response element binding protein (CREB) and IL-6 expression, VSMC migration and proliferation as well as MMP9 expression. Moreover, suppression of CREB level by siRNA inhibited TLR4-induced IL-6 production and VSMC migration. Inhibition of F-spondin siRNA on LPS-induced migration was restored by addition of exogenous recombinant mouse IL-6. We conclude that upon ligand binding, TLR4 activates PI3K/Akt signaling to induce F-spondin expression, subsequently control CREB-mediated IL-6 production to promote VSMC migration. These findings provide vital insights into the essential role of F-spondin in VSMC function and will be valuable for developing new therapeutic strategies against atherosclerosis. -- Highlights: •LPS-induced F-spondin expression of VSMCs is via a TLR4/PI3K/Akt signaling. •F-spondin is pivotal for LPS-induced CREB-mediated IL-6 production. •F-spondin is required for LPS-induced VSMC migration and proliferation.

  1. TLR4 induces CREB-mediated IL-6 production via upregulation of F-spondin to promote vascular smooth muscle cell migration

    International Nuclear Information System (INIS)

    Lee, Guan-Lin; Wu, Jing-Yiing; Yeh, Chang-Ching; Kuo, Cheng-Chin

    2016-01-01

    Toll-like receptor 4 (TLR4) is important in promoting inflammation and vascular smooth muscle cell (VSMC) migration, both of which contribute to atherosclerosis development and progression. But the mechanism underlying the regulation of TLR4 in VSMC migration remains unclear. Stimulation of VSMCs with LPS increased the cellular level of F-spondin which is associated with the regulation of proinflammatory cytokine production. The LPS-induced F-spondin expression depended on TLR4-mediated PI3K/Akt pathway. Suppression of F-spondin level by siRNA inhibited not only F-spondin expression but also LPS-induced phosphorylation of cAMP response element binding protein (CREB) and IL-6 expression, VSMC migration and proliferation as well as MMP9 expression. Moreover, suppression of CREB level by siRNA inhibited TLR4-induced IL-6 production and VSMC migration. Inhibition of F-spondin siRNA on LPS-induced migration was restored by addition of exogenous recombinant mouse IL-6. We conclude that upon ligand binding, TLR4 activates PI3K/Akt signaling to induce F-spondin expression, subsequently control CREB-mediated IL-6 production to promote VSMC migration. These findings provide vital insights into the essential role of F-spondin in VSMC function and will be valuable for developing new therapeutic strategies against atherosclerosis. -- Highlights: •LPS-induced F-spondin expression of VSMCs is via a TLR4/PI3K/Akt signaling. •F-spondin is pivotal for LPS-induced CREB-mediated IL-6 production. •F-spondin is required for LPS-induced VSMC migration and proliferation.

  2. Voltage dependent potassium channel remodeling in murine intestinal smooth muscle hypertrophy induced by partial obstruction.

    Science.gov (United States)

    Liu, Dong-Hai; Huang, Xu; Guo, Xin; Meng, Xiang-Min; Wu, Yi-Song; Lu, Hong-Li; Zhang, Chun-Mei; Kim, Young-chul; Xu, Wen-Xie

    2014-01-01

    Partial obstruction of the small intestine causes obvious hypertrophy of smooth muscle cells and motility disorder in the bowel proximate to the obstruction. To identify electric remodeling of hypertrophic smooth muscles in partially obstructed murine small intestine, the patch-clamp and intracellular microelectrode recording methods were used to identify the possible electric remodeling and Western blot, immunofluorescence and immunoprecipitation were utilized to examine the channel protein expression and phosphorylation level changes in this research. After 14 days of obstruction, partial obstruction caused obvious smooth muscle hypertrophy in the proximally located intestine. The slow waves of intestinal smooth muscles in the dilated region were significantly suppressed, their amplitude and frequency were reduced, whilst the resting membrane potentials were depolarized compared with normal and sham animals. The current density of voltage dependent potassium channel (KV) was significantly decreased in the hypertrophic smooth muscle cells and the voltage sensitivity of KV activation was altered. The sensitivity of KV currents (IKV) to TEA, a nonselective potassium channel blocker, increased significantly, but the sensitivity of IKv to 4-AP, a KV blocker, stays the same. The protein levels of KV4.3 and KV2.2 were up-regulated in the hypertrophic smooth muscle cell membrane. The serine and threonine phosphorylation levels of KV4.3 and KV2.2 were significantly increased in the hypertrophic smooth muscle cells. Thus this study represents the first identification of KV channel remodeling in murine small intestinal smooth muscle hypertrophy induced by partial obstruction. The enhanced phosphorylations of KV4.3 and KV2.2 may be involved in this process.

  3. Purification and partial characterization of glycosaminoglycans and proteoglycans from cultured rabbit smooth muscle cells

    International Nuclear Information System (INIS)

    Sabatino, R.D.

