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Sample records for a7r5 vascular smooth

  1. Erk1/2 MAPK and caldesmon differentially regulate podosome dynamics in A7r5 vascular smooth muscle cells

    International Nuclear Information System (INIS)

    We tested the hypothesis that the MEK/Erk/caldesmon phosphorylation cascade regulates PKC-mediated podosome dynamics in A7r5 cells. We observed the phosphorylation of MEK, Erk and caldesmon, and their translocation to the podosomes upon phorbol dibutyrate (PDBu) stimulation, together with the nuclear translocation of phospho-MEK and phospho-Erk. After MEK inhibition by U0126, Erk translocated to the interconnected actin-rich columns but failed to translocate to the nucleus, suggesting that podosomes served as a site for Erk phosphorylation. The interconnected actin-rich columns in U0126-treated, PDBu-stimulated cells contained α-actinin, caldesmon, vinculin, and metalloproteinase-2. Caldesmon and vinculin became integrated with F-actin at the columns, in contrast to their typical location at the ring of podosomes. Live-imaging experiments suggested the growth of these columns from podosomes that were slow to disassemble. The observed modulation of podosome size and life time in A7r5 cells overexpressing wild-type and phosphorylation-deficient caldesmon-GFP mutants in comparison to untransfected cells suggests that caldesmon and caldesmon phosphorylation modulate podosome dynamics in A7r5 cells. These results suggest that Erk1/2 and caldesmon differentially modulate PKC-mediated formation and/or dynamics of podosomes in A7r5 vascular smooth muscle cells

  2. Vasopressin stimulates action potential firing by protein kinase C-dependent inhibition of KCNQ5 in A7r5 rat aortic smooth muscle cells

    OpenAIRE

    Brueggemann, Lioubov I.; Moran, Christopher J.; Barakat, John A.; Yeh, Jay Z.; Cribbs, Leanne L.; Byron, Kenneth L.

    2006-01-01

    [Arg8]-vasopressin (AVP), at low concentrations (10–500 pM), stimulates oscillations in intracellular Ca2+ concentration (Ca2+ spikes) in A7r5 rat aortic smooth muscle cells. Our previous studies provided biochemical evidence that protein kinase C (PKC) activation and phosphorylation of voltage-sensitive K+ (Kv) channels are crucial steps in this process. In the present study, Kv currents (IKv) and membrane potential were measured using patch clamp techniques. Treatment of A7r5 cells with 100...

  3. Caveolin-3 promotes a vascular smooth muscle contractile phenotype

    Directory of Open Access Journals (Sweden)

    Jorge L. Gutierrez-Pajares

    2015-06-01

    Full Text Available Epidemiological studies have demonstrated the importance of cardiovascular diseases in Western countries. Among the cell types associated with a dysfunctional vasculature, smooth muscle cells are believed to play an essential role in the development of these illnesses. Vascular smooth muscle cells are key regulators of the vascular tone and also have an important function in the development of atherosclerosis and restenosis. While in the normal vasculature contractile smooth muscle cells are predominant, in atherosclerotic vascular lesions, synthetic cells migrate toward the neointima, proliferate, and synthetize extracellular matrix proteins. In the present study, we have examined the role of caveolin-3 in the regulation of smooth muscle cell phenotype. Caveolin-3 is expressed in vivo in normal arterial smooth muscle cells, but its expression appears to be lost in cultured smooth muscle cells. Our data show that caveolin-3 expression in the A7r5 smooth muscle cell line is associated with increased expression of contractility markers such as smooth muscle  actin, smooth muscle myosin heavy chain but decreased expression of the synthetic phenotype markers such as p-Elk and Klf4. Moreover, we also show that caveolin-3 expression can reduce proliferation upon treatment with LDL or PDGF. Finally, we show that caveolin-3-expressing smooth muscle cells are less sensitive to apoptosis than control cells upon treatment with oxidized LDL. Taken together, our data suggest that caveolin-3 can regulate the phenotypic switch between contractile and synthetic smooth muscle cells. A better understanding of the factors regulating caveolin-3 expression and function in this cell type will permit the development of a better comprehension of the factors regulating smooth muscle function in atherosclerosis and restenosis.

  4. Biophysical induction of vascular smooth muscle cell podosomes.

    Directory of Open Access Journals (Sweden)

    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.

  5. Enhanced antiproliferative effects of combination hexokinase II shRNA and NIS gene therapy on vascular smooth muscle cells

    International Nuclear Information System (INIS)

    Introduction: This study was designed to determine the antiproliferative effects of combination gene therapy using sodium iodide symporter (NIS)-based radioiodine and lentivirus-mediated short hairpin RNA (shRNA) against hexokinase II (HKII) on vascular smooth muscle cells (VSMCs). Methods: A7r5 rat VSMCs were stably transfected with a dual-expression vector of NIS and Fluc (A7r5-NL cells). Functional assessment was performed by radioiodine uptake assay, luciferase assay and confocal microscopy. After exposure to lentivirus-HKII-shRNA, the 18F-FDG uptake test and HK activity assay were performed. The effects of combination therapy with 131I and lentivirus-HKII-shRNA on VSMCs were assessed with an in vitro clonogenic assay. In vivo bioluminescence and nuclear imaging were undertaken using a xenografted mouse model. Results: In vitro functional assessment confirmed expression of NIS and Fluc genes in A7r5-NL, but not in parent A7r5 cells. Transfection of lentivirus-HKII-shRNA resulted in a significant decrease in messenger RNA expression of the HKII gene, 18F-FDG uptake and HK activity. The cell survival rate of A7r5-NL decreased to 61.9% and 90.5% by single therapy with 7.4 MBq of 131I or lentivirus-HKII-shRNA, respectively, and further decreased to 42.9% by combined therapy (P99mTc pertechnetate uptake at the site of A7r5-NL cell inoculation in nude mice. Conclusion: The enhanced antiproliferative effect on VSMCs was achieved by a combination of NIS-based radioiodine and lentivirus-mediated HKII shRNA gene therapy. Successful demonstration of in vivo dual reporter gene imaging assures the potential for further application in an animal model.

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

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    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.

  7. Effects of high glucose on expression of OPG and RANKL in rat aortic vascular smooth muscle cells

    Institute of Scientific and Technical Information of China (English)

    Hong-Juan Chang; Shao-Jiang Zheng; Tian-Fa Li; Jun-Li Guo; You-Ling Lan; Yue-Qiong Kong; Xin Meng; Xian-Ji Ma; Xiao-Ling Lu; Wei-Ying Lu

    2015-01-01

    Objective:To explore effect of high glucose on expression of osteoprotegerin (OPG) and receptor activator of NF-毷B ligand (RANKL) in rat aortic vascular smooth muscle cells. Methods: SD rats were intraperitoneally injected with streptozotocin, OPG and RANKL expression in rat thoracic aortas were detected by immunohistochemical staining. In cultured vascular smooth muscle cells (VSMCs) (A7r5), qRT-PCR and Western blot analysis were used to examine the mRNA and protein levels of OPG and RANKL.Results: Our results demonstrated that OPG expression was increased in hyperglycemic rat aortic VSMCs, while RANKL expression was decreased. Besides,in vitroexperiments high glucose induced OPG expression, but depressed RANKL expression by dose- and time-dependent manner in cultured A7r5.Conclusions: Our findings suggested that high glucose could promote the expression of OPG, and inhibit the expression of RANKL in VSMCs, which may be partly be the molecular mechanism of diabetic vascular calcification.

  8. Effects of high glucose on expression of OPG and RANKL in rat aortic vascular smooth muscle cells

    Institute of Scientific and Technical Information of China (English)

    Hong-Juan; Chang; Tian-Fa; Li; Jun-Li; Guo; You-Ling; Lan; Yue-Qiong; Kong; Xin; Meng; Xian-Ji; Ma; Xiao-Ling; Lu; Wei-Ying; Lu; Shao-Jiang; Zheng

    2015-01-01

    Objective:To explore effect of high glucose on expression of osteoprotegerin(OPG) and receptor activator of NF- κB ligand(RANKL) in rat aortic vascular smooth muscle cells.Methods:SD rats were intraperitoneally injected with streptozotocin,OPG and RANKL expression in rat thoracic aortas were detected by immunohistochemical staining.In cultured vascular smooth muscle cells(VSMCs)(A7r5),qRT-PCR and Western blot analysis were used to examine the mRNA and protein levels of OPG and RANKL.Results:Our results demonstrated that OPG expression was increased in hyperglycemic rat aortic VSMCs.while RANKL expression was decreased.Besides,in vitro experiments high glucose induced OPG expression,but depressed RANKL expression by dose- and time-dependent manner in cultured A7r5.Conclusions:Our findings suggested that high glucose could promote the expression of OPG,and inhibit the expression of RANKL in VSMCs,which may be partly be the molecular mechanism of diabetic vascular calcification.

  9. Microscopic study of ultrasound-mediated microbubble destruction effects on vascular smooth muscle cells

    Institute of Scientific and Technical Information of China (English)

    Bo Zhang; Yi-Rong Hou; Tian Chen; Bing Hu

    2015-01-01

    Objective: To observe vascular smooth muscle cell morphological changes induced by ultrasound combined with microbubbles by Atomic Force Acoustic Microscopy (AFAM). Methods: A7r5 rat aortic smooth muscle cells were divided into groups: control group (without ultrasonic irradiation, no micro bubbles) and US+MB group (45 kHz, 0.4 W/cm2 ultrasound irradiate for 20 seconds with a SonoVue™ concentration of [(56-140)×10 5/mL]. Cell micro-morphological changes (such as topographic and acoustic prognosis) were detected, before and after ultrasound destruction by AFAM. Results: In cell morphology, smooth muscle cells were spread o and connected to each another by fibers. At the center of the cell, the nuclear area had a rough surface and was significantly elevated from its surroundings. The cytoskeletal structure of the reticular nucleus and cytoplasm in the morphology of A7r5 cells (20μm×20μm) were clear before microbubble intervention. After acoustic exciting, the cell structure details of the acoustic image were improved with better resolution, showing the elasticity of different tissues. In the acoustic image, the nucleus was harder, more flexible and uneven compared with the cytoplasm. Many strong various-sized echo particles were stuck on the rough nuclear membrane’s substrate surface. The nuclear membrane did not have a continuous smooth surface; there were many obstructions (pores). After ultrasound-intervention was combined with microbubbles, the dark areas of the A7r5 cell images was increased in various sizes and degrees. The dark areas showed the depth or low altitudes of the lower regions, suggesting regional depressions. However, the location and scope of the acoustic image dark areas were not similar to those found in the topographic images. Therefore, it was likely that the dark areas, both from the topographic and acoustic images, were sound-holes. In addition, some cell nuclei become round in different degrees after irradiation. Conclusions: Atomic

  10. 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 rat aorta, brain, aortic smooth muscle cells (A7r5), VSMCs, and mesangial cells. Immunolabeling with an anti-alpha(1A) antibody was positive in acid-macerated, microdissected preglomerular vessels and in A7r5 cells. Patch-clamp experiments on aortic A7r5 cells showed 22+/-4% (n=6) inhibition of inward...

  11. SIRT1 inhibits angiotensin Ⅱ-induced vascular smooth muscle cell hypertrophy

    Institute of Scientific and Technical Information of China (English)

    Li Li; Chihchuan Liang; Peng Gao; Huina Zhang; Houzao Chen; Wei Zheng; Xiang Lv; Tingting Xu; Yusheng Wei; Depei Liu

    2011-01-01

    Angiotensin Ⅱ (Ang Ⅱ) stimulates vascular smooth muscle cell (VSMC) hypertrophy as a critical event in the development of vascular diseases such as atherosclerosis.Sirtuin (SIRT) 1, a nicotinamide adenine dinucleotide dependent deacetylase, has been demonstrated to exert protective effects in atherosclerosis by promoting endo-thelium-dependent vascular relaxation and reducing macrophage foam cell formation, but its role in VSMC hypertrophy remains unknown. In this study, we tried to investigate the effect of SIRTI on Ang Ⅱ-induced VSMC hypertrophy. Results showed that adenoviral-mediated over-expression of SIRT1 significantly inhibited Ang Ⅱ-induced VSMC hypertrophy, while knockdown of SIRT1 by RNAi resulted in an increased [3H]-leucine incorpor-ation of VSMC. Accordingly, nicotinamide adenine dinu-cleotide phosphate oxidase 1 (Nox1) expression induced by Ang Ⅱ was inhibited by SIRT1 in VSMCs. SIRT1 acti-vator resveratrol decreased, whereas endogenous SIRT1 inhibitor nicotinamide increased Nox1 expression in A7r5 VSMCs. Furthermore, transcription factor GATA-6 was involved in the down-regulation of Nox1 expression by SIRT1. These results provide new insight into SIRTI's anti-atherogenic properties by suppressing Ang Ⅱ-induced VSMC hypertrophy.

  12. Intracellular Angiotensin II and cell growth of vascular smooth muscle cells

    NARCIS (Netherlands)

    Filipeanu, CM; Henning, RH; de Zeeuw, D; Nelemans, A

    2001-01-01

    1 We recently demonstrated that intracellular application of Angiotensin II (Angiotensin IIintr) induces rat aorta contraction independent of plasma membrane Angiotensin II receptors. In this study we investigated the effects of Angiotensin IIintr on cell growth in A7r5 smooth muscle cells. 2 DNA-sy

  13. Functional preservation of vascular smooth muscle tissue

    Science.gov (United States)

    Alexander, W. C.; Hutchins, P. M.; Kimzey, S. L.

    1973-01-01

    The ionic and cellular feedback relationships operating to effect the vascular decompensatory modifications were examined to reveal procedures for implementing protective measures guarding against vascular collapse when returning from a weightless environment to that of the earth's gravity. The surgical procedures for preparing the rat cremaster, and the fixation methods are described. Abstracts of publications resulting from this research are included.

  14. Piperine Congeners as Inhibitors of Vascular Smooth Muscle Cell Proliferation.

    Science.gov (United States)

    Mair, Christina E; Liu, Rongxia; Atanasov, Atanas G; Wimmer, Laurin; Nemetz-Fiedler, Daniel; Sider, Nadine; Heiss, Elke H; Mihovilovic, Marko D; Dirsch, Verena M; Rollinger, Judith M

    2015-08-01

    Successful vascular healing after percutaneous coronary interventions is related to the inhibition of abnormal vascular smooth muscle cell proliferation and efficient re-endothelialization. In the search for vascular smooth muscle cell anti-proliferative agents from natural sources we identified piperine (1), the main pungent constituent of the fruits from Piper nigrum (black pepper). Piperine inhibited vascular smooth muscle cell proliferation with an IC50 of 21.6 µM, as quantified by a resazurin conversion assay. Investigations of ten piperamides isolated from black pepper fruits and 15 synthesized piperine derivatives resulted in the identification of three potent vascular smooth muscle cell proliferation inhibitors: the natural alkaloid pipertipine (4), and the two synthetic derivatives (2E,4E)-N,N-dibutyl-5-(3,5-dimethoxyphenyl)penta-2,4-dienamide (14) and (E)-N,N-dibutyl-3-(naphtho[2,3-d][1,3]dioxol-5-yl)acrylamide (20). They showed IC50 values of 3.38, 6.00, and 7.85 µM, respectively. Furthermore, the synthetic compound (2E,4E)-5-(4-fluorophenyl)-1-(piperidin-1-yl)penta-2,4-dien-1-one (12) was found to be cell type selective, by inhibiting vascular smooth muscle cell proliferation with an IC50 of 11.8 µM without influencing the growth of human endothelial cells. PMID:26132851

  15. Hypoxia, vascular smooth muscles and endothelium

    Directory of Open Access Journals (Sweden)

    Calvin K. Chan

    2013-02-01

    Full Text Available Hypoxia, or the lack of oxygen, has multiple impacts on the vascular system. The major molecular sensors for hypoxia at the cellular level are hypoxia inducible factor and heme oxygenase. Hypoxia also acts on the vasculature directly conveying its damaging effects through disruption of the control of vascular tone, particularly in the coronary circulation, enhancement of inflammatory responses and activation of coagulation pathways. These effects could be particularly detrimental under pathological conditions such as obstructive sleep apnea and other breathing disorders.

  16. Advance in molecular imaging research of vascular smooth muscle cells in the vascular diseases

    International Nuclear Information System (INIS)

    Vascular smooth muscle cells (VSMCs) are the primary cells within the vascular wall structure and maintain the tension of blood vessels, playing a key role in the restenosis, atherosclerosis and some other vascular diseases. With the development of molecular imaging, VSMCs cellular level of imaging studies is becoming more and more attention. The phenotype modulation, proliferation, migration and molecular imaging research progress of VSMCs in pathologic state were reviewed, to improve the management of vascular restenosis and atherosclerosis. (authors)

  17. [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.

  18. Monomethylarsonous acid, but not inorganic arsenic, is a mitochondria-specific toxicant in vascular smooth muscle cells.

    Science.gov (United States)

    Pace, Clare; Banerjee, Tania Das; Welch, Barrett; Khalili, Roxana; Dagda, Ruben K; Angermann, Jeff

    2016-09-01

    Arsenic exposure has been implicated as a risk factor for cardiovascular diseases, metabolic disorders, and cancer, yet the role mitochondrial dysfunction plays in the cellular mechanisms of pathology is largely unknown. To investigate arsenic-induced mitochondrial dysfunction in vascular smooth muscle cells (VSMCs), we exposed rat aortic smooth muscle cells (A7r5) to inorganic arsenic (iAs(III)) and its metabolite monomethylarsonous acid (MMA(III)) and compared their effects on mitochondrial function and oxidative stress. Our results indicate that MMA(III) is significantly more toxic to mitochondria than iAs(III). Exposure of VSMCs to MMA(III), but not iAs(III), significantly decreased basal and maximal oxygen consumption rates and concomitantly increased compensatory extracellular acidification rates, a proxy for glycolysis. Treatment with MMA(III) significantly increased hydrogen peroxide and superoxide levels compared to iAs(III). Exposure to MMA(III) resulted in significant decreases in mitochondrial ATP, aberrant perinuclear clustering of mitochondria, and decreased mitochondrial content. Mechanistically, we observed that mitochondrial superoxide and hydrogen peroxide contribute to mitochondrial toxicity, as treatment of cells with MnTBAP (a mitochondrial superoxide dismutase mimetic) and catalase significantly reduced mitochondrial respiration deficits and cell death induced by both arsenic compounds. Overall, our data demonstrates that MMA(III) is a mitochondria-specific toxicant that elevates mitochondrial and non-mitochondrial sources of ROS. PMID:27327130

  19. Monomethylarsonous acid, but not inorganic arsenic, is a mitochondria-specific toxicant in vascular smooth muscle cells.

    Science.gov (United States)

    Pace, Clare; Banerjee, Tania Das; Welch, Barrett; Khalili, Roxana; Dagda, Ruben K; Angermann, Jeff

    2016-09-01

    Arsenic exposure has been implicated as a risk factor for cardiovascular diseases, metabolic disorders, and cancer, yet the role mitochondrial dysfunction plays in the cellular mechanisms of pathology is largely unknown. To investigate arsenic-induced mitochondrial dysfunction in vascular smooth muscle cells (VSMCs), we exposed rat aortic smooth muscle cells (A7r5) to inorganic arsenic (iAs(III)) and its metabolite monomethylarsonous acid (MMA(III)) and compared their effects on mitochondrial function and oxidative stress. Our results indicate that MMA(III) is significantly more toxic to mitochondria than iAs(III). Exposure of VSMCs to MMA(III), but not iAs(III), significantly decreased basal and maximal oxygen consumption rates and concomitantly increased compensatory extracellular acidification rates, a proxy for glycolysis. Treatment with MMA(III) significantly increased hydrogen peroxide and superoxide levels compared to iAs(III). Exposure to MMA(III) resulted in significant decreases in mitochondrial ATP, aberrant perinuclear clustering of mitochondria, and decreased mitochondrial content. Mechanistically, we observed that mitochondrial superoxide and hydrogen peroxide contribute to mitochondrial toxicity, as treatment of cells with MnTBAP (a mitochondrial superoxide dismutase mimetic) and catalase significantly reduced mitochondrial respiration deficits and cell death induced by both arsenic compounds. Overall, our data demonstrates that MMA(III) is a mitochondria-specific toxicant that elevates mitochondrial and non-mitochondrial sources of ROS.

  20. Vasopressin-stimulated Ca2+ spiking in vascular smooth muscle cells involves phospholipase D.

    Science.gov (United States)

    Li, Y; Shiels, A J; Maszak, G; Byron, K L

    2001-06-01

    Physiological concentrations of [Arg(8)]vasopressin (AVP; 10-500 pM) stimulate oscillations of cytosolic free Ca2+ concentration (Ca2+ spikes) in A7r5 vascular smooth muscle cells. We previously reported that this effect of AVP was blocked by a putative phospholipase A2 (PLA2) inhibitor, ONO-RS-082 (5 microM). In the present study, the products of PLA2, arachidonic acid (AA), and lysophospholipids were found to be ineffective in stimulating Ca2+ spiking, and inhibitors of AA metabolism did not prevent AVP-stimulated Ca2+ spiking. Thin layer chromatography was used to monitor the release of AA and phosphatidic acid (PA), which are the products of PLA2 and phospholipase D (PLD), respectively. AVP (100 pM) stimulated both AA and PA formation, but only PA formation was inhibited by ONO-RS-082 (5 microM). Exogenous PLD (type VII; 2.5 U/ml) stimulated Ca2+ spiking equivalent to the effect of 100 pM AVP. AVP stimulated transphosphatidylation of 1-butanol (a PLD-catalyzed reaction) but not 2-butanol, and 1-butanol (but not 2-butanol) completely prevented AVP-stimulated Ca2+ spiking. Protein kinase C (PKC) inhibition, which completely prevents AVP-stimulated Ca2+ spiking, did not inhibit AVP-stimulated phosphatidylbutanol formation. These results suggest that AVP-stimulated Ca2+ spiking depends on activation of PLD rather than PLA2 and that PKC activation may be downstream of PLD in the signaling cascade.

  1. Crk-associated substrate, vascular smooth muscle and hypertension

    Institute of Scientific and Technical Information of China (English)

    Dale D. TANG

    2008-01-01

    Hypertension is characterized by vascular smooth muscle constriction and vascular remodeling involving cell migration, hypertrophy and growth. Crk-associated substrate (CAS), the first discovered member of the adapter protein CAS family, has been shown to be a critical cellular component that regulates various smooth muscle functions. In this review, the molecular structure and protein interactions of the CAS family members are summarized. Evidence for the role of CAS in the regu-lation of vascular smooth muscle contractility is pre-sented. Contraction stimulation induces CAS phosphor-ylation on Tyr-410 in arterial smooth muscle, creating the binding site for the Src homology (SH) 2/SH3 protein Crkll, which activates neuronal Wiskott-Aldrich syn-drome protein (N-WASP)-mediated actin assembly and force development. The functions of CAS in cell migra-tion, hypertrophy and growth are also summarized. Abelson tyrosine kinase (Abl), c-Src, focal adhesion kinase (FAK), proline-rich tyrosine kinase 2 (PYK2), pro-tein tyrosine phosphatase-proline, glutamate, serine and threonine sequence protein (PTP-PEST) and SHP-2 have been documented to coordinate the phosphorylation and dephosphorylation of CAS. The downstream signaling partners of CAS in the context of cell motility, hyper-trophy, survival and growth are also discussed. These new findings establish the important role of CAS in the modulation of vascular smooth muscle functions. Furthermore, the upstream regulators of CAS may be new biologic targets for the development of more effective and specific treatment of cardiovascular diseases such as hypertension.

  2. 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

  3. Poly(ethylene glycol-cholesterol inhibits L-type Ca2+ channel currents and augments voltage-dependent inactivation in A7r5 cells.

    Directory of Open Access Journals (Sweden)

    Rikuo Ochi

    Full Text Available Cholesterol distributes at a high density in the membrane lipid raft and modulates ion channel currents. Poly(ethylene glycol cholesteryl ether (PEG-cholesterol is a nonionic amphipathic lipid consisting of lipophilic cholesterol and covalently bound hydrophilic PEG. PEG-cholesterol is used to formulate lipoplexes to transfect cultured cells, and liposomes for encapsulated drug delivery. PEG-cholesterol is dissolved in the external leaflet of the lipid bilayer, and expands it to flatten the caveolae and widen the gap between the two leaflets. We studied the effect of PEG-cholesterol on whole cell L-type Ca(2+ channel currents (I(Ca,L recorded from cultured A7r5 arterial smooth muscle cells. The pretreatment of cells with PEG-cholesterol decreased the density of ICa,L and augmented the voltage-dependent inactivation with acceleration of time course of inactivation and negative shift of steady-state inactivation curve. Methyl-β-cyclodextrin (MβCD is a cholesterol-binding oligosaccharide. The enrichment of cholesterol by the MβCD:cholesterol complex (cholesterol (MβCD caused inhibition of I(Ca,L but did not augment voltage-dependent inactivation. Incubation with MβCD increased I(Ca,L, slowed the time course of inactivation and shifted the inactivation curve to a positive direction. Additional pretreatment by a high concentration of MβCD of the cells initially pretreated with PEG-cholesterol, increased I(Ca,L to a greater level than the control, and removed the augmented voltage-dependent inactivation. Due to the enhancement of the voltage-dependent inactivation, PEG-cholesterol inhibited window I(Ca,L more strongly as compared with cholesterol (MβCD. Poly(ethylene glycol conferred to cholesterol the efficacy to induce sustained augmentation of voltage-dependent inactivation of I(Ca,L.

  4. IP3 receptors regulate vascular smooth muscle contractility and hypertension

    Science.gov (United States)

    Lin, Qingsong; Zhao, Guiling; Fang, Xi; Peng, Xiaohong; Tang, Huayuan; Wang, Hong; Jing, Ran; Liu, Jie; Ouyang, Kunfu

    2016-01-01

    Inositol 1, 4, 5-trisphosphate receptor–mediated (IP3R-mediated) calcium (Ca2+) release has been proposed to play an important role in regulating vascular smooth muscle cell (VSMC) contraction for decades. However, whether and how IP3R regulates blood pressure in vivo remains unclear. To address these questions, we have generated a smooth muscle–specific IP3R triple-knockout (smTKO) mouse model using a tamoxifen-inducible system. In this study, the role of IP3R-mediated Ca2+ release in adult VSMCs on aortic vascular contractility and blood pressure was assessed following tamoxifen induction. We demonstrated that deletion of IP3Rs significantly reduced aortic contractile responses to vasoconstrictors, including phenylephrine, U46619, serotonin, and endothelin 1. Deletion of IP3Rs also dramatically reduced the phosphorylation of MLC20 and MYPT1 induced by U46619. Furthermore, although the basal blood pressure of smTKO mice remained similar to that of wild-type controls, the increase in systolic blood pressure upon chronic infusion of angiotensin II was significantly attenuated in smTKO mice. Taken together, our results demonstrate an important role for IP3R-mediated Ca2+ release in VSMCs in regulating vascular contractility and hypertension.

  5. Receptor for Advanced Glycation End-Products Signaling Interferes with the Vascular Smooth Muscle Cell Contractile Phenotype and Function.

    Directory of Open Access Journals (Sweden)

    Elie Simard

    Full Text Available Increased blood glucose concentrations promote reactions between glucose and proteins to form advanced glycation end-products (AGE. Circulating AGE in the blood plasma can activate the receptor for advanced end-products (RAGE, which is present on both endothelial and vascular smooth muscle cells (VSMC. RAGE exhibits a complex signaling that involves small G-proteins and mitogen activated protein kinases (MAPK, which lead to increased nuclear factor kappa B (NF-κB activity. While RAGE signaling has been previously addressed in endothelial cells, little is known regarding its impact on the function of VSMC. Therefore, we hypothesized that RAGE signaling leads to alterations in the mechanical and functional properties of VSMC, which could contribute to complications associated with diabetes. We demonstrated that RAGE is expressed and functional in the A7r5 VSMC model, and its activation by AGE significantly increased NF-κB activity, which is known to interfere with the contractile phenotype of VSMC. The protein levels of the contraction-related transcription factor myocardin were also decreased by RAGE activation with a concomitant decrease in the mRNA and protein levels of transgelin (SM-22α, a regulator of VSMC contraction. Interestingly, we demonstrated that RAGE activation increased the overall cell rigidity, an effect that can be related to an increase in myosin activity. Finally, although RAGE stimulation amplified calcium signaling and slightly myosin activity in VSMC challenged with vasopressin, their contractile capacity was negatively affected. Overall, RAGE activation in VSMC could represent a keystone in the development of vascular diseases associated with diabetes by interfering with the contractile phenotype of VSMC through the modification of their mechanical and functional properties.

  6. In vitro differentiation of porcine aortic vascular precursor cells to endothelial and vascular smooth muscle cells.

    Science.gov (United States)

    Zaniboni, Andrea; Bernardini, Chiara; Bertocchi, Martina; Zannoni, Augusta; Bianchi, Francesca; Avallone, Giancarlo; Mangano, Chiara; Sarli, Giuseppe; Calzà, Laura; Bacci, Maria Laura; Forni, Monica

    2015-09-01

    Recent findings suggest that progenitor and multipotent mesenchymal stromal cells (MSCs) are associated with vascular niches. Cells displaying mesenchymal properties and differentiating to whole components of a functional blood vessel, including endothelial and smooth muscle cells, can be defined as vascular stem cells (VSCs). Recently, we isolated a population of porcine aortic vascular precursor cells (pAVPCs), which have MSC- and pericyte-like properties. The aim of the present work was to investigate whether pAVPCs possess VSC-like properties and assess their differentiation potential toward endothelial and smooth muscle lineages. pAVPCs, maintained in a specific pericyte growth medium, were cultured in high-glucose DMEM + 10% FBS (long-term medium, LTM) or in human endothelial serum-free medium + 5% FBS and 50 ng/ml of hVEGF (endothelial differentiation medium, EDM). After 21 days of culture in LTM, pAVPCs showed an elongated fibroblast-like morphology, and they seem to organize in cord-like structures. qPCR analysis of smooth muscle markers [α-smooth muscle actin (α-SMA), calponin, and smooth muscle myosin (SMM) heavy chain] showed a significant increment of the transcripts, and immunofluorescence analysis confirmed the presence of α-SMA and SMM proteins. After 21 days of culture in EDM, pAVPCs displayed an endothelial cell-like morphology and revealed the upregulation of the expression of endothelial markers (CD31, vascular endothelial-cadherin, von Willebrand factor, and endothelial nitric oxide synthase) showing the CD31-typical pattern. In conclusion, pAVPCs could be defined as a VSC-like population considering that, if they are maintained in a specific pericyte medium, they express MSC markers, and they have, in addition to the classical mesenchymal trilineage differentiation potential, the capacity to differentiate in vitro toward the smooth muscle and the endothelial cell phenotypes. PMID:26135800

  7. Simulated Hypergravity Alters Vascular Smooth Muscle Cell Proliferation and Motility

    Science.gov (United States)

    Hunt, Shameka; Bettis, Barika; Harris-Hooker, Sandra; Sanford, Gary L.

    1997-01-01

    The cellular effects of gravity are poorly understood due to its constancy and nonavailability of altered gravitational models. Such an understanding is crucial for prolonged space flights. In these studies, we assessed the influence of centrifugation at 6G (HGrav) on vascular smooth muscle (SMC) mobility and proliferation. Cells were: (a) plated at low density and subjected to HGrav for 24-72 hr for proliferation studies, or (b) grown to confluency, subjected to HGrav, mechanically denuded and monitored for cell movement into the denuded area. Controls were maintained under normogravity. SMC showed a 50% inhibition of growth under HGrav and 10% serum; HGrav and low serum resulted in greater growth inhibition. The rate of movement of SMC into the denuded area was 2-3-fold higher under HGrav in low serum compared to controls, but similar in 10% serum. These studies show that HGrav has significant effects on SMC growth and mobility, which are dependent on serum levels.

  8. Calcium oscillations in human mesenteric vascular smooth muscle.

    Science.gov (United States)

    Navarro-Dorado, Jorge; Garcia-Alonso, Mauricio; van Breemen, Cornelis; Tejerina, Teresa; Fameli, Nicola

    2014-02-28

    Phenylephrine (PE)-induced oscillatory fluctuations in intracellular Ca(2+) concentration ([Ca(2+)]i) of vascular smooth muscle have been observed in many blood vessels isolated from a wide variety of mammals. Paradoxically, until recently similar observations in humans have proven elusive. In this study, we report for the first time observations of adrenergically-stimulated [Ca(2+)]i oscillations in human mesenteric artery smooth muscle. In arterial segments preloaded with Fluo-4 AM and mounted on a myograph on the stage of a confocal microscope, we observed PE-induced oscillations in [Ca(2+)]i, which initiated and maintained vasoconstriction. These oscillations present some variability, possibly due to compromised health of the tissue. This view is corroborated by our ultrastructural analysis of the cells, in which we found only (5 ± 2)% plasma membrane-sarcoplasmic reticulum apposition, markedly less than measured in healthy tissue from laboratory animals. We also partially characterized the oscillations by using the inhibitory drugs 2-aminoethoxydiphenyl borate (2-APB), cyclopiazonic acid (CPA) and nifedipine. After PE contraction, all drugs provoked relaxation of the vessel segments, sometimes only partial, and reduced or inhibited oscillations, except CPA, which rarely caused relaxation. These preliminary results point to a potential involvement of the sarcoplasmic reticulum Ca(2+) and inositol 1,4,5-trisphosphate receptor (IP3R) in the maintenance of the Ca(2+) oscillations observed in human blood vessels. PMID:24508261

  9. Arteriolar vascular smooth muscle cells: mechanotransducers in a complex environment.

    Science.gov (United States)

    Hill, Michael A; Meininger, Gerald A

    2012-09-01

    Contraction of small artery (diameters typically less than 250 μm) vascular smooth muscle cells (VSMCs) plays a critical role in local control of blood flow and arterial pressure through its affect on vascular caliber. Specifically, contraction of small arteries in response to increased intraluminal pressure is referred to as the myogenic response and represents an important role for mechanotransduction. Critical questions remain as to how changes in pressure are sensed by VSMCs and transduced across the cell membrane to tune the contractile state of the cell. Recent studies suggest a pivotal role for interactions between VSMCs and extracellular matrix (ECM) proteins. Thus, pressure-induced deformation of ECM proteins and their cell surface receptors (for example, integrins) may initiate contraction and cytoskeletal remodeling through modulation of ion channels, membrane depolarization, increased intracellular Ca(2+) and actomyosin crossbridge cycling. Importantly, it is argued that the contractile properties of small artery VSMCs reflect an intimate and integrated interaction with their extracellular environment and the three-dimensional structure of the vessel wall. PMID:22677491

  10. Calmodulin kinase II is required for angiotensin II-mediated vascular smooth muscle hypertrophy

    OpenAIRE

    Li, Hui; Li, Weiwei; Arun K Gupta; Mohler, Peter J.; Anderson, Mark E.; Grumbach, Isabella M.

    2009-01-01

    Despite our understanding that medial smooth muscle hypertrophy is a central feature of vascular remodeling, the molecular pathways underlying this pathology are still not well understood. Work over the past decade has illustrated a potential role for the multifunctional calmodulin-dependent kinase CaMKII in smooth muscle cell contraction, growth, and migration. Here we demonstrate that CaMKII is enriched in vascular smooth muscle (VSM) and that CaMKII inhibition blocks ANG II-dependent VSM c...

  11. Molecular mechanisms of decreased smooth muscle differentiation marker expression after vascular injury

    OpenAIRE

    Regan, Christopher P.; Adam, Paul J.; Madsen, Cort S.; Owens, Gary K.

    2000-01-01

    While it is well established that phenotypic modulation of vascular smooth muscle cells (VSMCs) contributes to the development and progression of vascular lesions, little is known regarding the molecular mechanisms of phenotypic modulation in vivo. Here we show that vascular injury reduces transcription of VSMC differentiation marker genes, and we identify cis regulatory elements that may mediate this decrease. Using a carotid wire-injury model in mice carrying transgenes for smooth muscle α-...

  12. Pasteur effect in vascular and intestinal smooth muscle.

    Science.gov (United States)

    Pettersson, G; Lundholm, L

    1985-01-01

    The increase in lactate production on changing from aerobic to anaerobic conditions, i.e. the Pasteur effect, has been reported to be small in vascular muscle and especially in aorta. It has been suggested that this may be an artefact caused by damage to the intimal endothelium. We have compared the Pasteur effect in different kinds of pig arteries, but also in rabbit colon. The aerobic lactate production in 60 min was 11-15 mumol/g in the aorta and the carotid artery, but 3 mumol/g in the mesenteric and renal arteries and 4 mumol/g in the rabbit colon. The increase in lactate production under anaerobic conditions was 12-20 mumol/g/60 min in the carotid artery, aorta and rabbit colon and 10 mumol/g/60 min in the mesenteric and renal arteries. When calculated in per cent, the Pasteur effect was greater in the mesenteric artery than in the aorta, but the actual rise in lactate production in mumol/g was higher in the aorta and carotid artery. The high aerobic lactate production of smooth muscle in vitro may be related to its low ability to oxidize glucose; some other substrates may be preferentially oxidized when present in vitro or in vivo.

  13. 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.

  14. 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) cel

  15. 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.

  16. An In Vitro Murine Model of Vascular Smooth Muscle Cell Mineralization.

    Science.gov (United States)

    Kelynack, Kristen J; Holt, Stephen G

    2016-01-01

    Vascular calcification (VC) is seen ubiquitously in aging blood vessels and prematurely in disease states like renal failure. It is thought to be driven by a number of systemic and local factors that lead to extra-osseous deposition of mineral in the vascular wall and valves as a common endpoint. The response of resident vascular smooth muscle cell to these dystrophic signals appears to be important in this process. Whilst in vivo models allow the observation of global changes in a pro-calcific environment, identifying the specific cells and mechanisms involved has been largely garnered from in vitro experiments, which provide added benefits in terms of reproducibility, cost, and convenience. Here we describe a 7-21 day cell culture model of calcification developed using immortalized murine vascular smooth muscle cells (MOVAS-1). This model provides a method by which vascular smooth muscle cell involvement and manipulation within a mineralizing domain can be studied.

  17. The effects of TSH on human vascular endothelial cells and smooth muscle cells

    Institute of Scientific and Technical Information of China (English)

    田利民

    2014-01-01

    Objective To study the effect of thyroid-stimulating hormone(TSH)on human vascular endothelial cells and smooth muscle cells and to explore the roles of TSH in the development of atherosclerosis.Methods Human vascular endothelial cells and smooth muscle cells were cultured in vitro.MTT method was used to assay the effect of TSH on cell viability.Real-time PCR was used

  18. Embryonic origins of human vascular smooth muscle cells: implications for in vitro modeling and clinical application

    OpenAIRE

    Sinha, Sanjay; Iyer, Dharini; Granata, Alessandra

    2014-01-01

    Vascular smooth muscle cells (SMCs) arise from multiple origins during development, raising the possibility that differences in embryological origins between SMCs could contribute to site-specific localization of vascular diseases. In this review, we first examine the developmental pathways and embryological origins of vascular SMCs and then discuss in vitro strategies for deriving SMCs from human embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs). We then review in detail...

  19. Sphingosine induces phospholipase D and mitogen activated protein kinase in vascular smooth muscle cells.

    Science.gov (United States)

    Taher, M M; Abd-Elfattah, A S; Sholley, M M

    1998-12-01

    The enzymes phospholipase D and diacylglycerol kinase generate phosphatidic acid which is considered to be a mitogen. Here we report that sphingosine produced a significant amount of phosphatidic acid in vascular smooth muscle cells from the rat aorta. The diacylglycerol kinase inhibitor R59 949 partially depressed sphingosine induced phosphatidic acid formation, suggesting that activation of phospholipase C and diacylglycerol kinase can not account for the bulk of phosphatidic acid produced and that additional pathways such as phospholipase D may contribute to this. Further, we have shown that phosphatidylethanol was produced by sphingosine when vascular smooth muscle cells were stimulated in the presence of ethanol. Finally, as previously shown for other cell types, sphingosine stimulated mitogen-activated protein kinase in vascular smooth muscle cells.

  20. Doxycycline Alters Vascular Smooth Muscle Cell Adhesion, Migration, and Reorganization of Fibrillar Collagen Matrices

    OpenAIRE

    Franco, Christopher; Ho, Bernard; Mulholland, Diane; Hou, Guangpei; Islam, Muzharul; Donaldson, Katey; Bendeck, Michelle Patricia

    2006-01-01

    Remodeling of injured blood vessels is dependent on smooth muscle cells and matrix metalloproteinase activity. Doxycycline is a broad spectrum matrix metalloproteinase inhibitor that is under investigation for the treatment of acute coronary syndromes and aneurysms. In the present study, we examine the mechanisms by which doxycycline inhibits smooth muscle cell responses using a series of in vitro assays that mimic critical steps in pathological vascular remodeling. Doxycycline treatment dram...

  1. Endothelin converting enzyme (ECE) activity in human vascular smooth muscle

    OpenAIRE

    Maguire, Janet J.; Johnson, Christopher M.; Mockridge, James W; Davenport, Anthony P

    1997-01-01

    We have characterized the human smooth muscle endothelin converting enzyme (ECE) present in the media of the endothelium-denuded human umbilical vein preparation.Endothelin-1 (ET-1) and ET-2 were potent constrictors of umbilical vein with EC50 values of 9.2 nM and 29.6 nM, respectively. ET-1 was at least 30 times more potent than ET-3 suggesting the presence of constrictor ETA receptors. Little or no response was obtained to the ETB-selective agonist sarafotoxin 6c. These data suggest that en...

  2. Resveratrol Induces Vascular Smooth Muscle Cell Differentiation through Stimulation of SirT1 and AMPK

    OpenAIRE

    Anne Marie Thompson; Martin, Kathleen A.; Rzucidlo, Eva M.

    2014-01-01

    Phenotypic plasticity in vascular smooth muscle cells (VSMC) is necessary for vessel maintenance, repair and adaptation to vascular changes associated with aging. De-differentiated VSMC contribute to pathologies including atherosclerosis and intimal hyperplasia. As resveratrol has been reported to have cardio- protective effects, we investigated its role in VSMC phenotypic modulation. We demonstrated the novel finding that resveratrol promoted VSMC differentiation as measured by contractile p...

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

    OpenAIRE

    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 viv...

  4. 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.

  5. Vascular smooth muscle cell spreading onto fibrinogen is regulated by calpains and phospholipase C.

    Science.gov (United States)

    Paulhe, F; Bogyo, A; Chap, H; Perret, B; Racaud-Sultan, C

    2001-11-01

    Fibrinogen deposition and smooth muscle cell migration are important causes of atherosclerosis and angiogenesis. Involvement of calpains in vascular smooth muscle cell adhesion onto fibrinogen was investigated. Using calpain inhibitors, we showed that activation of calpains was required for smooth muscle cell spreading. An increase of (32)P-labeled phosphatidic acid and phosphatidylinositol-3,4-bisphosphate, respective products of phospholipase C and phosphoinositide 3-kinase activities, was measured in adherent cells. Addition of the calpain inhibitor calpeptin strongly decreased phosphatidic acid and phosphatidylinositol-3,4-bisphosphate. However, smooth muscle cell spreading was prevented by the phospholipase C inhibitor U-73122, but poorly modified by phosphoinositide 3-kinase inhibitors wortmannin and LY-294002. Moreover, PLC was found to act upstream of the PI 3-kinase IA isoform. Thus, our data provide the first evidence that calpains are required for smooth muscle cell spreading. Further, phospholipase C activation is pointed as a key step of cell-spreading regulation by calpains.

  6. MFAP4 Promotes Vascular Smooth Muscle Migration, Proliferation and Accelerates Neointima Formation

    DEFF Research Database (Denmark)

    Schlosser, Anders; Pilecki, Bartosz; Hemstra, Line E.;

    2016-01-01

    OBJECTIVE: Arterial injury stimulates remodeling responses that, when excessive, lead to stenosis. These responses are influenced by integrin signaling in vascular smooth muscle cells (VSMCs). Microfibrillar-associated protein 4 (MFAP4) is an integrin ligand localized to extracellular matrix fibe...

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

    Science.gov (United States)

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

    2016-02-01

    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. 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.

  8. Cinematographic analysis of vascular smooth muscle cell interactions with extracellular matrix.

    Science.gov (United States)

    Absher, M; Baldor, L

    1991-01-01

    The interactions of vascular smooth muscle cells with growth modulators and extracellular matrix molecules may play a role in the proliferation and migration of these cells after vascular injury and during the development of atherosclerosis. Time-lapse cinematographic techniques have been used to study cell division and migration of bovine carotid artery smooth muscle cells in response to matrix molecules consisting of solubilized basement membrane (Matrigel) and type I collagen. When cells were grown adjacent to Matrigel, both migration and cell proliferation were increased and interdivision time was shortened. Cells grown in Matrigel or in type I collagen had markedly reduced migration rates but interdivision time was not altered. Further, diffusible components of the Matrigel were found to stimulate proliferation of the smooth muscle cells.

  9. Azelnidipine inhibits Msx2-dependent osteogenic differentiation and matrix mineralization of vascular smooth muscle cells.

    Science.gov (United States)

    Shimizu, Takehisa; Tanaka, Toru; Iso, Tatsuya; Kawai-Kowase, Keiko; Kurabayashi, Masahiko

    2012-01-01

    Vascular calcification is an active and regulated process that is similar to bone formation. While calcium channel blockers (CCBs) have been shown to improve outcomes in atherosclerotic vascular disease, it remains unknown whether CCBs have an effect on the process of vascular calcification. Here we investigated whether CCBs inhibit osteogenic differentiation and matrix mineralization of vascular smooth muscle cells induced by Msx2, a key factor of vascular calcification. Human aortic smooth muscle cells (HASMCs) were transduced with adenovirus expressing MSX2 and were treated with 3 distinct CCBs. Azelnidipine, a dihydropyridine subclass of CCBs, significantly decreased alkaline phosphatase (ALP) activity of Msx2-overexpressed HASMCs, whereas verapamil and diltiazem had no effect. Furthermore, azelnidipine, but not verapamil and diltiazem, significantly decreased matrix mineralization of Msx2-overexpressing HASMCs. Azelnidipine significantly attenuated the induction of ALP gene expression by Msx2, a key transcription factor in osteogenesis, while it did not reduce enzymatic activity of ALP. Furthermore, azelnidipine inhibited the ability of Msx2 to activate the ALP gene, but had no effect on Notch-induced Msx2 expression. Given that L-type calcium channels are equally blocked by these CCBs, our results suggest that azelnidipine inhibits the Msx2-dependent process of vascular calcification by mechanisms other than inhibition of calcium channel activity.

  10. Paraoxon Attenuates Vascular Smooth Muscle Contraction through Inhibiting Ca2+ Influx in the Rabbit Thoracic Aorta

    Directory of Open Access Journals (Sweden)

    Shouhong Zhou

    2010-01-01

    Full Text Available We investigated the effect of paraoxon on vascular contractility using organ baths in thoracic aortic rings of rabbits and examined the effect of paraoxon on calcium homeostasis using a whole-cell patch-clamp technique in isolated aortic smooth muscle cells of rabbits. The findings show that administration of paraoxon (30 μM attenuated thoracic aorta contraction induced by phenylephrine (1 μM and/or a high K+ environment (80 mM in both the presence and absence of thoracic aortic endothelium. This inhibitory effect of paraoxon on vasoconstrictor-induced contraction was abolished in the absence of extracellular Ca2+, or in the presence of the Ca2+ channel inhibitor, verapamil. But atropine had little effect on the inhibitory effect of paraoxon on phenylephrine-induced contraction. Paraoxon also attenuated vascular smooth muscle contraction induced by the cumulative addition of CaCl2 and attenuated an increase of intracellular Ca2+ concentration induced by K+ in vascular smooth muscle cells. Moreover, paraoxon (30 μM inhibited significantly L-type calcium current in isolated aortic smooth muscle cells of rabbits. In conclusion, our results demonstrate that paraoxon attenuates vasoconstrictor-induced contraction through inhibiting Ca2+ influx in the rabbits thoracic aorta.

  11. Embryonic origins of human vascular smooth muscle cells: implications for in vitro modeling and clinical application.

    Science.gov (United States)

    Sinha, Sanjay; Iyer, Dharini; Granata, Alessandra

    2014-06-01

    Vascular smooth muscle cells (SMCs) arise from multiple origins during development, raising the possibility that differences in embryological origins between SMCs could contribute to site-specific localization of vascular diseases. In this review, we first examine the developmental pathways and embryological origins of vascular SMCs and then discuss in vitro strategies for deriving SMCs from human embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs). We then review in detail the potential for vascular disease modeling using iPSC-derived SMCs and consider the pathological implications of heterogeneous embryonic origins. Finally, we touch upon the role of human ESC-derived SMCs in therapeutic revascularization and the challenges remaining before regenerative medicine using ESC- or iPSC-derived cells comes of age.

  12. 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.

  13. Transmembrane Protein 184A Is a Receptor Required for Vascular Smooth Muscle Cell Responses to Heparin.

    Science.gov (United States)

    Pugh, Raymond J; Slee, Joshua B; Farwell, Sara Lynn N; Li, Yaqiu; Barthol, Trista; Patton, Walter A; Lowe-Krentz, Linda J

    2016-03-01

    Vascular cell responses to exogenous heparin have been documented to include decreased vascular smooth muscle cell proliferation following decreased ERK pathway signaling. However, the molecular mechanism(s) by which heparin interacts with cells to induce those responses has remained unclear. Previously characterized monoclonal antibodies that block heparin binding to vascular cells have been found to mimic heparin effects. In this study, those antibodies were employed to isolate a heparin binding protein. MALDI mass spectrometry data provide evidence that the protein isolated is transmembrane protein 184A (TMEM184A). Commercial antibodies against three separate regions of the TMEM184A human protein were used to identify the TMEM184A protein in vascular smooth muscle cells and endothelial cells. A GFP-TMEM184A construct was employed to determine colocalization with heparin after endocytosis. Knockdown of TMEM184A eliminated the physiological responses to heparin, including effects on ERK pathway activity and BrdU incorporation. Isolated GFP-TMEM184A binds heparin, and overexpression results in additional heparin uptake. Together, these data support the identification of TMEM184A as a heparin receptor in vascular cells.

  14. Generation of vascular endothelial and smooth muscle cells from human pluripotent stem cells

    OpenAIRE

    Patsch, Christoph; Challet-Meylan, Ludivine; Eva C Thoma; Urich, Eduard; Heckel, Tobias; O’Sullivan, John F.; Grainger, Stephanie J.; Kapp, Friedrich G.; Sun, Lin; Christensen, Klaus; Xia, Yulei; Florido, Mary H. C.; He, Wei; Pan, Wei; Prummer, Michael

    2015-01-01

    The use of human pluripotent stem cells for in vitro disease modeling and clinical applications requires protocols that convert these cells into relevant adult cell types. Here, we report the rapid and efficient differentiation of human pluripotent stem cells into vascular endothelial and smooth muscle cells. We found that GSK3 inhibition and BMP4 treatment rapidly committed pluripotent cells to a mesodermal fate and subsequent exposure to VEGF or PDGF-BB resulted in the differentiation of ei...

  15. Antagonism by lipophilic quaternary ions of the K+ channel opener, levcromakalim, in vascular smooth muscle.

    OpenAIRE

    McPherson, G. A.; Piekarska, A. E.

    1994-01-01

    1. The aim of this study was to characterize the interaction between the K+ channel opener levcromakalim (LKM) and several quaternary ions, in vascular smooth muscle, in vitro. Segments of isolated, thoracic aorta of the rat were suspended in organ baths filled with Krebs solution at 37 degrees C. Cumulative concentration-response curves to LKM were obtained in the absence and in the presence of increasing concentrations of quaternary ions using a number of agents to pre-constrict the vessel....

  16. Vascular smooth muscle cell-derived adiponectin: a paracrine regulator of contractile phenotype

    OpenAIRE

    Ding, Min; Carrao, Ana Catarina; Wagner, Robert J.; Xie, Yi; Jin, Yu; Rzucidlo, Eva M.; Yu, Jun; Li, Wei; Tellides, George; Hwa, John; Aprahamian, Tamar R.; Martin, Kathleen A.

    2011-01-01

    Adiponectin is a cardioprotective adipokine derived predominantly from visceral fat. We recently demonstrated that exogenous adiponectin induces vascular smooth muscle cell (VSMC) differentiation via repression of mTORC1 and FoxO4. Here we report for the first time that VSMC express and secrete adiponectin, which acts in an autocrine and paracrine manner to regulate VSMC contractile phenotype. Adiponectin was found to be expressed in human coronary artery and mouse aortic VSMC. Importantly, s...

  17. miR-599 Inhibits Vascular Smooth Muscle Cells Proliferation and Migration by Targeting TGFB2

    OpenAIRE

    Baodong Xie; Chunfeng Zhang; Kai Kang; Shulin Jiang

    2015-01-01

    Aberrant proliferation and migration of vascular smooth muscle cells (VSMCs) play a crucial role in the pathogenesis of cardiovascular diseases including coronary heart disease, restenosis and atherosclerosis. MicroRNAs are a class of small, non-coding and endogenous RNAs that play critical roles in VSMCs function. In this study, we showed that PDGF-bb, as a stimulant, promoted VSMCs proliferation and suppressed the expression of miR-599. Moreover, overexpression of miR-599 inhibited VSMCs pr...

  18. Smooth Muscle Cell Mineralocorticoid Receptors Are Mandatory for Aldosterone–Salt to Induce Vascular Stiffness

    OpenAIRE

    Galmiche, Guillaume; Pizard, Anne; Gueret, Alexandre; El Moghrabi, Soumaya; Ouvrard-Pascaud, Antoine; Berger, Stefan; Challande, Pascal; Jaffe, Iris Z.; Labat, Carlos; Lacolley, Patrick; Jaisser, Frédéric

    2013-01-01

    Arterial stiffness is recognized as a risk factor for many cardiovascular diseases. Aldosterone via its binding to and activation of the mineralocorticoid receptors (MRs) is a main regulator of blood pressure by controlling renal sodium reabsorption. Although both clinical and experimental data indicate that MR activation by aldosterone is involved in arterial stiffening, the molecular mechanism is not known. In addition to the kidney, MR is expressed in both endothelial and vascular smooth m...

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

    OpenAIRE

    Son, Dong Ju; Kim, Soo Yeon; Han, Seong Su; Kim, Chan Woo; Kumar, Sandeep; Park, Byeoung Soo; Lee, Sung Eun; Yun, Yeo Pyo; Jo, Hanjoong; Park, Young Hyun

    2012-01-01

    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 significant...

  20. 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...

  1. The vascular smooth muscle alpha-actin gene is reactivated during cardiac hypertrophy provoked by load.

    OpenAIRE

    Black, F M; Packer, S E; Parker, T G; Michael, L H; Roberts, R; R J Schwartz; Schneider, M D

    1991-01-01

    Cardiac hypertrophy triggered by mechanical load possesses features in common with growth factor signal transduction. A hemodynamic load provokes rapid expression of the growth factor-inducible nuclear oncogene, c-fos, and certain peptide growth factors specifically stimulate the "fetal" cardiac genes associated with hypertrophy, even in the absence of load. These include the gene encoding vascular smooth muscle alpha-actin, the earliest alpha-actin expressed during cardiac myogenesis; howeve...

  2. The Fat1 cadherin integrates vascular smooth muscle cell growth and migration signals

    OpenAIRE

    Hou, Rong; Liu, Liming; Anees, Syed; Hiroyasu, Shungo; Sibinga, Nicholas E. S.

    2006-01-01

    The significance of cadherin superfamily proteins in vascular smooth muscle cell (VSMC) biology is undefined. Here we describe recent studies of the Fat1 protocadherin. Fat1 expression in VSMCs increases significantly after arterial injury or growth factor stimulation. Fat1 knockdown decreases VSMC migration in vitro, but surprisingly, enhances cyclin D1 expression and proliferation. Despite limited similarity to classical cadherins, the Fat1 intracellular domain (Fat1IC) interacts with β-cat...

  3. 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.

  4. 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.

  5. Matrix Metalloproteinase 2 as a Potential Mediator of Vascular Smooth Muscle Cell Migration and Chronic Vascular Remodeling in Hypertension.

    Science.gov (United States)

    Belo, V A; Guimarães, Danielle A; Castro, Michele Mazzaron

    2015-01-01

    For vascular remodeling in hypertension, it is essential that vascular smooth muscle cells (VSMCs) reshape in order to proliferate and migrate. The extracellular matrix (ECM) needs to be degraded to favor VSMC migration. Many proteases, including matrix metalloproteinases (MMPs), contribute to ECM proteolysis and VSMC migration. Bioactive peptides, hemodynamic forces and reactive oxygen-nitrogen species regulate MMP-2 expression and activity. Increased MMP-2 activity contributes to hypertension-induced maladaptive arterial changes and sustained hypertension. New ECM is synthesized to supply VSMCs with bioactive mediators, which stimulate hypertrophy. MMP-2 stimulates the interaction of VSMCs with newly formed ECM, which triggers intracellular signaling via integrins to induce a phenotypic switch and persistent migration. VSMCs switch from a contractile to a synthetic phenotype in order to migrate and contribute to vascular remodeling in hypertension. MMPs also disrupt growth factors bound to ECM, thus contributing to their capacity to regulate VSMC migration. This review sheds light on the proteolytic effects of MMP-2 on ECM and non-ECM substrates in the vasculature and how these effects contribute to VSMC migration in hypertension. The inhibition of MMP activity as a therapeutic target may make it possible to reduce arterial maladaptation caused by hypertension and prevent the resulting fatal cardiovascular events. PMID:26731549

  6. Human vascular smooth muscle cells both express and respond to heparin-binding growth factor I (endothelial cell growth factor).

    OpenAIRE

    Winkles, J A; Friesel, R; Burgess, W H; Howk, R; Mehlman, T; Weinstein, R.; T. MACIAG

    1987-01-01

    The control of vascular endothelial and smooth muscle cell proliferation is important in such processes as tumor angiogenesis, wound healing, and the pathogenesis of atherosclerosis. Class I heparin-binding growth factor (HBGF-I) is a potent mitogen and chemoattractant for human endothelial cells in vitro and will induce angiogenesis in vivo. RNA gel blot hybridization experiments demonstrate that cultured human vascular smooth muscle cells, but not human umbilical vein endothelial cells, exp...

  7. Rosiglitzone Suppresses Angiotensin II-Induced Production of KLF5 and Cell Proliferation in Rat Vascular Smooth Muscle Cells

    OpenAIRE

    Gao, Dengfeng; Hao, Guanghua; Meng, Zhe; Ning, Ning; Yang, Guang; Liu, Zhongwei; Dong, Xin; Niu, Xiaolin

    2015-01-01

    Krüppel-like factor (KLF) 5, which initiates vascular smooth muscle cell (VSMC) proliferation, also participates in Angiotensin (Ang) II-induced vascular remodeling. The protective effect of rosiglitazone on vascular remodeling may be due to their impact on VSMC proliferation. However, the underlying mechanisms involved remain unclear. This study was designed to investigate whether the antiproliferation effects of rosiglitazone are mediated by regulating Ang II/KLF5 response. We found that, i...

  8. Impaired Peroxisome Proliferator-activated Receptor-γ Contributes to Phenotypic Modulation of Vascular Smooth Muscle Cells during Hypertension*

    OpenAIRE

    Zhang, Lili; Xie, Peng; Wang, Jingzhou; Yang, Qingwu; Fang, Chuanqin; Zhou, Shuang; Li, Jingcheng

    2010-01-01

    The phenotypic modulation of vascular smooth muscle cells (VSMCs) plays a pivotal role in hypertension-induced vascular changes including vascular remodeling. The precise mechanisms underlying VSMC phenotypic modulation remain elusive. Here we test the role of peroxisome proliferator-activated receptor (PPAR)-γ in the VSMC phenotypic modulation during hypertension. Both spontaneously hypertensive rat (SHR) aortas and SHR-derived VSMCs exhibited reduced PPAR-γ expression and excessive VSMC phe...

  9. Intercellular ultrafast Ca(2+) wave in vascular smooth muscle cells: numerical and experimental study.

    Science.gov (United States)

    Quijano, J C; Raynaud, F; Nguyen, D; Piacentini, N; Meister, J J

    2016-01-01

    Vascular smooth muscle cells exhibit intercellular Ca(2+) waves in response to local mechanical or KCl stimulation. Recently, a new type of intercellular Ca(2+) wave was observed in vitro in a linear arrangement of smooth muscle cells. The intercellular wave was denominated ultrafast Ca(2+) wave and it was suggested to be the result of the interplay between membrane potential and Ca(2+) dynamics which depended on influx of extracellular Ca(2+), cell membrane depolarization and its intercel- lular propagation. In the present study we measured experimentally the conduction velocity of the membrane depolarization and performed simulations of the ultrafast Ca(2+) wave along coupled smooth muscle cells. Numerical results reproduced a wide spectrum of experimental observations, including Ca(2+) wave velocity, electrotonic membrane depolarization along the network, effects of inhibitors and independence of the Ca(2+) wave speed on the intracellular stores. The numerical data also provided new physiological insights suggesting ranges of crucial model parameters that may be altered experimentally and that could significantly affect wave kinetics allowing the modulation of the wave characteristics experimentally. Numerical and experimental results supported the hypothesis that the propagation of membrane depolarization acts as an intercellular messenger mediating intercellular ultrafast Ca(2+) waves in smooth muscle cells. PMID:27507785

  10. Intercellular ultrafast Ca2+ wave in vascular smooth muscle cells: numerical and experimental study

    Science.gov (United States)

    Quijano, J. C.; Raynaud, F.; Nguyen, D.; Piacentini, N.; Meister, J. J.

    2016-01-01

    Vascular smooth muscle cells exhibit intercellular Ca2+ waves in response to local mechanical or KCl stimulation. Recently, a new type of intercellular Ca2+ wave was observed in vitro in a linear arrangement of smooth muscle cells. The intercellular wave was denominated ultrafast Ca2+ wave and it was suggested to be the result of the interplay between membrane potential and Ca2+ dynamics which depended on influx of extracellular Ca2+, cell membrane depolarization and its intercel- lular propagation. In the present study we measured experimentally the conduction velocity of the membrane depolarization and performed simulations of the ultrafast Ca2+ wave along coupled smooth muscle cells. Numerical results reproduced a wide spectrum of experimental observations, including Ca2+ wave velocity, electrotonic membrane depolarization along the network, effects of inhibitors and independence of the Ca2+ wave speed on the intracellular stores. The numerical data also provided new physiological insights suggesting ranges of crucial model parameters that may be altered experimentally and that could significantly affect wave kinetics allowing the modulation of the wave characteristics experimentally. Numerical and experimental results supported the hypothesis that the propagation of membrane depolarization acts as an intercellular messenger mediating intercellular ultrafast Ca2+ waves in smooth muscle cells. PMID:27507785

  11. Cytotoxicity of some oxysterols on human vascular smooth muscle cells was mediated by apoptosis.

    Science.gov (United States)

    Miyashita, Y; Shirai, K; Ito, Y; Watanabe, J; Urano, Y; Murano, T; Tomioka, H

    1997-01-01

    A decrease in smooth muscle cells is observed in advanced atherosclerotic lesion. To understand this mechanism, we selected oxysterols as candidates for toxic lipid, and examined their cytotoxicity on human cultured vascular smooth muscle cells, together with the manner of cell death. In the presence of 7-ketocholesterol or 7 beta-hydroxycholesterol (50 mumol/L), the percentage of detached cells increased significantly with dose dependency, and an increase in detached cell number and DNA nick detected by terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling study (TUNEL) preceded an increase in lactate dehydrogenase released into the medium. DNA extracted from smooth muscle cells incubated with 7-ketocholesterol or 7 beta-hydroxycholesterol showed a laddering pattern on agarose electrophoresis. In the presence of 7-ketocholesterol or 7 beta-hydroxycholesterol, fragmented DNA quantified by the quantitative sandwich enzyme immunoassay was significantly increased. From these results, it is proposed that 7-ketocholesterol and 7 beta-hydroxycholesterol are toxic to smooth muscle cells, and that this cytotoxicity is mediated by apoptosis. PMID:9638517

  12. 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

    ) 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......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...

  13. 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

    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......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....../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...

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

    OpenAIRE

    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-pol...

  15. 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.

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

    International Nuclear Information System (INIS)

    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 3H-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

  17. Activating transcription factor-4 promotes mineralization in vascular smooth muscle cells

    Science.gov (United States)

    Masuda, Masashi; Miyazaki-Anzai, Shinobu; Keenan, Audrey L.; Shiozaki, Yuji; Okamura, Kayo; Chick, Wallace S.; Williams, Kristina; Zhao, Xiaoyun; Rahman, Shaikh Mizanoor; Tintut, Yin; Adams, Christopher M.

    2016-01-01

    Emerging evidence indicates that upregulation of the ER stress–induced pro-osteogenic transcription factor ATF4 plays an important role in vascular calcification, a common complication in patients with aging, diabetes, and chronic kidney disease (CKD). In this study, we demonstrated the pathophysiological role of ATF4 in vascular calcification using global Atf4 KO, smooth muscle cell–specific (SMC-specific) Atf4 KO, and transgenic (TG) mouse models. Reduced expression of ATF4 in global ATF4-haplodeficient and SMC-specific Atf4 KO mice reduced medial and atherosclerotic calcification under normal kidney and CKD conditions. In contrast, increased expression of ATF4 in SMC-specific Atf4 TG mice caused severe medial and atherosclerotic calcification. We further demonstrated that ATF4 transcriptionally upregulates the expression of type III sodium-dependent phosphate cotransporters (PiT1 and PiT2) by interacting with C/EBPβ. These results demonstrate that the ER stress effector ATF4 plays a critical role in the pathogenesis of vascular calcification through increased phosphate uptake in vascular SMCs. PMID:27812542

  18. 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.

  19. Increased expression of osteoprotegerin in vascular smooth muscle cells from spontaneously hypertensive rats

    Institute of Scientific and Technical Information of China (English)

    Yongshan MOU; Tianhua LEI; Luning ZHAO; Xiaojun ZHU; Mingui FU; Yuqing E CHEN

    2004-01-01

    Background Osteoprotegerin (OPG) is a secreted protein of the tumor necrosis factor receptor family, which regulates bone mass by inhibiting osteoclast differentiation and activation. Although OPG is expressed ubiquitously and abundantly in many tissues and cell types including vascular cells, the role of OPG in other tissues is unknown.Our previous studies demonstrated that OPG was highly expressed in vascular smooth muscle cells (VSMC) and upregulated during vascular lesion formation. Methods and Results We documented, by Northern blot analysis,that the expression of OPG was more prevalent in the aorta and cultured VSMC from spontaneously hypertensive rats (SI-IR) compared to Wistar-Kyoto rats (WKY). In addition, we found that the expression of Angiotensin II (Ang II)type I receptor (AT1R) in SHR VSMC was at significantly increased levels than in WKY VSMC. Furthermore, Ang II potently induced the expression of OPG in VSMC in a time- and dose-dependent manner through the AT1R signaling pathway. Conclusions OPG expression was substantially greater in SHR VSMC, suggesting that OPG may be an important determinant of vascular remodeling in SHR.

  20. Effect of Caspase Inhibitor Ac-DEVD-CHO on Apoptosis of Vascular Smooth Muscle Cells Induced by Artesunate

    Directory of Open Access Journals (Sweden)

    Jingwen Zhang

    2014-05-01

    Full Text Available Numerous studies have shown that the proliferation and apoptosis of vascular smooth muscle cells play a key role in restenosis. Artesunate is a triterpenoid with a peroxide structure and its antimalarial, antitumor, and antiangiogenetic activities can inhibit the proliferation and apoptosis of multifarious cells. Apoptosis is caused by the activation of a series of intracellular proteolytic enzymes, among which caspase-dependent apoptosis was the earliest to be recognized. The purpose of this article is to study the effects of caspase-3 inhibitor Ac-DEVD-CHO on proliferation and apoptosis of vascular smooth muscle cells induced by Artesunate and to explore the mechanism of Artesunate-induced apoptosis of vascular smooth muscle cells. By using the method based on methyl thiazolyl tetrazolium to observe the effects of Artesunate on the growth and proliferation of vascular smooth muscle cells; observing the change in cell shape before and after Artesunate administration by transmission electron microscopy; detecting the changes in cell cycle and apoptosis rates before and after drug administration by flow cytometry; detecting the activity of caspase-3 in the caspase apoptosis pathway by the Western Blot method, we found that Artesunate inhibits the growth and proliferation of vascular smooth muscle cells in a dose- and time-dependent manner within the concentration range of 7.5–120 μg/mL, and the inhibition rate of Artesunate can be as high as 89.49 % at a concentration of 120 μg/mL after acting for 72 hours; vascular smooth muscle cells show a typical apoptosis peak due to the effects of higher concentration of Artesunate. Compared with the control group, the higher-concentration group shows major variability, Ac-DEVD-CHO, however, can significantly decrease this induction; it has been detected by Western Blot that Artesunate can induce caspase-3 activity dramatically in vascular smooth muscle cells, but this activation may be remarkably

  1. Matrine inhibits the expression of adhesion molecules in activated vascular smooth muscle cells.

    Science.gov (United States)

    Liu, Jun; Zhang, Lihua; Ren, Yingang; Gao, Yanli; Kang, Li; Lu, Shaoping

    2016-03-01

    Atherosclerosis is a chronic inflammatory disease associated with increased expression of adhesion molecules in vascular smooth muscle cells (VSMCs). Matrine is a main active ingredient of Sophora flavescens roots, which are used to treat inflammatory diseases. However, the effects of matrine on the expression of adhesion molecules in VSMCs have largely remained elusive. Therefore, the present study investigated the effects of matrine on the expression of adhesion molecules in tumor necrosis factor (TNF)‑α‑stimulated human aortic smooth muscle cells (HASMCs). The results showed that matrine inhibited the expression of vascular cell adhesion molecule‑1 (VCAM‑1) and intercellular adhesion molecule‑1 (ICAM‑1) in TNF‑α‑stimulated HASMCs. Matrine markedly inhibited the TNF‑α‑induced expression of nuclear factor (NF)‑κB p65 and prevented the TNF‑α‑caused degradation of inhibitor of NF‑κB; it also inhibited TNF‑α‑induced activation of mitogen‑activated protein kinases (MAPKs). Furthermore, matrine inhibited the production of intracellular reactive oxygen species (ROS) in TNF‑α‑stimulated HASMCs. In conclusion, the results of the present study demonstrated that matrine inhibited the expression of VCAM‑1 and ICAM‑1 in TNF‑α‑stimulated HASMCs via the suppression of ROS production as well as NF‑κB and MAPK pathway activation. Therefore, matrine may have a potential therapeutic use for preventing the advancement of atherosclerotic lesions.

  2. 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.

  3. Activity of sap from Croton lechleri on rat vascular and gastric smooth muscles.

    Science.gov (United States)

    Froldi, G; Zagotto, G; Filippini, R; Montopoli, M; Dorigo, P; Caparrotta, L

    2009-08-01

    The effects of red sap from Croton lechleri (SdD), Euphorbiaceae, on vascular and gastric smooth muscles were investigated. SdD, from 10 to 1000 microg/ml, induced concentration-dependent vasoconstriction in rat caudal arteries, which was endothelium-independent. In arterial preparations pre-constricted by phenylephrine (0.1 microM) or KCl (30 mM), SdD also produced concentration-dependent vasoconstriction. To study the mechanisms implicated in this effect we used selective inhibitors such as prazosin (0.1 microM), an antagonist of alpha(1)-adrenoceptors, atropine (0.1 microM), an antagonist of muscarinic receptors, and ritanserin (50 nM), a 5-HT(2A) antagonist; none of these influenced vasoconstriction caused by SdD. Likewise, nifedipine (50 nM), an inhibitor of L-type calcium channels, did not modify the action of SdD. Capsaicin (100 nM), an agonist of vanilloid receptors, also did not affect vasoconstriction by SdD. We also investigated the action of SdD (10-1000 microg/ml) on rat gastric fundus; per se the sap slightly increased contractile tension. When the gastric fundus was pre-treated with SdD (100 microg/ml) the contraction induced by carbachol (1 microM) was increased, whereas that by KCl (60mM) or capsaicin (100 nM) were unchanged. The data shows that SdD increased contractile tension in a concentration-dependent way, both on vascular and gastric smooth muscles. The vasoconstriction is unrelated to alpha(1), M, 5-HT(2A) and vanilloid receptors as well as L-type calcium channels. SdD increased also contraction by carbachol on rat gastric fundus. Thus for the first time, experimental data provides evidence that sap from C. lechleri owns constricting activity on smooth muscles. PMID:19406630

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

    International Nuclear Information System (INIS)

    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 (Δψm) 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.

  5. 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.

  6. Calphostin-C induction of vascular smooth muscle cell apoptosis proceeds through phospholipase D and microtubule inhibition.

    Science.gov (United States)

    Zheng, Xi-Long; Gui, Yu; Du, Guangwei; Frohman, Michael A; Peng, Dao-Quan

    2004-02-20

    Calphostin-C, a protein kinase C inhibitor, induces apoptosis of cultured vascular smooth muscle cells. However, the mechanisms are not completely defined. Because apoptosis of vascular smooth muscle cells is critical in several proliferating vascular diseases such as atherosclerosis and restenosis after angioplasty, we decided to investigate the mechanisms underlying the calphostin-C-induced apoptotic pathway. We show here that apoptosis is inhibited by the addition of exogenous phosphatidic acid, a metabolite of phospholipase D (PLD), and that calphostin-C inhibits completely the activities of both isoforms of PLD, PLD1 and PLD2. Overexpression of either PLD1 or PLD2 prevented the vascular smooth muscle cell apoptosis induced by serum withdrawal but not the calphostin-C-elicited apoptosis. These data suggest that PLDs have anti-apoptotic effects and that complete inhibition of PLD activity by calphostin-C induces smooth muscle cell apoptosis. We also report that calphostin-C induced microtubule disruption and that the addition of exogenous phosphatidic acid inhibits calphostin-C effects on microtubules, suggesting a role for PLD in stabilizing the microtubule network. Overexpressing PLD2 in Chinese hamster ovary cells phenocopies this result, providing strong support for the hypothesis. Finally, taxol, a microtubule stabilizer, not only inhibited the calphostin-C-induced microtubule disruption but also inhibited apoptosis. We therefore conclude that calphostin-C induces apoptosis of cultured vascular smooth muscle cells through inhibiting PLD activity and subsequent microtubule polymerization. PMID:14660552

  7. Calphostin-C induction of vascular smooth muscle cell apoptosis proceeds through phospholipase D and microtubule inhibition.

    Science.gov (United States)

    Zheng, Xi-Long; Gui, Yu; Du, Guangwei; Frohman, Michael A; Peng, Dao-Quan

    2004-02-20

    Calphostin-C, a protein kinase C inhibitor, induces apoptosis of cultured vascular smooth muscle cells. However, the mechanisms are not completely defined. Because apoptosis of vascular smooth muscle cells is critical in several proliferating vascular diseases such as atherosclerosis and restenosis after angioplasty, we decided to investigate the mechanisms underlying the calphostin-C-induced apoptotic pathway. We show here that apoptosis is inhibited by the addition of exogenous phosphatidic acid, a metabolite of phospholipase D (PLD), and that calphostin-C inhibits completely the activities of both isoforms of PLD, PLD1 and PLD2. Overexpression of either PLD1 or PLD2 prevented the vascular smooth muscle cell apoptosis induced by serum withdrawal but not the calphostin-C-elicited apoptosis. These data suggest that PLDs have anti-apoptotic effects and that complete inhibition of PLD activity by calphostin-C induces smooth muscle cell apoptosis. We also report that calphostin-C induced microtubule disruption and that the addition of exogenous phosphatidic acid inhibits calphostin-C effects on microtubules, suggesting a role for PLD in stabilizing the microtubule network. Overexpressing PLD2 in Chinese hamster ovary cells phenocopies this result, providing strong support for the hypothesis. Finally, taxol, a microtubule stabilizer, not only inhibited the calphostin-C-induced microtubule disruption but also inhibited apoptosis. We therefore conclude that calphostin-C induces apoptosis of cultured vascular smooth muscle cells through inhibiting PLD activity and subsequent microtubule polymerization.