    1985-01-01

    Glycosaminoglycans synthesized by cultured rabbit smooth muscle cells were isolated after incorporation of [ 3 H]-glucosamine into glycosaminoglycans in the presence or absence of 10% fetal bovine serum. Glycosaminoglycans were quantitated by two-dimensional electrophoresis after proteolytic digestion of the cell layers and media. The results show that the presence of serum has no effect on the chondroitin sulfate, heparan sulfate and dermatan sulfate content of the cell layers. The incorporation of [ 3 H]-glucosamine into hyaluronic acid of the cell layers was three times higher in the presence of serum. In the medium , the quantity of hyaluronic was two times higher in the presence of serum while the other glycosaminoglycans remained unchanged. The incorporation of [ 3 H]-glucosamine into hyaluronic acid was unaffected by the presence of serum. Specific proteoglycans were isolated from medium after with [ 35 S]-sulfate and [ 3 H]-serine by isopycnic ultracentrifugation and chromatography on Sepharose CL-4B and DEAE-cellulose. Preparations contained a chondroitin sulfate proteoglycan, a condroitin sulfate-dermatan sulfate proteoglycan and a heparan sulfate proteoglycan. Glycosaminoglycans and proteoglycans synthesized by rabbit aorta smooth muscle cells are similar to those from human aorta

  4. MicroRNA expression profile and functional analysis reveal their roles in contact inhibition and its disruption switch of rat vascular smooth muscle cells.

    Science.gov (United States)

    Sun, Ye-Ying; Qin, Shan-Shan; Cheng, Yun-Hui; Wang, Chao-Yun; Liu, Xiao-Jun; Liu, Ying; Zhang, Xiu-Li; Zhang, Wendy; Zhan, Jia-Xin; Shao, Shuai; Bian, Wei-Hua; Luo, Bi-Hui; Lu, Dong-Feng; Yang, Jian; Wang, Chun-Hua; Zhang, Chun-Xiang

    2018-05-01

    Contact inhibition and its disruption of vascular smooth muscle cells (VSMCs) are important cellular events in vascular diseases. But the underlying molecular mechanisms are unclear. In this study we investigated the roles of microRNAs (miRNAs) in the contact inhibition and its disruption of VSMCs and the molecular mechanisms involved. Rat VSMCs were seeded at 30% or 90% confluence. MiRNA expression profiles in contact-inhibited confluent VSMCs (90% confluence) and non-contact-inhibited low-density VSMCs (30% confluence) were determined. We found that multiple miRNAs were differentially expressed between the two groups. Among them, miR-145 was significantly increased in contact-inhibited VSMCs. Serum could disrupt the contact inhibition as shown by the elicited proliferation of confluent VSMCs. The contact inhibition disruption accompanied with a down-regulation of miR-145. Serum-induced contact inhibition disruption of VSMCs was blocked by overexpression of miR-145. Moreover, downregulation of miR-145 was sufficient to disrupt the contact inhibition of VSMCs. The downregulation of miR-145 in serum-induced contact inhibition disruption was related to the activation PI3-kinase/Akt pathway, which was blocked by the PI3-kinase inhibitor LY294002. KLF5, a target gene of miR-145, was identified to be involved in miR-145-mediated effect on VSMC contact inhibition disruption, as it could be inhibited by knockdown of KLF5. In summary, our results show that multiple miRNAs are differentially expressed in contact-inhibited VSMCs and in non-contact-inhibited VSMCs. Among them, miR-145 is a critical gene in contact inhibition and its disruption of VSMCs. PI3-kinase/Akt/miR-145/KLF5 is a critical signaling pathway in serum-induced contact inhibition disruption. Targeting of miRNAs related to the contact inhibition of VSMCs may represent a novel therapeutic approach for vascular diseases.

  5. Nuclear localization of CPI-17, a protein phosphatase-1 inhibitor protein, affects histone H3 phosphorylation and corresponds to proliferation of cancer and smooth muscle cells

    Energy Technology Data Exchange (ETDEWEB)

    Eto, Masumi, E-mail: masumi.eto@jefferson.edu [Department of Molecular Physiology and Biophysics, and Kimmel Cancer Center, Thomas Jefferson University, 1020 Locust Street, PA 19107 (United States); Kirkbride, Jason A.; Chugh, Rishika; Karikari, Nana Kofi [Department of Molecular Physiology and Biophysics, and Kimmel Cancer Center, Thomas Jefferson University, 1020 Locust Street, PA 19107 (United States); Kim, Jee In [Department of Molecular Physiology and Biophysics, and Kimmel Cancer Center, Thomas Jefferson University, 1020 Locust Street, PA 19107 (United States); Cardiovascular Research Institute, Kyungpook National University School of Medicine, Daegu 700-422 (Korea, Republic of)

    2013-04-26

    Highlights: •Non-canonical roles of the myosin phosphatase inhibitor (CPI-17) were studied. •CPI-17 is localized in the nucleus of hyperplastic cancer and smooth muscle cells. •CPI-17 Ser12 phosphorylation may regulate the nuclear import. •CPI-17 regulates histone H3 phosphorylation and cell proliferation. •The nuclear CPI-17-PP1 axis plays a proliferative role in cells. -- Abstract: CPI-17 (C-kinase-activated protein phosphatase-1 (PP1) inhibitor, 17 kDa) is a cytoplasmic protein predominantly expressed in mature smooth muscle (SM) that regulates the myosin-associated PP1 holoenzyme (MLCP). Here, we show CPI-17 expression in proliferating cells, such as pancreatic cancer and hyperplastic SM cells. Immunofluorescence showed that CPI-17 was concentrated in nuclei of human pancreatic cancer (Panc1) cells. Nuclear accumulation of CPI-17 was also detected in the proliferating vascular SM cell culture and cells at neointima of rat vascular injury model. The N-terminal 21-residue tail domain of CPI-17 was necessary for the nuclear localization. Phospho-mimetic Asp-substitution of CPI-17 at Ser12 attenuated the nuclear import. CPI-17 phosphorylated at Ser12 was not localized at nuclei, suggesting a suppressive role of Ser12 phosphorylation in the nuclear import. Activated CPI-17 bound to all three isoforms of PP1 catalytic subunit in Panc1 nuclear extracts. CPI-17 knockdown in Panc1 resulted in dephosphorylation of histone H3 at Thr3, Ser10 and Thr11, whereas it had no effects on the phosphorylation of myosin light chain and merlin, the known targets of MLCP. In parallel, CPI-17 knockdown suppressed Panc1 proliferation. We propose that CPI-17 accumulated in the nucleus through the N-terminal tail targets multiple PP1 signaling pathways regulating cell proliferation.

  6. Voltage dependent potassium channel remodeling in murine intestinal smooth muscle hypertrophy induced by partial obstruction.

    Directory of Open Access Journals (Sweden)

    Dong-Hai Liu

    Full Text Available Partial obstruction of the small intestine causes obvious hypertrophy of smooth muscle cells and motility disorder in the bowel proximate to the obstruction. To identify electric remodeling of hypertrophic smooth muscles in partially obstructed murine small intestine, the patch-clamp and intracellular microelectrode recording methods were used to identify the possible electric remodeling and Western blot, immunofluorescence and immunoprecipitation were utilized to examine the channel protein expression and phosphorylation level changes in this research. After 14 days of obstruction, partial obstruction caused obvious smooth muscle hypertrophy in the proximally located intestine. The slow waves of intestinal smooth muscles in the dilated region were significantly suppressed, their amplitude and frequency were reduced, whilst the resting membrane potentials were depolarized compared with normal and sham animals. The current density of voltage dependent potassium channel (KV was significantly decreased in the hypertrophic smooth muscle cells and the voltage sensitivity of KV activation was altered. The sensitivity of KV currents (IKV to TEA, a nonselective potassium channel blocker, increased significantly, but the sensitivity of IKv to 4-AP, a KV blocker, stays the same. The protein levels of KV4.3 and KV2.2 were up-regulated in the hypertrophic smooth muscle cell membrane. The serine and threonine phosphorylation levels of KV4.3 and KV2.2 were significantly increased in the hypertrophic smooth muscle cells. Thus this study represents the first identification of KV channel remodeling in murine small intestinal smooth muscle hypertrophy induced by partial obstruction. The enhanced phosphorylations of KV4.3 and KV2.2 may be involved in this process.

  7. Matrix Metalloproteinases: Inflammatory Regulators of Cell Behaviors in Vascular Formation and Remodeling

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    Qishan Chen

    2013-01-01

    Full Text Available Abnormal angiogenesis and vascular remodeling contribute to pathogenesis of a number of disorders such as tumor, arthritis, atherosclerosis, restenosis, hypertension, and neurodegeneration. During angiogenesis and vascular remodeling, behaviors of stem/progenitor cells, endothelial cells (ECs, and vascular smooth muscle cells (VSMCs and its interaction with extracellular matrix (ECM play a critical role in the processes. Matrix metalloproteinases (MMPs, well-known inflammatory mediators are a family of zinc-dependent proteolytic enzymes that degrade various components of ECM and non-ECM molecules mediating tissue remodeling in both physiological and pathological processes. MMPs including MMP-1, MMP-2, MMP-3, MMP-7, MMP-8, MMP-9, MMP-12, and MT1-MMP, are stimulated and activated by various stimuli in vascular tissues. Once activated, MMPs degrade ECM proteins or other related signal molecules to promote recruitment of stem/progenitor cells and facilitate migration and invasion of ECs and VSMCs. Moreover, vascular cell proliferation and apoptosis can also be regulated by MMPs via proteolytically cleaving and modulating bioactive molecules and relevant signaling pathways. Regarding the importance of vascular cells in abnormal angiogenesis and vascular remodeling, regulation of vascular cell behaviors through modulating expression and activation of MMPs shows therapeutic potential.