  8. Endoplasmic reticulum stress induced by Thapsigargin in vascular smooth muscle cells of rat coronary artery

    Institute of Scientific and Technical Information of China (English)

    CHEN Xiao-yan; DENG Chun-yu; JIANG Li

    2016-01-01

    AIM:To establish the endoplasmic reticulum stress ( ERS) cell model in vascular smooth muscle cells ( VSMCs) of Sprague-Dawley (SD) rats.METHODS:Under sterile condition, the coronary arteries were isolated from SD rats .The primary VSMCs were cultured by tissue-sticking method , and observed the basic morphological characteristics under optical microscope .The marker proteins of VSMCs including α-smooth muscle actin (α-SMA) and smooth muscle myosin heavy chain ( SM-MHC) were identified by immuno-fluorescence technique .VSMCs were treated with thapsigargin (0.5, 1 and 2 μmol/L) for 24 h, and the expression levels of binding immunoglobulin protein (BiP) and C/EBP homologus protein (CHOP), the marker molecules of ERS, were detected using Western blotting.RESULTS:VSMCs climbed out from coronary artery tissues after about six days , and the cells had a nice state and formed the VSMC-like typical "peak valley".The results of immunofluorescence technique show that the marker proteins of VSMCs ,α-SMA and SM-MHC were expressed significantly .The results of Western blotting show that the protein expression levels of BiP and CHOP were increased by thapsigargin in a dose-dependent manner .CONCLUSION:VSMCs can be successfully cultured by tissue-sticking method and built the ERS model induced by thapsigargin .

  9. Anti-Proliferative Effect of an Aqueous Extract of Prunella vulgaris in Vascular Smooth Muscle Cells

    Directory of Open Access Journals (Sweden)

    Sun Mi Hwang

    2013-01-01

    Full Text Available The abnormal proliferation of vascular smooth muscle cells (VSMCs in arterial walls is an important pathogenic factor of vascular disorders such as diabetic atherosclerosis. We have reported the anti-inflammatory effect of an aqueous extract from Prunella vulgaris (APV in vascular endothelial cell. In the present study, APV exhibited inhibitory effects on high glucose-stimulated VSMC proliferation, migration, and invasion activities, inducing G1 cell cycle arrest with downregulation of cyclins and CDKs and upregulation of the CKIs, p21waf1/cip1 and p27kip1. Furthermore, APV dose dependently suppressed the high glucose-induced matrix metalloproteinase activity. High glucose-induced phosphorylation of ERK, p38 MAPK, was decreased by the pretreatment of APV. NF-κB activation by high glucose was attenuated by APV, as an antioxidant. APV attenuated the high glucose-induced decrease of nuclear factor E2-related factor-2 (Nrf2 translocation and heme oxygenase-1 (HO-1 expression. Intracellular cGMP level was also increased by APV treatment. These results demonstrate that APV may inhibit VSMC proliferation via downregulating ROS/NF-κB /ERK/p38 MAPK pathways. In addition, APV has a beneficial effect by the interaction of Nrf2-mediated NO/cGMP with HO-1, suggesting that Prunella vulgaris may be useful in preventing diabetic atherosclerosis.

  10. Mesenchymal stem cells-derived vascular smooth muscle cells release abundant levels of osteoprotegerin

    Directory of Open Access Journals (Sweden)

    M Vaccarezza

    2009-03-01

    Full Text Available Although several studies have shown that the serum levels of osteoprotegerin (OPG are significantly elevated in patients affected with atherosclerotic lesions in coronary and peripheral arteries, the cellular source and the role of OPG in the physiopathology of atherosclerosis are not completely defined. Therefore, we aimed to investigate the potential contribution of mesenchymal stem cells in the production/release of OPG. OPG was detectable by immunohistochemistry in aortic and coronary atherosclerotic plaques, within or in proximity of intimal vascular smooth muscle cells (SMC. In addition, bone marrow mesenchymal stem cell (MSC-derived vascular SMC as well as primary aortic SMC released in the culture supernatant significantly higher levels of OPG with respect to MSCderived endothelial cells (EC or primary aortic EC. On the other hand, in vitro exposure to full-length human recombinant OPG significantly increased the proliferation rate of aortic SMC cultures, as monitored by bromodeoxyuridine incorporation. Taken together, these data suggest that OPG acts as an autocrine/paracrine growth factor for vascular SMC, which might contribute to the progression of atherosclerotic lesions.

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

    Science.gov (United States)

    Whitesell, Thomas R; Kennedy, Regan M; Carter, Alyson D; Rollins, Evvi-Lynn; Georgijevic, Sonja; Santoro, Massimo M; Childs, Sarah J

    2014-01-01

    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.

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

    Directory of Open Access Journals (Sweden)

    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.

  13. Unexpected role of the copper transporter ATP7A in PDGF-induced vascular smooth

    Energy Technology Data Exchange (ETDEWEB)

    Ashino, T.; Varadarajan, S.; Urao, N.; Oshikawa, J.; Chen, G. -F.; Wang, H.; Huo, Y.; Finney, L.; Vogt, S.; McKinney, R. D.; Maryon, E. B.; Kaplan, J. H.; Ushio-Fukai, M.; Fukai, T. (Biosciences Division); ( XSD); ( PSC-USR); (Univ. of Illinois at Chicago); (Univ. of Minnesota)

    2010-09-09

    Copper, an essential nutrient, has been implicated in vascular remodeling and atherosclerosis with unknown mechanism. Bioavailability of intracellular copper is regulated not only by the copper importer CTR1 (copper transporter 1) but also by the copper exporter ATP7A (Menkes ATPase), whose function is achieved through copper-dependent translocation from trans-Golgi network (TGN). Platelet-derived growth factor (PDGF) promotes vascular smooth muscle cell (VSMC) migration, a key component of neointimal formation. To determine the role of copper transporter ATP7A in PDGF-induced VSMC migration. Depletion of ATP7A inhibited VSMC migration in response to PDGF or wound scratch in a CTR1/copper-dependent manner. PDGF stimulation promoted ATP7A translocation from the TGN to lipid rafts, which localized at the leading edge, where it colocalized with PDGF receptor and Rac1, in migrating VSMCs. Mechanistically, ATP7A small interfering RNA or CTR small interfering RNA prevented PDGF-induced Rac1 translocation to the leading edge, thereby inhibiting lamellipodia formation. In addition, ATP7A depletion prevented a PDGF-induced decrease in copper level and secretory copper enzyme precursor prolysyl oxidase (Pro-LOX) in lipid raft fraction, as well as PDGF-induced increase in LOX activity. In vivo, ATP7A expression was markedly increased and copper accumulation was observed by synchrotron-based x-ray fluorescence microscopy at neointimal VSMCs in wire injury model. These findings suggest that ATP7A plays an important role in copper-dependent PDGF-stimulated VSMC migration via recruiting Rac1 to lipid rafts at the leading edge, as well as regulating LOX activity. This may contribute to neointimal formation after vascular injury. Our findings provide insight into ATP7A as a novel therapeutic target for vascular remodeling and atherosclerosis.

  14. AMPK induces vascular smooth muscle cell senescence via LKB1 dependent pathway

    Energy Technology Data Exchange (ETDEWEB)

    Sung, Jin Young; Woo, Chang-Hoon [Department of Pharmacology, College of Medicine, Yeungnam University, Daegu 705-717 (Korea, Republic of); Aging-associated Vascular Disease Research Center, College of Medicine, Yeungnam University, Daegu 705-717 (Korea, Republic of); Kang, Young Jin; Lee, Kwang Youn [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); Aging-associated Vascular Disease Research Center, College of Medicine, Yeungnam University, Daegu 705-717 (Korea, Republic of)

    2011-09-16

    Highlights: {yields} An aging model was established by stimulating VSMC with adriamycin. {yields} Adriamycin increased p-LKB1, p-AMPK, p53 and p21 expressions. {yields} Inhibition of AMPK diminished SA-{beta}-gal staining and restored VSMC proliferation. {yields} p53 and p21 siRNA attenuated adriamycin-induced SA-{beta}-gal staining in VSMC. {yields} p53-p21 pathway is a mediator of LKB1/AMPK induced VSMC senescence. -- Abstract: Vascular cells have a limited lifespan with limited cell proliferation and undergo cellular senescence. The functional changes associated with cellular senescence are thought to contribute to age-related vascular disorders. AMP-activated protein kinase (AMPK) has been discussed in terms of beneficial or harmful effects for aging-related diseases. However, the detailed functional mechanisms of AMPK are largely unclear. An aging model was established by stimulating vascular smooth muscle cell (VSMC) with adriamycin. Adriamycin progressively increased the mRNA and protein expressions of AMPK. The phosphorylation levels of LKB1 and acetyl-CoA carboxylase (ACC), the upstream and downstream of AMPK, were dramatically increased by adriamycin stimulation. The expressions of p53 and p21, which contribute to vascular senescence, were also increased. Inhibition of AMPK diminished senescence-associated {beta}-galactosidase (SA-{beta}-gal) staining, and restored VSMC proliferation. Cytosolic translocation of LKB1 by adriamycin could be a mechanism for AMPK activation in senescence. Furthermore, p53 siRNA and p21 siRNA transfection attenuated adriamycin-induced SA-{beta}-gal staining. These results suggest that LKB1 dependent AMPK activation elicits VSMC senescence and p53-p21 pathway is a mediator of LKB1/AMPK-induced senescence.

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

    International Nuclear Information System (INIS)

    Objective: To evaluate the influence of 103Pd 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 103Pd radioactive stent for preventing restenosis after angioplasty. Methods: Fifty male New Zealand rabbits were randomized into stent group and 103Pd 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 103Pd stent group was less than that of stent group. It was most obvious at the 56th day (P103Pd 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)% (P103Pd 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)% (P103Pd 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 103Pd 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 (P103Pd 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

  16. Human vascular smooth muscle cells and endothelial cells cocultured on polyglycolic acid (70/30) scaffold in tissue engineered vascular graft

    Institute of Scientific and Technical Information of China (English)

    WEN Shao-jun; ZHAO Li-min; WANG Shen-guo; LI Jing-xing; CHEN Hua-ying; LIU Jie-lin; LIU Ya; LUO Yi; Roo Changizi

    2007-01-01

    Background Current prosthetic, small diameter vascular grafts showing poor long term patency rates have led to the pursuit of other biological materials. Biomaterials that successfully integrate into surrounding tissue should match not only the mechanical properties of tissues, but also topography. Polyglycolic acid (70/30) has been used as synthetic grafts to determine whether human vascular smooth muscle cells and endothelial cells attach, survive and secrete endothelin and 6-keto-prostaglandin F1α (6-keto-PGF1α).Methods Endothelial cells and smooth muscle cells were isolated from adult human great saphenous vein. They were seeded on polyglycolic acid scaffold in vitro separately to grow vascular patch (Groups A and B respectively) and cocultured in vitro to grow into vascular patch (Group C). Smooth muscle cells and endothelial cells were identified by immunohistochemical analysis and growth of cells on polyglycolic acid was investigated using scanning electron microscopy. The levels of endothelin and 6-keto-PGF1α in the culturing solutions were examined by radioimmunology to measure endothelial function.Results Seed smooth muscle cells adhered to polyglycolic acid scaffold and over 28 days grew in the interstices to form a uniform cell distribution throughout the scaffold. Then seed endothelial cells formed a complete endothelial layer on the smooth muscle cells. The levels of endothelin and 6-keto-prostaglandin F1 alpha in the culturing solution were (234±29) pg/ml and (428+98) pg/ml respectively in Group C and (196+30) pg/ml and (346±120) pg/ml in Group B; both significantly higher than in Groups A and D (blank control group, all P<0.05 ).Conclusions Cells could be grown successfully on polyglycolic acid and retain functions of secretion. Our next step is to use human saphenous vein smooth muscle cells and endothelial cells to grow tubular vascular grafts in vitro.

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

    International Nuclear Information System (INIS)

    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 Ca2+ and then causes an increasing in the inner calcium concentration [Ca2]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 Emax 1 Gy maxc 3 Gy max 5 Gy max 7 Gy. There is an important shift on irradiated rats (3, 5, 7 Gy) where the maximum effect of Acetylcholine (Emax) can be obtained in lower concentrations of Acetylcholine. These results mean that irradiation activates the inward flux of Ca2+ 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. (author)

  18. Involvement of phospholipase D in store-operated calcium influx in vascular smooth muscle cells.

    Science.gov (United States)

    Walter, M; Tepel, M; Nofer, J R; Neusser, M; Assmann, G; Zidek, W

    2000-08-11

    In non-excitable cells, sustained intracellular Ca2+ increase critically depends on influx of extracellular Ca2+. Such Ca2+ influx is thought to occur by a 'store-operated' mechanism, i.e. the signal for Ca2+ entry is believed to result from the initial release of Ca2+ from inositol 1,4,5-trisphosphate-sensitive intracellular stores. Here we show that the depletion of cellular Ca2+ stores by thapsigargin or bradykinin is functionally linked to a phosphoinositide-specific phospholipase D (PLD) activity in cultured vascular smooth muscle cells (VSMC), and that phosphatidic acid formed via PLD enhances sustained calcium entry in this cell type. These results suggest a regulatory role for PLD in store-operated Ca2+ entry in VSMC.

  19. STUDY ON THE INHIBITORY EFFECT OF ANTISENSE ETAR OLIGODEOXYNUCLEOTIDES ON THE PROLIFERATION OF VASCULAR SMOOTH CELLS

    Institute of Scientific and Technical Information of China (English)

    张岚; 张柏根; 张纪蔚; 钱济先; 张皓; 黄晓钟

    2002-01-01

    Objective To study the inhibitory effect of antisense endothelin receptor A (ETAR) on the proliferation of the vascular smooth muscle cells. Methods The sense, antisense and mismatched ODNs for ETAR were designed and synthetized. The study was carried out using MTT method and binding assays.Results ETAR-ODNs could move successfully across VSMC membranes. Photo-absorption in the MTT test was reduced significantly (P<0.05) in the antisense group at 5μmol/L; the reduction of CPM also occurred in the 125I-ET-1 specific binding assay; and the sense and mismatched ODNs groups did not show this reduction. Conclusion Our study suggested that the antisense oligomers inhibited the proliferation of VSMCs by hindering the translation of target mRNA and by reducing the production of related protein.

  20. Local electromechanical properties of different phenotype models of vascular smooth muscle cells using force microscopy

    Science.gov (United States)

    Thompson, Gary; Reukov, Vladimir; Nikiforov, Maxim; Guo, Senli; Ovchinnikov, Oleg; Jesse, Stephen; Kalinin, Sergei; Vertegel, Alexey

    2010-03-01

    Vascular smooth muscle cells (VSMCs) exist as a spectrum of diverse phenotypes raning between contractile and synthetic, the latter being associated with disease states. Different VSMC phenotypes, modeled using serum-starvation, exhibit characteristic electromechanical responses that can be distinguished using band excitation piezoresponse force microscopy (BEPFM), which maps information at the same rate as the atomic force microscope (AFM) scan performed simultaneously. BEPFM image formation mechanism in the culture medium is determined using excitation steps from 1 mV to 100 V. High voltage improves contrast between cells and collagen-coated substrates. Viscoelasticity from AFM stress relaxation experiments and local elasticity from force maps correlate to BEPFM data providing a map of local mechanical properties on different VSMCs.

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

    International Nuclear Information System (INIS)

    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 PDGFBB (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

  2. Mechanisms of Vascular Smooth Muscle NADPH oxidase 1 (Nox1) Contribution to Injury-induced Neointimal Formation

    OpenAIRE

    Lee, M. Y.; San Martin, Alejandra; Mehta, P. K.; Dikalova, A. E.; Garrido, A. M.; Datla, S. R.; Lyons, Erin; Krause, Karl-Heinz; Banfi, Botond; Lambeth, J D; Lassegue, Bernard; Griendling, K. K.

    2009-01-01

    OBJECTIVE: Vascular NADPH oxidases (Noxes) have been implicated in cardiovascular diseases; however, the importance of individual Nox homologues remains unclear. Here, the role of the vascular smooth muscle cell (VSMC) Nox1 in neointima formation was studied using genetically modified animal models. METHODS AND RESULTS: Wire injury-induced neointima formation in the femoral artery, along with proliferation and apoptosis, was reduced in Nox1(y/-) mice, but there was little difference in Tg(SMC...

  3. Differential effects of formoterol on thrombin- and PDGF-induced proliferation of human pulmonary arterial vascular smooth muscle cells

    OpenAIRE

    Goncharova Elena A; Khavin Irene S; Goncharov Dmitry A; Krymskaya Vera P

    2012-01-01

    Abstract Background Increased pulmonary arterial vascular smooth muscle (PAVSM) cell proliferation is a key pathophysiological component of pulmonary vascular remodeling in pulmonary arterial hypertension (PH). The long-acting β2-adrenergic receptor (β2AR) agonist formoterol, a racemate comprised of (R,R)- and (S,S)-enantiomers, is commonly used as a vasodilator in chronic obstructive pulmonary disease (COPD). PH, a common complication of COPD, increases patients’ morbidity and reduces surviv...

  4. Differential effects of formoterol on thrombin- and PDGF-induced proliferation of human pulmonary arterial vascular smooth muscle cells

    OpenAIRE

    Goncharova, Elena A.; Khavin, Irene S; Goncharov, Dmitry A; Vera P Krymskaya

    2012-01-01

    Background Increased pulmonary arterial vascular smooth muscle (PAVSM) cell proliferation is a key pathophysiological component of pulmonary vascular remodeling in pulmonary arterial hypertension (PH). The long-acting β2-adrenergic receptor (β2AR) agonist formoterol, a racemate comprised of (R,R)- and (S,S)-enantiomers, is commonly used as a vasodilator in chronic obstructive pulmonary disease (COPD). PH, a common complication of COPD, increases patients’ morbidity and reduces survival. Recen...

  5. Pharmacological inhibition of PHOSPHO1 suppresses vascular smooth muscle cell calcification.

    Science.gov (United States)

    Kiffer-Moreira, Tina; Yadav, Manisha C; Zhu, Dongxing; Narisawa, Sonoko; Sheen, Campbell; Stec, Boguslaw; Cosford, Nicholas D; Dahl, Russell; Farquharson, Colin; Hoylaerts, Marc F; Macrae, Vicky E; Millán, José Luis

    2013-01-01

    Medial vascular calcification (MVC) is common in patients with chronic kidney disease, obesity, and aging. MVC is an actively regulated process that resembles skeletal mineralization, resulting from chondro-osteogenic transformation of vascular smooth muscle cells (VSMCs). Here, we used mineralizing murine VSMCs to study the expression of PHOSPHO1, a phosphatase that participates in the first step of matrix vesicles-mediated initiation of mineralization during endochondral ossification. Wild-type (WT) VSMCs cultured under calcifying conditions exhibited increased Phospho1 gene expression and Phospho1(-/-) VSMCs failed to mineralize in vitro. Using natural PHOSPHO1 substrates, potent and specific inhibitors of PHOSPHO1 were identified via high-throughput screening and mechanistic analysis and two of these inhibitors, designated MLS-0390838 and MLS-0263839, were selected for further analysis. Their effectiveness in preventing VSMC calcification by targeting PHOSPHO1 function was assessed, alone and in combination with a potent tissue-nonspecific alkaline phosphatase (TNAP) inhibitor MLS-0038949. PHOSPHO1 inhibition by MLS-0263839 in mineralizing WT cells (cultured with added inorganic phosphate) reduced calcification in culture to 41.8% ± 2.0% of control. Combined inhibition of PHOSPHO1 by MLS-0263839 and TNAP by MLS-0038949 significantly reduced calcification to 20.9% ± 0.74% of control. Furthermore, the dual inhibition strategy affected the expression of several mineralization-related enzymes while increasing expression of the smooth muscle cell marker Acta2. We conclude that PHOSPHO1 plays a critical role in VSMC mineralization and that "phosphatase inhibition" may be a useful therapeutic strategy to reduce MVC.

  6. 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.

  7. In vascular smooth muscle cells paricalcitol prevents phosphate-induced Wnt/β-catenin activation.

    Science.gov (United States)

    Martínez-Moreno, Julio M; Muñoz-Castañeda, Juan R; Herencia, Carmen; Oca, Addy Montes de; Estepa, Jose C; Canalejo, Rocio; Rodríguez-Ortiz, Maria E; Perez-Martinez, Pablo; Aguilera-Tejero, Escolástico; Canalejo, Antonio; Rodríguez, Mariano; Almadén, Yolanda

    2012-10-15

    The present study investigates the differential effect of two vitamin D receptor agonists, calcitriol and paricalcitol, on human aortic smooth muscle cells calcification in vitro. Human vascular smooth muscle cells were incubated in a high phosphate (HP) medium alone or supplemented with either calcitriol 10(-8)M (HP + CTR) or paricalcitol 3·10(-8) M (HP + PC). HP medium induced calcification, which was associated with the upregulation of mRNA expression of osteogenic factors such as bone morphogenetic protein 2 (BMP2), Runx2/Cbfa1, Msx2, and osteocalcin. In these cells, activation of Wnt/β-catenin signaling was evidenced by the translocation of β-catenin into the nucleus and the increase in the expression of direct target genes as cyclin D1, axin 2, and VCAN/versican. Addition of calcitriol to HP medium (HP + CTR) further increased calcification and also enhanced the expression of osteogenic factors together with a significant elevation of nuclear β-catenin levels and the expression of cyclin D1, axin 2, and VCAN. By contrast, the addition of paricalcitol (HP + PC) not only reduced calcification but also downregulated the expression of BMP2 and other osteoblastic phenotype markers as well as the levels of nuclear β-catenin and the expression of its target genes. The role of Wnt/β-catenin on phosphate- and calcitriol-induced calcification was further demonstrated by the inhibition of calcification after addition of Dickkopf-related protein 1 (DKK-1), a specific natural antagonist of the Wnt/β-catenin signaling pathway. In conclusion, the differential effect of calcitriol and paricalcitol on vascular calcification appears to be mediated by a distinct regulation of the BMP and Wnt/β-catenin signaling pathways.

  8. Sustained diacylglycerol formation from inositol phospholipids in angiotensin II-stimulated vascular smooth muscle cells

    Energy Technology Data Exchange (ETDEWEB)

    Griendling, K.K.; Rittenhouse, S.E.; Brock, T.A.; Ekstein, L.S.; Gimbrone, M.A. Jr.; Alexander, R.W.

    1986-05-05

    Angiotensin II acts on cultured rat aortic vascular smooth muscle cells to stimulate phospholipase C-mediated hydrolysis of membrane phosphoinositides and subsequent formation of diacylglycerol and inositol phosphates. In intact cells, angiotensin II induces a dose-dependent increase in diglyceride which is detectable after 5 s and sustained for at least 20 min. Angiotensin II (100 nM)-stimulated diglyceride formation is biphasic, peaking at 15 s (227 +/- 19% control) and at 5 min (303 +/- 23% control). Simultaneous analysis of labeled inositol phospholipids shows that at 15 s phosphatidylinositol 4,5-bisphosphate (PIP2) and phosphatidylinositol 4-phosphate (PIP) decline to 52 +/- 6% control and 63 +/- 5% control, respectively, while phosphatidylinositol (PI) remains unchanged. In contrast, at 5 min, PIP2 and PIP have returned toward control levels (92 +/- 2 and 82 +/- 4% control, respectively), while PI has decreased substantially (81 +/- 2% control). The calcium ionophore ionomycin (15 microM) stimulates diglyceride accumulation but does not cause PI hydrolysis. 4 beta-Phorbol 12-myristate 13-acetate, an activator of protein kinase C, inhibits early PIP and PIP2 breakdown and diglyceride formation, without inhibiting late-phase diglyceride accumulation. Thus, angiotensin II induces rapid transient breakdown of PIP and PIP2 and delayed hydrolysis of PI. The rapid attenuation of polyphosphoinositide breakdown is likely caused by a protein kinase C-mediated inhibition of PIP and PIP2 hydrolysis. While in vascular smooth muscle stimulated with angiotensin II inositol 1,4,5-trisphosphate formation is transient, diglyceride production is biphasic, suggesting that initial and sustained diglyceride formation from the phosphoinositides results from different biochemical and/or cellular processes.

  9. Assays for in vitro monitoring of proliferation of human airway smooth muscle (ASM) and human pulmonary arterial vascular smooth muscle (VSM) cells.

    Science.gov (United States)

    Goncharova, Elena A; Lim, Poay; Goncharov, Dmitry A; Eszterhas, Andrew; Panettieri, Reynold A; Krymskaya, Vera P

    2006-01-01

    Vascular and airway remodeling, which are characterized by airway smooth muscle (ASM) and pulmonary arterial vascular smooth muscle (VSM) proliferation, contribute to the pathology of asthma, pulmonary hypertension, restenosis and atherosclerosis. To evaluate the proliferation of VSM and ASM cells in response to mitogens, we perform a [3H]thymidine incorporation assay. The proliferation protocol takes approximately 48 h and includes stimulating cells synchronized in G0/G1 phase of the cell cycle with agonists, labeling cells with [3H]thymidine and examining levels of [3H]thymidine incorporation by scintillation counting. Although using radiolabeled [3H]thymidine incorporation is a limitation, the greatest benefit of the assay is providing reliable and statistically significant data. PMID:17406550

  10. Insulin induces PKC-dependent proliferation of mesenteric vascular smooth muscle cells from hypertensive patients

    Institute of Scientific and Technical Information of China (English)

    Xukai WANG; Yan WANG; Chenming YANG; Ying WAN; Xianwen JI

    2006-01-01

    Background and objectives Proliferation of human vascular smooth muscle cells (VSMCs) induced by hyperinsulinemia is a very common clinical pathology. Extensive research has focused on PKC (Protein kinase C)-MAPK (mitogen-activated protein kinase)intracellular signal transduction and the phenotypic modulation accompanied by reorganization of intracellular F-actins in VSMCs.Methods DNA synthesis, signaling of ERK1/2 MAPKs, and changes in α-smooth muscle (SM) actin and F-actin were studied in hypertensive and normotensive human arterial VSMCs exposed to insulin and PMA with and without the PKC inhibitor, GF109203X.Results Differences among cell types in MAPK signaling, α-SM actin, and F-actin isoforms in VSMCs harvested from the arteries of patients with essential hypertension (EH) and normotension (NT) were identified in response to insulin treatment. Proliferation and activation of MAPK were more pronounced in EH VSMCs than in NEH VSMCs. Insulin exposure decreased expression of α-SM actin and was accompanied by rearrangement of intracellular F-actins in VSMCs, especially in the EH group. These effects were reversed by treatment with the PKC inhibitor. Conclusions Human mesenteric VSMCs of EH and NT patients differed in proliferation, MAPK signaling, and degree of changes in α-SM actin and F-actin isoforms immediately following insulin exposure in vitro.

  11. Cell-Cell Interactions Mediate the Response of Vascular Smooth Muscle Cells to Substrate Stiffness

    Science.gov (United States)

    Sazonova, Olga V.; Lee, Kristen L.; Isenberg, Brett C.; Rich, Celeste B.; Nugent, Matthew A.; Wong, Joyce Y.

    2011-01-01

    The vessel wall experiences progressive stiffening with age and the development of cardiovascular disease, which alters the micromechanical environment experienced by resident vascular smooth muscle cells (VSMCs). In vitro studies have shown that VSMCs are sensitive to substrate stiffness, but the exact molecular mechanisms of their response to stiffness remains unknown. Studies have also shown that cell-cell interactions can affect mechanotransduction at the cell-substrate interface. Using flexible substrates, we show that the expression of proteins associated with cell-matrix adhesion and cytoskeletal tension is regulated by substrate stiffness, and that an increase in cell density selectively attenuates some of these effects. We also show that cell-cell interactions exert a strong effect on cell morphology in a substrate-stiffness dependent manner. Collectively, the data suggest that as VSMCs form cell-cell contacts, substrate stiffness becomes a less potent regulator of focal adhesion signaling. This study provides insight into the mechanisms by which VSMCs respond to the mechanical environment of the blood vessel wall, and point to cell-cell interactions as critical mediators of VSMC response to vascular injury. PMID:21806930

  12. Activation of the Retinoid X Receptor Modulates Angiotensin II-Induced Smooth Muscle Gene Expression and Inflammation in Vascular Smooth Muscle Cells

    OpenAIRE

    Lehman, Allison M.B.; Montford, John R.; Horita, Henrick; Ostriker, Allison C.; Weiser-Evans, Mary C. M.; Nemenoff, Raphael A.; Furgeson, Seth B.

    2014-01-01

    The retinoid X receptor (RXR) partners with numerous nuclear receptors, such as the peroxisome proliferator activated receptor (PPAR) family, liver X receptors (LXRs), and farnesoid X receptor (FXR). Although each heterodimer can be activated by specific ligands, a subset of these receptors, defined as permissive nuclear receptors, can also be activated by RXR agonists known as rexinoids. Many individual RXR heterodimers have beneficial effects in vascular smooth muscle cells (SMCs). Because ...

  13. Loss of Notch3 Signaling in Vascular Smooth Muscle Cells Promotes Severe Heart Failure Upon Hypertension.

    Science.gov (United States)

    Ragot, Hélène; Monfort, Astrid; Baudet, Mathilde; Azibani, Fériel; Fazal, Loubina; Merval, Régine; Polidano, Evelyne; Cohen-Solal, Alain; Delcayre, Claude; Vodovar, Nicolas; Chatziantoniou, Christos; Samuel, Jane-Lise

    2016-08-01

    Hypertension, which is a risk factor of heart failure, provokes adaptive changes at the vasculature and cardiac levels. Notch3 signaling plays an important role in resistance arteries by controlling the maturation of vascular smooth muscle cells. Notch3 deletion is protective in pulmonary hypertension while deleterious in arterial hypertension. Although this latter phenotype was attributed to renal and cardiac alterations, the underlying mechanisms remained unknown. To investigate the role of Notch3 signaling in the cardiac adaptation to hypertension, we used mice with either constitutive Notch3 or smooth muscle cell-specific conditional RBPJκ knockout. At baseline, both genotypes exhibited a cardiac arteriolar rarefaction associated with oxidative stress. In response to angiotensin II-induced hypertension, the heart of Notch3 knockout and SM-RBPJκ knockout mice did not adapt to pressure overload and developed heart failure, which could lead to an early and fatal acute decompensation of heart failure. This cardiac maladaptation was characterized by an absence of media hypertrophy of the media arteries, the transition of smooth muscle cells toward a synthetic phenotype, and an alteration of angiogenic pathways. A subset of mice exhibited an early fatal acute decompensated heart failure, in which the same alterations were observed, although in a more rapid timeframe. Altogether, these observations indicate that Notch3 plays a major role in coronary adaptation to pressure overload. These data also show that the hypertrophy of coronary arterial media on pressure overload is mandatory to initially maintain a normal cardiac function and is regulated by the Notch3/RBPJκ pathway. PMID:27296994

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

    International Nuclear Information System (INIS)

    Highlights: ► Protocatechuic aldehyde (PCA) inhibits ROS production in VSMCs. ► PCA inhibits proliferation and migration in PDGF-induced VSMCs. ► PCA has anti-platelet effects in ex vivo rat whole blood. ► 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′-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 μM). 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.

  15. Regulation of SIRT1 in vascular smooth muscle cells from streptozotocin-diabetic rats.

    Directory of Open Access Journals (Sweden)

    Alice Toniolo

    Full Text Available Sirtuins enzymes are a conserved family of nicotinamide adenine dinucleotide (NAD-dependent deacetylases and ADP-ribosyltransferases that mediate responses to oxidative stress, fasting and dietary restriction in mammals. Vascular smooth muscle cells (VSMCs are involved in many mechanisms that regulate vascular biology in vivo but the role of SIRT1 has not been explored in much detail. Therefore, we investigated the regulation of SIRT1 in cultured VSMCs under various stress conditions including diabetes. Sprague-Dawley rats were made diabetic by injecting a single dose of streptozotocin (65 mg/Kg, and aortic VSMCs were isolated after 4 weeks. Immunocytochemistry showed that SIRT1 was localized predominantly in the nucleus, with lower staining in VSMCs from STZ-diabetic as compared with normoglycemic rats. Previous diabetes induction in vivo and high glucose concentrations in vitro significantly downregulated SIRT1 amounts as detected in Western blot assays, whereas TNF-α (30 ng/ml stimulation failed to induce significant changes. Because estrogen signaling affects several pathways of oxidative stress control, we also investigated SIRT1 modulation by 17β-estradiol. Treatment with the hormone (10 nM or a selective estrogen receptor-α agonist decreased SIRT1 levels in VSMCs from normoglycemic but not in those from STZ-diabetic animals. 17β-estradiol treatment also enhanced activation of AMP-dependent kinase, which partners with SIRT1 in a signaling axis. SIRT1 downregulation by 17β-estradiol could be observed as well in human peripheral blood mononuclear cells, a cell type in which SIRT1 downregulation is associated with insulin resistance and subclinical atherosclerosis. These data suggest that SIRT1 protein levels are regulated by diverse cellular stressors to a variable extent in VSMCs from diabetic and normoglycemic rats, warranting further investigation on SIRT1 as a modulator of VSMC activity in settings of vascular inflammation.

  16. 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.

  17. 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.

  18. Effect of L-Arginine on Pulmonary Artery Smooth Muscle Cell Apoptosis in Rats with Hypoxic Pulmonary Vascular Structural Remodeling

    Institute of Scientific and Technical Information of China (English)

    Ingrid Karmane SUMOU; Jun-Bao DU; Bing WEI; Chun-Yu ZHANG; Jian-Guang QI; Chao-Shu TANG

    2006-01-01

    This study investigated the effect of L-arginine (L-Arg) on the apoptosis of pulmonary artery smooth muscle cells (PASMC) in rats with hypoxic pulmonary vascular structural remodeling, and its mechanisms. Seventeen Wistar rats were randomly divided into a control group (n=5), a hypoxia group (n=7), and a hypoxia+L-Arg group (n=5). The morphologic changes of lung tissues were observed under optical microscope. Using the terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphatebiotin nick end labeling assay, the apoptosis of PASMC was examined. Fas expression in PASMC was examined using immunohistochemistry. The results showed that the percentage of muscularized artery in small pulmonary vessels, and the relative medial thickness and relative medial area of the small and median pulmonary muscularized arteries in the hypoxic group were all significantly increased. Pulmonary vascular structural remodeling developed after hypoxia. Apoptotic smooth muscle cells of the small and median pulmonary arteries in the hypoxia group were significantly less than those in the control group. After 14 d of hypoxia, Fas expression by smooth muscle cells of median and small pulmonary arteries was significantly inhibited. L-Arg significantly inhibited hypoxic pulmonary vascular structural remodeling in association with an augmentation of apoptosis of smooth muscle cells as well as Fas expression in PASMC. These results showed that L-Arg could play an important role in attenuating hypoxic pulmonary vascular structural remodeling by upregulating Fas expression in PASMC, thus promoting the apoptosis of PASMC.

  19. 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.

  20. Connective tissue growth factor and vascular endothelial growth factor from airway smooth muscle interact with the extracellular matrix

    NARCIS (Netherlands)

    Burgess, Janette K; Ge, Qi; Poniris, Maree H; Boustany, Sarah; Twigg, Stephen M; Black, Judith L; Johnson, Peter R A

    2006-01-01

    Airway remodeling describes the structural changes that occur in the asthmatic airway that include airway smooth muscle hyperplasia, increases in vascularity due to angiogenesis, and thickening of the basement membrane. Our aim in this study was to examine the effect of transforming growth factor-be

  1. Human vascular smooth muscle cells both express and respond to heparin-binding growth factor I (endothelial cell growth factor)

    Energy Technology Data Exchange (ETDEWEB)

    Winkles, J.A.; Friesel, R.; Burgess, W.H.; Howk, R.; Mehlman, T.; Weinstein, R.; Maciag, T.

    1987-10-01

    The control of vascular endothelial and muscle cell proliferation is important in such processes as tumor angiogenesis, wound healing, and the pathogenesis of atherosclerosis. Class I heparin-binding growth factor (HBGF-I) is a potent mitogen and chemoattractant for human endothelial cells in vitro and will induce angiogenesis in vivo. RNA gel blot hybridization experiments demonstrate that cultured human vascular smooth muscle cells, but not human umbilical cells also synthesize an HBGF-I mRNA. Smooth muscle cells also synthesize an HBGF-I-like polypeptide since (i) extract prepared from smooth muscle cells will compete with /sup 125/I-labeled HBGF-I for binding to the HBGF-I cell surface receptor, and (ii) the competing ligand is eluted from heparin-Sepharose affinity resin at a NaCl concentration similar to that required by purified bovine brain HBGF-I and stimulates endothelial cell proliferation in vitro. Furthermore, like endothelial cells, smooth muscle cells possess cell-surface-associated HBGF-I receptors and respond to HBGF-I as a mitogen. These results indicate the potential for an additional autocrine component of vascular smooth muscle cell growth control and establish a vessel wall source of HBGF-I for endothelial cell division in vivo.

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

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    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

  3. Chemerin Stimulates Vascular Smooth Muscle Cell Proliferation and Carotid Neointimal Hyperplasia by Activating Mitogen-Activated Protein Kinase Signaling

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    Xiong, Wei; Luo, Yu; Wu, Lin; Liu, Feng; Liu, Huadong; Li, Jianghua; Liao, Bihong; Dong, Shaohong

    2016-01-01

    Vascular neointimal hyperplasia and remodeling arising from local inflammation are characteristic pathogeneses of proliferative cardiovascular diseases, such as atherosclerosis and post angioplasty restenosis. The molecular mechanisms behind these pathological processes have not been fully determined. The adipokine chemerin is associated with obesity, metabolism, and control of inflammation. Recently, chemerin has gained increased attention as it was found to play a critical role in the development of cardiovascular diseases. In this study, we investigated the effects of chemerin on the regulation of vascular smooth muscle cells and carotid neointimal formation after angioplasty. We found that circulating chemerin levels increased after carotid balloon injury, and that knockdown of chemerin significantly inhibited the proliferative aspects of vascular smooth muscle cells induced by platelet-derived growth factor-BB and pro-inflammatory chemokines in vitro as well as prohibited carotid neointimal hyperplasia and pro-inflammatory chemokines in vivo after angioplasty. Additionally, inhibition of chemerin down-regulated the expression of several proteins, including phosphorylated p38 mitogen-activated protein kinase, phosphorylated extracellular signal regulated kinase 1/2, nuclear factor-kappa B p65, and proliferation cell nuclear antigen. The novel finding of this study is that chemerin stimulated vascular smooth muscle cells proliferation and carotid intimal hyperplasia through activation of the mitogen-activated protein kinase signaling pathway, which may lead to vascular inflammation and remodeling, and is relevant to proliferative cardiovascular diseases. PMID:27792753

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

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    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

  5. Cadmium Toxicity on Arterioles Vascular Smooth Muscle Cells of Spontaneously Hypertensive Rats

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    Elbert L. Myles

    2006-12-01

    Full Text Available Cadmium (Cd is frequently used in various industrial applications and is a ubiquitous environmental toxicant, also present in tobacco smoke. An important route of exposure is the circulatory system whereas blood vessels are considered to be main stream organs of Cd toxicity. Our previous results indicate that cadmium chloride (CdCl2 affects mean arterial blood pressure in hypertensive rats. We hypothesized that Cd alters the intracellular calcium transient mechanism, by cadmium-induced stimulation of MAPKs (ERK 1 & 2 which is mediated partially through calcium-dependent PKC mechanism. To investigate this hypothesis, we exposed primary cultures of vascular smooth muscle cells (VSMCs from wistar kyoto (WKY and spontaneously hypertensive rats (SHR to increased concentrations of CdCl2 on cell viability, expression of mitogen-activated protein kinases (MAPKs/ERK 1 & 2, and protein kinase C (PKC which are activated by Cd in several cell types. The results from these studies indicate that CdCl2 decreased cell viability of both SHR and WKY VSMCs in a concentration dependent-manner. Viability of both cell types decreased 33±5.3 (SHR and 39±2.3% (WKY when exposed to 1 μM CdCl2, whereas, 8 and 16 μM reduced viability by 66±3.1 and 62±4.5% in SHR cells. CdCl2 increased ERK 1 & 2 in a biphasic manner with maximum increase occurring when cells are exposed to 1 and 4 μM in SHR VSMCs, whereas, a reduction in ERK 1 and 2 is observed when WKY cells are treated with 2 μM. The results also indicate that CdCl2 increased PKC a/ß in both SHR and WKY VSMCs with a greater increase in expression in SHR VSMCs. In addition, the [Ca2+]i chelator, BAPTA, suppressed the CdCl2 effect, whereas, the PKC inhibitor, GF109203X, reduced the CdCl2 induced-effect on PKC expression. The present studies support the hypothesis that Cd can be a risk factor of hypertension through dysfunction of vascular smooth muscle cells

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

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    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

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

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    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.

  8. Critical Parameters of the In Vitro Method of Vascular Smooth Muscle Cell Calcification.

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    Luis Hortells

    Full Text Available Vascular calcification (VC is primarily studied using cultures of vascular smooth muscle cells. However, the use of very different protocols and extreme conditions can provide findings unrelated to VC. In this work we aimed to determine the critical experimental parameters that affect calcification in vitro and to determine the relevance to calcification in vivo.Rat VSMC calcification in vitro was studied using different concentrations of fetal calf serum, calcium, and phosphate, in different types of culture media, and using various volumes and rates of change. The bicarbonate content of the media critically affected pH and resulted in supersaturation, depending on the concentration of Ca2+ and Pi. Such supersaturation is a consequence of the high dependence of bicarbonate buffers on CO2 vapor pressure and bicarbonate concentration at pHs above 7.40. Such buffer systems cause considerable pH variations as a result of minor experimental changes. The variations are more critical for DMEM and are negligible when the bicarbonate concentration is reduced to ¼. Particle nucleation and growth were observed by dynamic light scattering and electron microscopy. Using 2mM Pi, particles of ~200nm were observed at 24 hours in MEM and at 1 hour in DMEM. These nuclei grew over time, were deposited in the cells, and caused osteogene expression or cell death, depending on the precipitation rate. TEM observations showed that the initial precipitate was amorphous calcium phosphate (ACP, which converts into hydroxyapatite over time. In blood, the scenario is different, because supersaturation is avoided by a tightly controlled pH of 7.4, which prevents the formation of PO43--containing ACP.The precipitation of ACP in vitro is unrelated to VC in vivo. The model needs to be refined through controlled pH and the use of additional procalcifying agents other than Pi in order to reproduce calcium phosphate deposition in vivo.

  9. Nuclear envelope proteins modulate proliferation of vascular smooth muscle cells during cyclic stretch application.

    Science.gov (United States)

    Qi, Ying-Xin; Yao, Qing-Ping; Huang, Kai; Shi, Qian; Zhang, Ping; Wang, Guo-Liang; Han, Yue; Bao, Han; Wang, Lu; Li, Hai-Peng; Shen, Bao-Rong; Wang, Yingxiao; Chien, Shu; Jiang, Zong-Lai

    2016-05-10

    Cyclic stretch is an important inducer of vascular smooth muscle cell (VSMC) proliferation, which is crucial in vascular remodeling during hypertension. However, the molecular mechanism remains unclear. We studied the effects of emerin and lamin A/C, two important nuclear envelope proteins, on VSMC proliferation in hypertension and the underlying mechano-mechanisms. In common carotid artery of hypertensive rats in vivo and in cultured cells subjected to high (15%) cyclic stretch in vitro, VSMC proliferation was increased significantly, and the expression of emerin and lamin A/C was repressed compared with normotensive or normal (5%) cyclic stretch controls. Using targeted siRNA to mimic the repressed expression of emerin or lamin A/C induced by 15% stretch, we found that VSMC proliferation was enhanced under static and 5%-stretch conditions. Overexpression of emerin or lamin A/C reversed VSMC proliferation induced by 15% stretch. Hence, emerin and lamin A/C play critical roles in suppressing VSMC hyperproliferation induced by hyperstretch. ChIP-on-chip and MOTIF analyses showed that the DNAs binding with emerin contain three transcription factor motifs: CCNGGA, CCMGCC, and ABTTCCG; DNAs binding with lamin A/C contain the motifs CVGGAA, GCCGCYGC, and DAAGAAA. Protein/DNA array proved that altered emerin or lamin A/C expression modulated the activation of various transcription factors. Furthermore, accelerating local expression of emerin or lamin A/C reversed cell proliferation in the carotid artery of hypertensive rats in vivo. Our findings establish the pathogenetic role of emerin and lamin A/C repression in stretch-induced VSMC proliferation and suggest mechanobiological mechanism underlying this process that involves the sequence-specific binding of emerin and lamin A/C to specific transcription factor motifs.

  10. Calcineurin-NFAT signaling is involved in phenylephrine-induced vascular smooth muscle cell proliferation

    Institute of Scientific and Technical Information of China (English)

    Xiao PANG; Ning-ling SUN

    2009-01-01

    Aim: Catecholamine-induced vascular smooth muscle cell (VSMC) proliferation is one of the major events in the pathogenesis of atherosclerosis and vascular remodeling. The calcineurin-NFAT pathway plays a role in regulating growth and differentiation in various cell types. We investigated whether the calcineurin-NFAT pathway was involved in the regulation of phenylephrine-induced VSMC proliferation.Methods: Proliferation of VSMC was measured using an MTT assay and cell counts. Localization of NFATcl was detected by immunofluorescence staining. NFATcl-DNA binding was determined by EMSA and luciferase activity analyses.NFATcl and calcineurin levels were assayed by immunoprecipitation.Results: Phenylephrine (PE, an α1-adrenoceptor agonist) increased VSMC proliferation and cell number. Prazosin (an α1-adrenoceptor antagonist), cyclosporin A (CsA, an inhibitor of calcineurin) and chelerythrine (an inhibitor of PKC)decreased PE-induced proliferation and cell number. Additional treatment of VSMC with CsA or chelerythrine further inhibited proliferation and cell number in the chelerythrine-pretreatment group and the CsA-pretreatment group. CsA and chelerythrine alone had no effect on either absorbance or cell number. CsA decreased PE-induced calcineurin levels and activity. NFATc1 was translocated from the cytoplasm to the nucleus upon treatment with PE. This translocation was reversed by CsA. CsA decreased the PE-induced NFATc1 level in the nucleus. PE increased NFAT's DNA binding activity and NFAT-dependent reporter gene expression. CsA blocked these effects.Conclusion: CsA partially suppresses PE-induced VSMC proliferation by inhibiting calcineurin activity and NFATc1 nuclear translocation. The calcineurin-NFATc1 pathway is involved in the hyperplastic growth of VSMC induced by phenylephrine.

  11. Autophagy inhibits PDGF-BB-induced calcification in vascular smooth muscle cells

    Institute of Scientific and Technical Information of China (English)

    PEI Qian-qian; MEI Han; ZHANG Xu-hui; DONG Li-hua

    2016-01-01

    AIM:To investigate the relationship between autophagy and calcification in vascular smooth muscle cells ( VSMCs) after platelet-derived growth factor (PDGF)-BB stimulation.METHODS:Cultured VSMCs were stimulated with PDGF-BB for different time, the expression of vascular calcification-related proteins and autophagy-related proteins were detected by Western blot .The interaction be-tween Beclin1 and PI3KC3 was detected by co-immunoprecipitation.RESULTS: The expression of BMP2 and ALP showed a trend from decline to rise.ALP slumped at 12 h, and BMP2 slumped at 6 h.Moreover, the expression of Beclin-1 showed a trend from rise to decline, and peaked at 12 h.The conversion of LC3-ⅠtoⅡincreased in a time-dependent manner , and peaked at 24 h.The ex-pression of BMP2 and ALP was increased in VSMCs incubated with PDGF-BB and autophagy inhibitor 3-MA, compared with PDGF-BB-stimulated VSMCs.Furthermore, the interaction between Beclin1 and PI3KC3 was enhanced at 6 h after PDGF-BB stimulated, peaked at 12 h, and kept in high level at 24 h.Moreover, the phosphorylation level of Beclin 1 was enhanced by PDGF-BB stimulation, and peaked at 6 h.CONCLUSION:Our findings demonstrate that PDGF-BB-induced autophagy inhibits VSMC calcification by en-hancing Beclin1 phosphorylation and interaction between Beclin 1 and PI3KC3.

  12. Vascular Smooth Muscle Sirtuin-1 Protects Against Diet-Induced Aortic Stiffness.

    Science.gov (United States)

    Fry, Jessica L; Al Sayah, Leona; Weisbrod, Robert M; Van Roy, Isabelle; Weng, Xiang; Cohen, Richard A; Bachschmid, Markus M; Seta, Francesca

    2016-09-01

    Arterial stiffness, a major cardiovascular risk factor, develops within 2 months in mice fed a high-fat, high-sucrose (HFHS) diet, serving as a model of human metabolic syndrome, and it is associated with activation of proinflammatory and oxidant pathways in vascular smooth muscle (VSM) cells. Sirtuin-1 (SirT1) is an NAD(+)-dependent deacetylase regulated by the cellular metabolic status. Our goal was to study the effects of VSM SirT1 on arterial stiffness in the context of diet-induced metabolic syndrome. Overnight fasting acutely decreased arterial stiffness, measured in vivo by pulse wave velocity, in mice fed HFHS for 2 or 8 months, but not in mice lacking SirT1 in VSM (SMKO). Similarly, VSM-specific genetic SirT1 overexpression (SMTG) prevented pulse wave velocity increases induced by HFHS feeding, during 8 months. Administration of resveratrol or S17834, 2 polyphenolic compounds known to activate SirT1, prevented HFHS-induced arterial stiffness and were mimicked by global SirT1 overexpression (SirT1 bacterial artificial chromosome overexpressor), without evident metabolic improvements. In addition, HFHS-induced pulse wave velocity increases were reversed by 1-week treatment with a specific, small molecule SirT1 activator (SRT1720). These beneficial effects of pharmacological or genetic SirT1 activation, against HFHS-induced arterial stiffness, were associated with a decrease in nuclear factor kappa light chain enhancer of activated B cells (NFκB) activation and vascular cell adhesion molecule (VCAM-1) and p47phox protein expressions, in aorta and VSM cells. In conclusion, VSM SirT1 activation decreases arterial stiffness in the setting of obesity by stimulating anti-inflammatory and antioxidant pathways in the aorta. SirT1 activators may represent a novel therapeutic approach to prevent arterial stiffness and associated cardiovascular complications in overweight/obese individuals with metabolic syndrome. PMID:27432859

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

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    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.

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

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    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.

  15. 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

  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-X(L) and Bax and the expression of bcl-2 and bcl-x(L), 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. Inhibitors of soluble epoxide hydrolase attenuate vascular smooth muscle cell proliferation

    Science.gov (United States)

    Davis, Benjamin B.; Thompson, David A.; Howard, Laura L.; Morisseau, Christophe; Hammock, Bruce D.; Weiss, Robert H.

    2002-02-01

    Atherosclerosis, in its myriad incarnations the foremost killer disease in the industrialized world, is characterized by aberrant proliferation of vascular smooth muscle (VSM) cells in part as a result of the recruitment of inflammatory cells to the blood vessel wall. The epoxyeicosatrienoic acids are synthesized from arachidonic acid in a reaction catalyzed by the cytochrome P450 system and are vasoactive substances. Metabolism of these compounds by epoxide hydrolases results in the formation of compounds that affect the vasculature in a pleiotropic manner. As an outgrowth of our observations that urea inhibitors of the soluble epoxide hydrolase (sEH) reduce blood pressure in spontaneously hypertensive rats as well as the findings of other investigators that these compounds possess antiinflammatory actions, we have examined the effect of sEH inhibitors on VSM cell proliferation. We now show that the sEH inhibitor 1-cyclohexyl-3-dodecyl urea (CDU) inhibits human VSM cell proliferation in a dose-dependent manner and is associated with a decrease in the level of cyclin D1. In addition, cis-epoxyeicosatrienoic acid mimics the growth-suppressive activity of CDU; there is no evidence of cellular toxicity or apoptosis in CDU-treated cells when incubated with 20 μM CDU for up to 48 h. These results, in light of the antiinflammatory and antihypertensive properties of these compounds that have been demonstrated already, suggest that the urea class of sEH inhibitors may be useful for therapy for diseases such as hypertension and atherosclerosis characterized by exuberant VSM cell proliferation and vascular inflammation.

  18. 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.

  19. Vascular smooth muscle desensitization in rabbit epigastric and mesenteric arteries during hemorrhagic shock.

    Science.gov (United States)

    Ratz, P H; Miner, A S; Huang, Y; Smith, C A; Barbee, R W

    2016-07-01

    The decompensatory phase of hemorrhage (shock) is caused by a poorly defined phenomenon termed vascular hyporeactivity (VHR). VHR may reflect an acute in vivo imbalance in levels of contractile and relaxant stimuli favoring net vascular smooth muscle (VSM) relaxation. Alternatively, VHR may be caused by intrinsic VSM desensitization of contraction resulting from prior exposure to high levels of stimuli that temporarily adjusts cell signaling systems. Net relaxation, but not desensitization, would be expected to resolve rapidly in an artery segment removed from the in vivo shock environment and examined in vitro in a fresh solution. Our aim was to 1) induce shock in rabbits and apply an in vitro mechanical analysis on muscular arteries isolated pre- and postshock to determine whether VHR involves intrinsic VSM desensitization, and 2) identify whether net VSM relaxation induced by nitric oxide and cyclic nucleotide-dependent protein kinase activation in vitro can be sustained for some time after relaxant stimulus washout. The potencies of phenylephrine- and histamine-induced contractions in in vitro epigastric artery removed from rabbits posthemorrhage were decreased by ∼0.3 log units compared with the control contralateral epigastric artery removed prehemorrhage. Moreover, a decrease in KCl-induced tonic, relative to phasic, tension of in vitro mesenteric artery correlated with the degree of shock severity as assessed by rates of lactate and K(+) accumulation. VSM desensitization was also caused by tyramine in vivo and PE in vitro, but not by relaxant agents in vitro. Together, these results support the hypothesis that VHR during hemorrhagic decompensation involves contractile stimulus-induced long-lasting, intrinsic VSM desensitization. PMID:27199133

  20. miRNA-146a induces vascular smooth muscle cell apoptosis in a rat model of coronary heart disease via NF-κB pathway.

    Science.gov (United States)

    Wu, Z W; Liu, Y F; Wang, S; Li, B

    2015-12-29

    The aim of this study was to investigate the role of miRNA-146a in modulating the function of vascular smooth muscle cells in a rat model of coronary heart disease. Vascular smooth muscle cells were isolated and cultured from the rat coronary heart disease model and normal rats (controls). miRNA-146a levels were measured in vascular smooth muscle cells obtained from rats with coronary heart disease and control rats. The proliferation, growth, apoptosis, and activation of the NF-κB pathway in the vascular smooth muscle cells were detected using the MTT assay and flow cytometry, respectively. The role of the NF-κB pathway in modulating the apoptosis of vascular smooth muscle cells was investigated by measuring the reactivity of the cells to an NF-κB pathway inhibitor (TPCA-1). Vascular smooth muscle cells from the disease model exhibited higher levels of miRNA-146a than that by the normal controls (P = 0.0024). The vascular smooth muscle cells obtained from rats with coronary heart disease showed decreased proliferation and growth and increased apoptosis. miRNA-146a overexpression elevated the rate of cell apoptosis. The NF-κB pathway was activated in vascular smooth muscle cells obtained from rats with coronary heart disease. Inhibition of the NF- κB pathway significantly decreased the rate of vascular smooth muscle cell apoptosis in coronary heart disease rats (P = 0.0038). In conclusion, miRNA- 146a was found to induce vascular smooth muscle cell apoptosis in rats with coronary heart disease via the activation of the NF-κB signal pathway.

  1. Differential Mitochondrial Adaptation in Primary Vascular Smooth Muscle Cells from a Diabetic Rat Model.

    Science.gov (United States)

    Keller, Amy C; Knaub, Leslie A; McClatchey, P Mason; Connon, Chelsea A; Bouchard, Ron; Miller, Matthew W; Geary, Kate E; Walker, Lori A; Klemm, Dwight J; Reusch, Jane E B

    2016-01-01

    Diabetes affects more than 330 million people worldwide and causes elevated cardiovascular disease risk. Mitochondria are critical for vascular function, generate cellular reactive oxygen species (ROS), and are perturbed by diabetes, representing a novel target for therapeutics. We hypothesized that adaptive mitochondrial plasticity in response to nutrient stress would be impaired in diabetes cellular physiology via a nitric oxide synthase- (NOS-) mediated decrease in mitochondrial function. Primary smooth muscle cells (SMCs) from aorta of the nonobese, insulin resistant rat diabetes model Goto-Kakizaki (GK) and the Wistar control rat were exposed to high glucose (25 mM). At baseline, significantly greater nitric oxide evolution, ROS production, and respiratory control ratio (RCR) were observed in GK SMCs. Upon exposure to high glucose, expression of phosphorylated eNOS, uncoupled respiration, and expression of mitochondrial complexes I, II, III, and V were significantly decreased in GK SMCs (p diabetes phenotype. Overall, nutrient stress in GK SMCs caused a persistent decline in eNOS and mitochondrial function and disrupted mitochondrial plasticity, illustrating eNOS and mitochondria as potential therapeutic targets. PMID:27034743

  2. Effects of caffeic acid phenethyl ester on proliferation of vascular smooth muscle cells in rats

    Institute of Scientific and Technical Information of China (English)

    Gang Yang; Chao Chang; YuQing Wang; Yibo Feng; ShuLing Rong

    2006-01-01

    Objective: To investigate the inhibitory effect of caffeic acid phenethyl ester(CAPE) on the proliferation of vascular smooth muscle cells (VSMC) activated by lipopolysaccharide (LPS) and to clarify its mechanism. Methods: VSMC activated by LPS (1 mg·L-1) were treated with CAPE at different concentrations. The inhibitory effects of CAPE on the proliferation of VSMC were determined by methabenzthiazuron(MTT) colorimetry. The effects of CAPE on the expression of proliferating cell nuclear antigen (PCNA) and Survivin protein in VSMC were evaluated by immunocytochemistry staining technique (SABC method). Cell cycle was analyzed by flow cytometry(FCM) with propidium iodide (PI) labeling method. The relative expression level of Survivin mRNA was measured with real-time quantified RT-PCR technique. Results: CAPE exerted significant inhibitory effects on. proliferation of VSMC at concentrations ranging from 5 mg·L-1 to 80 mg·L-1, decreased the rate of cells positive for PCNA and Survivin protein and repressed the expression of Survivin mRNA in a dose- and time-dependent manner (P < 0.05).FCM analysis displayed that CAPE up-regulated the ratio of G0/G1 stages and reduced the percentage of VSMC in S stage (P <0.05). Conclusion: CAPE can significantly inhibit the proliferation of VSMC activated by LPS in a dose- and time-dependent manner, which may be carried out through regulating cell cycle and repressing the expression of PCNA and Survivin.

  3. Vascular smooth muscle cell apoptosis promotes transplant arteriosclerosis through inducing the production of SDF-1α.

    Science.gov (United States)

    Li, J; Liu, S; Li, W; Hu, S; Xiong, J; Shu, X; Hu, Q; Zheng, Q; Song, Z

    2012-08-01

    Transplant arteriosclerosis is a leading cause of late allograft loss. Medial smooth muscle cell (SMC) apoptosis is considered to be an important event in transplant arteriosclerosis. However, the precise contribution of medial SMC apoptosis to transplant arteriosclerosis and the underlying mechanisms remain unclear. We transferred wild-type p53 to induce apoptosis of cultured SMCs. We found that apoptosis induces the production of SDF-1α from apoptotic and neighboring viable cells, resulting in increased SDF-1α in the culture media. Conditioned media from Ltv-p53-transferred SMCs activated PI3K/Akt/mTOR and MAPK/Erk signaling in a SDF-1α-dependent manner and thereby promoted mesenchymal stem cell (MSC) migration and proliferation. In a rat aorta transplantation model, lentivirus-mediated BclxL transfer selectively inhibits medial SMC apoptosis in aortic allografts, resulting in a remarkable decrease of SDF-1α both in allograft media and in blood plasma, associated with diminished recruitment of CD90(+)CD105(+) double-positive cells and impaired neointimal formation. Systemic administration of rapamycin or PD98059 also attenuated MSC recruitment and neointimal formation in the aortic allografts. These results suggest that medial SMC apoptosis is critical for the development of transplant arteriosclerosis through inducing SDF-1α production and that MSC recruitment represents a major component of vascular remodeling, constituting a relevant target and mechanism for therapeutic interventions.

  4. Increased proliferation of explanted vascular smooth muscle cells: a marker presaging atherogenesis.

    Science.gov (United States)

    Absher, P M; Schneider, D J; Baldor, L C; Russell, J C; Sobel, B E

    1997-06-01

    The JCR:LA-cp homozygous cp/cp corpulent rat is genetically predisposed to develop atherosclerosis evident after 9 and 18 months of age in males and females and to manifest metabolic derangements resembling those seen in type II diabetes in humans (hyperinsulinemia, insulin resistance, hyperglycemia and hypertriglyceridemia). The present study was undertaken to determine whether vascular smooth muscle cells (SMCs) explanted from vessels destined to become atherosclerotic later in life exhibit intrinsic properties ex vivo that presage atherogenesis to provide a means for evaluating promptly intervention designed to modify it. SMCs were cultured from aortic explants of JCR:LA-cp corpulent (cp/cp) and lean control (+/+) rats of 4, 5, 6, and 9 months of age. Compared with SMCs from controls, SMCs from cp/cp rats exhibited increased proliferation, higher saturation density, increased augmentation of proliferation in response to selected mitogens and greater adherence to extracellular matrix proteins. The increased proliferative activity ex vivo anteceded by several months the development of atherosclerotic lesions in vivo. Thus, it is a promising marker in assessments of the efficacy of interventions designed to retard or prevent atherosclerosis.

  5. Molecular Pathways Regulating Macrovascular Pathology and Vascular Smooth Muscle Cells Phenotype in Type 2 Diabetes

    Directory of Open Access Journals (Sweden)

    Sara Casella

    2015-10-01

    Full Text Available Type 2 diabetes mellitus (T2DM is a disease reaching a pandemic proportion in developed countries and a major risk factor for almost all cardiovascular diseases and their adverse clinical manifestations. T2DM leads to several macrovascular and microvascular alterations that influence the progression of cardiovascular diseases. Vascular smooth muscle cells (VSMCs are fundamental players in macrovascular alterations of T2DM patients. VSMCs display phenotypic and functional alterations that reflect an altered intracellular biomolecular scenario of great vessels of T2DM patients. Hyperglycemia itself and through intraparietal accumulation of advanced glycation-end products (AGEs activate different pathways, in particular nuclear factor-κB and MAPKs, while insulin and insulin growth-factor receptors (IGFR are implicated in the activation of Akt and extracellular-signal-regulated kinases (ERK 1/2. Nuclear factor-κB is also responsible of increased susceptibility of VSMCs to pro-apoptotic stimuli. Down-regulation of insulin growth-factor 1 receptors (IGFR-1R activity in diabetic vessels also influences negatively miR-133a levels, so increasing apoptotic susceptibility of VSMCs. Alterations of those bimolecular pathways and related genes associate to the prevalence of a synthetic phenotype of VSMCs induces extracellular matrix alterations of great vessels. A better knowledge of those biomolecular pathways and related genes in VSMCs will help to understand the mechanisms leading to macrovascular alterations in T2DM patients and to suggest new targeted therapies.

  6. Inhibition Mechanism of Emodin on Rabbit Vascular Smooth Muscle Cells Proliferation

    Institute of Scientific and Technical Information of China (English)

    1999-01-01

    The proliferation of vascular smooth muscle cells (VSMCs) contributes to the pathogenesis of atherosclerosis and restenosis after angioplasty and vein graft.In this study, MTT colormetry was used to test the effective scope of emodin to inhibit VSMCs proliferation.Flow cytometry and confocal image were adopted to investigate its inhibitive mechanism.The results show that emodin could inhibit the growth and proliferation of VSMCs and the inhibition rate of emodin on VSMCs is 24.6%-94.58%, which is time - and concentration - dependent.Emodin could reduce S phase entry, increase the apoptosis of VSMCs, and reduce the intensity of[Ca2+]i in hPDGF B/B stimulated VSMCs.This research provides theoretical basis for medical application of emodin.It is concluded that emodin could inhibit the growth and proliferation of VSMCs effectively.Decreasing the DNA synthesis, increasing the cell apoptosis and reducing the intensity of[Ca2+]i in hPDGF B/B stimulated VSMCs may be the inhibitive mechanism of emodin against VSMCs proliferation.

  7. Differential Mitochondrial Adaptation in Primary Vascular Smooth Muscle Cells from a Diabetic Rat Model

    Directory of Open Access Journals (Sweden)

    Amy C. Keller

    2016-01-01

    Full Text Available Diabetes affects more than 330 million people worldwide and causes elevated cardiovascular disease risk. Mitochondria are critical for vascular function, generate cellular reactive oxygen species (ROS, and are perturbed by diabetes, representing a novel target for therapeutics. We hypothesized that adaptive mitochondrial plasticity in response to nutrient stress would be impaired in diabetes cellular physiology via a nitric oxide synthase- (NOS- mediated decrease in mitochondrial function. Primary smooth muscle cells (SMCs from aorta of the nonobese, insulin resistant rat diabetes model Goto-Kakizaki (GK and the Wistar control rat were exposed to high glucose (25 mM. At baseline, significantly greater nitric oxide evolution, ROS production, and respiratory control ratio (RCR were observed in GK SMCs. Upon exposure to high glucose, expression of phosphorylated eNOS, uncoupled respiration, and expression of mitochondrial complexes I, II, III, and V were significantly decreased in GK SMCs (p<0.05. Mitochondrial superoxide increased with high glucose in Wistar SMCs (p<0.05 with no change in the GK beyond elevated baseline concentrations. Baseline comparisons show persistent metabolic perturbations in a diabetes phenotype. Overall, nutrient stress in GK SMCs caused a persistent decline in eNOS and mitochondrial function and disrupted mitochondrial plasticity, illustrating eNOS and mitochondria as potential therapeutic targets.

  8. 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.

  9. Critical role of exogenous nitric oxide in ROCK activity in vascular smooth muscle cells.

    Directory of Open Access Journals (Sweden)

    Tatsuya Maruhashi

    Full Text Available Rho-associated kinase (ROCK signaling pathway has been shown to mediate various cellular functions including cell proliferation, migration, adhesion, apoptosis, and contraction, all of which may be involved in pathogenesis of atherosclerosis. Endogenous nitric oxide (NO is well known to have an anti-atherosclerotic effect, whereas the exogenous NO-mediated cardiovascular effect still remains controversial. The purpose of this study was to evaluate the effect of exogenous NO on ROCK activity in vascular smooth muscle cells (VSMCs in vitro and in vivo.VSMCs migration was evaluated using a modified Boyden chamber assay. ROCK activities were measured by Western blot analysis in murine and human VSMCs and aorta of mice treated with or without angiotensin II (Ang II and/or sodium nitroprusside (SNP, an NO donor.Co-treatment with SNP inhibited the Ang II-induced cell migration and increases in ROCK activity in murine and human VSMCs. Similarly, the increased ROCK activity 2 weeks after Ang II infusion in the mouse aorta was substantially inhibited by subcutaneous injection of SNP.These findings suggest that administration of exogenous NO can inhibit ROCK activity in VSMCs in vitro and in vivo.

  10. 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.

  11. Cyclic strain increases protease-activated receptor-1 expression in vascular smooth muscle cells

    Science.gov (United States)

    Nguyen, K. T.; Frye, S. R.; Eskin, S. G.; Patterson, C.; Runge, M. S.; McIntire, L. V.

    2001-01-01

    Cyclic strain regulates many vascular smooth muscle cell (VSMC) functions through changing gene expression. This study investigated the effects of cyclic strain on protease-activated receptor-1 (PAR-1) expression in VSMCs and the possible signaling pathways involved, on the basis of the hypothesis that cyclic strain would enhance PAR-1 expression, reflecting increased thrombin activity. Uniaxial cyclic strain (1 Hz, 20%) of cells cultured on elastic membranes induced a 2-fold increase in both PAR-1 mRNA and protein levels. Functional activity of PAR-1, as assessed by cell proliferation in response to thrombin, was also increased by cyclic strain. In addition, treatment of cells with antioxidants or an NADPH oxidase inhibitor blocked strain-induced PAR-1 expression. Preincubation of cells with protein kinase inhibitors (staurosporine or Ro 31-8220) enhanced strain-increased PAR-1 expression, whereas inhibitors of NO synthase, tyrosine kinase, and mitogen-activated protein kinases had no effect. Cyclic strain in the presence of basic fibroblast growth factor induced PAR-1 mRNA levels beyond the effect of cyclic strain alone, whereas no additive effect was observed between cyclic strain and platelet-derived growth factor-AB. Our findings that cyclic strain upregulates PAR-1 mRNA expression but that shear stress downregulates this gene in VSMCs provide an opportunity to elucidate signaling differences by which VSMCs respond to different mechanical forces.

  12. CD146 expression on mesenchymal stem cells is associated with their vascular smooth muscle commitment.

    Science.gov (United States)

    Espagnolle, Nicolas; Guilloton, Fabien; Deschaseaux, Frédéric; Gadelorge, Mélanie; Sensébé, Luc; Bourin, Philippe

    2014-01-01

    Bone marrow mesenchymal stem cells (MSCs) are plastic adherent cells that can differentiate into various tissue lineages, including osteoblasts, adipocytes and chondrocytes. However, this progenitor property is not shared by all cells within the MSC population. In addition, MSCs vary in their proliferation capacity and expression of markers. Because of heterogeneity of CD146 expression in the MSC population, we compared CD146(-/Low) and CD146(High) cells under clonal conditions and after sorting of the non-clonal cell population to determine whether this expression is associated with specific functions. CD146(-/Low) and CD146(High) bone marrow MSCs did not differ in colony-forming unit-fibroblast number, osteogenic, adipogenic and chondrogenic differentiation or in vitro haematopoietic-supportive activity. However, CD146(-/Low) clones proliferated slightly but significantly faster than did CD146(High) clones. In addition, a strong expression of CD146 molecule was associated with a commitment to a vascular smooth muscle cell (VSMC) lineage characterized by a strong up-regulation of calponin-1 and SM22α expression and an ability to contract collagen matrix. Thus, within a bone marrow MSC population, certain subpopulations characterized by high expression of CD146, are committed towards a VSMC lineage.

  13. Killing effect of coexpressing cytosine deaminase and thymidine kinase on rat vascular smooth muscle cells

    Institute of Scientific and Technical Information of China (English)

    曹慧青; 孟宪敏; 刘冬青; 赵秀文; 丁金凤

    2004-01-01

    Background Vascular smooth muscle cell (VSMC) proliferation following arterial injury plays a critical role in a variety of vascular proliferative disorders, such as atherosclerosis and restenosis after balloon angioplasty. Herpes simplex virus-thymidine kinase (HSV-TK)/ganciclovir (GCV) and E.coli cytosine deaminase (CD)/5-fluorocytosine (5-Fc) suicide gene systems have been successfully employed in cardiovascular gene therapy, respectively. We reasoned that coexpression of both HSV-TK with CD suicide genes would lead to increased cell killing. To test this imagine, the adenoviral vectors expressing TK and/or CD genes were developed and tested on vascular smooth muscle cells. Methods Adenoviral vectors, including Ad-EF1α-CD-cytomegolovirus (CMV)-TK coexpressing both CD and TK double suicide genes, Ad-EF1α-CD and Ad-CMV-TK expressing CD and TK respectively, and control vector Ad-CMV-LacZ, were constructed and prepared with homologous recombination in RecA+E.coli cells. Integration and expression of CD and/or TK gene were identified by PCR and Western blot. Primary cultured VSMCs were infected at a multiplicity of infection (MOI) of 20 with exposure to their matching prodrugs 5-Fc and GCV. Cell mortality was measured by methyl thiazolyl tetrazolium (MTT) assays. Flow cytometry analysis was used to detect cell death. Apoptotic cells were analyzed using Hoechst 33342 fluorescence dye as a DNA probe. Genomic DNA cleavage of apoptotic VSMCs was tested by agarose gel electrophoresis. Results Recombinant adenovirus expressing CD and/or TK suicide genes were successfully constructed. Both single and double suicide genes could be integrated into adenoviral genome and expressed. Cytotoxic effects of Ad-EF1α-CD-CMV-TK double suicide genes combined with 5-Fc and GCV were higher than those of Ad-CMV-TK and Ad-EF1α-CD single gene groups. The rate of cell survival was only (9±3)% in the Ad-EF1α-CD-CMV-TK group, but (37±3)% in the Ad-CMV-TK and (46±4)% in the Ad-EF1

  14. Conditional deletion of Dicer in vascular smooth muscle cells leads to the developmental delay and embryonic mortality

    Energy Technology Data Exchange (ETDEWEB)

    Pan, Yaoqian [Department of Physiology, University of Tennessee Health Science Center, Memphis, TN 38163 (United States); Center for Cancer Research, University of Tennessee Health Science Center, Memphis, TN 38163 (United States); Balazs, Louisa [Department of Pathology, University of Tennessee Health Science Center, Memphis, TN 38163 (United States); Tigyi, Gabor [Department of Physiology, University of Tennessee Health Science Center, Memphis, TN 38163 (United States); Yue, Junming, E-mail: yue@uthsc.edu [Department of Physiology, University of Tennessee Health Science Center, Memphis, TN 38163 (United States); Center for Cancer Research, University of Tennessee Health Science Center, Memphis, TN 38163 (United States)

    2011-05-13

    Highlights: {yields} Deletion of Dicer in vascular smooth muscle cells(VSMCs) leads to embryonic mortality. {yields} Loss of Dicer in VSMCs leads to developmental delay. {yields} Loss of Dicer in VSMCs leads to hemorrhage in various organs including brain, skin and liver. {yields} Loss of Dicer in VSMCs leads to vascular wall remodeling. {yields} Loss of Dicer in VSMCs dysregulates the expression of miRNA and VSMC marker genes. -- Abstract: Dicer is a RNAase III enzyme that cleaves double stranded RNA and generates small interfering RNA (siRNA) and microRNA (miRNA). The goal of this study is to examine the role of Dicer and miRNAs in vascular smooth muscle cells (VSMCs). We deleted Dicer in VSMCs of mice, which caused a developmental delay that manifested as early as embryonic day E12.5, leading to embryonic death between E14.5 and E15.5 due to extensive hemorrhage in the liver, brain, and skin. Dicer KO embryos showed dilated blood vessels and a disarray of vascular architecture between E14.5 and E15.5. VSMC proliferation was significantly inhibited in Dicer KOs. The expression of VSMC marker genes were significantly downregulated in Dicer cKO embryos. The vascular structure of the yolk sac and embryo in Dicer KOs was lost to an extent that no blood vessels could be identified after E15.5. Expression of most miRNAs examined was compromised in VSMCs of Dicer KO. Our results indicate that Dicer is required for vascular development and regulates vascular remodeling by modulating VSMC proliferation and differentiation.

  15. Synergistic effects of matrix nanotopography and stiffness on vascular smooth muscle cell function.

    Science.gov (United States)

    Chaterji, Somali; Kim, Peter; Choe, Seung H; Tsui, Jonathan H; Lam, Christoffer H; Ho, Derek S; Baker, Aaron B; Kim, Deok-Ho

    2014-08-01

    Vascular smooth muscle cells (vSMCs) retain the ability to undergo modulation in their phenotypic continuum, ranging from a mature contractile state to a proliferative, secretory state. vSMC differentiation is modulated by a complex array of microenvironmental cues, which include the biochemical milieu of the cells and the architecture and stiffness of the extracellular matrix. In this study, we demonstrate that by using UV-assisted capillary force lithography (CFL) to engineer a polyurethane substratum of defined nanotopography and stiffness, we can facilitate the differentiation of cultured vSMCs, reduce their inflammatory signature, and potentially promote the optimal functioning of the vSMC contractile and cytoskeletal machinery. Specifically, we found that the combination of medial tissue-like stiffness (11 MPa) and anisotropic nanotopography (ridge width_groove width_ridge height of 800_800_600 nm) resulted in significant upregulation of calponin, desmin, and smoothelin, in addition to the downregulation of intercellular adhesion molecule-1, tissue factor, interleukin-6, and monocyte chemoattractant protein-1. Further, our results allude to the mechanistic role of the RhoA/ROCK pathway and caveolin-1 in altered cellular mechanotransduction pathways via differential matrix nanotopography and stiffness. Notably, the nanopatterning of the stiffer substrata (1.1 GPa) resulted in the significant upregulation of RhoA, ROCK1, and ROCK2. This indicates that nanopatterning an 800_800_600 nm pattern on a stiff substratum may trigger the mechanical plasticity of vSMCs resulting in a hypercontractile vSMC phenotype, as observed in diabetes or hypertension. Given that matrix stiffness is an independent risk factor for cardiovascular disease and that CFL can create different matrix nanotopographic patterns with high pattern fidelity, we are poised to create a combinatorial library of arterial test beds, whether they are healthy, diseased, injured, or aged. Such

  16. Orphan nuclear receptor small heterodimer partner inhibits angiotensin II-stimulated PAI-1 expression in vascular smooth muscle cells

    OpenAIRE

    Lee, Kyeong-Min; Seo, Hye-Young; Kim, Mi-Kyung; Min, Ae-Kyung; Ryu, Seong-Yeol; Kim, Yoon-Nyun; Park, Young Joo; Choi, Hueng-Sik; Lee, Ki-Up; Park, Wan-Ju; Park, Keun-Gyu; Lee, In-Kyu

    2009-01-01

    Angiotensin II is a major effector molecule in the development of cardiovascular disease. In vascular smooth muscle cells (VSMCs), angiotensin II promotes cellular proliferation and extracellular matrix accumulation through the upregulation of plasminogen activator inhibitor-1 (PAI-1) expression. Previously, we demonstrated that small heterodimer partner (SHP) represses PAI-1 expression in the liver through the inhibition of TGF-β signaling pathways. Here, we investigated whether SHP inhibite...