  8. Enhanced expressions of microvascular smooth muscle receptors after focal cerebral ischemia occur via the MAPK MEK/ERK pathway

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    Edvinsson Lars

    2008-09-01

    Full Text Available Abstract Background MEK1/2 is a serine/threonine protein that phosphorylates extracellular signal-regulated kinase (ERK1/2. Cerebral ischemia results in enhanced expression of cerebrovascular contractile receptors in the middle cerebral artery (MCA leading to the ischemic region. Here we explored the role of the MEK/ERK pathway in receptor expression following ischemic brain injury using the specific MEK1 inhibitor U0126. Methods and result Rats were subjected to a 2-h middle cerebral artery occlusion (MCAO followed by reperfusion for 48-h and the ischemic area was calculated. The expression of phosphorylated ERK1/2 and Elk-1, and of endothelin ETA and ETB, angiotensin AT1, and 5-hydroxytryptamine 5-HT1B receptors were analyzed with immunohistochemistry using confocal microscopy in cerebral arteries, microvessels and in brain tissue. The expression of endothelin ETB receptor was analyzed by quantitative Western blot. We demonstrate that there is an increase in the number of contractile smooth muscle receptors in the MCA and in micro- vessels within the ischemic region. The enhanced expression occurs in the smooth muscle cells as verified by co-localization studies. This receptor upregulation is furthermore associated with enhanced expression of pERK1/2 and of transcription factor pElk-1 in the vascular smooth muscle cells. Blockade of transcription with the MEK1 inhibitor U0126, given at the onset of reperfusion or as late as 6 hours after the insult, reduced transcription (pERK1/2 and pElk-1, the enhanced vascular receptor expression, and attenuated the cerebral infarct and improved neurology score. Conclusion Our results show that MCAO results in upregulation of cerebrovascular ETB, AT1 and 5-HT1B receptors. Blockade of this event with a MEK1 inhibitor as late as 6 h after the insult reduced the enhanced vascular receptor expression and the associated cerebral infarction.

  9. The Effects of Environmental Factors on Smooth Muscle Cells Differentiation from Adipose-Derived Stem Cells and Esophagus Tissues Engineering

    DEFF Research Database (Denmark)

    Wang, Fang

    Adipose-derived stem cells (ASCs) are increasingly being used for regenerative medicine and tissue engineering. Smooth muscle cells (SMCs) can be differentiated from ASCs. Oxygen is a key factor influencing the stem cell differentiation. Tissue engineered esophagus has been a preferred solution...... of esophagus was studied. Our results showed that both SMCs and ASCs could attach on the porcine esophageal acellular matrix (EAM) scaffold in vitro after 24 hours and survive until 7 days. Thus ASCs might be a substitute for SMCs in the construction of tissue engineered esophageal muscle layer....

  10. Uric acid stimulates proliferative pathways in vascular smooth muscle cells through the activation of p38 MAPK, p44/42 MAPK and PDGFRβ.

    Science.gov (United States)

    Kırça, M; Oğuz, N; Çetin, A; Uzuner, F; Yeşilkaya, A

    2017-04-01

    Hyperuricemia and angiotensin II (Ang II) may have a pathogenetic role in the development of hypertension and atherosclerosis as well as cardiovascular disease (CVD) and its prognosis. The purpose of this study was to investigate whether uric acid can induce proliferative pathways of vascular smooth muscle cell (VSMC) that are thought to be responsible for the development of CVD. The phosphorylation of p38 mitogen-activated protein kinase (p38 MAPK), p44/42 mitogen-activated protein kinase (p44/42 MAPK) and platelet-derived growth factor receptor β (PDGFRβ) was measured by Elisa and Western blot techniques to determine the activation of proliferative pathways in primary cultured VSMCs from rat aorta. Results demonstrated that uric acid can stimulate p38 MAPK, p44/42 MAPK and PDGFRβ phosphorylation in a time- and concentration-dependent manner. Furthermore, treatment of VSMCs with the angiotensin II type I receptor (AT1R) inhibitor losartan suppressed p38 MAPK and p44/42 MAPK induction by uric acid. The stimulatory effect of uric acid on p38 MAPK was higher compared to that of Ang II. The results of this study show for the first time that uric acid-induced PDGFRβ phosphorylation plays a crucial role in the development of CVDs and that elevated uric acid levels could be a potential therapeutical target in CVD patients.