  17. Inhibition of apoptotic signaling in spermine-treated vascular smooth muscle cells by a novel glutathione precursor

    OpenAIRE

    Sinha-Hikim, Indrani; Shen, Ruoqing; Kovacheva, Ekaterina; Crum, Albert; Vaziri, Nosratola D.; Norris, Keith C.

    2010-01-01

    Chronic kidney disease (CKD) is a public health problem, mediated by hemodynamic and non-hemodynamic events including oxidative stress. We investigated the effect of two glutathione (GSH) precursors, N-acetyl-cysteine (NAC) and cystine as the physiologic carrier of cysteine in GSH with added selenomethionine (F1) in preventing spermine (uremic toxin) induced apoptosis in cultured human aortic vascular smooth muscle cells (VSMC). VSMCs exposed to spermine (15 μM) with or without antioxidants (...

  18. Atrophin Proteins Interact with the Fat1 Cadherin and Regulate Migration and Orientation in Vascular Smooth Muscle Cells*S⃞

    OpenAIRE

    Hou, Rong; Sibinga, Nicholas E. S.

    2009-01-01

    Fat1, an atypical cadherin induced robustly after arterial injury, has significant effects on mammalian vascular smooth muscle cell (VSMC) growth and migration. The related Drosophila protein Fat interacts genetically and physically with Atrophin, a protein essential for development and control of cell polarity. We hypothesized that interactions between Fat1 and mammalian Atrophin (Atr) proteins might contribute to Fat1 effects on VSMCs. Like Fat1, mammalian Atr expres...

  19. TLR4-Activated MAPK-IL-6 Axis Regulates Vascular Smooth Muscle Cell Function

    Science.gov (United States)

    Lee, Guan-Lin; Wu, Jing-Yiing; Tsai, Chien-Sung; Lin, Chih-Yuan; Tsai, Yi-Ting; Lin, Chin-Sheng; Wang, Yi-Fu; Yet, Shaw-Fang; Hsu, Yu-Juei; Kuo, Cheng-Chin

    2016-01-01

    Migration of vascular smooth muscle cells (VSMCs) into the intima is considered to be a vital event in the pathophysiology of atherosclerosis. Despite substantial evidence supporting the pathogenic role of Toll-like receptor 4 (TLR4) in the progression of atherogenesis, its function in the regulation of VSMC migration remains unclear. The goal of the present study was to elucidate the mechanism by which TLR4 regulates VSMC migration. Inhibitor experiments revealed that TLR4-induced IL-6 secretion and VSMC migration were mediated via the concerted actions of MyD88 and TRIF on the activation of p38 MAPK and ERK1/2 signaling. Neutralizing anti-IL-6 antibodies abrogated TLR4-driven VSMC migration and F-actin polymerization. Blockade of p38 MAPK or ERK1/2 signaling cascade inhibited TLR4 agonist-mediated activation of cAMP response element binding protein (CREB). Moreover, siRNA-mediated suppression of CREB production repressed TLR4-induced IL-6 production and VSMC migration. Rac-1 inhibitor suppressed TLR4-driven VSMC migration but not IL-6 production. Importantly, the serum level of IL-6 and TLR4 endogenous ligand HMGB1 was significantly higher in patients with coronary artery diseases (CAD) than in healthy subjects. Serum HMGB1 level was positively correlated with serum IL-6 level in CAD patients. The expression of both HMGB1 and IL-6 was clearly detected in the atherosclerotic tissue of the CAD patients. Additionally, there was a positive association between p-CREB and HMGB1 in mouse atherosclerotic tissue. Based on our findings, we concluded that, upon ligand binding, TLR4 activates p38 MAPK and ERK1/2 signaling through MyD88 and TRIF in VSMCs. These signaling pathways subsequently coordinate an additive augmentation of CREB-driven IL-6 production, which in turn triggers Rac-1-mediated actin cytoskeleton to promote VSMC migration. PMID:27563891

  20. Drug packaging and delivery using perfluorocarbon nanoparticles for targeted inhibition of vascular smooth muscle cells

    Institute of Scientific and Technical Information of China (English)

    Zhao-xiong ZHOU; Bai-gen ZHANG; Hao ZHANG; Xiao-zhong HUANG; Ya-li HU; Li SUN; Xiao-min WANG; Ji-wei ZHANG

    2009-01-01

    Aim: To investigate the in vitro release profile of drugs encapsulated within perfluorocarbon (PFC) nanoparticles (NPs) and their ability to inhibit the activity of vascular smooth muscle cells (SMCs).Methods: Dexamethasone phosphate (DxP) or dexamethasone acetate (DxA) was encapsulated into PFC nanoparticles using a high-pressure homogenous method. The morphology and size of the NPs were examined using scanning electron microscopy (SEM) and a laser particle size analyzer. Drug loading and in vitro release were assessed by high-performance liquid chromatography (HPLC). The impact of NP capsules on SMC proliferation, migration and apoptosis in vitro was assessed using cell counting kit-8, transwell cell migration and flow cytometry assays.Results: The sizes of DxP-NPs and DxA-NPs were 224±6 nm and 236±9 nm, respectively. The encapsulation efficiency (EE) of DxP-NPs was 66.4%±1.0%, with an initial release rate of 77.2%, whereas the EE of DxA-NPs was 95.3%±1.3%, with an initial release rate of 23.6%. Both of the NP-coated drugs could be released over 7 d. Human umbilical artery SMCs were harvested and cultured for four to six passages. Compared to free DxP, SMCs treated with tissue factor (TF)-directed DxP-NPs showed significant differences in the inhibition of proliferation, migration and apoptosis (P<0.05).Conclusion: The results collectively suggest that PFC nanoparticles will be beneficial for targeted drug delivery because of the sustained drug release and effective inhibition of SMC proliferation and migration.

  1. 45Ca distribution and transport in saponin skinned vascular smooth muscle

    International Nuclear Information System (INIS)

    45Ca distribution and transport were studied in chemically skinned strips of caudal artery from Kyoto Wistar rats. Sarcolemmal membranes were made hyperpermeable by exposure for 60 min to solutions containing 0.1 mg/ml of saponin. Skinned helical strips responded with graded contractions to changes in ethylene glycol bis-(beta-aminoethyl ether)-N,N'-tetraacetic acid buffered free Ca solutions (10(-7) to 10(-5) M) and were sensitive to the Mg-ATP concentration. Tissues loaded in the presence of 10(-7) M Ca contracted in response to 10 mM caffeine. These experiments indicate the strips are skinned and possess a functional regulatory and contractile system and an intact Ca sequestering system. 45Ca distributes in three compartments in skinned caudal artery strips. The Ca contents of two components are linear functions of the Ca-ethylene glycol bis-(beta-aminoethyl ether)-N,N'-tetraacetic acid concentration and desaturate at rapid rates. They correspond to the extracellular and cytoplasmic spaces. A significantly smaller component releases Ca at comparatively slower rates. 45Ca uptake by the slow component consists of an ATP-dependent and an ATP-independent fraction. The 45Ca content of the ATP-dependent fraction is a function of the free Ca concentration and is independent of the Ca-ethylene glycol bis-(beta-aminoethyl ether)-N,N'-tetraacetic acid concentration. Its content was enhanced by oxalate and was abolished by Triton X-100 skinning solutions. The ATP-independent component was not affected by Triton X-100 skinning and may represent Ca binding to cytoplasmic molecules and structures. The sequestered Ca was released with caffeine or Ca but not by epinephrine. The observations indicate that the sarcoplasmic reticulum and mitochondria of vascular smooth muscle strips skinned with saponin retain their functional integrity after saponin skinning

  2. Yangxueqingnao particles inhibit rat vascular smooth muscle cell proliferation induced by lysophosphatidic acid

    Institute of Scientific and Technical Information of China (English)

    CAI Wei; XU Yi; CHEN Jun-zhu; HUANG Shu-ru; LU Zhen-ya; WANG Zhan-kun

    2005-01-01

    Objective: To observe the effect of Yangxueqingnao particles on rat vascular smooth muscle cell (VSMC) proliferation induced by lysophosphatidic acid (LPA). Methods: The amount of3H-TdR (3H-thymidine) admixed in cultured rat VSMC was measured and mitogen-activated protein kinase (MAPK) activity and lipid peroxidation end product malondialdehyde (MDA)content of the VSMC were assayed. Results: 1×10-9, 1×10-8, 1×10-7 mol/L LPA in a concentration dependent manner, induced the amount of 3H-TdR admixed, MAP kinase activity, and MDA content of the cultured rat VSMC to increase. However, 5%, 10%,and 15% Yangxueqingnao serum preincubation resulted in a decrease of 23.0%, 42.0%, and 52.0% (P<0.01) respectively in the amount of 3H-TdR admixed, a decline in VSMC MAP kinase activity of 13.9% (P<0.05), 29.6% (P<0.01), and 48.9% (P<0.01)respectively, and also, a decrease in MDA content of VSMC of 19.4%, 24.7%, and 43.2% (P<0.01) respectively, in the 1×10-7mol/L LPA-treated VSMC. Conclusions: LPA activates the proliferation and lipid peroxidation of VSMC in a concentration dependent manner. The LPA-induced VSMC proliferation is related to the activity of MAP kinases, enzymes involved in an intracellular signalling pathway. The results of the present study showed that Yangxueqingnao particles can effectively inhibit LPA-induced VSMC proliferation, MAP kinase activation, and reduce lipid peroxidative lesion.

  3. Microarray Analysis of Human Vascular Smooth Muscle Cell Responses to Bacterial Lipopolysaccharide

    Directory of Open Access Journals (Sweden)

    Joe Minta

    2007-01-01

    Full Text Available Accumulating evidence suggest a causal role of bacterial and viral infections in atherogenesis. Bacterial lipopolysaccharide (LPS has been shown to stimulate resting vascular smooth muscle cells (SMC with the production of inflammatory cytokines and modulation of quiescent cells to the proliferative and synthetic phenotype. To comprehensively identify biologically important genes associated with LPS-induced SMC phenotype modulation, we compared the transcriptomes of quiescent human coronary artery SMC and cells treated with LPS for 4 and 22 h. The SMCs responded robustly to LPS treatment by the differential regulation of several genes involved in chromatin remodeling, transcription regulation, translation, signal transduction, metabolism, host defense, cell proliferation, apoptosis, matrix formation, adhesion and motility and suggest that the induction of clusters of genes involved in cell proliferation, migration and ECM production may be the main force that drives the LPS-induced phenotypic modulation of SMC rather than the differential expression of a single gene or a few genes. An interesting observation was the early and dramatic induction of four tightly clustered interferon-induced genes with tetratricopeptide repeats (IFIT1, 2, 4, 5. siRNA knock-down of IFIT1 in SMC was found to be associated with a remarkable up-regulation of TP53, CDKN1A and FOS, suggesting that IFIT1 may play a role in cell proliferation. Our data provide a comprehensive list of genes involved in LPS biology and underscore the important role of LPS in SMC activation and phenotype modulation which is a pivotal event in the onset of atherogenesis.

  4. Lipogenesis in arterial wall and vascular smooth muscular cells: regulation and abnormalities in insulin-resistance

    Directory of Open Access Journals (Sweden)

    Feugier Patrick

    2009-12-01

    Full Text Available Abstract Background Vascular smooth muscular cells (VSMC express lipogenic genes. Therefore in situ lipogenesis could provide fatty acids for triglycerides synthesis and cholesterol esterification and contribute to lipid accumulation in arterial wall with aging and during atheroma. Methods We investigated expression of lipogenic genes in human and rat arterial walls, its regulation in cultured VSMC and determined if it is modified during insulin-resistance and diabetes, situations with increased risk for atheroma. Results Zucker obese (ZO and diabetic (ZDF rats accumulated more triglycerides in their aortas than their respective control rats, and this triglycerides content increased with age in ZDF and control rats. However the expression in aortas of lipogenic genes, or of genes involved in fatty acids uptake, was not higher in ZDF and ZO rats and did not increase with age. Expression of lipogenesis-related genes was not increased in human arterial wall (carotid endarterectomy of diabetic compared to non-diabetic patients. In vitro, glucose and adipogenic medium (ADM stimulated moderately the expression and activity of lipogenesis in VSMC from control rats. LXR agonists, but not PXR agonist, stimulated also lipogenesis in VSMC but not in arterial wall in vivo. Lipogenic genes expression was lower in VSMC from ZO rats and not stimulated by glucose or ADM. Conclusion Lipogenic genes are expressed in arterial wall and VSMC; this expression is stimulated (VSMC by glucose, ADM and LXR agonists. During insulin-resistance and diabetes, this expression is not increased and resists to the actions of glucose and ADM. It is unlikely that this metabolic pathway contribute to lipid accumulation of arterial wall during insulin-resistance and diabetes and thus to the increased risk of atheroma observed in these situations.

  5. Taurine antagonized oxidative stress injury induced by homocysteine in rat vascular smooth muscle cells

    Institute of Scientific and Technical Information of China (English)

    Lin CHANG; Jian-xin XU; Jing ZHAO; Yong-zheng PANG; Chao-shu TANG; Yong-fen QI

    2004-01-01

    AIM: To observe protective effects of taurine on reactive oxygen species generation induced by homocysteine in rat vascular smooth muscle cells (VSMC). METHODS: Rat VSMC was incubated with various concentrations of homocysteine and taurine. The lactate dehydrogenase (LDH) activity which released into culture medium was elevated as an indicator for VSMC injury. The reactive oxygen species (ROS) - hydrogen peroxide (H2O2) and superoxide anion (O2- )were measured with luminol or lucigenin chemiluminescences method, and the mitochondria Mn-superoxide dismutase (Mn-SOD) and catalase (CAT) were also measured in treated VSMC. RESULTS: LDH leakage from cultured VSMC treated with homocystenie, was increased (P<0.01 vs control), and it was markedly inhibited when co-incubated with taurine (P<0.01). Homocysteine induced H2O2 generation from VSMC in a concentration dependent manner (P<0.01 vs control). However, taurine (5, 10, and 20 mmol/L) significantly antagonized 0.5 mmol/L homocysteine-induced H2O2 generation in VSMC in a concentration dependent manner (P<0.01 vs homocysteine alone group), although taurine itself did not alter the H2O2 generation in VSMC (P>0.05 vs control).In this study, the superoxide anion in VSMC was not detectable by chemiluminent method. In addition, treatment of VSMC with taurine increased mitochondria Mn-SOD and CAT activity in a concentration dependent manner (P<0.05), but homocysteine decreased mitochondria Mn-SOD and CAT activity (P<0.01 vs control). In addition,co-administration of taurine markedly ameliorated homocysteine-induced inhibition of Mn-SOD and CAT activity in VSMC (P<0.01 vs homocysteine alone group). CONCLUSION: Taurine antagonized the effects of homocysteine on ROS generation and anti-oxidant enzyme activities in rat VSMC in vitro.

  6. Batroxobin reduces intracellular calcium concentration and inhibits proliferation of vascular smooth muscle cells

    Institute of Scientific and Technical Information of China (English)

    SONG Qing-bin 宋清斌; WEI Min-jie 魏敏杰; DUAN Zhi-quan 段志泉; ZHANG Hai-qiang 张海强; LB Schwartz; XIN Shi-jie 辛世杰

    2004-01-01

    Background Batroxobin (BX), a serine protease used in defibrinogenation and thrombolysis, also has an effect on c-fos gene and growth factor. This study attempted to determine the effects of BX on the proliferation of vascular smooth muscle cells (VSMCs) and calcium metabolism. Methods VSMCs were treated with BX at concentrations of 0.1, 0.3, or 1.0 mmol/L and cell numbers were determined at 0, 24, 48, and 72 hours. Intracellular calcium concentration ([Ca2+]I) was measured using direct fluorescence methods. Results BX was found to suppress proliferation of VSMCs in a dose-dependent fashion with inhibition rates of 18% and 31% by 48 and 72 hours, respectively. In addition, BX decreases basal [Ca2+]I significantly. The basal level in untreated cells was 162.7±33.8 nmol/L, and decreased to 131.5±27.7 nmol/L, 128.3±28.5 nmol/L, and 125.6±34.3 nmol/L with the three concentrations of BX, respectively. Noradrenaline (NE)-induced [Ca2+]I stimulation was also attenuated by BX (0.1 mmol/L BX, 20%±8% inhibition; 0.3 mmol/L BX, 54%±11% inhibition; 1.0 mmol/L BX, 62%±15% inhibition). The ability of NE to stimulate [Ca2+]I was attenuated in cultures in Ca2+-free medium, as was the ability of BX to blunt NE-induced stimulation. Conclusion These findings demonstrate that BX can effectively inhibit proliferation of VSMCs, probably by blocking the release and uptake of Ca2+, thus influencing [Ca2+]I.

  7. Urotensin Ⅱ increases endothelin production by vascular smooth muscle cells in rats

    Institute of Scientific and Technical Information of China (English)

    2002-01-01

    The aim of this study was to observe the effects of urotensin Ⅱ (UII) on production of endothelin (ET) in rat aortic vascular smooth muscle cells (VSMC). Cultured VSMCs incubated with various concentrations of UII were used to measure the VSMC 3H-TdR incorporation, the amount of ET mRNA and ET production in VSMCs. In this work we found that UII (10-10-10-8 mol/L) promoted VSMC 3H-TdR incorporation (47%-83%, P < 0.01) and increased the amount of ET mRNA by 17.1% (P < 0.05) to 112.8% (P < 0.01), respectively, in a concentration dependent manner compared with control. After 4 and 8 h incubation, 10-10-10-8 mol/L of UII elevated the ET synthesis and release in a concentration dependent manner. After 4 h incubation, the content of ET in medium was 4.9, 5.36 and 7.12 pg/mL (P < 0.01). After 8 h incubation, the ET content released from VSMCs was 12.6, 12.07 and 17.17 pg/mL (P < 0.01). In addition, it was found that BQ123, a specific ETA receptor antagonist, obviously decreased the VSMC DNA synthesis induced by UII. The results of this study showed that UII could stimulate the ET mRNA expression and ET production in VSMC. The effects of UII on VSMC DNA synthesis were partly mediated by ET autocrine pathway. It suggests that the interaction between UII and ET plays an important biological regulating role as endogenous active peptides.

  8. 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.

  9. The tight junction protein ZO-2 and Janus kinase 1 mediate intercellular communications in vascular smooth muscle cells

    Energy Technology Data Exchange (ETDEWEB)

    Tkachuk, Natalia; Tkachuk, Sergey; Patecki, Margret [Department of Nephrology, Hannover Medical School, Hannover D-30625 (Germany); Kusch, Angelika [Department of Nephrology and Intensive Care Medicine, Charite Campus Virchow-Klinikum, Berlin D-13353 (Germany); Korenbaum, Elena; Haller, Hermann [Department of Nephrology, Hannover Medical School, Hannover D-30625 (Germany); Dumler, Inna, E-mail: dumler.inna@mh-hannover.de [Department of Nephrology, Hannover Medical School, Hannover D-30625 (Germany)

    2011-07-08

    Highlights: {yields} The tight junction protein ZO-2 associates with Jak1 in vascular smooth muscle cells via ZO-2 N-terminal fragment. {yields} Jak1 mediates ZO-2 tyrosine phosphorylation and ZO-2 localization to the sites of homotypic intercellular contacts. {yields} The urokinase receptor uPAR regulates ZO-2/Jak1 functional association. {yields} The ZO-2/Jak1/uPAR signaling complex is required for vascular smooth muscle cells functional network formation. -- Abstract: Recent evidence points to a multifunctional role of ZO-2, the tight junction protein of the MAGUK (membrane-associated guanylate kinase-like) family. Though ZO-2 has been found in cell types lacking tight junction structures, such as vascular smooth muscle cells (VSMC), little is known about ZO-2 function in these cells. We provide evidence that ZO-2 mediates specific homotypic cell-to-cell contacts between VSMC. Using mass spectrometry we found that ZO-2 is associated with the non-receptor tyrosine kinase Jak1. By generating specific ZO-2 constructs we further found that the N-terminal fragment of ZO-2 molecule is responsible for this interaction. Adenovirus-based expression of Jak1 inactive mutant demonstrated that Jak1 mediates ZO-2 tyrosine phosphorylation. By means of RNA silencing, expression of Jak1 mutant form and fluorescently labeled ZO-2 fusion protein we further specified that active Jak1, but not Jak1 inactive mutant, mediates ZO-2 localization to the sites of intercellular contacts. We identified the urokinase receptor uPAR as a pre-requisite for these cellular events. Functional requirement of the revealed signaling complex for VSMC network formation was confirmed in experiments using Matrigel and in contraction assay. Our findings imply involvement of the ZO-2 tight junction independent signaling complex containing Jak1 and uPAR in VSMC intercellular communications. This mechanism may contribute to vascular remodeling in occlusive cardiovascular diseases and in arteriogenesis.

  10. Impaired Peroxisome Proliferator-activated Receptor-γ Contributes to Phenotypic Modulation of Vascular Smooth Muscle Cells during Hypertension*

    Science.gov (United States)

    Zhang, Lili; Xie, Peng; Wang, Jingzhou; Yang, Qingwu; Fang, Chuanqin; Zhou, Shuang; Li, Jingcheng

    2010-01-01

    The phenotypic modulation of vascular smooth muscle cells (VSMCs) plays a pivotal role in hypertension-induced vascular changes including vascular remodeling. The precise mechanisms underlying VSMC phenotypic modulation remain elusive. Here we test the role of peroxisome proliferator-activated receptor (PPAR)-γ in the VSMC phenotypic modulation during hypertension. Both spontaneously hypertensive rat (SHR) aortas and SHR-derived VSMCs exhibited reduced PPAR-γ expression and excessive VSMC phenotypic modulation identified by reduced contractile proteins, α-smooth muscle actin (α-SMA) and smooth muscle 22α (SM22α), and enhanced proliferation and migration. PPAR-γ overexpression rescued the expression of α-SMA and SM22α, and inhibited the proliferation and migration in SHR-derived VSMCs. In contrast, PPAR-γ silencing exerted the opposite effect. Activating PPAR-γ using rosiglitazone in vivo up-regulated aortic α-SMA and SM22α expression and attenuated aortic remodeling in SHRs. Increased activation of phosphoinositide 3-kinase (PI3K)/protein kinase B (AKT) signaling was observed in SHR-derived VSMCs. PI3K inhibitor LY294002 rescued the impaired expression of contractile proteins, and inhibited proliferation and migration in VSMCs from SHRs, whereas constitutively active PI3K mutant had the opposite effect. Overexpression or silencing of PPAR-γ inhibited or excited PI3K/Akt activity, respectively. LY294002 counteracted the PPAR-γ silencing induced proliferation and migration in SHR-derived VSMCs, whereas active PI3K mutant had the opposite effect. In contrast, reduced proliferation and migration by PPAR-γ overexpression were reversed by the active PI3K mutant, and further inhibited by LY294002. We conclude that PPAR-γ inhibits VSMC phenotypic modulation through inhibiting PI3K/Akt signaling. Impaired PPAR-γ expression is responsible for VSMC phenotypic modulation during hypertension. These findings highlight an attractive therapeutic target for

  11. Graded effects of unregulated smooth muscle myosin on intestinal architecture, intestinal motility and vascular function in zebrafish.

    Science.gov (United States)

    Abrams, Joshua; Einhorn, Zev; Seiler, Christoph; Zong, Alan B; Sweeney, H Lee; Pack, Michael

    2016-05-01

    Smooth muscle contraction is controlled by the regulated activity of the myosin heavy chain ATPase (Myh11). Myh11 mutations have diverse effects in the cardiovascular, digestive and genitourinary systems in humans and animal models. We previously reported a recessive missense mutation, meltdown (mlt), which converts a highly conserved tryptophan to arginine (W512R) in the rigid relay loop of zebrafish Myh11. The mlt mutation disrupts myosin regulation and non-autonomously induces invasive expansion of the intestinal epithelium. Here, we report two newly identified missense mutations in the switch-1 (S237Y) and coil-coiled (L1287M) domains of Myh11 that fail to complement mlt Cell invasion was not detected in either homozygous mutant but could be induced by oxidative stress and activation of oncogenic signaling pathways. The smooth muscle defect imparted by the mlt and S237Y mutations also delayed intestinal transit, and altered vascular function, as measured by blood flow in the dorsal aorta. The cell-invasion phenotype induced by the three myh11 mutants correlated with the degree of myosin deregulation. These findings suggest that the vertebrate intestinal epithelium is tuned to the physical state of the surrounding stroma, which, in turn, governs its response to physiologic and pathologic stimuli. Genetic variants that alter the regulation of smooth muscle myosin might be risk factors for diseases affecting the intestine, vasculature, and other tissues that contain smooth muscle or contractile cells that express smooth muscle proteins, particularly in the setting of redox stress. PMID:26893369

  12. Onychin inhibits proliferation of vascular smooth muscle cells by regulating cell cycle

    Institute of Scientific and Technical Information of China (English)

    Ming YANG; Hong-lin HUANG; Bing-yang ZHU; Qin-hui TUO; Duan-fang LIAO

    2005-01-01

    Aim: To investigate the effects of onychin on the proliferation of cultured rat artery vascular smooth muscle cells (VSMCs) in the presence of 10% new-borncalf serum (NCS). Methods: Rat VSMCs were incubated with onychin 1-50 μmol/L or genistein 10 μmol/L in the presence of 10% NCS for 24 h. The proliferation of VSMCs was measured by cell counting and MTS/PMS colorimetric assays. Cell cycle progression was evaluated by flow cytometry. Retinoblastoma (Rb) phosphorylation, and expression of cyclin D1 and cyclin E were measured by Western blot assays. The tyrosine phosphorylation of ERK1/2 was examined by immunoprecipitation techniques using anti-phospho-tyrosine antibodies. Results: The proliferation of VSMCs was accelerated significantly in the presence of 10% NCS. Onychin reduced the metabolic rate of MTS and the cell number of VSMCs in the presence of 10% NCS in a dose-dependent manner. Flow cytometry analy sis revealed that the G1-phase fraction ratio in the onychin group was higher than that in the 10% NCS group (85.2% vs 70.0%, P<0.01), while the S-phase fraction ratio in the onychin group was lower than that in 10% NCS group (4.3% vs 16.4%, P<0.01). Western blot analysis showed that onychin inhibited Rb phos phorylation and reduced the expression of cyclin D1 and cyclin E. The effects of onychin on proliferation, the cell cycle and the expression of cyclins in VSMCs were similar to those of genistein, an inhibitor of tyrosine kinase. Furthermore immunoprecipitation studies showed that both onychin and genistein markedly inhibited the tyrosine phosphorylation of ERK1/2 induced by 10% NCS.Conclusion: Onychin inhibits the proliferation of VSMCs through G1 phase cell cycle arrest by decreasing the tyrosine phosphorylation of ERK1/2, and the expression of cyclin D1 and cyclin E, and sequentially inhibiting Rb phosphorylation.

  13. Extravillous trophoblast cells-derived exosomes promote Vascular Smooth Muscle Cell Migration

    Directory of Open Access Journals (Sweden)

    Carlos eSalomon

    2014-08-01

    Full Text Available Background: Vascular smooth muscle cells (VSMCs migration is a critical process during human uterine spiral artery (SpA remodeling and a successful pregnancy. Extravillous trophoblast cells (EVT interact with VSMC and enhance their migration, however, the mechanisms by which EVT remodel SpA remain to be fully elucidated. We hypothesize that exosomes released from EVT promote VSMC migration.Methods: JEG-3 and HTR-8/SVneo cell lines were used as models for EVT. Cells were cultured at 37 0C and humidified under an atmosphere of 5% CO2-balanced N2 to obtain 8% O2. Cell-conditioned media were collected and exosomes (exo-JEG-3 and exo- HTR-8/SVneo isolated by differential and buoyant density centrifugation. The effects of exo-EVT on VSMC migration were established using a real-time, live-cell imaging system (Incucyte™. Exosomal proteins where identified by mass spectrometry and submitted to bioinformatic pathway analysis (Ingenuity software .Results: HTR-8/SVneo cells were significantly more (~30% invasive than JEG-3 cells. HTR-8/SVneo cells released 2.6-fold more exosomes (6.39 x 108 ± 2.5 x108 particles/106 cells compared to JEG-3 (2.86 x 108 ± 0.78 x108 particles/106 cells. VSMC migration was significantly increased in the presence of exo-JEG-3 and exo-HTR-8/SVneo compared to control (-exosomes (21.83 ± 0.49 h and 15.57 ± 0.32, respectively, versus control 25.09 ± 0.58 h, p<0.05. Sonication completely abolished the effect of exosomes on VSMC migration. Finally, mass spectrometry analysis identified unique exosomal proteins for each EVT cell line-derived exosomes.Conclusion: The data obtained in this study are consistent with the hypothesis that the release, content and bioactivity of exosomes derived from EVT-like cell lines is cell origin-dependent and differentially regulates VSMC migration. Thus, an EVT exosomal signaling pathway may contribute to SpA remodeling by promoting the migration of VSMC out of the vessel walls.

  14. 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

  15. Vascular progenitor cells isolated from human embryonic stem cells give rise to endothelial and smooth muscle like cells and form vascular networks in vivo.

    Science.gov (United States)

    Ferreira, Lino S; Gerecht, Sharon; Shieh, Hester F; Watson, Nicki; Rupnick, Maria A; Dallabrida, Susan M; Vunjak-Novakovic, Gordana; Langer, Robert

    2007-08-01

    We report that human embryonic stem cells contain a population of vascular progenitor cells that have the ability to differentiate into endothelial-like and smooth muscle (SM)-like cells. Vascular progenitor cells were isolated from EBs grown in suspension for 10 days and were characterized by expression of the endothelial/hematopoietic marker CD34 (CD34+ cells). When these cells are subsequently cultured in EGM-2 (endothelial growth medium) supplemented with vascular endothelial growth factor-165 (50 ng/mL), they give rise to endothelial-like cells characterized by a cobblestone cell morphology, expression of endothelial markers (platelet endothelial cell-adhesion molecule-1, CD34, KDR/Flk-1, vascular endothelial cadherin, von Willebrand factor), incorporation of acetylated low-density lipoprotein, and formation of capillary-like structures when placed in Matrigel. In contrast, when CD34+ cells are cultured in EGM-2 supplemented with platelet-derived growth factor-BB (50 ng/mL), they give rise to SM-like cells characterized by spindle-shape morphology, expression of SM cell markers (alpha-SM actin, SM myosin heavy chain, calponin, caldesmon, SM alpha-22), and the ability to contract and relax in response to common pharmacological agents such as carbachol and atropine but rarely form capillary-like structures when placed in Matrigel. Implantation studies in nude mice show that both cell types contribute to the formation of human microvasculature. Some microvessels contained mouse blood cells, which indicates functional integration with host vasculature. Therefore, the vascular progenitors isolated from human embryonic stem cells using methods established in the present study could provide a means to examine the mechanisms of endothelial and SM cell development, and they could also provide a potential source of cells for vascular tissue engineering.

  16. 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.

  17. Andrographolide Inhibits Nuclear Factor-κB Activation through JNK-Akt-p65 Signaling Cascade in Tumor Necrosis Factor-α-Stimulated Vascular Smooth Muscle Cells

    OpenAIRE

    Yu-Ying Chen; Ming-Jen Hsu; Cheng-Ying Hsieh; Lin-Wen Lee; Zhih-Cherng Chen; Joen-Rong Sheu

    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 proinfl...

  18. Effects of Raloxifene on Caveolin-1 mRNA and Protein Expressions in Vascular Smooth Muscle Cells

    Institute of Scientific and Technical Information of China (English)

    Fa-Lin YANG; Hong HE; Xian-Xi LIU; Bing TU; Xian-Wei ZENG; Ji-Xin SU; Xin WANG; Qin HU

    2006-01-01

    Caveolin-1 is regulated by estrogen in vascular smooth muscle cells. Raloxifene, a selectiveestrogen receptor modulator that possibly has cardioprotective properties without an increased risk of c ancer or other side effects of estrogen, may be used in women with risk of coronary artery disease. However, the relationship between raloxifene and caveolin-1 is still unknown. Therefore, this study was designed to see whether raloxifene regulates caveolin- 1 expression and if so, whether such regulation is mediated by estrogen receptor. Rat aortic smooth muscle cells were cultured in the absence or presence of raloxifene (10-8 to 10-6 M) for 12 or 24 h. Both mRNA and protein levels of caveolin-1 were increased significantly after 24 h treatment with raloxifene. These increases were inhibited by estrogen receptor antagonist ICI 182780 (10-5 M). Results of this study suggest that raloxifene stimulates caveolin- 1 transcription and translation through estrogen receptor mediated mechanisms.

  19. UDP acts as a growth factor for vascular smooth muscle cells by activation of P2Y(6) receptors

    DEFF Research Database (Denmark)

    Hou, Mingyan; Harden, T Kendall; Kuhn, Cynthia M;

    2002-01-01

    Mitogenic effects of the extracellular nucleotides ATP and UTP are mediated by P2Y(1), P2Y(2), and P2Y(4) receptors. However, it has not been possible to examine the highly expressed UDP-sensitive P2Y(6) receptor because of the lack of stable, selective agonists. In rat aorta smooth muscle cells...... (vascular smooth muscle cells; VSMC), UDP and UTP stimulated (3)H-labeled thymidine incorporation with similar pEC(50) values (5.96 and 5.69). Addition of hexokinase did not reduce the mitogenic effect of UDP. In cells transfected with P2Y receptors the stable pyrimidine agonist uridine 5'-O-(2...

  20. 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; P<0.05) at all examined time points (2 to 24 hours). mRNA half-life studies showed that this response was not due to increased mRNA instability. tPA mRNA expression was decreased (to 10% of stationary control; P<0.05) by low shear stress after 12 hours of exposure and was increased (to 250% of stationary control; P<0.05) after 24 hours at high shear stress. The same trends in PAR-1 mRNA levels were observed in rat smooth muscle 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.

  1. Graded effects of unregulated smooth muscle myosin on intestinal architecture, intestinal motility and vascular function in zebrafish

    Directory of Open Access Journals (Sweden)

    Joshua Abrams

    2016-05-01

    Full Text Available Smooth muscle contraction is controlled by the regulated activity of the myosin heavy chain ATPase (Myh11. Myh11 mutations have diverse effects in the cardiovascular, digestive and genitourinary systems in humans and animal models. We previously reported a recessive missense mutation, meltdown (mlt, which converts a highly conserved tryptophan to arginine (W512R in the rigid relay loop of zebrafish Myh11. The mlt mutation disrupts myosin regulation and non-autonomously induces invasive expansion of the intestinal epithelium. Here, we report two newly identified missense mutations in the switch-1 (S237Y and coil-coiled (L1287M domains of Myh11 that fail to complement mlt. Cell invasion was not detected in either homozygous mutant but could be induced by oxidative stress and activation of oncogenic signaling pathways. The smooth muscle defect imparted by the mlt and S237Y mutations also delayed intestinal transit, and altered vascular function, as measured by blood flow in the dorsal aorta. The cell-invasion phenotype induced by the three myh11 mutants correlated with the degree of myosin deregulation. These findings suggest that the vertebrate intestinal epithelium is tuned to the physical state of the surrounding stroma, which, in turn, governs its response to physiologic and pathologic stimuli. Genetic variants that alter the regulation of smooth muscle myosin might be risk factors for diseases affecting the intestine, vasculature, and other tissues that contain smooth muscle or contractile cells that express smooth muscle proteins, particularly in the setting of redox stress.

  2. Modification of intracellular free calcium in cultured A10 vascular smooth muscle cells by exogenous phosphatidic acid.

    Science.gov (United States)

    Bhugra, Praveen; Xu, Yan-Jun; Rathi, Satyajeet; Dhalla, Naranjan S

    2003-06-15

    Exogenous phosphatidic acid (PA) was observed to produce a concentration-dependent increase in [Ca(2+)](i) in cultured A10 vascular smooth muscle cells. Preincubation of cells with sarcoplasmic reticulum Ca(2+)-ATPase inhibitors (cyclopiazonic acid and thapsigargin), a phospholipase C inhibitor (2-nitro-4-carboxyphenyl-N,N-diphenylcarbamate), inositol 1,4,5-trisphosphate receptor antagonists (2-aminoethoxydiphenyl borate and xestospongin), and an activator of protein kinase C (PKC) (phorbol 12-myristate 13-acetate) depressed the PA-evoked increase in [Ca(2+)](i). Although EGTA, an extracellular Ca(2+) chelator, decreased the PA-induced increase in [Ca(2+)](i), sarcolemmal Ca(2+)-channel blockers (verapamil or diltiazem) did not alter the action of PA. On the other hand, inhibitors of PKC (bisindolylmaleimide I) and G(i)-protein (pertussis toxin) potentiated the increase in [Ca(2+)](i) evoked by PA significantly. These results suggest that the PA-induced increase in [Ca(2+)](i) in vascular smooth muscle cells may occur upon the activation of phospholipase C and the subsequent release of Ca(2+) from the inositol 1,4,5-trisphosphate-sensitive Ca(2+) pool in the sarcoplasmic reticulum. This action of PA may be mediated through the involvement of PKC. PMID:12787890

  3. Modification of intracellular free calcium in cultured A10 vascular smooth muscle cells by exogenous phosphatidic acid.