  11. Characterization of vascular endothelial progenitor cells from chicken bone marrow

    Directory of Open Access Journals (Sweden)

    Bai Chunyu

    2012-05-01

    Full Text Available Abstract Background Endothelial progenitor cells (EPC are a type of stem cell used in the treatment of atherosclerosis, vascular injury and regeneration. At present, most of the EPCs studied are from human and mouse, whereas the study of poultry-derived EPCs has rarely been reported. In the present study, chicken bone marrow-derived EPCs were isolated and studied at the cellular level using immunofluorescence and RT-PCR. Results We found that the majority of chicken EPCs were spindle shaped. The growth-curves of chicken EPCs at passages (P 1, -5 and -9 were typically “S”-shaped. The viability of chicken EPCs, before and after cryopreservation was 92.2% and 81.1%, respectively. Thus, cryopreservation had no obvious effects on the viability of chicken EPCs. Dil-ac-LDL and FITC-UAE-1 uptake assays and immunofluorescent detection of the cell surface markers CD34, CD133, VEGFR-2 confirmed that the cells obtained in vitro were EPCs. Observation of endothelial-specific Weibel-Palade bodies using transmission electron microscopy further confirmed that the cells were of endothelial lineage. In addition, chicken EPCs differentiated into endothelial cells and smooth muscle cells upon induction with VEGF and PDGF-BB, respectively, suggesting that the chicken EPCs retained multipotency in vitro. Conclusions These results suggest that chicken EPCs not only have strong self-renewal capacity, but also the potential to differentiate into endothelial and smooth muscle cells. This research provides theoretical basis and experimental evidence for potential therapeutic application of endothelial progenitor cells in the treatment of atherosclerosis, vascular injury and diabetic complications.

  12. Nodular smooth muscle metaplasia in multiple peritoneal endometriosis

    OpenAIRE

    Kim, Hyun-Soo; Yoon, Gun; Ha, Sang Yun; Song, Sang Yong

    2015-01-01

    We report here an unusual presentation of peritoneal endometriosis with smooth muscle metaplasia as multiple protruding masses on the lateral pelvic wall. Smooth muscle metaplasia is a common finding in rectovaginal endometriosis, whereas in peritoneal endometriosis, smooth muscle metaplasia is uncommon and its nodular presentation on the pelvic wall is even rarer. To the best of our knowledge, this is the first case of nodular smooth muscle metaplasia occurring in peritoneal endometriosis. A...

  13. Cyclic GMP alters Ca exchange in vascular smooth muscle

    International Nuclear Information System (INIS)

    Magliola, L.; Bailey, B.; Jones, A.W.

    1986-01-01

    Contraction and 42 K efflux from vascular smooth muscle stimulated either by norepinephrine (NE) or by K-depolarization is dependent on an increase in cytosolic Ca concentration. The purpose of this study was to determine if cyclic GMP (cGMP) inhibited these processes and if inhibition was secondary to the action of cGMP on Ca movements. Basal cGMP content of rat aorta was 1.2 fmol/mg wet wt. Sodium nitroprusside (NP) increased cGMP ∼2-fold at 1 nM and ∼750-fold at 1 μM with no effect on cAMP levels. A 5 min pretreatment with NP (1 μM) completely prevented tension development induced by 3 μM NE. The same concentration of NP also inhibited NE-stimulated 42 K and 45 Ca efflux > 90 and > 80%, respectively. Removal of NP in the continued presence of NE (3 μM) caused recovery of the 42 K efflux response to ∼75% of control with a half-time of ∼2.5 min. NP (1 μM) also caused a rapid relaxation of aorta contracted with 3 μM NE and a loss of the 42 K efflux response with half-times of 2-3 min. In contrast, 100 μM NP produced only a 50% inhibition of contraction induced by high K (55 mM). Also, NP (1 μM) inhibited K-stimulated 42 K efflux only ∼25%. These results demonstrate both a concentration- and a time-dependent relationship between increases in cGMP induced by NP and decreases in NE-stimulated contraction, 42 K and 45 Ca effluxes. They also indicate that the sensitivity of NE-induced contraction and 42 K efflux to NP is greater than that induced by high K. These studies suggest that cGMP modulates the control sites for Ca exchange in the plasma membrane and sarcoplasmic reticulum

  14. Camptothecin inhibits platelet-derived growth factor-BB-induced proliferation of rat aortic vascular smooth muscle cells through inhibition of PI3K/Akt signaling pathway

    Energy Technology Data Exchange (ETDEWEB)

    Park, Eun-Seok [Department of Applied Biochemistry, Division of Life Science, College of Health and Biomedical Science, Konkuk University, Chungju, Chungbuk (Korea, Republic of); Kang, Shin-il [College of Pharmacy Medical Research Center, Chungbuk National University, Cheongju (Korea, Republic of); Yoo, Kyu-dong [Hazardous Substances Analysis Division, Gwangju Regional Food and Drug Administration, Gwangju (Korea, Republic of); Lee, Mi-Yea [Department of Nursing Kyungbok University, Pocheon (Korea, Republic of); Yoo, Hwan-Soo; Hong, Jin-Tae [College of Pharmacy Medical Research Center, Chungbuk National University, Cheongju (Korea, Republic of); Shin, Hwa-Sup [Department of Applied Biochemistry, Division of Life Science, College of Health and Biomedical Science, Konkuk University, Chungju, Chungbuk (Korea, Republic of); Kim, Bokyung [Department of Physiology, Konkuk Medical School, Konkuk University, Chungju, Chungbuk (Korea, Republic of); Yun, Yeo-Pyo, E-mail: ypyun@chungbuk.ac.kr [College of Pharmacy Medical Research Center, Chungbuk National University, Cheongju (Korea, Republic of)

    2013-04-15

    The abnormal proliferation of vascular smooth muscle cells (VSMCs) in arterial wall is a major cause of vascular disorders such as atherosclerosis and restenosis after angioplasty. In this study, we investigated not only the inhibitory effects of camptothecin (CPT) on PDGF-BB-induced VSMC proliferation, but also its molecular mechanism of this inhibition. CPT significantly inhibited proliferation with IC50 value of 0.58 μM and the DNA synthesis of PDGF-BB-stimulated VSMCs in a dose-dependent manner (0.5–2 μM ) without any cytotoxicity. CPT induced the cell cycle arrest at G0/G1 phase. Also, CPT decreased the expressions of G0/G1-specific regulatory proteins including cyclin-dependent kinase (CDK)2, cyclin D1 and PCNA in PDGF-BB-stimulated VSMCs. Pre-incubation of VSMCs with CPT significantly inhibited PDGF-BB-induced Akt activation, whereas CPT did not affect PDGF-receptor beta phosphorylation, extracellular signal-regulated kinase (ERK) 1/2 phosphorylation and phospholipase C (PLC)-γ1 phosphorylation in PDGF-BB signaling pathway. Our data showed that CPT pre-treatment inhibited VSMC proliferation, and that the inhibitory effect of CPT was enhanced by LY294002, a PI3K inhibitor, on PDGF-BB-induced VSMC proliferation. In addition, inhibiting the PI3K/Akt pathway by LY294002 significantly enhanced the suppression of PCNA expression and Akt activation by CPT. These results suggest that the anti-proliferative activity of CPT is mediated in part by downregulating the PI3K/Akt signaling pathway. - Highlights: ► CPT inhibits proliferation of PDGF-BB-induced VSMC without cytotoxicity. ► CPT arrests the cell cycle in G0/G1 phase by downregulation of cyclin D1 and CDK2. ► CPT significantly attenuates Akt phosphorylation in PDGF-BB signaling pathway. ► LY294002 enhanced the inhibitory effect of CPT on VSMC proliferation. ► Thus, CPT is mediated by downregulating the PI3K/Akt signaling pathway.

  15. Camptothecin inhibits platelet-derived growth factor-BB-induced proliferation of rat aortic vascular smooth muscle cells through inhibition of PI3K/Akt signaling pathway

    International Nuclear Information System (INIS)

    Park, Eun-Seok; Kang, Shin-il; Yoo, Kyu-dong; Lee, Mi-Yea; Yoo, Hwan-Soo; Hong, Jin-Tae; Shin, Hwa-Sup; Kim, Bokyung; Yun, Yeo-Pyo

    2013-01-01

    The abnormal proliferation of vascular smooth muscle cells (VSMCs) in arterial wall is a major cause of vascular disorders such as atherosclerosis and restenosis after angioplasty. In this study, we investigated not only the inhibitory effects of camptothecin (CPT) on PDGF-BB-induced VSMC proliferation, but also its molecular mechanism of this inhibition. CPT significantly inhibited proliferation with IC50 value of 0.58 μM and the DNA synthesis of PDGF-BB-stimulated VSMCs in a dose-dependent manner (0.5–2 μM ) without any cytotoxicity. CPT induced the cell cycle arrest at G0/G1 phase. Also, CPT decreased the expressions of G0/G1-specific regulatory proteins including cyclin-dependent kinase (CDK)2, cyclin D1 and PCNA in PDGF-BB-stimulated VSMCs. Pre-incubation of VSMCs with CPT significantly inhibited PDGF-BB-induced Akt activation, whereas CPT did not affect PDGF-receptor beta phosphorylation, extracellular signal-regulated kinase (ERK) 1/2 phosphorylation and phospholipase C (PLC)-γ1 phosphorylation in PDGF-BB signaling pathway. Our data showed that CPT pre-treatment inhibited VSMC proliferation, and that the inhibitory effect of CPT was enhanced by LY294002, a PI3K inhibitor, on PDGF-BB-induced VSMC proliferation. In addition, inhibiting the PI3K/Akt pathway by LY294002 significantly enhanced the suppression of PCNA expression and Akt activation by CPT. These results suggest that the anti-proliferative activity of CPT is mediated in part by downregulating the PI3K/Akt signaling pathway. - Highlights: ► CPT inhibits proliferation of PDGF-BB-induced VSMC without cytotoxicity. ► CPT arrests the cell cycle in G0/G1 phase by downregulation of cyclin D1 and CDK2. ► CPT significantly attenuates Akt phosphorylation in PDGF-BB signaling pathway. ► LY294002 enhanced the inhibitory effect of CPT on VSMC proliferation. ► Thus, CPT is mediated by downregulating the PI3K/Akt signaling pathway

  16. Clathrin-dependent internalization of the angiotensin II AT₁A receptor links receptor internalization to COX-2 protein expression in rat aortic vascular smooth muscle cells.