    Science.gov (United States)

    Bhugra, Praveen; Xu, Yan-Jun; Rathi, Satyajeet; Dhalla, Naranjan S

    2003-06-15

    Exogenous phosphatidic acid (PA) was observed to produce a concentration-dependent increase in [Ca(2+)](i) in cultured A10 vascular smooth muscle cells. Preincubation of cells with sarcoplasmic reticulum Ca(2+)-ATPase inhibitors (cyclopiazonic acid and thapsigargin), a phospholipase C inhibitor (2-nitro-4-carboxyphenyl-N,N-diphenylcarbamate), inositol 1,4,5-trisphosphate receptor antagonists (2-aminoethoxydiphenyl borate and xestospongin), and an activator of protein kinase C (PKC) (phorbol 12-myristate 13-acetate) depressed the PA-evoked increase in [Ca(2+)](i). Although EGTA, an extracellular Ca(2+) chelator, decreased the PA-induced increase in [Ca(2+)](i), sarcolemmal Ca(2+)-channel blockers (verapamil or diltiazem) did not alter the action of PA. On the other hand, inhibitors of PKC (bisindolylmaleimide I) and G(i)-protein (pertussis toxin) potentiated the increase in [Ca(2+)](i) evoked by PA significantly. These results suggest that the PA-induced increase in [Ca(2+)](i) in vascular smooth muscle cells may occur upon the activation of phospholipase C and the subsequent release of Ca(2+) from the inositol 1,4,5-trisphosphate-sensitive Ca(2+) pool in the sarcoplasmic reticulum. This action of PA may be mediated through the involvement of PKC.

  4. Chinese Yellow Wine Inhibit Production of Homocysteine-induced Extracellular Matrix Metalloproteinase-2 in Cultured Rat Vascular Smooth Muscle Cells

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    Objectives Regular consumption of moderate amounts of Chinese yellow wine is associated with a reduced risk of coronary disease.Matrix metalloproteinases (MMPs) that participate in extracellular matrix degradation have been involved in atherosclerotic plaque growth and instability. The present research aimed to study the effects of Chinese yellow wine on the production of homocysteineinduced extracellular MMP-2 in cultured rats' vascular smooth muscle cells. Methods The effects of different homocysteine levels (0-1000 μmol/l) on MMP-2 production, and the effects of Chinese yellow wine with low alcohol concentrations (12-19% ) on homocysteine-induced MMP-2 in cultured rat vascular smooth muscle cells (VSMCs) were examined using gelatin zymography and western blotting. The changes of MMP-2 under various treatments for 12h, 24h and 48 h were further compared. Results Homocysteine (50-1000 μmol/l) increased the production of MMP-2 significantly in a dose-dependent manner. Increased production of MMP-2 induced by homocysteine was reduced by extracellularly added Chinese yellow wine.Production of MMP-2 under various treatments for 48 h increased more than 12 h and 24 h. Conclusions Extracellularly added Chinese yellow wine decreased homocysteine-induced MMP-2 secretion. The inhibitory effect of yellow wine on the activation of MMP-2 might contribute to their beneficial effects on the cardiovascular system.

  5. Rosiglitzone suppresses angiotensin II-induced production of KLF5 and cell proliferation in rat vascular smooth muscle cells.

    Directory of Open Access Journals (Sweden)

    Dengfeng Gao

    Full Text Available Krüppel-like factor (KLF 5, which initiates vascular smooth muscle cell (VSMC proliferation, also participates in Angiotensin (Ang II-induced vascular remodeling. The protective effect of rosiglitazone on vascular remodeling may be due to their impact on VSMC proliferation. However, the underlying mechanisms involved remain unclear. This study was designed to investigate whether the antiproliferation effects of rosiglitazone are mediated by regulating Ang II/KLF5 response. We found that, in aortas of Ang II-infused rats, vascular remodeling and KLF5 expression were markedly increased, and its target gene cyclin D1 was overexpressed. Co-treatment with rosiglitazone diminished these changes. In growth-arrested VSMCs, PPAR-γ agonists (rosiglitazone and 15d-PGJ2 dose-dependently inhibited Ang II-induced cell proliferation and expression of KLF5 and cyclin D1. Moreover, these effects were attenuated by the PPAR-γ antagonists GW9662, bisphenol A diglycidyl ether and PPAR-γ specific siRNA. Furthermore, rosiglitazone inhibited Ang II-induced phosphorylation of protein kinase C (PKC ζ and extracellular signal-regulated kinase (ERK 1/2 and activation of early growth response protein (Egr. In conclusion, in Ang II-stimulated VSMCs, rosiglitazone might have an antiproliferative effect through mechanisms that include reducing KLF5 expression, and a crosstalk between PPAR-γ and PKCζ/ERK1/2/Egr may be involved in. These findings not only provide a previously unrecognized mechanism by which PPAR-γ agonists inhibit VSMC proliferation, but also document a novel evidence for the beneficial vascular effect of PPAR-γ activation.

  6. Rosiglitzone Suppresses Angiotensin II-Induced Production of KLF5 and Cell Proliferation in Rat Vascular Smooth Muscle Cells

    Science.gov (United States)

    Gao, Dengfeng; Hao, Guanghua; Meng, Zhe; Ning, Ning; Yang, Guang; Liu, Zhongwei; Dong, Xin; Niu, Xiaolin

    2015-01-01

    Krüppel-like factor (KLF) 5, which initiates vascular smooth muscle cell (VSMC) proliferation, also participates in Angiotensin (Ang) II-induced vascular remodeling. The protective effect of rosiglitazone on vascular remodeling may be due to their impact on VSMC proliferation. However, the underlying mechanisms involved remain unclear. This study was designed to investigate whether the antiproliferation effects of rosiglitazone are mediated by regulating Ang II/KLF5 response. We found that, in aortas of Ang II-infused rats, vascular remodeling and KLF5 expression were markedly increased, and its target gene cyclin D1 was overexpressed. Co-treatment with rosiglitazone diminished these changes. In growth-arrested VSMCs, PPAR-γ agonists (rosiglitazone and 15d-PGJ2) dose-dependently inhibited Ang II-induced cell proliferation and expression of KLF5 and cyclin D1. Moreover, these effects were attenuated by the PPAR-γ antagonists GW9662, bisphenol A diglycidyl ether and PPAR-γ specific siRNA. Furthermore, rosiglitazone inhibited Ang II-induced phosphorylation of protein kinase C (PKC) ζ and extracellular signal-regulated kinase (ERK) 1/2 and activation of early growth response protein (Egr). In conclusion, in Ang II-stimulated VSMCs, rosiglitazone might have an antiproliferative effect through mechanisms that include reducing KLF5 expression, and a crosstalk between PPAR-γ and PKCζ/ERK1/2/Egr may be involved in. These findings not only provide a previously unrecognized mechanism by which PPAR-γ agonists inhibit VSMC proliferation, but also document a novel evidence for the beneficial vascular effect of PPAR-γ activation. PMID:25874449

  7. Doxorubicin-induced vasomotion and [Ca~(2+)]_i elevation in vascular smooth muscle cells from C57BL/6 mice

    Institute of Scientific and Technical Information of China (English)

    Bing SHEN; Chun-ling YE; Kai-he YE; Lan ZHUANG; Jia-hua JIANG

    2009-01-01

    Aim: To explore the action of doxorubicin on vascular smooth muscle cells.Methods: Isometric tension of denuded or intact thoracic aortic vessels was recorded and [Ca~(2+)]_i in isolated aortic smooth muscle cells was measured by using Fluo-3.Results: Doxorubicin induced phasic and tonic contractions in denuded vessels and increased levels of [Ca~(2+)]_i in single muscle cells. Treatment with 10 μmol/L ryanodine had no effect on basal tension, but it did abolish doxorubicin-induced phasic contraction. Treatment with 10 mmol/L caffeine induced a transient phasic contraction only, and the effect was not significantly altered by ryanodine, the omission of extracellular Ca~(2+) or both. Phenylephrine induced rhythmic contraction (RC) in intact vessels. Treatment with 100 μmol/L doxorubicin enhanced RC amplitude, but 1 mmol/L doxorubicin abolished RC, with an increase in maximal tension. Caffeine at 100 μmol/L increased the frequency of the RC only. In the presence of 100 μmol/L caffeine, however, 100 μmol/L doxorubicin abolished the RC and decreased its maximal tension. Treatment with 10 μmol/L ryanodine abolished the RC, with an increase in the maximal tension. In Ca~(2+)-free solution, doxorubicin induced a transient [Ca~(2+)]_i increase that could be abolished by ryanodine pretreatment in single muscle cells. The doxorubicin-induced increase in [Ca~(2+)]_i was suppressed by nifedipine and potentiated by ryanodine and cha-rybdotoxin.Conclusion: Doxorubicin not only releases Ca~(2+) from the sarcoplasmic reticulum but also promotes the entry of extracellular Ca~(2+) into vascular smooth muscle cells.

  8. Leptin augments recruitment of IRF-1 and CREB to thrombospondin-1 gene promoter in vascular smooth muscle cells in vitro.

    Science.gov (United States)

    Sahu, Soumyadip; Ganguly, Rituparna; Raman, Priya

    2016-08-01

    We previously reported that high pathophysiological concentrations of leptin, the adipocyte-secreted peptide, upregulate the expression of a potent proatherogenic matricellular protein, thrombospondin-1 (TSP-1), in vascular smooth muscle cells. Moreover, this regulation was found to occur at the level of transcription; however, the underlying molecular mechanisms remain unknown. The goal of the present study was to investigate the specific transcriptional mechanisms that mediate upregulation of TSP-1 expression by leptin. Primary human aortic smooth muscle cell cultures were transiently transfected with different TSP-1 gene (THBS1) promoter-linked luciferase reporter constructs, and luciferase activity in response to leptin (100 ng/ml) was assessed. We identified a long THBS1 promoter (-1270/+750) fragment with specific leptin response elements that are required for increased TSP-1 transcription by leptin. Promoter analyses, protein/DNA array and gel shift assays demonstrated activation and association of transcription factors, interferon regulatory factor-1 (IRF-1) and cAMP response element-binding protein (CREB), to the distal fragment of the THBS1 promoter in response to leptin. Supershift, chromatin immunoprecipitation, and coimmunoprecipitation assays revealed formation of a single complex between IRF-1 and CREB in response to leptin; importantly, recruitment of this complex to the THBS1 promoter mediated leptin-induced TSP-1 transcription. Finally, binding sequence decoy oligomer and site-directed mutagenesis revealed that regulatory elements for both IRF-1 (-1019 to -1016) and CREB (-1198 to -1195), specific to the distal THBS1 promoter, were required for leptin-induced TSP-1 transcription. Taken together, these findings demonstrate that leptin promotes a cooperative association between IRF-1 and CREB on the THBS1 promoter driving TSP-1 transcription in vascular smooth muscle cells. PMID:27281481

  9. An Egr-1-specific DNAzyme regulates Egr-1 and proliferating cell nuclear antigen expression in rat vascular smooth muscle cells

    Science.gov (United States)

    ZHANG, JUNBIAO; GUO, CHANGLEI; WANG, RAN; HUANG, LULI; LIANG, WANQIAN; LIU, RUNNAN; SUN, BING

    2013-01-01

    The aim of the present study was to transfect rat aortic smooth muscle cells with an early growth response factor-1 (Egr-1)-specific DNAzyme (ED5), to observe its effect on Egr-1 and proliferating cell nuclear antigen (PCNA) expression and to elucidate the mechanism of ED5-mediated inhibition of vascular smooth muscle cell (VSMC) proliferation. VSMCs in primary culture obtained by tissue block adhesion were identified by morphological observation and α smooth muscle actin (α-SM-actin) immunocytochemistry. The cells were then transfected with ED5 or scrambled ED5 (ED5SCR). The three groups of cells used in the present study were the control group, ED5 group and ED5SCR group. The expression levels of Egr-1 and PCNA protein were detected following transfection by analyzing and calculating the integral optical density value in each group. Primary culture of VSMCs and transfection of ED5 and ED5SCR were successfully accomplished. Following stimulation with 10% fetal calf serum, the Egr-1 protein was expressed most strongly at 1 h and demonstrated a declining trend over time; the expression of PCNA protein began at 4 h, peaked at 24 h and then demonstrated a slightly declining trend over time. Compared with the control group and the ED5SCR group, ED5 inhibited the expression of Egr-1 and PCNA (P<0.05). ED5 was able to inhibit the expression of Egr-1 and PCNA proteins in VSMCs to a certain extent and VSMC proliferation in vitro. DNAzyme gene therapy may be useful as a new method for treating vascular proliferative diseases, including atherosclerosis and restenosis. PMID:23737882

  10. Activation of the retinoid X receptor modulates angiotensin II-induced smooth muscle gene expression and inflammation in vascular smooth muscle cells.

    Science.gov (United States)

    Lehman, Allison M B; Montford, John R; Horita, Henrick; Ostriker, Allison C; Weiser-Evans, Mary C M; Nemenoff, Raphael A; Furgeson, Seth B

    2014-11-01

    The retinoid X receptor (RXR) partners with numerous nuclear receptors, such as the peroxisome proliferator activated receptor (PPAR) family, liver X receptors (LXRs), and farnesoid X receptor (FXR). Although each heterodimer can be activated by specific ligands, a subset of these receptors, defined as permissive nuclear receptors, can also be activated by RXR agonists known as rexinoids. Many individual RXR heterodimers have beneficial effects in vascular smooth muscle cells (SMCs). Because rexinoids can potently activate multiple RXR pathways, we hypothesized that treating SMCs with rexinoids would more effectively reverse the pathophysiologic effects of angiotensin II than an individual heterodimer agonist. Cultured rat aortic SMCs were pretreated with either an RXR agonist (bexarotene or 9-cis retinoic acid) or vehicle (dimethylsulfoxide) for 24 hours before stimulation with angiotensin II. Compared with dimethylsulfoxide, bexarotene blocked angiotensin II-induced SM contractile gene induction (calponin and smooth muscle-α-actin) and protein synthesis ([(3)H]leucine incorporation). Bexarotene also decreased angiotensin II-mediated inflammation, as measured by decreased expression of monocyte chemoattractant protein-1 (MCP-1). Activation of p38 mitogen-activated protein (MAP) kinase but not extracellular signal-related kinase (ERK) or protein kinase B (Akt) was also blunted by bexarotene. We compared bexarotene to five agonists of nuclear receptors (PPARα, PPARγ, PPARδ, LXR, and FXR). Bexarotene had a greater effect on calponin reduction, MCP-1 inhibition, and p38 MAP kinase inhibition than any individual agonist. PPARγ knockout cells demonstrated blunted responses to bexarotene, indicating that PPARγ is necessary for the effects of bexarotene. These data demonstrate that RXR is a potent modulator of angiotensin II-mediated responses in the vasculature, partially through inhibition of p38. PMID:25169989

  11. Characterization of vascular smooth muscle cell phenotype in long-term culture.

    Science.gov (United States)

    Absher, M; Woodcock-Mitchell, J; Mitchell, J; Baldor, L; Low, R; Warshaw, D

    1989-02-01

    Studies of bovine carotid artery smooth muscle cells, during long-term in vitro subcultivation (up to 100 population doublings), have revealed phenotypic heterogeneity among cells, as characterized by differences in proliferative behavior, cell morphology, and contractile-cytoskeletal protein profiles. In vivo, smooth muscle cells were spindle-shaped and expressed desmin and alpha-smooth muscle actin (50% of total actin) as their predominant cytoskeletal and contractile proteins. Within 24 h of culture, vimentin rather than desmin was the predominant intermediate filament protein, with little change in alpha-actin content. Upon initial subcultivation, all cells were flattened and fibroblastic in appearance with a concomitant fivefold reduction in alpha-actin content, whereas the beta and gamma nonmuscle actins predominated. In three out of four cell lines studied, fluctuations in proliferative activity were observed during the life span of the culture. These spontaneous fluctuations in proliferation were accompanied by coordinated changes in morphology and contractile-cytoskeletal protein profiles. During periods of enhanced proliferation a significant proportion of cells reverted to their original spindle-shaped morphology with a simultaneous increase in alpha-actin content (20 to 30% of total actin). These results suggest that in long-term culture smooth muscle cells undergo spontaneous modulations in cell phenotype and may serve as a useful model for studying the regulation of intracellular protein expression.

  12. Static pressure drives proliferation of vascular smooth muscle cells via caveolin-1/ERK1/2 pathway

    Energy Technology Data Exchange (ETDEWEB)

    Luo, Di-xian, E-mail: luodixian_2@163.com [Division of Pharmacoproteomics, Institute of Pharmacy and Pharmacology, Research Center of Life Science, University of South China, Hengyang, Hunan 421001 (China); Department of Pharmacology, School of Pharmaceutics, Central South University, Changsha, Hunan 410083 (China); The First People' s Hospital of Chenzhou City, Chenzhou, Hunan 421001 (China); Cheng, Jiming [Internal Medicine and SimmonsCooper Cancer Institute, Southern Illinois University School of Medicine, 911 N. Rutledge Street, Springfield, IL 62794-9626 (United States); Suzhou Health College of Technology, 20 Shuyuanxiang, Suzhou, Jiangsu 215002 (China); Xiong, Yan [Department of Pharmacology, School of Pharmaceutics, Central South University, Changsha, Hunan 410083 (China); Li, Junmo [Division of Pharmacoproteomics, Institute of Pharmacy and Pharmacology, Research Center of Life Science, University of South China, Hengyang, Hunan 421001 (China); Xia, Chenglai [Division of Pharmacoproteomics, Institute of Pharmacy and Pharmacology, Research Center of Life Science, University of South China, Hengyang, Hunan 421001 (China); School of Pharmaceutics, Southern Medical University, Guangzhou, Guangdong 510515 (China); Xu, Canxin; Wang, Chun; Zhu, Bingyang [Division of Pharmacoproteomics, Institute of Pharmacy and Pharmacology, Research Center of Life Science, University of South China, Hengyang, Hunan 421001 (China); Hu, Zhuowei [Institute of Materia Medical, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100730 (China); Liao, Duan-fang, E-mail: dfliao66@yahoo.com.cn [Division of Pharmacoproteomics, Institute of Pharmacy and Pharmacology, Research Center of Life Science, University of South China, Hengyang, Hunan 421001 (China)

    2010-01-22

    Intimal hyperplasia plays an important role in various types of vascular remodeling. Mechanical forces derived from blood flow are associated with the proliferation of vascular smooth muscle cells (VSMC). This contributes to many vascular disorders such as hypertension, atherosclerosis and restenosis after percutaneous transluminal angioplasty (PTA). In this study, we show that static pressure induces the proliferation of VSMC and activates its related signal pathway. VSMC from a rat aorta were treated with different pressures (0, 60, 90, 120, 150 and 180 mm Hg) in a custom-made pressure incubator for 24 h. The most active proliferation of VSMC was detected at a pressure of 120 mm Hg. VSMC was also incubated under a static pressure of 120 mm Hg for different time intervals (0, 2, 4, 8, 12 and 24 h). We found that static pressure significantly stimulates VSMC proliferation. Extracellular signal-regulated kinases 1/2 (ERK1/2) activation showed a peak at the pressure of 120 mm Hg at 4-h time point. Moreover, caveolin-1 expression was significantly inhibited by rising static pressure. Downregulation of VSMC proliferation could be found after PD98059 (ERK1/2 phosphorylation inhibitor) treatment. Our data also showed that a siRNA-mediated caveolin-1 knock down increased ERK1/2 phosphorylation and VSMC proliferation. These results demonstrate that static pressure promotes VSMC proliferation via the Caveolin-1/ERK1/2 pathway.

  13. Mechanical stretch increases MMP-2 production in vascular smooth muscle cells via activation of PDGFR-β/Akt signaling pathway.

    Directory of Open Access Journals (Sweden)

    Kyo Won Seo

    Full Text Available Increased blood pressure, leading to mechanical stress on vascular smooth muscle cells (VSMC, is a known risk factor for vascular remodeling via increased activity of matrix metalloproteinase (MMP within the vascular wall. This study aimed to identify cell surface mechanoreceptors and intracellular signaling pathways that influence VSMC to produce MMP in response to mechanical stretch (MS. When VSMC was stimulated with MS (0-10% strain, 60 cycles/min, both production and gelatinolytic activity of MMP-2, but not MMP-9, were increased in a force-dependent manner. MS-enhanced MMP-2 expression and activity were inhibited by molecular inhibition of Akt using Akt siRNA as well as by PI3K/Akt inhibitors, LY293002 and AI, but not by MAPK inhibitors such as PD98059, SP600125 and SB203580. MS also increased Akt phosphorylation in VSMC, which was attenuated by AG1295, a PDGF receptor (PDGFR inhibitor, but not by inhibitors for other receptor tyrosine kinase including EGF, IGF, and FGF receptors. Although MS activated PDGFR-α as well as PDGFR-β in VSMC, MS-induced Akt phosphorylation was inhibited by molecular deletion of PDGFR-β using siRNA, but not by inhibition of PDGFR-α. Collectively, our data indicate that MS induces MMP-2 production in VSMC via activation of Akt pathway, that is mediated by activation of PDGFR-β signaling pathways.

  14. Influences on vascular wall smooth muscle cells with novel short-duration thermal angioplasty

    Science.gov (United States)

    Kunio, M.; Shimazaki, N.; Arai, T.; Sakurada, M.

    2012-02-01

    We investigated the influences on smooth muscle cells after our novel short-duration thermal angioplasty, Photo-thermo Dynamic Balloon Angioplasty (PTDBA), to reveal the mechanism that can suppress neo-intimal hyperplasia after PTDBA. We obtained the sufficient arterial dilatations by short-duration heating (angioplasty in vivo. The measured neo-intimal hyperplasia occupancy rate was less than 20% after PTDBA in vivo. We prospect that the inhibition of the growth factor's expression by stretch-fixing may result to suppress the neo-intimal hyperplasia. In addition, the decrease of smooth muscle cells' density in the vessel media by heating might be another reason for the neo-intimal hyperplasia suppression.

  15. Differential Mitochondrial Adaptation in Primary Vascular Smooth Muscle Cells from a Diabetic Rat Model

    OpenAIRE

    Keller, Amy C.; Knaub, Leslie A; P. Mason McClatchey; Chelsea A. Connon; Ron Bouchard; Miller, Matthew W.; Kate E. Geary; Walker, Lori A.; Klemm, Dwight J.; Reusch, Jane E. B.

    2016-01-01

    Diabetes affects more than 330 million people worldwide and causes elevated cardiovascular disease risk. Mitochondria are critical for vascular function, generate cellular reactive oxygen species (ROS), and are perturbed by diabetes, representing a novel target for therapeutics. We hypothesized that adaptive mitochondrial plasticity in response to nutrient stress would be impaired in diabetes cellular physiology via a nitric oxide synthase- (NOS-) mediated decrease in mitochondrial function. ...

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

    International Nuclear Information System (INIS)

    Mercury exposure is known to increase cardiovascular risk but the underlying cellular mechanisms remain undetermined. We analyzed whether chronic exposure to HgCl2 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 HgCl2 (first dose 4.6 mg kg−1, subsequent doses 0.07 mg kg−1 day−1, 30 days) and cultured aortic VSMC stimulated with HgCl2 (0.05–5 μg/ml) were used. Treatment of rats with HgCl2 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 HgCl2: 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 HgCl2. Blockade of ERK1/2 and p38 signaling pathways abolished the HgCl2-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 HgCl2 exposure induces vascular remodeling. ► HgCl2 induces proliferation and decreased cell size in vascular smooth muscle cells. ► HgCl2 induces MAPK activation, oxidative stress and COX-2 expression. ► Inhibition of

  17. 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

  18. MAPKK-dependent growth factor-induced upregulation of P2Y2 receptors in vascular smooth muscle cells

    DEFF Research Database (Denmark)

    Hou, M; Möller, S; Edvinsson, L;

    1999-01-01

    The ATP- and UTP-sensitive P2Y2 receptor which mediates both contractile and mitogenic effects has recently been shown to be upregulated in the synthetic phenotype of the vascular smooth muscle cell (VSMC). Using a competitive RT-PCR we demonstrate that the P2Y2 receptor mRNA is increased by fetal...... calf serum and other growth factors in a MAPKK-dependent way. This was confirmed at the functional level by examining UTP-stimulated release of intracellular Ca2+. Furthermore, the P2Y2 receptor mRNA is positively autoregulated by ATP and the mRNA is rapidly degraded with only 26% remaining after 1 h...... in the presence of actinomycin D. Our results indicate growth factor regulation and rapid turnover of the P2Y2 receptor mRNA, which may be of importance in atherosclerosis and neointima formation after balloon angioplasty....

  19. [Influence of curcumin--loaded poly (lactide-co-glycolide) films on the proliferation of vascular smooth muscle cells].

    Science.gov (United States)

    Ren, Ling; Wang, Jin; Tang, Jiaju; Pan, Changjiang; Huang, Nan

    2008-08-01

    In-stent restenosis is the major problem of percutaneous coronary interventions. Drug-eluting stent became a landmark in the treatment of coronary disease. Curcumin could be used for drug-eluting stent due to its antithrombogenity and antiproliferative properties. In this paper, 3-(4, 5-Dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assays were performed to decide the optimal concentration of curcumin for inhibiting the proliferation of vascular smooth muscle cells (VSMC). The result disclosed that more than 80% of VSMC were inhibited when the concentration of curcumin ranged from 2.5 microg/ml to 10 microg/ml (P 316 stainless steel (SS). Therefore, these films may be used for stent coating to inhibit the in-stent restenosis induced by VSMC proliferation. PMID:18792454

  20. Inhibitory Effect of Diabetes on Proliferation of Vascular Smooth Muscle After Balloon Injury in Rat Aorta

    OpenAIRE

    Dahlfors, Gunilla; Chen, Yun; Gustafsson, Bertil; Arnqvist, Hans J.

    2000-01-01

    The effect of streptozotocin-induced diabetes on cell proliferation in rat aortic intima-media, as well as on local gene expression of transforming growth factor-β1 (TGF-β1) was studied. TGF-β1 mRNA was measured by solution hybridization and TGF-β1 protein by ELISA. Proliferation was measured by bromodeoxyuridine incorporation into DNA two days after balloon injury. All BrdU-labelled cells observed were smooth muscle cells. After a diabetes duration of 2 and 4 weeks, labelled cells were signi...

  1. Cyclosporin A inhibits PGE2 release from vascular smooth muscle cells

    OpenAIRE

    Kurtz, Armin; Pfeilschifter, J.; Kühn, K; KOCH, K M

    1987-01-01

    The influence of the fungoid undecapeptide cyclosporin A (CyA) on PGE2 release from cultured rat aortic smooth muscle cells was investigated in this study. We found that CyA time and concentration dependently (ED50:500 ng/ml) inhibited PGE2 release from the cells. CyA attenuated both basal and PGE2 release evoked by angiotensin II (10(-10)-10(-6) M), arginine vasopressin (10(-10)-10(-6) M) and ionomycin (10(-9)-10(-6) M). CyA (1 microgram/ml) did not affect the conversion of exogenous arachid...

  2. Andrographolide, a Novel NF-κB Inhibitor, Induces Vascular Smooth Muscle Cell Apoptosis via a Ceramide-p47phox-ROS Signaling Cascade

    OpenAIRE

    Yu-Ying Chen; Ming-Jen Hsu; Joen-Rong Sheu; Lin-Wen Lee; Cheng-Ying Hsieh

    2013-01-01

    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- κ B inhibitor, is the most active and critical constituent isolated from the lea...

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

    OpenAIRE

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

    2014-01-01

    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 ...

  4. Potassium channels in vascular smooth muscle and essential hypertension%血管平滑肌钾通道与原发性高血压

    Institute of Scientific and Technical Information of China (English)

    周述芝; 魏宗德

    2003-01-01

    Essential hypertension(EH)is characterized by an increased total peripheral resistance.There are four types of potassium channels in vascular smooth muscle cells,including Kca,Kv,Kir,KATP,which play an important role in regulating the diameter of vascular.The change of potassium channels may have something to do with the pathogenesis of hypertension.This article reviews the characters of potassium channels and their roles in EH.

  5. Phosphatidylcholine is a major source of phosphatidic acid and diacylglycerol in angiotensin II-stimulated vascular smooth-muscle cells.

    Science.gov (United States)

    Lassègue, B; Alexander, R W; Clark, M; Akers, M; Griendling, K K

    1993-06-01

    In cultured vascular smooth-muscle cells, angiotensin II produces a sustained formation of diacylglycerol (DG) and phosphatidic acid (PtdOH). Since the fatty acid composition of these molecules is likely to determine their efficacy as second messengers, it is important to ascertain the phospholipid precursors and the biochemical pathways from which they are produced. Our experiments suggest that phospholipase D (PLD)-mediated phosphatidylcholine (PtdCho) hydrolysis is the major source of both DG and PtdOH during the late signalling phase. First, in cells labelled with [3H]myristate, which preferentially labels PtdCho, formation of [3H]PtdOH precedes formation of [3H]DG. Second, in contrast with phospholipase C (PLC) activation, DG mass accumulation is dependent on extracellular Ca2+. Similarly, DG mass accumulation is not attenuated by protein kinase C activation, which we have previously shown to inhibit the phosphoinositide-specific PLC. Third, the fatty acid composition of late-phase DG and PtdOH more closely resembles that of PtdCho than that of phosphatidylinositol. Finally, in cells labelled for a short time with [3H]glycerol, the radioactivity incorporated into [3H]DG and PtdOH was greater than that incorporated into PtdIns, but not into PtdCho. We found no evidence that synthesis de novo or phosphatidylethanolamine breakdown contributes to sustained DG and PtdOH formation. Thus, in angiotensin II-stimulated cultured vascular smooth-muscle cells, PLD-mediated PtdCho hydrolysis is the major source of sustained DG and PtdOH, whereas phosphoinositide breakdown is a minor contributor. Furthermore, PtdOH phosphohydrolase, which determines the relative levels of DG and PtdOH, appears to be regulated by protein kinase C. These results have important implications for the role of these second messengers in growth and contraction.

  6. Preparation of liposome-coated oligonucleotide labeled with 99mTc and its uptake in vascular smooth muscle cells

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    To explore the preparation method of liposome-coated 99mTc-labeled antisense oligonucleotide (ASON),targeteing the proliferating cell nuclear antigen (PCNA), and to explore the biological characteristics and the uptake kinetics of a radiolabeled probe in vascular smooth muscle cells, an 18-base single-stranded antisense oligonucleotide targeting PCNA mRNA and the complementary strand (sense oligonucleotide, SON) were synthesized. The ASON (SON) was labeled with 99mTc, by conjugating the bifunctional chelator (hydrazino nicotinamide, HYNIC), and purified through a gel filtration column of Sephadex G-25. The product was then encapsulated in cationic liposome (oligofectamineTM). The radiolabeling efficiency, radiochemical purity, stability of the liposome-coated 99mTc-HYNIC-ASON in a phosphate buffered solution (PBS), and fresh human serum and its uptake rate were studied. There was no significant difference between the 99mTc radiolabeling efficiencies of HYNIC-ASON and HYNIC-SON, which were 60.04% ± 1.92% and 59.60% ± 2.53%, respectively (P > 0.05, n = 5). The radiochemical purity of the liposome-coated 99mTc-HYNIC-ASON was 94.70% ± 1.90% (n = 5). And after incubation with PBS and fresh human seAt 90 min after transfection, the uptake rate of the liposome-coated 99mTc-HYNIC-ASON reached its peak of 83.8% ±5.92% in vascular smooth muscle cells (VSMCs) and was much higher than that of the nonliposome-coated 99mTc-HYNIC-ASON, which was 11.16% ± 0.54% (P < 0.01, n = 4). The labeling method of PCNA ASON (SON) conjugated by HYNIC has been proved successful. The liposome was able to enhance the ASON (SON) uptake in VSMCs,and could be widely used as a safe, convenient, effective gene transfer carrier.

  7. YAP is oppositely regulated in iPSC-induced cardiovascular progenitor cell and vascular smooth muscle cell differentiation

    Institute of Scientific and Technical Information of China (English)

    WANG Yong-yu; FAN Xiao-fang; DING Lu; CHEN Dan-yang; ZHAO Ru; LI Lan; GONG Yong-sheng

    2016-01-01

    AIM:To explore whether YAP protein is important in induced pluripotent stem cell ( iPSC)-induced cardiovascular progenitor cell and/or vascular smooth muscle differentiation .METHODS:Using episomal vector based reprogramming , we generated human iPSCs from donor fibroblasts .We used both this iPSCs and human H 1 embryonic stem cells to differentiate into vascular smooth muscle cells (VSMCs) through cardiovascular progenitor cells (CVPC).Western blotting, qPCR and immunofluorescence microscopy were used to check the expression of YAP and related genes during this differentiation process .RESULTS:The results showed that iPSCs expressed pluripotent stem cell markers, such as Oct4, Nanog, Sox2, TRA-1-60 and SSEA3, and could form teratoma in SCID mice.YAP was highly expressed in pluripotent stem cells , but dramatically decreased when CVPC differentiation started .YAP gradually increased dur-ing CVPC three-day differentiation.The TAZ and YAP binding partner TEAD1, but not TEAD2 and TEAD4, have similar expression pattern in CVPC differentiation .Immunofluorescence result confirmed that YAP was activated and accumulated in nucleus .Interesting-ly, both YAP and phosphorylated YAP expression decreased to very low level after CVPC differentiated into VSMCs in 7 days.TEAD4 and TAZ also decreased, while TEAD1, TEAD2 and TEAD3 expression did not change during VSMC differentiation .CONCLU-SION:YAP and TEAD1 expression increased during CVPC differentiation , while YAP and TEAD4 expression decreased from CVPC to VSMCs differentiation , which suggested YAP might have different function during diverse cell differentiation .

  8. Effects of nitrendipine on growth activity in cultured vascular smooth muscle cells.

    Science.gov (United States)

    Absher, M P; Baldor, L; Warshaw, D M

    1988-01-01

    Proliferation and migration of smooth muscle cells (SMCs) in the arterial wall may play a role in the development of atherosclerosis and hypertension. If cell migration and proliferation are dependent on extracellular calcium, then treatment with calcium channel blockers such as nitrendipine may alter these cellular responses. In the studies reported here, proliferation and migration activities were assessed in cultured bovine carotid artery smooth muscle cells exposed to nitrendipine. SMCs in long-term culture are characterized by periods of either stable or enhanced proliferative activity. During the stable periods, 1 microM nitrendipine has no effect on proliferation, but during periods of enhanced proliferation, 1 microM nitrendipine augments growth by approximately 20%. SMC migration rates and interdivision times were determined from analysis of time-lapse cinematography films. During stable periods of growth, cell migration rate was inversely related to interdivision time (i.e., fast migrating cells had the shortest interdivision times). Treatment with 1 microM nitrendipine abolished the relationship between migration rate and interdivision time and prolonged interdivision times. These data suggest that the ability of nitrendipine to alter SMC proliferation, interdivision time, and migration is dependent upon the overall proliferative state of the culture.

  9. MiR-21 inhibits c-Ski signaling to promote the proliferation of rat vascular smooth muscle cells.

    Science.gov (United States)

    Li, Jun; Zhao, Li; He, Xie; Yang, Ting; Yang, Kang

    2014-04-01

    Previously, we reported that the decrease of endogenous c-Ski expression is implicated in the progression of vascular smooth muscle cell (VSMC) proliferation after arterial injury. However, the molecular mechanism of the down-regulation of c-Ski is not clear. In this study, a potential miR-21 recognition element was identified in the 3'-untranslated region (UTR) of rat c-Ski mRNA. A reporter assay revealed that miR-21 could recognize the miR-21 recognition element of c-Ski mRNA. In A10 rat aortic smooth muscle cells, overexpression of miR-21 significantly inhibited the expression of c-Ski protein and promoted cell proliferation, which could be blocked by inhibition of miR-21 or overexpression of c-Ski. Further investigation demonstrated that the effect of miR-21 on VSMC proliferation resulted from negative regulation of c-Ski to suppress p38-p21/p27 signaling, the downstream pathway of c-Ski in VSMCs. These results indicate that c-Ski is a target gene of miR-21. miR-21 specifically binds to the 3'-untranslated region of c-Ski and negatively regulates c-Ski expression to diminish the protective effects of c-Ski and stimulate VSMC proliferation in the progression of arterial injury.

  10. Eukaryotic Expression of Human Arresten Gene and Its Effect on the Proliferation of Vascular Smooth Muscle Cells

    Institute of Scientific and Technical Information of China (English)

    SHANG Dan; ZHENG Qichang; SONG Zifang; LI Yiqing; WANG Xiedan; GUO Xingjun

    2006-01-01

    The eukaryotic expression of human arresten geneand its effect on the proliferation of in vitro cultured vascular smooth cells (VSMCs) in vitro were investigated. COS-7 cells were transfected with recombinant eukaryotic expression plasmid pSecTag2-AT or control plasmid pSecTag2 mediated by liposome. Forty-eight h after transfection, reverse transcription-polymerase chain reaction (RT-PCR) was used to detect the expression of arresten mRNA in the cells,while Western blot assay was applied to detect the expression of arresten protein in concentrated supernatant. Primary VSMCs from thoracic aorta of male Sprague-Dawley rats were cultured using the tissue explant method, and identified by immunohistochemical staining with a smooth muscle-specific anti-αactin monoclonal antibody before serial subcultivation. VSMCs were then co-cultured with the concentrated supernatant and their proliferation was detected using Cell Counting Kit-8 (CCK-8) in vitro. The results showed that RT-PCR revealed that the genome of arresten-transfected cells contained a 449 bp specific fragment of arresten gene, suggesting the successful transfection. Successful protein expression in supernatants was confirmed by Western blot. CCK-8 assay showed that the proliferation of VSMCs were inhibited significantly by arresten protein as compared with control cells (F=40.154, P<0.01). It was concluded that arresten protein expressed in eukaryotic cells can inhibit proliferation of VSMCs effectively in vitro, which would provide possibility to the animal experiments.

  11. 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

    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.

  12. Structural properties of lipid reconstructs and lipid composition of normotensive and hypertensive rat vascular smooth muscle cell membranes

    Directory of Open Access Journals (Sweden)

    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.

  13. 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. PMID:26706847

  14. Biomimetic control of vascular smooth muscle cell morphology and phenotype for functional tissue-engineered small-diameter blood vessels.