    Science.gov (United States)

    Morinelli, Thomas A; Walker, Linda P; Velez, Juan Carlos Q; Ullian, Michael E

    2015-02-05

    The major effects of Angiotensin II (AngII) in vascular tissue are mediated by AngII AT1A receptor activation. Certain effects initiated by AT1A receptor activation require receptor internalization. In rat aortic vascular smooth muscle cells (RASMC), AngII stimulates cyclooxygenase 2 protein expression. We have previously shown this is mediated by β-arrestin-dependent receptor internalization and NF-κB activation. In this study, a specific inhibitor of clathrin-mediated endocytosis (CME), pitstop-2, was used to test the hypothesis that clathrin-dependent internalization of activated AT1A receptor mediates NF-κB activation and subsequent cyclooxygenase 2 expression. Radioligand binding assays, real time qt-PCR and immunoblotting were used to document the effects of pitstop-2 on AngII binding and signaling in RASMC. Laser scanning confocal microscopy (LSCM) was used to image pitstop-2׳s effects on AT1 receptor/GFP internalization in HEK-293 cells and p65 NF-κB nuclear localization in RASMC. Pitstop-2 significantly inhibited internalization of AT1A receptor (44.7% ± 3.1% Control vs. 13.2% ± 8.3% Pitstop-2; n=3) as determined by radioligand binding studies in RASMC. Studies utilizing AT1A receptor/GFP expressed in HEK 293 cells and LSCM confirmed these findings. Pitstop-2 significantly inhibited AngII-induced p65 NF-κB phosphorylation and nuclear localization, COX-2 message and protein expression in RASMC without altering activation of p42/44 ERK or TNFα signaling. Pitstop-2, a specific inhibitor of clathrin-mediated endocytosis, confirms that internalization of activated AT1A receptor mediates AngII activation of cyclooxygenase 2 expression in RASMC. These data provide support for additional intracellular signaling pathways activated through β-arrestin mediated internalization of G protein-coupled receptors, such as AT1A receptors. Copyright © 2014 Elsevier B.V. All rights reserved.

  17. EFFECT OF STENT ABSORBED c-myc ANTISENSE OLIGODEOXYNUCLEOTIDE ON SMOOTH MUSCLE CELLS APOPTOSIS IN RABBIT CAROTID ARTERY

    Institute of Scientific and Technical Information of China (English)

    张新霞; 崔长琮; 李江; 崔翰斌; 徐仓宝; 朱参战

    2002-01-01

    Objective To investigate the effect of gelatin coated Platinium-Iridium stent absorbed c-myc antisense oligodeoxynucleotide (ASODN) on smooth muscle cells apoptosis in a normal rabbit carotid arteries. Methods Gelatin coated Platinium-Iridium stents were implanted in the right carotid arteries of 32 rabbits under vision. Animals were randomly divided into control group and treated group receiving c-myc ASODN (n=16, respectively). On 7, 14, 30 and 90 days following the stenting procedure ,morphometry for caculation of neointimal area and mean neointimal thickness were performed.The expression of c-myc protein was detected by immunohistochemical method. Apoptotic smooth muscle cells was detected by terminal deoxynucleotidyl transferase mediated dUTP nick end labeling (TUNEL). Results At 7 and 14 days after stenting,there were no detectable apoptotic cells in both groups. The apoptotic cells occurred in the neointima 30 and 90 days after stenting, and the number of apoptotic cells at 30 days were less [4.50±1.29 vs 25.75±1.89 (number/0.1mm2)] than that at 90 days [13.50±1.91 vs 41.50±6.46 (number/0.1mm2)]. Meanwhile c-myc ASODN induced more apoptotic cells than the control group(P<0.0001). c-myc protein expression was weak positive or negative in treated group and positive in control group.Conclusion c-myc ASODN can induce smooth muscle cells apoptosis after stenting in normal rabbit carotid arteries,and it can be used to prevent in-stent restenosis.

  18. Regulation of the sodium/potassium/chloride cotransporter by calcium and cyclic AMP in cultured vascular smooth muscle cells

    International Nuclear Information System (INIS)

    Higgins, B.L.; Smith, L.; Smith, J.B.

    1987-01-01

    The activity of the Na/K/Cl cotransporter in smooth muscle cells cultured from rat aorta was assayed by measuring the initial rate of furosemide-inhibitable 86 Rb influx or efflux. Five uM furosemide or 0.2 uM bumetanide inhibited influx by 50%. Furosemide-inhibitable 86 Rb influx depended on the presence of all 3 ions in the external medium. The dependence on Na and K was hyperbolic with apparent Km values of 45 and 5 mM, respectively. The dependence on Cl was sigmoidal. Assuming a stoichiometry of 1:1:2 for Na:K:Cl, a Km for Cl of 60 mM was obtained from a Hofstee plot of the data. Rapidly growing cells had 3 fold higher cotransport activity than quiescent cells. Angiotensin II (ANG) stimulated furosemide-inhibitable 86 Rb efflux by 2 fold. An ANG receptor antagonist prevented ANG from increasing cotransport activity. Two calcium ionophores, A23187 and ionomycin, increased cotransport activity by 2 fold. Phorbol myristate acetate had no effect on cotransport activity. Isoproterenol, dibutyryl cyclic AMP, cholera toxin, or methylisobutylxanthine inhibited furosemide-sensitive 86 Rb influx by 35 to 50%. From these findings they conclude that increasing cytoplasmic free calcium stimulates cotransport activity, whereas increasing cellular cyclic AMP inhibits the cotransporter

  19. microRNA expression profile in human coronary smooth muscle cell-derived microparticles is a source of biomarkers.

    Science.gov (United States)

    de Gonzalo-Calvo, David; Cenarro, Ana; Civeira, Fernando; Llorente-Cortes, Vicenta

    2016-01-01

    microRNA (miRNA) expression profile of extracellular vesicles is a potential tool for clinical practice. Despite the key role of vascular smooth muscle cells (VSMC) in cardiovascular pathology, there is limited information about the presence of miRNAs in microparticles secreted by this cell type, including human coronary artery smooth muscle cells (HCASMC). Here, we tested whether HCASMC-derived microparticles contain miRNAs and the value of these miRNAs as biomarkers. HCASMC and explants from atherosclerotic or non-atherosclerotic areas were obtained from coronary arteries of patients undergoing heart transplant. Plasma samples were collected from: normocholesterolemic controls (N=12) and familial hypercholesterolemia (FH) patients (N=12). Both groups were strictly matched for age, sex and cardiovascular risk factors. Microparticle (0.1-1μm) isolation and characterization was performed using standard techniques. VSMC-enriched miRNAs expression (miR-21-5p, -143-3p, -145-5p, -221-3p and -222-3p) was analyzed using RT-qPCR. Total RNA isolated from HCASMC-derived microparticles contained small RNAs, including VSMC-enriched miRNAs. Exposition of HCASMC to pathophysiological conditions, such as hypercholesterolemia, induced a decrease in the expression level of miR-143-3p and miR-222-3p in microparticles, not in cells. Expression levels of miR-222-3p were lower in circulating microparticles from FH patients compared to normocholesterolemic controls. Microparticles derived from atherosclerotic plaque areas showed a decreased level of miR-143-3p and miR-222-3p compared to non-atherosclerotic areas. We demonstrated for the first time that microparticles secreted by HCASMC contain microRNAs. Hypercholesterolemia alters the microRNA profile of HCASMC-derived microparticles. The miRNA signature of HCASMC-derived microparticles is a source of cardiovascular biomarkers. Copyright © 2016 Sociedad Española de Arteriosclerosis. Publicado por Elsevier España, S.L.U. All rights

  20. Sirtuins, Cell Senescence, and Vascular Aging.

    Science.gov (United States)

    Kida, Yujiro; Goligorsky, Michael S

    2016-05-01

    The sirtuins (SIRTs) constitute a class of proteins with nicotinamide adenine dinucleotide-dependent deacetylase or adenosine diphosphate-ribosyltransferase activity. Seven SIRT family members have been identified in mammals, from SIRT1, the best studied for its role in vascular aging, to SIRT7. SIRT1 and SIRT2 are localized in the nucleus and cytoplasm. SIRT3, SIRT4, and SIRT5 are mitochondrial, and SIRT6 and SIRT7 are nuclear. Extensive studies have clearly revealed that SIRT proteins regulate diverse cell functions and responses to stressors. Vascular aging involves the aging process (senescence) of endothelial and vascular smooth muscle cells. Two types of cell senescence have been identified: (1) replicative senescence with telomere attrition; and (2) stress-induced premature senescence without telomere involvement. Both types of senescence induce vascular cell growth arrest and loss of vascular homeostasis, and contribute to the initiation and progression of cardiovascular diseases. Previous mechanistic studies have revealed in detail that SIRT1, SIRT3, and SIRT6 show protective functions against vascular aging, and definite vascular function of other SIRTs is under investigation. Thus, direct SIRT modulation and nicotinamide adenine dinucleotide stimulation of SIRT are promising candidates for cardiovascular disease therapy. A small number of pilot studies have been conducted to assess SIRT modulation in humans. These clinical studies have not yet provided convincing evidence that SIRT proteins alleviate morbidity and mortality in patients with cardiovascular diseases. The outcomes of multiple ongoing clinical trials are awaited to define the efficacy of SIRT modulators and SIRT activators in cardiovascular diseases, along with the potential adverse effects of chronic SIRT modulation. Copyright © 2016 Canadian Cardiovascular Society. Published by Elsevier Inc. All rights reserved.