    Science.gov (United States)

    Chan-Park, Mary B; Shen, Jin Ye; Cao, Ye; Xiong, Yun; Liu, Yunxiao; Rayatpisheh, Shahrzad; Kang, Gavin Chun-Wei; Greisler, Howard P

    2009-03-15

    Small-diameter blood vessel substitutes are urgently needed for patients requiring replacements of their coronary and below-the-knee vessels and for better arteriovenous dialysis shunts. Circulatory diseases, especially those arising from atherosclerosis, are the predominant cause of mortality and morbidity in the developed world. Current therapies include the use of autologous vessels or synthetic materials as vessel replacements. The limited availability of healthy vessels for use as bypass grafts and the failure of purely synthetic materials in small-diameter sites necessitate the development of a biological substitute. Tissue engineering is such an approach and has achieved promising results, but reconstruction of a functional vascular tunica media, with circumferentially oriented contractile smooth muscle cells (SMCs) and extracellular matrix, appropriate mechanical properties, and vasoactivity has yet to be demonstrated. This review focuses on strategies to effect the switch of SMC phenotype from synthetic to contractile, which is regarded as crucial for the engineering of a functional vascular media. The synthetic SMC phenotype is desired initially for cell proliferation and tissue remodeling, but the contractile phenotype is then necessary for sufficient vasoactivity and inhibition of neointima formation. The factors governing the switch to a more contractile phenotype with in vitro culture are reviewed.

  15. N-Cadherin Induction by ECM Stiffness and FAK Overrides the Spreading Requirement for Proliferation of Vascular Smooth Muscle Cells

    Directory of Open Access Journals (Sweden)

    Keeley L. Mui

    2015-03-01

    Full Text Available In contrast to the accepted pro-proliferative effect of cell-matrix adhesion, the proliferative effect of cadherin-mediated cell-cell adhesion remains unresolved. Here, we studied the effect of N-cadherin on cell proliferation in the vasculature. We show that N-cadherin is induced in smooth muscle cells (SMCs in response to vascular injury, an in vivo model of tissue stiffening and proliferation. Complementary experiments performed with deformable substrata demonstrated that stiffness-mediated activation of a focal adhesion kinase (FAK-p130Cas-Rac signaling pathway induces N-cadherin. Additionally, by culturing paired and unpaired SMCs on microfabricated adhesive islands of different areas, we found that N-cadherin relaxes the spreading requirement for SMC proliferation. In vivo SMC deletion of N-cadherin strongly reduced injury-induced cycling. Finally, SMC-specific deletion of FAK inhibited proliferation after vascular injury, and this was accompanied by reduced induction of N-cadherin. Thus, a stiffness- and FAK-dependent induction of N-cadherin connects cell-matrix to cell-cell adhesion and regulates the degree of cell spreading needed for cycling.

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

    International Nuclear Information System (INIS)

    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: → Mechanical stretch augments insulin-induced VSMC proliferation via IGF-1 receptor. → Src/EGFR-mediated ERK and Akt phosphorylation are augmented in stretched VSMCs. → Similar to in vitro experiment, IGF-1 receptor is increased in hypertensive rats. → Results provide possible mechanisms of vascular remodeling in hypertension with DM.

  17. WISP1 overexpression promotes proliferation and migration of human vascular smooth muscle cells via AKT signaling pathway.

    Science.gov (United States)

    Lu, Shun; Liu, Hao; Lu, Lihe; Wan, Heng; Lin, Zhiqi; Qian, Kai; Yao, Xingxing; Chen, Qing; Liu, Wenjun; Yan, Jianyun; Liu, Zhengjun

    2016-10-01

    Proliferation and migration of vascular smooth muscle cells (VSMCs) play crucial roles in the development of vascular restenosis. Our previous study showed that CCN4, namely Wnt1 inducible signaling pathway protein 1 (WISP1), significantly promotes proliferation and migration of rat VSMCs, but its mechanism remains unclear. This study aims to investigate whether and how WISP1 stimulates proliferation and migration of human VSMCs. Western blot analysis showed that FBS treatment increased WISP1 protein levels in human VSMCs in a dose-dependent manner. Overexpression of WISP1 using adenovirus encoding WISP1 (AD-WISP1) significantly increased proliferation rate of human VSMCs by 2.98-fold compared with empty virus (EV)-transfected cells, shown by EdU incorporation assay. Additionally, Scratch-induced wound healing assay revealed that adenovirus-mediated overexpression of WISP1 significantly increased cell migration compared with EV-transfected cells from 6h (4.56±1.14% vs. 11.23±2.25%, PMigration Assay confirmed that WISP1 overexpression significantly promoted human VSMC migration by 2.25-fold compared with EV. Furthermore, WISP1 overexpression stimulated Akt signaling activation in human VSMCs. Blockage of Akt signaling by Akt inhibitor AZD5363 or PI3K inhibitor LY294002, led to an inhibitory effect of WISP1-induced proliferation and migration in human VSMCs. Moreover, we found that WISP1 overexpression stimulated GSK3α/β phosphorylation, and increased expression of cyclin D1 and MMP9 in human VSMCs, and this effect was abolished by AZD5363. Collectively, we demonstrated that Akt signaling pathway mediates WISP1-induced migration and proliferation of human VSMCs, suggesting that WISP1 may act as a novel potential therapeutic target for vascular restenosis.

  18. 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.

  19. Testosterone delays vascular smooth muscle cell senescence and inhibits collagen synthesis via the Gas6/Axl signaling pathway.

    Science.gov (United States)

    Chen, Yan-Qing; Zhao, Jing; Jin, Cheng-Wei; Li, Yi-Hui; Tang, Meng-Xiong; Wang, Zhi-Hao; Zhang, Wei; Zhang, Yun; Li, Li; Zhong, Ming

    2016-06-01

    Testosterone deficiency is associated with a higher incidence of cardiovascular diseases in men. However, its effect on cell senescence, which plays a causal role in vascular aging, remains unclear. Here, we tested the hypothesis that testosterone alleviated vascular smooth muscle cell (VSMC) senescence and collagen synthesis via growth arrest-specific protein 6 (Gas6)/Axl- and Akt/FoxO1a-dependent pathways. Testosterone significantly ameliorated angiotensin II-induced VSMC senescence and collagen overexpression. In addition, testosterone inhibited angiotensin II-induced matrix metalloproteinase-2 (MMP-2) activity, which played a pivotal role in facilitating age-related collagen deposition. Testosterone increased the expression of tissue inhibitor of metalloproteinase-2 but decreased the expression of MMP-2 and membrane type-1 metalloproteinase which contributed to increase MMP-2 activity. The effects on VSMCs senescence and collagen synthesis were mediated by restoration of angiotensin II-induced downregulation of Gas6 and Axl expression and a subsequent reduction of Akt and FoxO1a phosphorylation. The effects of testosterone were reversed by a Gas6 blocker, Axl-Fc, and a specific inhibitor of Axl, R428. Treatment of VSMCs with PI3K inhibitor LY294002 abrogated the downregulating effect of testosterone on MMP-2 activity. Furthermore, when FoxO1a expression was silenced by using a specific siRNA, the inhibitory effect of testosterone on MMP-2 activity was revered as well, that indicated this process was Akt/FoxO1a dependence. Taken together, Gas6/Axl and Akt/FoxO1a were involved in protective effects of testosterone on VSMCs senescence and collagen synthesis. Our results provide a novel mechanism underlying the protective effect of testosterone on vascular aging and may serve as a theoretical basis for testosterone replacement therapy. PMID:27206970

  20. Influence of cholesterol and fish oil dietary intake on nitric oxide-induced apoptosis in vascular smooth muscle cells.

    Science.gov (United States)

    Perales, Sonia; Alejandre, Ma José; Palomino-Morales, Rogelio; Torres, Carolina; Linares, Ana

    2010-04-01

    Apoptosis of vascular smooth muscle cells (SMC) is critically involved in the progression of atherosclerosis. We previously reported that dietary cholesterol intake induces changes in SMC at molecular and gene expression levels. The objectives of the present study were to investigate the differential response to nitric oxide of vascular SMC obtained from chicks after cholesterol and fish oil dietary intake and to examine effects on the main pro-apoptotic and anti-apoptotic genes. Dietary cholesterol intake reduced the Bcl-2/Bax (anti-apoptotic/pro-apoptotic) protein ratio in SMC, making them more susceptible to apoptosis. When cholesterol was withdrawn and replaced with a fish oil-enriched diet, the Bcl-xl/Bax protein ratio significantly increased, reversing the changes induced by cholesterol. The decrease in c-myc gene expression after apoptotic stimuli and the increase in Bcl-xl/Bax ratio indicate that fish oil has a protective role against apoptosis in SMC. Nitroprussiate-like nitric oxide donors exerted an intensive action on vascular SMC cultures. However, SMC-C (isolated from animals fed with control diet) and SMC-Ch (isolated from animals fed with cholesterol-enriched diet) responded differently to nitric oxide, especially in their bcl-2 and bcl-xl gene expression. SMC isolated from animals fed with cholesterol-enriched and then fish oil-enriched diet (SMC-Ch-FO cultures) showed an intermediate apoptosis level (Bcl-2/Bax ratio) between SMC-C and SMC-Ch, induction of c-myc expression and elevated p53 expression. These findings indicate that fish oil protects SMC against apoptosis.

  1. Piperine inhibits platelet-derived growth factor-BB-induced proliferation and migration in vascular smooth muscle cells.

    Science.gov (United States)

    Lee, Kang Pa; Lee, Kwan; Park, Won-Hwan; Kim, Hyuck; Hong, Heeok

    2015-02-01

    The proliferation and migration of vascular smooth muscle cells (VSMCs) in blood vessels are important in the pathogenesis of vascular disorders such as atherosclerosis and restenosis. Piperine, a major component of black pepper, has antioxidant, anticancer, and anti-inflammatory activity. However, the antiatherosclerotic effects of piperine have not been investigated. In this study, the effects of piperine on platelet-derived growth factor (PDGF)-BB-induced proliferation and migration of VSMCs were investigated. The antiproliferative effects of piperine were determined using MTT assays, cell counting, real-time polymerase chain reaction, and western blots. Our results showed that piperine significantly attenuated the proliferation of VSMCs by increasing the expression of p27(kip1), regulating the mRNA expression of cell cycle enzymes (cyclin D, cyclin E, and PCNA), and decreasing the phosphorylation of extracellular signal-regulated kinase (ERK)1/2 in a noncytotoxic concentration-dependent manner (30-100 μM). Moreover, we examined the effects of piperine on the migration of PDGF-BB-stimulated VSMCs, as determined by the Boyden chamber assay, H2DCFDA staining, and western blots. Our results showed that 100 μM piperine decreased cell migration, the production of reactive oxygen species (ROS), and phosphorylation of the p38 mitogen-activated protein kinase (MAPK). Taken together, our results suggest that piperine inhibits PDGF-BB-induced proliferation and the migration of VSMCs by inducing cell cycle arrest and suppressing MAPK phosphorylation and ROS. These findings suggest that piperine may be beneficial for the treatment of vascular-related disorders and diseases.

  2. MicroRNA-34a Induces Vascular Smooth Muscle Cells Senescence by SIRT1 Downregulation and Promotes the Expression of Age-Associated Pro-inflammatory Secretory Factors.

    Science.gov (United States)

    Badi, Ileana; Burba, Ilaria; Ruggeri, Clarissa; Zeni, Filippo; Bertolotti, Matteo; Scopece, Alessandro; Pompilio, Giulio; Raucci, Angela

    2015-11-01

    Arterial aging is a major risk factor for the occurrence of cardiovascular diseases. The aged artery is characterized by endothelial dysfunction and vascular smooth muscle cells altered physiology together with low-grade chronic inflammation. MicroRNA-34a (miR-34a) has been recently implicated in cardiac, endothelial, and endothelial progenitor cell senescence; however, its contribution to aging-associated vascular smooth muscle cells phenotype has not been explored so far. We found that miR-34a was highly expressed in aortas isolated from old mice. Moreover, its well-known target, the longevity-associated protein SIRT1, was significantly downregulated during aging in both endothelial cells and vascular smooth muscle cells. Increased miR-34a as well as decreased SIRT1 expression was also observed in replicative-senescent human aortic smooth muscle cells. miR-34a overexpression in proliferative human aortic smooth muscle cells caused cell cycle arrest along with enhanced p21 protein levels and evidence of cell senescence. Furthermore, miR-34a ectopic expression induced pro-inflammatory senescence-associated secretory phenotype molecules. Finally, SIRT1 protein significantly decreased upon miR-34a overexpression and restoration of its levels rescued miR-34a-dependent human aortic smooth muscle cells senescence, but not senescence-associated secretory phenotype factors upregulation. Taken together, our findings suggest that aging-associated increase of miR-34a expression levels, by promoting vascular smooth muscle cells senescence and inflammation through SIRT1 downregulation and senescence-associated secretory phenotype factors induction, respectively, may lead to arterial dysfunctions.

  3. 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.

  4. Comparison of the effects of elevated inorganic phosphate on primary human vascular smooth muscle cells and the pre-osteoblastic cell line MC3T3-E1

    DEFF Research Database (Denmark)

    Pedersen, Lasse Ebdrup

    into the role of Pi on vascular mineralization has revealed that vascular smooth muscle cells (VSMCs) mineralize in vitro when cultured in hyperphosphatemic media in a manner that is dependent on the type III sodium-dependent Pi transporter, PiT1, and that Pi causes regulation of gene expression, e...... the existence of various inhibitors of vascular mineralization, which are expressed by VSMCs. Together these observations suggest that vascular mineralization is not a passive deposition of calcium-phosphate, as believed for many years, but that VSMCs play a role in the process. In this thesis, I have.......g., upregulation of the osteoblastic transcription factor Runx2, demonstrating that Pi is a signaling molecule. Other similarities with mineralizing osteoblasts have also often been observed in vascular mineralization. In addition, in the past two decades, a number of studies on knockout mice have demonstrated...

  5. Inhibition of Collagen Synthesis and Regulation of Cell Motility in Vascular Smooth Muscle Cells by Suppression of Connective Tissue growth Factor Expression Using RNA Interference

    Institute of Scientific and Technical Information of China (English)

    Xiao-Jing LIU; Huai-Qing CHEN

    2005-01-01

    @@ 1 Introduction Vascular smooth muscle cell (VSMC) hyperplasia plays an important role in both chronic and acute pathologies including atherosclerosis and restenosis. Recent studies have shown that connective tissue growth factor (CTGF) is a novel growth factor involved in the development and progression of atherosclerosis.

  6. MARCKS Signaling Differentially Regulates Vascular Smooth Muscle and Endothelial Cell Proliferation through a KIS-, p27kip1- Dependent Mechanism.

    Directory of Open Access Journals (Sweden)

    Dan Yu

    Full Text Available Overexpression of the myristolated alanine-rich C kinase substrate (MARCKS occurs in vascular proliferative diseases such as restenosis after bypass surgery. MARCKS knockdown results in arrest of vascular smooth muscle cell (VSMC proliferation with little effect on endothelial cell (EC proliferation. We sought to identify the mechanism of differential regulation by MARCKS of VSMC and EC proliferation in vitro and in vivo.siRNA-mediated MARCKS knockdown in VSMCs inhibited proliferation and prevented progression from phase G0/G1 to S. Protein expression of the cyclin-dependent kinase inhibitor p27kip1, but not p21cip1 was increased by MARCKS knockdown. MARCKS knockdown did not affect proliferation in VSMCs derived from p27kip1-/- mice indicating that the effect of MARCKS is p27kip1-dependent. MARCKS knockdown resulted in decreased phosphorylation of p27kip1 at threonine 187 and serine 10 as well as, kinase interacting with stathmin (KIS, cyclin D1, and Skp2 expression. Phosphorylation of p27kip1 at serine 10 by KIS is required for nuclear export and degradation of p27kip1. MARCKS knockdown caused nuclear trapping of p27kip1. Both p27kip1 nuclear trapping and cell cycle arrest were released by overexpression of KIS, but not catalytically inactive KIS. In ECs, MARCKS knockdown paradoxically increased KIS expression and cell proliferation. MARCKS knockdown in a murine aortic injury model resulted in decreased VSMC proliferation determined by bromodeoxyuridine (BrdU integration assay, and inhibition of vascular wall thickening. MARCKS knockdown increased the rate of re-endothelialization.MARCKS knockdown arrested VSMC cell cycle by decreasing KIS expression. Decreased KIS expression resulted in nuclear trapping of p27kip1 in VSMCs. MARCKS knockdown paradoxically increased KIS expression in ECs resulting in increased EC proliferation. MARCKS knockdown significantly attenuated the VSMC proliferative response to vascular injury, but accelerated

  7. Induction of cyclooxygenase-2 by insulin in cultured rat vascular smooth muscle cells

    Institute of Scientific and Technical Information of China (English)

    LI Hong-tao; PAN Xiao-ming; FAN Min; WU Zong-gui; LU Hui-qi; TU Xiao-qing; GAO Chun-fang

    2004-01-01

    To evaluate cyclooxygenase-2 (COX-2) expression induced by insulin in cultured rat vascularmuscle smooth cells (VSMCs). Methods: The rat VSMCs were isolated and cultured in 60 mm plates, pre-treated with in-sulin 10, 100, 500, 1 000 mU/L for 6 h, and 100 mU/L for 3, 6, 12, and 24 h, respectively. RT-PCR was performed tomeasure COX-2 mRNA induction, and Western-blot was used to measure protein expression. In regard to the COX-2 enzymeactivity, after stimulation for a specified duration, the culture medium was collected; Prostaglandin E2 (PGE2) released byVSMCs was measured with an EIA kit. Results: Three hours after insulin (100 mU/L) stimulation, the induction of COX-2mRNA was detectable and increased with the time and dosage of insulin. Insulin induced COX-2 protein expression occurredin a time- and dose-dependent manner. In rat VSMCs, COX-2 induced by insulin resulted in PGE2 production, which wasincreased with insulin action. Conclusion: Insulin could induce COX-2 expression and PGE2 production in VSMCs in time-and dose-dependent manners.

  8. Vascular cell responses to ECM produced by smooth muscle cells on TiO2 nanotubes

    International Nuclear Information System (INIS)

    Graphical abstract: - Highlights: • TiO2 nanotubes with the tube diameter of 30 nm via anodic oxidation was prepared. • SMCs on TiO2 nanotubes presented enhanced extracellular matrix secreting. • ECM prepared via decellularization retained the components: Fn, Ln and collagen. • ECM-covered TiO2 nanotubes significantly improved the proliferation of ECs. - Abstract: There is an increasing interest in developing new methods to promote biocompatibility of biomedical materials. The TiO2 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 TiO2 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 TiO2 nanotubes. The ECM secreted from HUASMC was successfully deposited onto the 30 nm TiO2 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 TiO2 nanotubes significantly improved the proliferation of HUVECs in vitro. This suggested that the ECM secreted from HUASMCs on the TiO2 nanotubular surface could further improve the HUVECs adhesion and proliferation

  9. Messenger molecules of the phospholipase signaling system have dual effects on vascular smooth muscle contraction.

    Science.gov (United States)

    Vidulescu, Cristina; Mironneau, J.; Mironneau, Chantal; Popescu, L. M.

    2000-01-01

    Background and methods. In order to investigate the role of phospholipases and their immediately derived messengers in agonist-induced contraction of portal vein smooth muscle, we used the addition in the organ bath of exogenous molecules such as: phospholipases C, A(2), and D, diacylglycerol, arachidonic acid, phosphatidic acid, choline. We also used substances modulating activity of downstream molecules like protein kinase C, phosphatidic acid phosphohydrolase, or cyclooxygenase. Results. a) Exogenous phospholipases C or A(2), respectively, induced small agonist-like contractions, while exogenous phospholipase D did not. Moreover, phospholipase D inhibited spontaneous contractions. However, when added during noradrenaline-induced plateau, phospholipase D shortly potentiated it. b) The protein kinase C activator, phorbol dibutyrate potentiated both the exogenous phospholipase C-induced contraction and the noradrenaline-induced plateau, while the protein kinase C inhibitor 1-(-5-isoquinolinesulfonyl)-2-methyl-piperazine relaxed the plateau. c) When added before noradrenaline, indomethacin inhibited both phasic and tonic contractions, but when added during the tonic contraction shortly potentiated it. Arachidonic acid strongly potentiated both spontaneous and noradrenaline-induced contractions, irrespective of the moment of its addition. d) In contrast, phosphatidic acid inhibited spontaneous contractile activity, nevertheless it was occasionally capable of inducing small contractions, and when repetitively added during the agonist-induced tonic contraction, produced short potentiations of the plateau. Pretreatment with propranolol inhibited noradrenaline-induced contractions and further addition of phosphatidic acid augmented this inhibition. Choline augmented the duration and amplitude of noradrenaline-induced tonic contraction and final contractile oscillations. Conclusions. These data suggest that messengers produced by phospholipase C and phospholipase A(2

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

    Institute of Scientific and Technical Information of China (English)

    LUO Quan-Yong; ZHU Jun; LU Han-Kui; ZHU Rui-Sen

    2003-01-01

    This study aimed at the effect of γ -emitting radionuclide 103Pd on the proliferation and apoptosis ofvascular SMCs (smooth muscle cells) in vitro. The cavy aortic SMCs were cultured with culture medium M-199. Theexperiments were carried out in two groups, one for proliferation test and the other for apoptosis test. In each group,103Pd 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. 3H-thymidine incorporation test and liquid scin-tillator were used to detect the effect of 103Pd on the proliferation of SMCs. Flow cytometer was used to detect theapoptotic SMCs. The inhibition rate of SMCs proliferation by 1.85 MBq 103Pd solution was 2.3%, which was not sig-nificant, while the inhibition rate increased from 41.6% to 91.3% as the 103Pd activity increased from 7.40 MBq to 37MBq. The apoptosis rate of SMCs was extremely low (less than 4.0%) by 103Pd with activity from 1.85 MBq to 37MBq. The results suggest that the proliferation of SMCs can be repressed effectively in a dose-dependent fashion by103Pd in vitro. The mechanism of its inhibiting over neointima proliferation is likely to inhibite SMCs proliferationrather than to induce its apoptosis by 103Pd. 103Pd can be used as a γ -emitting intravascular brachytherapy radionu-clide to inhibit SMCs proliferation.

  11. Differential effects of formoterol on thrombin- and PDGF-induced proliferation of human pulmonary arterial vascular smooth muscle cells

    Directory of Open Access Journals (Sweden)

    Goncharova Elena A

    2012-11-01

    Full Text Available Abstract Background Increased pulmonary arterial vascular smooth muscle (PAVSM cell proliferation is a key pathophysiological component of pulmonary vascular remodeling in pulmonary arterial hypertension (PH. The long-acting β2-adrenergic receptor (β2AR agonist formoterol, a racemate comprised of (R,R- and (S,S-enantiomers, is commonly used as a vasodilator in chronic obstructive pulmonary disease (COPD. PH, a common complication of COPD, increases patients’ morbidity and reduces survival. Recent studies demonstrate that formoterol has anti-proliferative effects on airway smooth muscle cells and bronchial fibroblasts. The effects of formoterol and its enantiomers on PAVSM cell proliferation are not determined. The goals of this study were to examine effects of racemic formoterol and its enantiomers on PAVSM cell proliferation as it relates to COPD-associated PH. Methods Basal, thrombin-, PDGF- and chronic hypoxia-induced proliferation of primary human PAVSM cells was examined by DNA synthesis analysis using BrdU incorporation assay. ERK1/2, mTORC1 and mTORC2 activation were determined by phosphorylation levels of ERK1/2, ribosomal protein S6 and S473-Akt using immunoblot analysis. Results We found that (R,R and racemic formoterol inhibited basal, thrombin- and chronic hypoxia-induced proliferation of human PAVSM cells while (S,S formoterol had lesser inhibitory effect. The β2AR blocker propranolol abrogated the growth inhibitory effect of formoterol. (R,R, but not (S,S formoterol attenuated basal, thrombin- and chronic hypoxia-induced ERK1/2 phosphorylation, but had little effect on Akt and S6 phosphorylation levels. Formoterol and its enantiomers did not significantly affect PDGF-induced DNA synthesis and PDGF-dependent ERK1/2, S473-Akt and S6 phosphorylation in human PAVSM cells. Conclusions Formoterol inhibits basal, thrombin-, and chronic hypoxia-, but not PDGF-induced human PAVSM cell proliferation and ERK1/2, but has little effect on

  12. Spontaneous Ca2+ oscillations in subcellular compartments of vascular smooth muscle cells rely on different Ca2+ pools

    Institute of Scientific and Technical Information of China (English)

    2004-01-01

    Spontaneous Ca2+ oscillations in vascular smooth muscle cells have been modeled using a single Ca2+ pool. This report describes spontaneous Ca2+ oscillations dependent on two separate Ca2+ sources for the nuclear versus cytoplasmic compartments. Changes in free intracellular Ca2+ were monitored with ratiometric Ca2+- fluorophores using confocal microscopy. On average, spontaneous oscillations developed in 79% of rat aortic smooth muscle cells that were synchronous between the cytoplasm and nucleus. Reduction of extracellular Ca2+ (< 1 μM) decreased the frequency and amplitude of the cytoplasmic oscillations with 48% of the oscillations asynchronous between the nuclear and cytoplasmic compartments. Similar results were obtained with the Ca2+ channel blockers, nimodipine and diltiazem.Arg-vasopressin (AVP) induced a rapid release of intracellular Ca2+ stores that was greater in the nuclear compartment (4.20 ± 0.23 ratio units, n = 56) than cytoplasm (2.54 ± 0.28) in cells that had spontaneously developed prior oscillations.Conversely, cells in the same conditions lacking oscillations had a greater AVP-induced Ca2+ transient in the cytoplasm (4.99 ± 0.66, n = 17) than in the nucleus (2.67 ± 0.29). Pre-treatment with Ca2+ channel blockers depressed the AVP responses in both compartments with the cytoplasmic Ca2+ most diminished. Depletion of internal Ca2+ stores prior to AVP exposure blunted the nuclear response, mimicking the response of cells that lacked prior oscillations. Spontaneous oscillating cells had a greater sarcoplasmic reticulum network than cells that did not oscillate. We propose that spontaneous nuclear oscillations rely on perinuclear sarcoplasmic reticulum stores, while the cytoplasmic oscillations rely on Ca2+ influx.

  13. K channel activation by nucleotide diphosphates and its inhibition by glibenclamide in vascular smooth muscle cells.

    Science.gov (United States)

    Beech, D J; Zhang, H; Nakao, K; Bolton, T B

    1993-10-01

    1. Whole-cell and inside-out patch recordings were made from single smooth muscle cells that had been isolated enzymatically and mechanically from the rabbit portal vein. 2. In whole-cells the inclusion in the recording pipette solution of nucleotide diphosphates (NDPs), but not tri- or monophosphates, induced a K-current that developed gradually over 5 to 15 min. Intracellular 1 mM guanosine 5'-diphosphate (GDP) induced a slowly developing outward K-current at -37 mV that reached a maximum on average of 72 +/- 4 pA (n = 40). Half maximal effect was estimated to occur with about 0.2 mM GDP. Except for ADP, other NDPs had comparable effects. At 0.1 mM, ADP was equivalent to GDP but at higher concentration ADP was less effective. ADP induced its maximum effect at 1 mM but had almost no effect at 10 mM. 3. In 14% of inside-out patches exposed to 1 mM GDP at the intracellular surface, characteristic K channel activity was observed which showed long (> 1 s) bursts of openings separated by longer closed periods. The current-voltage relationship for the channel was linear in a 60 mM:130 mM K-gradient and the unitary conductance was 24 pS. 4. Glibenclamide applied via the extracellular solution was found to be a potent inhibitor of GDP-induced K-current (IK(GDP)) in the whole-cell. The Kd was 25 nM and the inhibition was fully reversible on wash-out. 5. IK(GDP) was not evoked if Mg ions were absent from the pipette solution. In contrast the omission of extracellular Mg ions had no effect on outward or inward IK(GDP). 6. Inclusion of 1 mM ATP in the recording pipette solution reduced IK(GDP) and also attenuated its decline during long (25 min) recordings. 7. When perforated-patch whole-cell recording was used, metabolic poisoning with cyanide and 2-deoxy-D-glucose induced a glibenclamide-sensitive K-current. This current was not observed when conventional whole-cell recording was used. Possible reasons for this difference are discussed. 8. These K channels appear similar to

  14. c-Ski inhibits autophagy of vascular smooth muscle cells induced by oxLDL and PDGF.

    Directory of Open Access Journals (Sweden)

    Jun Li

    Full Text Available Autophagy is increasingly being recognized as a critical determinant of vascular smooth muscle cell (VSMC biology. Previously, we have demonstrated that c-Ski inhibits VSMC proliferation stimulated by transforming growth factor β (TGF-β, but it is not clear whether c-Ski has the similar protective role against other vascular injury factors and whether regulation of autophagy is involved in its protective effects on VSMC. Accordingly, in this study, rat aortic A10 VSMCs were treated with 40 µg/ml oxidized low-density lipoprotein (oxLDL or 20 ng/ml platelet-derived growth factor (PDGF, both of which were autophagy inducers and closely related to the abnormal proliferation of VSMCs. Overexpression of c-Ski in A10 cells significantly suppressed the oxLDL- and PDGF- induced autophagy. This action of c-Ski resulted in inhibiting the cell proliferation, the decrease of contractile phenotype marker α-SMA expression while the increase of synthetic phenotype marker osteopontin expression stimulated by oxLDL or PDGF. Inversely, knockdown of c-Ski by RNAi enhanced the stimulatory effects of oxLDL or PDGF on A10 cell growth and phenotype transition. And further investigation found that inhibition of AKT phosphorylation to downregulate proliferating cell nuclear antigen (PCNA expression, was involved in the regulation of autophagy and associated functions by c-Ski in the oxLDL- and PDGF-stimulated VSMCs. Collectively, c-Ski may play an important role in inhibiting autophagy to protect VSMCs against some harsh stress including oxLDL and PDGF.

  15. c-Ski inhibits autophagy of vascular smooth muscle cells induced by oxLDL and PDGF.

    Science.gov (United States)

    Li, Jun; Zhao, Li; Yang, Ting; Zeng, Yi-Jun; Yang, Kang

    2014-01-01

    Autophagy is increasingly being recognized as a critical determinant of vascular smooth muscle cell (VSMC) biology. Previously, we have demonstrated that c-Ski inhibits VSMC proliferation stimulated by transforming growth factor β (TGF-β), but it is not clear whether c-Ski has the similar protective role against other vascular injury factors and whether regulation of autophagy is involved in its protective effects on VSMC. Accordingly, in this study, rat aortic A10 VSMCs were treated with 40 µg/ml oxidized low-density lipoprotein (oxLDL) or 20 ng/ml platelet-derived growth factor (PDGF), both of which were autophagy inducers and closely related to the abnormal proliferation of VSMCs. Overexpression of c-Ski in A10 cells significantly suppressed the oxLDL- and PDGF- induced autophagy. This action of c-Ski resulted in inhibiting the cell proliferation, the decrease of contractile phenotype marker α-SMA expression while the increase of synthetic phenotype marker osteopontin expression stimulated by oxLDL or PDGF. Inversely, knockdown of c-Ski by RNAi enhanced the stimulatory effects of oxLDL or PDGF on A10 cell growth and phenotype transition. And further investigation found that inhibition of AKT phosphorylation to downregulate proliferating cell nuclear antigen (PCNA) expression, was involved in the regulation of autophagy and associated functions by c-Ski in the oxLDL- and PDGF-stimulated VSMCs. Collectively, c-Ski may play an important role in inhibiting autophagy to protect VSMCs against some harsh stress including oxLDL and PDGF.

  16. Expression and Suppressive Effects of Interleukin-19 on Vascular Smooth Muscle Cell Pathophysiology and Development of Intimal Hyperplasia

    Science.gov (United States)

    Tian, Ying; Sommerville, Laura J.; Cuneo, Anthony; Kelemen, Sheri E.; Autieri, Michael V.

    2008-01-01

    Anti-inflammatory cytokines may play a protective role in the progression of vascular disease. The purpose of this study was to characterize interleukin (IL)-19 expression and function in the development of intimal hyperplasia, and discern a potential mechanism of its direct effects on vascular smooth muscle cells (VSMCs). IL-19 is an immunomodulatory cytokine, the expression of which is reported to be restricted to inflammatory cells. In the present study, we found that IL-19 is not expressed in quiescent VSMCs or normal arteries but is induced in human arteries by injury and in cultured human VSMCs by inflammatory cytokines. Recombinant IL-19 significantly reduced VSMC proliferation (37.1 ± 4.8 × 103 versus 72.2 ± 6.1 × 103 cells/cm2) in a dose-dependent manner. IL-19 adenoviral gene transfer significantly reduced proliferation and neointimal formation in balloon angioplasty-injured rat carotid arteries (0.172 ± 29.9, versus 0.333 ± 71.9, and 0.309 ± 56.6 μm2). IL-19 induced activation of STAT3 as well as the expression of the suppressor of cytokine signaling 5 (SOCS5) in VSMCs. IL-19 treatment significantly reduced the activation of p44/42 and p38 MAPKs in stimulated VSMCs. Additionally, SOCS5 was found to interact with both p44/42 and p38 MAPKs in IL-19-treated human VSMCs. This is the first description of the expression of both IL-19 and SOCS5 in VSMCs and of the functional interaction between SOCS5 and MAPKs. We propose that through induction of SOCS5 and inhibition of signal transduction, IL-19 expression in VSMCs may represent a novel, protective, autocrine response of VSMCs to inflammatory stimuli. PMID:18669613

  17. 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.

  18. Aldosterone increases oxidant stress to impair guanylyl cyclase activity by cysteinyl thiol oxidation in vascular smooth muscle cells.

    Science.gov (United States)

    Maron, Bradley A; Zhang, Ying-Yi; Handy, Diane E; Beuve, Annie; Tang, Shiow-Shih; Loscalzo, Joseph; Leopold, Jane A

    2009-03-20

    Hyperaldosteronism is associated with impaired endothelium-dependent vascular reactivity owing to increased reactive oxygen species and decreased bioavailable nitric oxide (NO(.)); however, the effects of aldosterone on vasodilatory signaling pathways in vascular smooth muscle cells (VSMC) remain unknown. Soluble guanylyl cyclase (GC) is a heterodimer that is activated by NO(.) to convert cytosolic GTP to cGMP, a second messenger required for normal VSMC relaxation. Here, we show that aldosterone (10(-9)-10(-7) mol/liter) diminishes GC activity by activating NADPH oxidase in bovine aortic VSMC to increase reactive oxygen species levels and induce oxidative posttranslational modification(s) of Cys-122, a beta(1)-subunit cysteinyl residue demonstrated previously to modulate NO(.) sensing by GC. In VSMC treated with aldosterone, Western immunoblotting detected evidence of GC beta(1)-subunit disulfide bonding, whereas mass spectrometry analysis of a homologous peptide containing the Cys-122-bearing sequence exposed to conditions of increased oxidant stress confirmed cysteinyl sulfinic acid (m/z 435), sulfonic acid (m/z 443), and disulfide (m/z 836) bond formation. The functional effect of these modifications was examined by transfecting COS-7 cells with wild-type GC or mutant GC containing an alanine substitution at Cys-122 (C122A). Exposure to aldosterone or hydrogen peroxide (H(2)O(2)) significantly decreased cGMP levels in cells expressing wild-type GC. In contrast, aldosterone or H(2)O(2) did not influence cGMP levels in cells expressing the mutant C122A GC, confirming that oxidative modification of Cys-122 specifically impairs GC activity. These findings demonstrate that pathophysiologically relevant concentrations of aldosterone increase oxidant stress to convert GC to an NO(.)-insensitive state, resulting in disruption of normal vasodilatory signaling pathways in VSMC.

  19. Neuronal chemorepellent Slit2 inhibits vascular smooth muscle cell migration by suppressing small GTPase Rac1 activation.

    Science.gov (United States)

    Liu, Dong; Hou, Jie; Hu, Xing; Wang, Xuerong; Xiao, Yan; Mou, Yongshan; De Leon, Hector

    2006-03-01

    The Slits are secreted proteins with roles in axonal guidance and leukocyte migration. On binding to Robo receptors, Slit2 repels developing axons and inhibits leukocyte chemotaxis. Slit2 is cleaved into Slit2-N, a protein tightly binding to cell membranes, and Slit2-C, a diffusible fragment. In the present study, we characterized the functional role of Slit2-N in vascular smooth muscle cells (VSMCs) and the cell association properties of 2 truncated versions of Slit2-N. Here, we document for the first time that Slit2-N is a chemorepellent of VSMCs. Intact blood vessels expressed Slit2 and Robo receptors as demonstrated by immunohistochemistry and quantitative real time PCR. Recombinant Slit2-N prevented the platelet-derived growth factor (PDGF)-stimulated migration of VSMCs. Slit2-N also abrogated PDGF-mediated activation of small guanosine triphosphatase (GTPase) Rac1, a member of the Rho GTPase superfamily of proteins involved in regulating the actin cytoskeleton. Furthermore, Slit2-N inhibited the PDGF-induced formation of lamellipodia, a crucial cytoskeletal reorganization event for cell motility. Slit2-N had no effect on the PDGF-mediated increase in DNA synthesis determined by [3H]thymidine uptake, suggesting that VSMC growth is unaffected by Slit2. Analysis of 2 engineered Slit2-N fragments (Slit2-N/1118 and Slit2-N/1121) indicated that 3 amino acids upstream of the putative cleavage site (Arg1121, Thr1122) are involved in the association of Slit2-N to the cell membrane. Our data assign a novel functional role to Slit2 in vascular function and show that cell guidance mechanisms that operate in the developing central nervous system are conserved in VSMCs.

  20. Glycogen synthase kinase 3 beta positively regulates Notch signaling in vascular smooth muscle cells: role in cell proliferation and survival.

    Science.gov (United States)

    Guha, Shaunta; Cullen, John P; Morrow, David; Colombo, Alberto; Lally, Caitríona; Walls, Dermot; Redmond, Eileen M; Cahill, Paul A

    2011-09-01

    The role of glycogen synthase kinase 3 beta (GSK-3β) in modulating Notch control of vascular smooth muscle cell (vSMC) growth (proliferation and apoptosis) was examined in vitro under varying conditions of cyclic strain and validated in vivo following changes in medial tension and stress. Modulation of GSK-3β in vSMC following ectopic expression of constitutively active GSK-3β, siRNA knockdown and pharmacological inhibition with SB-216763 demonstrated that GSK-3β positively regulates Notch intracellular domain expression, CBF-1/RBP-Jκ transactivation and downstream target gene mRNA levels, while concomitantly promoting vSMC proliferation and inhibiting apoptosis. In contrast, inhibition of GSK-3β attenuated Notch signaling and decreased vSMC proliferation and survival. Exposure of vSMC to cyclic strain environments in vitro using both a Flexercell™ Tension system and a novel Sylgard™ phantom vessel following bare metal stent implantation revealed that cyclic strain inhibits GSK-3β activity independent of p42/p44 MAPK and p38 activation concomitant with reduced Notch signaling and decreased vSMC proliferation and survival. Exposure of vSMC to changes in medial strain microenvironments in vivo following carotid artery ligation revealed that enhanced GSK-3β activity was predominantly localized to medial and neointimal vSMC concomitant with increased Notch signaling, proliferating nuclear antigen and decreased Bax expression, respectively, as vascular remodeling progressed. GSK-3β is an important modulator of Notch signaling leading to altered vSMC cell growth where low strain/tension microenvironments prevail.

  1. Phenotypic Modulation of Mesenteric Vascular Smooth Muscle Cells from Type 2 Diabetic Rats is Associated with Decreased Caveolin-1 Expression

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    Maria Alicia Carrillo-Sepulveda

    2014-10-01

    Full Text Available Aims: Diabetes-induced vascular complications are associated with vascular smooth muscle cell (VSMC phenotypic modulation, switching from a contractile to a synthetic-proliferative phenotype. Loss of caveolin-1 is involved with proliferation of VSMCs. We tested the hypothesis that mesenteric VSMCs from type 2 diabetic Goto-Kakizaki (GK rat undergo phenotypic modulation and it is linked to decreased caveolin-1 expression. Methods: VSMCs were isolated from mesenteric arteries from GK rats and age-matched control Wistar rats. Western blotting was used to determine expression of target proteins such as caveolin-1, calponin (marker of differentiation, and proliferating cell nuclear antigen (PCNA, marker of proliferation. In addition, we measured intracellular reactive oxygen species (ROS production using H2DCF-DA and activation of extracellular signal-regulated kinase (ERK1/2 by western blotting in VSMCs from GK stimulated with lipopolysaccharide (LPS, an endotoxin upregulated in diabetes. Results: Mesenteric VSMCs from diabetic GK rats exhibited decreased caveolin-1 and calponin expression and increased PCNA expression compared to control. Increased levels of ROS and phospho-ERK1/2 expression were also found in GK VSMCs. LPS augmented ROS and phosphorylated ERK1/2 levels to a greater extent in GK VSMCs than in control. Likewise, high glucose decreased caveolin-1 and calponin expression, increased PCNA expression and augmented ROS production in control mesenteric VSMCs. Conclusion: These results suggest that mesenteric VSMCs from diabetic GK rats undergo phenotypic modulation and it is associated with decreased caveolin-1 expression. These alterations may be due to enhanced inflammatory stimuli and glucose levels present in diabetic milieu.

  2. 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

  3. 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.

  4. High sodium augments angiotensin II-induced vascular smooth muscle cell proliferation through the ERK 1/2-dependent pathway.

    Science.gov (United States)

    Liu, Gang; Hitomi, Hirofumi; Rahman, Asadur; Nakano, Daisuke; Mori, Hirohito; Masaki, Tsutomu; Ma, Hong; Iwamoto, Takahiro; Kobori, Hiroyuki; Nishiyama, Akira

    2014-01-01

    Angiotensin II (Ang II)-induced vascular injury is exacerbated by high-salt diets. This study examined the effects of high-sodium level on Ang II-induced cell proliferation in rat vascular smooth muscle cells (VSMCs). The cells were cultured in a standard medium containing 137.5 mmol l(-1) of sodium. The high-sodium medium (140 mmol l(-1)) contained additional sodium chloride. Extracellular signal-regulated kinase (ERK) 1/2 phosphorylation was determined by western blot analysis. Cell proliferation was evaluated by [(3)H]-thymidine incorporation. Ang II (100 nmol l(-1)) significantly increased ERK 1/2 phosphorylation and cell proliferation in the both medium containing standard sodium and high sodium. High-sodium level augmented Ang II-induced ERK 1/2 phosphorylation and cell proliferation compared with standard sodium. Pre-treatment with candesartan (1 μmol l(-1), Ang II type 1 receptor blocker) or PD98095 (10 μmol l(-1), ERK kinase iinhibitor) abolished the proliferative effect induced by high sodium/Ang II. Pre-treatment with 5-N,N-hexamethylene amiloride (30 μmol l(-1), Na(+)/H(+) exchanger type 1 (NHE-1) inhibitor), but not SN-6 (10 μmol l(-1), Na(+)/Ca(2+) exchanger inhibitor) or ouabain (1 mmol l(-1), Na(+)/K(+)-ATPase inhibitor) attenuated ERK 1/2 phosphorylation or cell proliferation. Osmotic pressure or chloride had no effect on Ang II-induced proliferative changes. High-sodium level did not affect Ang II receptor expression. Ang II increased intracellular pH via NHE-1 activation, and high-sodium level augmented the pH increase induced by Ang II. These data suggest that high-sodium level directly augments Ang II-induced VSMC proliferation through NHE-1- and ERK 1/2-dependent pathways and may offer new insights into the mechanisms of vascular remodeling by high-sodium/Ang II.

  5. Ginsenoside Rg1 inhibits proliferation of vascular smooth muscle cells stimulated by tumor necrosis factor-α

    Institute of Scientific and Technical Information of China (English)

    Zeng-chun MA; Yue GAO; Yu-guang WANG; Hong-ling TAN; Cheng-rong XIAO; Sheng-qi WANG

    2006-01-01

    Aim: To investigate the proliferation of vascular smooth muscle cells (VSMC) affected by ginsenoside Rg1 and further explore the molecular mechanism of ginsenoside Rg1 using proteomics. Methods: The proliferation of VSMC was measured by MTS assay kit and flow cytometry. Proteomic alterations were analyzed using two-dimensional electrophoresis and peptide mass fingerprinting. Differential proteins found in proteomics were confirmed by RT-PCR. Results: The proliferation of VSMC was enhanced significantly after tumor necrosis factor-α (TNF-α) treatment, and ginsenoside Rg1 treatment inhibited proliferation in a dose-dependent manner. Proteomic analysis showed 24 protein spots were changed, including 17 spots that were increased and 7 spots that were decreased. Ginsenoside Rg1 could restore the expression levels of these proteins, at least partly, to basic levels of untreated cells. The expression of G-protein coupled receptor kinase, protein kinase C (PKC)-ζ, N-ras protein were decreased, while cycle related protein p21 was increased by ginsenoside Rg1 in TNF-α treated VSMC. Conclusion: PKC-ζ, and p21 pathway might be the mechanism for inhibitory effects of ginsenoside Rg1 on proliferation of VSMC.

  6. Effect of hammerhead ribozyme that specifically cleaves c-myc mRNA on proliferation of vascular smooth muscle cells

    Institute of Scientific and Technical Information of China (English)

    栾荣华; 贾国良; 李伟; 贾战生; 连建奇

    2003-01-01

    Ojective: To investigate the effect of hammerhead ribozyme that specifically cleaves c-myc mRNA on the proliferation of vascular smooth muscle cells (VSMCs). Methods: Based on the computer analysis of the secondary structure of c-myc mRNA, nt 2029 in rat c-myc oncogene was selected as a cleaving site for hammerhead ribozyme and the ribozyme was designed. With automatic DNA synthesizer, the two complementary DNA strands of the ribozyme were synthesized. The ribozyme gene was cloned into pGEM3Zf (+) vector and subcloned into eukaryotic expression pcDNA3 vector. The recombinant pcDNA-Rz was transfected into the cultured rat VSMCs by lipofectAMINE mediated DNA transfection protocol and individual cell clones were selected by G418. Results: The sequence of ribozyme gene inserted in pGEM3Zf (+) vector was proved to be perfectly correct. In VSMCs transfected with recombinant pcDNA-Rz, flow cytometry analysis showed that the S phase and G2/M fractions were decreased significantly and cell proliferation stagnated in the G0/G1 phase. Conclusion: The results suggest that hammerhead ribozyme that specifically cleaves c-myc mRNA can significantly inhibit the proliferation of VSMCs.

  7. Diminished Lipid Raft SNAP23 Increases Blood Pressure by Inhibiting the Membrane Fluidity of Vascular Smooth-Muscle Cells.

    Science.gov (United States)

    Yoon, Mi So; Won, Kyung-Jong; Kim, Do-Yoon; Hwang, Dae Il; Yoon, Seok Won; Jung, Seung Hyo; Lee, Kang Pa; Jung, Dongju; Choi, Wahn Soo; Kim, Bokyung; Lee, Hwan Myung

    2015-01-01

    Synaptosomal-associated protein 23 (SNAP23) is involved in microvesicle trafficking and exocytosis in various cell types, but its functional role in blood pressure (BP) regulation has not yet been defined. Here, we found that lipid raft SNAP23 expression was much lower in vascular smooth-muscle cells (VSMCs) from spontaneously hypertensive rats (SHR) than in those from normotensive Wistar-Kyoto (WKY) rats. This led us to investigate the hypothesis that this lower expression may be linked to the spontaneous hypertension found in SHR. The expression level of lipid raft SNAP23 and the fluidity in the plasma membrane of VSMCs were lower in SHR than in WKY rats. Cholesterol content in the VSMC membrane was higher, but the secreted cholesterols found in VSMC-conditioned medium and in the blood serum were lower in SHR than in WKY rats. SNAP23 knockdown in WKY rat VSMCs reduced the membrane fluidity and increased the membrane cholesterol level. Systemic overexpression of SNAP23 in SHR resulted in an increase of cholesterol content in their serum, a decrease in cholesterol in their aorta and the reduction of their BP. Our findings suggest that the low expression of the lipid raft SNAP23 in VSMCs might be a potential cause for the characteristic hypertension of SHR.

  8. 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.

  9. Regulation of angiotensinⅡon Gaq/11 protein of vascular smooth muscle cell and its underlying mechanism

    Institute of Scientific and Technical Information of China (English)

    2002-01-01

    To study the regulation of angiotensin Ⅱ(Ang Ⅱ) on Gαq/11 protein of vascular smooth muscle cell (VSMC) and its underlying mechanism, the protein synthesis was detected by [3H]-leucine incorporation. G?q/11 expression was measured by Western blot in cultured VSMC of rat aorta. The results showed that the level of G?q/11 was downregulated after stimulated by AngⅡ for 1-6 h, while it was upregulated significantly by 12-24 h stimulation (P < 0.01) in VSMC. The [3H]-leucine incorporation of VSMC was increased after 24 h Ang Ⅱ stimulation. The biphase regulation of Ang Ⅱ on G?q/11 protein was blocked by the Ang Ⅱ type Ⅰ receptor (AT1) specific antagnist losartan or PLC inhibitor U73122, while PD98059 did not have this effect. These data indicated that Ang Ⅱ contributed to VSMC hypertrophy by regulating the level of G?q/11, and this effect was mediated mainly through AT1 receptor-PLC signal transduction pathway.

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

    Directory of Open Access Journals (Sweden)

    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.

  11. 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.

  12. INHIBITORY EFFECT OF ANGIOTENSIN Ⅱ TYPE 1 RECEPTOR-ASSOCIATED PROTEIN ON VASCULAR SMOOTH MUSCLE CELL GROWTH AND NEOINTIMAL FORMATION

    Institute of Scientific and Technical Information of China (English)

    Zhen Li; Zhong-gao Wang; Xiu Chen; Xiao-dong Chen

    2007-01-01

    Objective To investigate the mechanism of a novel angiotensin n type 1 receptor-associated protein (ATRAP) interfering with angiotensin Ⅱ type 1 (AT1) receptor-mediated vascular smooth muscle cell (VSMC) growth and neointimal formation. Methods VSMCs isolated from thoracic aorta of adult Sprague-Dawley ( SD) rats were used in this study. ATRAP Cdna was subcloned into pcDNA3 vector and then transfected into VSMCs. DNA synthesis and extracellular signal-regulated kinase (ERK) and phospho-ERK expressions in VSMCs were assayed by measurement of 3H thymidine incorporation and Western blotting, respectively. Morphological changes were observed in the balloon injured artery with or without transfection of ATRAP Cdna using 12-week-old male SD rats. Results ATRAP overexpression in VSMCs inhibited angiotensin Ⅱ (Ang Ⅱ) -induced 3H thymidine incorporation 48 hours after Ang Ⅱ stimulation (P < 0.05). In VSMC, Ang Ⅱ stimulation increased the phosphorylation of ERK, which reached the peak around 60 minutes. The activation of phospho-ERK was significantly decreased by ATRAP (P < 0.05). Neointimal formation was markedly inhibited by ATRAP overexpression in injuried arteries. Conclusions The AT1 receptor-derived activation of ERK plays an essential role in Ang Ⅱ-induced VSMC growth. The growth inhibitory effects of ATRAP might be due to interfering with AT1 receptor-mediated activation of ERK in VSMC growth and neointimal formation.

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

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    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.

  14. 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.

  15. Paeonol Inhibits the Proliferation, Invasion, and Inflammatory Reaction Induced by TNF-α in Vascular Smooth Muscle Cells.

    Science.gov (United States)

    Meng, Liang; Xu, Weidong; Guo, Lihong; Ning, Wenqi; Zeng, Xiandong

    2015-11-01

    The aim of this study was to evaluate the effect of paeonol on the proliferation, migration, and inflammation induced by tumor necrosis factor (TNF-α) of rat vascular smooth muscle cells (VSMCs). Primary rat VSMCs were identified by immunofluorescence assay. The inhibition of VSMCs proliferation induced by TNF-α was observed after paeonol treatment in a dose-dependent manner. Treatment with 100 μM paeonol significantly reduced the expression of proliferating cell nuclear antigen (PCNA). On the other hand, transwell assay showed that treatment with paeonol suppressed the invasion of TNF-α-induced VSMCs and the production of inflammation factors stimulated by TNF-α. For apoptosis induced by paeonol, Western blot analysis showed that cleaved caspase-3 and -9 were detected, and pro-apoptotic protein Bax was up-regulated, whereas anti-apoptotic protein Bcl-2 was down-regulated by paeonol in TNF-α-stimulated VSMCs. ELISA analysis data showed that both levels of IL-1β and IL-6 produced by the stimulation of TNF-α were decreased by paeonol in a dose-dependent manner in VSMCs. These results suggest that paeonol can effectively inhibit the proliferation through apoptotic induction through caspase pathway in VSMCs induced by TNF-α. Also, paeonol significantly reduced the invasion and the inflammation stimulated by TNF-α in VSMCs. PMID:27352344

  16. High glucose decreases the expression of ATP-binding cassette transporter G1 in human vascular smooth muscle cells

    Institute of Scientific and Technical Information of China (English)

    Jiahong Xue; Zuyi Yuan; Yue Wu; Yan Zhao; Zhaofei Wan

    2008-01-01

    Objective:ATP-binding cassette transporters(ABC) A1 and G1 play an important role in mediating cholesterol efflux and preventing macrophage foam cell formation. In this study, we examined the regulation of ABC transporters by high glucose in human vascular smooth muscle cells(VSMCs), the other precursor of foam cells. Methods:Incubation of human VSMCs with D-ghicose(5 to 30 mM) for 1 to 7 days in the presence or absence of antioxidant and nuclear factor(NF)-kB inhibitors, the expressions of ABCA1 and ABCG1 were analyzed by real time PCR and Western blotting. Results:High glucose decreased ABCG1 mRNA and protein expression in cultured VSMCs, whereas the expression of ABCA1 was not significantly decreased. Down-regulation of ABCG1 mRNA expression by high glucose was abolished by antioxidant N-acetyl-L-cysteine(NAC) and NF-kB inhibitors, BAY 11-7085 and tosyl-phenylalanine chloromethyl-ketone(TPCK). Conclusion:High glucose suppresses the expression of ABCG1 in VSMCs, which is the possible mechanism of VSMC derived foam cell transformation.

  17. Cultured rat vascular smooth muscle cells are resistant to methylamine toxicity: no correlation to semicarbazide-sensitive amine oxidase

    Science.gov (United States)

    Langford, S. D.; Trent, M. B.; Boor, P. J.

    2001-01-01

    Methylamine (MA), a component of serum and a metabolite of nicotine and certain insecticides and herbicides, is metabolized by semicarbazide-sensitive amine oxidase (SSAO). MA is toxic to cultured human umbilical vein and calf pulmonary artery endothelial cells. Endothelial cells, which do not exhibit endogenous SSAO activity, are exposed to SSAO circulating in serum. In contrast, vascular smooth muscle cells (VSMC) do exhibit innate SSAO activity both in vivo and in vitro. This property, together with the critical localization of VSMC within the arterial wall, led us to investigate the potential toxicity of MA to VSMC. Cultured rat VSMC were treated with MA (10-5 to 1 M). In some cultures, SSAO was selectively inhibited with semicarbazide or MDL-72145 [(E)-2-(3,4-dimethoxyphenyl)-3-fluoroallylamine]. Cytotoxicity was measured via MTT, vital dye exclusion, and clonogenic assays. MA proved to be toxic to VSMC only at relatively high concentrations (LC(50) of 0.1 M). The inhibition of SSAO with semicarbazide or MDL-72145 did not increase MA toxicity, suggesting that the production of formaldehyde via tissue-bound, SSAO-mediated MA metabolism does not play a role in the minimal toxicity observed in isolated rat VSMC. The omission of fetal calf serum (FCS), which contains high SSAO activity, from media similarly showed little effect on cytotoxicity. We conclude that VSMC--in contrast to previous results in endothelial cells--are relatively resistant to MA toxicity, and SSAO does not play a role in VSMC injury by MA.

  18. Artesunate Reduces Proliferation, Interferes DNA Replication and Cell Cycle and Enhances Apoptosis in Vascular Smooth Muscle Cells

    Institute of Scientific and Technical Information of China (English)

    2005-01-01

    This study examined the effect of artesunate (Art) on the proliferation, DNA replication, cell cycles and apoptosis of vascular smooth muscle cells (VSMCs). Primary cultures of VSMCs were established from aortas of mice and artesunate of different concentrations was added into the medium. The number of VSMCs was counted and the curve of cell growth was recorded.The activity of VSMCs was assessed by using MTT method and inhibitory rate was calculated.DNA replication was evaluated by [3 H]-TdR method and apoptosis by DNA laddering and HE staining. Flowmetry was used for simultaneous analysis of cell apoptosis and cell cycles. Compared with the control group, VSMCs proliferation in Art interfering groups were inhibited and [3H]-TdR incorprating rate were decreased as well as cell apoptosis was induced. The progress of cell cycle was blocked in G0/G1 by Art in a dose-dependent manner. It is concluded that Art inhibits VSMCs proliferation by disturbing DNA replication, inducing cell apoptosis and blocking cell cycle in G0/G1 phase.

  19. 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.

  20. Effects of low-intensity laser irradiation on the apoptosis of rabbit vascular smooth muscle cells in culture

    Science.gov (United States)

    Li, S. D.; Chen, P.; Zhang, C. P.; Wen, J. X.; Liang, J.; Kang, H. X.; Gao, R. L.; Fu, X. B.

    2011-11-01

    Restenosis is a major complication after coronary intervention therapy. Excessive proliferation of vascular smooth muscle cells (VSMCs) and a decline in their apoptosis, which eventually leads to excessive neointimal thickening in coronary arteries, are the main causes of restenosis. Induction of the apoptosis of VSMCs and inhibition of excessive proliferation of VSMCs are therefore crucial for the prevention of restenosis, and low-intensity laser irradiation of coronary arteries may play a promising role in keeping this in balance. In this study, we used in vitro cultured rabbit VSMCs to investigate the effects of low-intensity laser irradiation at a wavelength of 532 nm on the apoptosis of VSMCs via morphological observation and molecular biology. The results showed that apoptotic bodies and obvious intranuclear apoptosis-positive particles formed within VSMCs 24 h after laser irradiation, suggesting that low-intensity laser irradiation at certain doses can inhibit the proliferation of VSMCs by promoting their apoptosis. This experiment provides evidences for further animal experiments and clinical trials on prevention and treatment of restenosis by intracoronary low-intensity laser irradiation at a wavelength of 532 nm.

  1. Effects and underlying mechanisms of curcumin on the proliferation of vascular smooth muscle cells induced by Chol:MβCD

    International Nuclear Information System (INIS)

    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.

  2. Induction of indoleamine 2,3-dioxygenase in vascular smooth muscle cells by interferon-gamma contributes to medial immunoprivilege.

    Science.gov (United States)

    Cuffy, Madison C; Silverio, Amanda M; Qin, Lingfeng; Wang, Yinong; Eid, Raymond; Brandacher, Gerald; Lakkis, Fadi G; Fuchs, Dietmar; Pober, Jordan S; Tellides, George

    2007-10-15

    Atherosclerosis and graft arteriosclerosis are characterized by leukocytic infiltration of the vessel wall that spares the media. The mechanism(s) for medial immunoprivilege is unknown. In a chimeric humanized mouse model of allograft rejection, medial immunoprivilege was associated with expression of IDO by vascular smooth muscle cells (VSMCs) of rejecting human coronary artery grafts. Inhibition of IDO by 1-methyl-tryptophan (1-MT) increased medial infiltration by allogeneic T cells and increased VSMC loss. IFN-gamma-induced IDO expression and activity in cultured human VSMCs was considerably greater than in endothelial cells (ECs) or T cells. IFN-gamma-treated VSMCs, but not untreated VSMCs nor ECs with or without IFN-gamma pretreatment, inhibited memory Th cell alloresponses across a semipermeable membrane in vitro. This effect was reversed by 1-MT treatment or tryptophan supplementation and replicated by the absence of tryptophan, but not by addition of tryptophan metabolites. However, IFN-gamma-treated VSMCs did not activate allogeneic memory Th cells, even after addition of 1-MT or tryptophan. Our work extends the concept of medial immunoprivilege to include immune regulation, establishes the compartmentalization of immune responses within the vessel wall due to distinct microenvironments, and demonstrates a duality of stimulatory EC signals versus inhibitory VSMC signals to artery-infiltrating T cells that may contribute to the chronicity of arteriosclerotic diseases.

  3. Inhibition of the cystathionine-γ-lyase/hydrogen sulfide pathway in rat vascular smooth muscle cells by cobalt-60 gamma radiation

    Institute of Scientific and Technical Information of China (English)

    ZHONG Guang-zhen; YANG Xin-chun; JIA Li-ping; CHEN Feng-rong; CUI Ming

    2009-01-01

    Background Radiation is a promising treatment for in stent restenosis and restenosis following percutaneous transluminal coronary angioplasty, which has troubled interventional cardiologists for a long time. It inhibits neointima hyperplasia, vascular remodeling, and increases the mean luminal diameter. The mechanism of intracoronary brachytherapy for restenosis is not well understood. Endogenous gaseous transmitters including nitric oxide and carbon monoxide are closely related to restenosis. Hydrogen sulfide, a new endogenous gaseous transmitter, is able to inhibit the proliferation of vascular smooth muscle cells and vascular remodeling. This study aimed to clarify the effect of radiation on cystathionine-y-lyase/hydrogen sulfide pathway in rat smooth muscle cells.Methods We studied the effect of radiation on the cystathionine-γ-lyase/hydrogen sulfide pathway. Rat vascular smooth muscle cells were radiated with 60Co y at doses of 14 Gy and 25 Gy respectively. Then the mRNA level of cystathionine-γ-lyase was studied by quantitative reverse-transcription competitive polymerase chain reaction. Hydrogen sulfide concentration in culture medium was determined by methylene blue spectrophotometry. Cystathionine-γ-lyase activity in vascular smooth muscle cells was also studied.Results 60Co y radiation at a dose of 1 Gy did not affect the cystathionine-γ-lyase/hydrogen sulfide pathway significantly. However, 60Co y radiation at doses of 14 Gy and 25 Gy decreased the hydrogen sulfide synthesis by 21.9% (P <0.05) and 26.8% (P <0.01 ) respectively. At the same time, they decreased the cystathionine-γ-lyase activity by 15.1% (P <0.05) and 20.5% (P <0.01) respectively, and cystathionine-γ-lyase mRNA expression by 29.3% (P <0.01 ) and 38.2% (P <0.01) respectively.Conclusion Appropriate 60Co γ radiation inhibits the H2S synthesis by inhibiting the gene expression of cystathionine-γ-lyase and the cystathionine-y-lyase activity.

  4. 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.

  5. Change in vascular smooth muscle response to 5-HT due to short- or long-term endothelial denudation of the bovine digital vein.

    Science.gov (United States)

    Punzi, Simona; Belloli, Chiara; Gogny, Marc; Desfontis, Jean-Claude; Mallem, Mohamed Y

    2016-01-01

    Several chronic progressive vascular diseases, such as laminitis, show vasocontractile dysfunction that might evolve into reperfusion injury and/or vessel structural remodelling, which may be traced back to aberrant endothelial function. In the present study, the vasomotor responses of bovine digital veins (BDVs) to 5-hydroxytryptamine (5-HT) were investigated in blood vessels, with and without endothelium present, and in samples deprived of endothelium before or after overnight incubation in tissue culture medium, to evaluate the effects of short- and long-term endothelial damage on vascular smooth muscle (VSM) reactivity. No significant effects were observed in the blood vessels tested immediately after the removal of endothelium. In contrast, a significant increase in VSM reactivity to 5-HT was seen in vessels incubated without endothelium. This long-term change in smooth muscle reactivity was prevented by exposure to the nitric oxide (NO) donor nitroprusside (P digital veins in animals affected with laminitis. PMID:26670334

  6. Magnesium inhibits Wnt/β-catenin activity and reverses the osteogenic transformation of vascular smooth muscle cells.

    Directory of Open Access Journals (Sweden)

    Addy Montes de Oca

    Full Text Available 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

  7. Hyperglycemia Enhances IGF-I–Stimulated Src Activation via Increasing Nox4-Derived Reactive Oxygen Species in a PKCζ-Dependent Manner in Vascular Smooth Muscle Cells

    OpenAIRE

    Xi, Gang; Shen, Xinchun; Maile, Laura A.; Wai, Christine; Gollahon, Katherine; Clemmons, David R.

    2011-01-01

    IGF-I–stimulated sarcoma viral oncogene (Src) activation during hyperglycemia is required for propagating downstream signaling. The aim of the current study was to determine the mechanism by which hyperglycemia enhances IGF-I–stimulated Src activation and the role of NADPH oxidase 4 (Nox4) and protein kinase C ζ (PKCζ) in mediating this response in vascular smooth muscle cells (VSMCs). Nox4 expression was analyzed in VSMCs exposed to hyperglycemia. The role of Nox4-derived reactive oxygen spe...

  8. Contractile effect of Sclerocarya birrea (A Rich) Hochst (Anacardiaceae) (Marula) leaf aqueous extract on rat and rabbit isolated vascular smooth muscles

    OpenAIRE

    Mawoza, Tariro; Ojewole, John AO; Owira, Peter MO

    2012-01-01

    Backround Sclerocarya birrea (Anacardiaceae) is traditionally used for treating hypertension. The pharmacological effects of S birrea leaf aqueous extract (SBE) on rabbit and rat vascular smooth muscles were investigated in this study. Methods Fresh S birrea leaves (1 kg) were air dried at 26 ± 1°C, milled, macerated in 2.5 l of distilled water for 48 hours, filtered, and the filtrate was concentrated in a rotary evaporator. Rat isolated portal vein preparations, as well as rabbit isolated en...

  9. Natriuretic peptide receptor-3 underpins the disparate regulation of endothelial and vascular smooth muscle cell proliferation by C-type natriuretic peptide

    OpenAIRE

    Khambata, Rayomand S.; Panayiotou, Catherine M; Hobbs, Adrian J

    2011-01-01

    BACKGROUND AND PURPOSE C-type natriuretic peptide (CNP) is an endothelium-derived vasorelaxant, exerting anti-atherogenic actions in the vasculature and salvaging the myocardium from ischaemic injury. The cytoprotective effects of CNP are mediated in part via the Gi-coupled natriuretic peptide receptor (NPR)3. As GPCRs are well-known to control cell proliferation, we investigated if NPR3 activation underlies effects of CNP on endothelial and vascular smooth muscle cell mitogenesis. EXPERIMENT...

  10. Lithium Chloride Inhibits Vascular Smooth Muscle Cell Proliferation and Migration and Alleviates Injury-Induced Neointimal Hyperplasia via Induction of PGC-1α

    OpenAIRE

    Zhuyao Wang; Xiwen Zhang; Siyu Chen; Danfeng Wang; Jun Wu; Tingming Liang; Chang Liu

    2013-01-01

    The proliferation and migration of vascular smooth muscle cells (VSMCs) contributes importantly to the development of in-stent restenosis. Lithium has recently been shown to have beneficial effects on the cardiovascular system, but its actions in VSMCs and the direct molecular target responsible for its action remains unknown. On the other hand, PGC-1α is a transcriptional coactivator which negatively regulates the pathological activation of VSMCs. Therefore, the purpose of the present study ...

  11. Inhibition of Collagen Synthesis and Regulation of Cell Motility in Vascular Smooth Muscle Cells by Suppression of Connective Tissue growth Factor Expression Using RNAInterference

    Institute of Scientific and Technical Information of China (English)

    2005-01-01

    1 IntroductionVascular smooth muscle cell (VSMC) hyperplasia plays an important role in both chronic and acute pathologies including atherosclerosis and restenosis. Recent studies have shown that connective tissue growth factor (CTGF) is a novel growth factor involved in the development and progression of atherosclerosis. However, previous data about the role of CTGF on the VSMC is conflicting. Hishikawa et al demonstrated that CTGF could act as a growth inhibitor in human VSMC; but some others' reports (Fa...

  12. Distinct Mechanisms of Receptor and Nonreceptor Tyrosine Kinase Activation by Reactive Oxygen Species in Vascular Smooth Muscle Cells: Role of Metalloprotease and Protein Kinase C-δ

    OpenAIRE

    Frank, Gerald D.; Mifune, Mizuo; Inagami, Tadashi; Ohba, Motoi; Sasaki, Terukatsu; Higashiyama, Shigeki; Dempsey, Peter J; Eguchi, Satoru

    2003-01-01

    Reactive oxygen species (ROS) are implicated in cardiovascular diseases. ROS, such as H2O2, act as second messengers to activate diverse signaling pathways. Although H2O2 activates several tyrosine kinases, including the epidermal growth factor (EGF) receptor, JAK2, and PYK2, in vascular smooth muscle cells (VSMCs), the intracellular mechanism by which ROS activate these tyrosine kinases remains unclear. Here, we identified two distinct signaling pathways required for receptor and nonreceptor...

  13. Andrographolide Enhances Nuclear Factor-κB Subunit p65 Ser536 Dephosphorylation through Activation of Protein Phosphatase 2A in Vascular Smooth Muscle Cells*

    OpenAIRE

    Hsieh, Cheng Y.; Hsu, Ming J.; Hsiao, George; Wang, Yi H.; Huang, Chi W.; Chen, Shiuan W.; Jayakumar, Thanasekaran; Chiu, Pei T.; Chiu, Yi H.; Sheu, Joen R

    2010-01-01

    Recent studies have demonstrated that transcription factor nuclear factor (NF)-κB inhibition may contribute to the protective anti-inflammatory actions of andrographolide, an abundant component of plants of the genus Andrographis. However, the precise mechanism by which andrographolide inhibits NF-κB signaling remains unclear. We thus investigated the mechanism involved in andrographolide suppression of NF-κB signaling in rat vascular smooth muscle cells (VSMCs) exposed to proinflammatory sti...

  14. Angiotensin II induces Fat1 expression/activation and vascular smooth muscle cell migration via Nox1-dependent reactive oxygen species generation

    OpenAIRE

    Bruder-Nascimento, T; Chinnasamy, P; Riascos-Bernal, DF; Cau, SB; Callera, GE; Touyz, RM; Tostes, RC; Sibinga, NES

    2014-01-01

    Fat1 is an atypical cadherin that controls vascular smooth muscle cell (VSMC) proliferation and migration. Nicotinamide adenine dinucleotide phosphate (NADPH) oxidase 1 (Nox1) is an important source of reactive oxygen species (ROS) in VSMCs. Angiotensin II (Ang II) induces the expression and/or activation of both Fat1 and Nox1 proteins. This study tested the hypothesis that Ang II-induced Fat1 activation and VSMC migration are mediated by Nox1-dependent ROS generation and redox signaling. Stu...

  15. Increased atherosclerosis and vascular smooth muscle cell activation in AIF-1 transgenic mice fed a high-fat diet.

    Science.gov (United States)

    Sommerville, Laura J; Kelemen, Sheri E; Ellison, Stephen P; England, Ross N; Autieri, Michael V

    2012-01-01

    Allograft inflammatory factor-1 (AIF-1) is a cytoplasmic, scaffold signal transduction protein constitutively expressed in inflammatory cells, but inducible in vascular smooth muscle cells (VSMCs) in response to injury or inflammatory stimuli. Although several basic science and population studies have reported increased AIF-1 expression in human and experimental atherosclerosis, a direct causal effect of AIF-1 expression on development of atherosclerosis has not been reported. The purpose of this study is to establish a direct relationship between AIF-1 expression and development of atherosclerosis. AIF-1 expression is detected VSMC in atherosclerotic lesions from ApoE(-/-) mice, but not normal arteries from wild-type mice. AIF-1 expression can be induced in cultured VSMC by stimulation with oxidized LDL (ox-LDL). Transgenic mice in which AIF-1 expression is driven by the G/C modified SM22 alpha promoter to restrict AIF-1 expression to VSMC develop significantly increased atherosclerosis compared with wild-type control mice when fed a high-fat diet (P=0.022). Cultured VSMC isolated from Tg mice demonstrated significantly increased migration in response to ox-LDL compared with matched controls (P<0.001). VSMC isolated from Tg mice and cultured human VSMC which over express AIF-1 demonstrated increased expression of MMP-2 and MMP-9 mRNA and protein and increased NF-κB activation in response to ox-LDL as compared with wild-type control mice. VSMC which over express AIF-1 have significantly increased uptake of ox-LDL, and increased CD36 expression. Together, these data suggest a strong association between AIF-1 expression, NF-κB activation, and development of experimental atherosclerosis.

  16. The Ang II-induced growth of vascular smooth muscle cells involves a phospholipase D-mediated signaling mechanism.

    Science.gov (United States)

    Freeman, E J

    2000-02-15

    Angiotensin (Ang) II acts as a mitogen in vascular smooth muscle cells (VSMC) via the activation of multiple signaling cascades, including phospholipase C, tyrosine kinase, and mitogen-activated protein kinase pathways. However, increasing evidence supports signal-activated phospholipases A(2) and D (PLD) as additional mechanisms. Stimulation of PLD results in phosphatidic acid (PA) formation, and PA has been linked to cell growth. However, the direct involvement of PA or its metabolite diacylglycerol (DAG) in Ang II-induced growth is unclear. PLD activity was measured in cultured rat VSMC prelabeled with [(3)H]oleic acid, while the incorporation of [(3)H]thymidine was used to monitor growth. We have previously reported the Ang II-dependent, AT(1)-coupled stimulation of PLD and growth in VSMC. Here, we show that Ang II (100 nM) and exogenous PLD (0.1-100 units/mL; Streptomyces chromofuscus) stimulated thymidine incorporation (43-208% above control). PA (100 nM-1 microM) also increased thymidine incorporation to 135% of control. Propranolol (100 nM-10 microM), which inhibits PA phosphohydrolase, blocked the growth stimulated by Ang II, PLD, or PA by as much as 95%, an effect not shared by other beta-adrenergic antagonists. Propranolol also increased the production of PA in the presence of Ang II by 320% and reduced DAG and arachidonic acid (AA) accumulation. The DAG lipase inhibitor RHC-80267 (1-10 microM) increased Ang II-induced DAG production, while attenuating thymidine incorporation and release of AA. Thus, it appears that activation of PLD, formation of PA, conversion of PA to DAG, and metabolism of DAG comprise an important signaling cascade in Ang II-induced growth of VSMC.

  17. Benefits of Synchrotron Microangiography for Dynamic Studies of Smooth Muscle and Endothelial Roles in the Pathophysiology of Vascular Disease

    Science.gov (United States)

    Pearson, James T.; Schwenke, Daryl O.; Jenkins, Mathew J.; Edgley, Amanda J.; Sonobe, Takashi; Ishibashi-Ueda, Hatsue; Umetani, Keiji; Eppel, Gabriela A.; Evans, Roger G.; Okura, Yasuhiko; Shirai, Mikiyasu

    2010-07-01

    Changes in endothelial and smooth muscle function compromise organ perfusion in the chronic disease states of diabetes, atherosclerosis and hypertension. Moreover, vascular dysfunction increases the likelihood of lethal acute events such as myocardial infarction and stroke, which are now leading causes of adult mortality. Many circulating and local tissue factors in these disease states contribute to impaired vasomotor regulation of the arterial vessels, leading to spasm, chronic constriction and eventually vessel remodelling. X-ray contrast absorption imaging allows assessment of vessel lumen diameter and the factors contributing to steady-state vessel calibre, however, conventional clinical devices (>200 μm resolution) are not adequate to detect microvessels or accurately assess function in real time. Using synchrotron imaging we are now able to detect small vessel calibres (˜30 μm) and quantify regional differences in calibre even under conditions of high heart rate (>500 bpm). Herein we describe recent experiments that were conducted at the Japanese Synchrotron, SPring-8 using anaesthetised Sprague-Dawley rats and C57Bl/6 mice and a synchrotron radiation contrast angiography (single narrow energy bandwidth) approach based on selective arterial injection of iodine contrast agents. Application of this approach to imaging of the heart and other vasculatures are described. Our studies show that within-animal comparisons of 3-4 branching orders of arterial vessels are possible using small bolus contrast injections and appropriate contrast washout times (15-30 min) in many organ systems. Determination of relative calibre changes before and after any treatment allows us to evaluate the contributions of different endogenous factors and ligand-receptor pathways in the maintenance of vasomotor tone. Finally, we will present our findings relating to novel therapies to prevent endothelial dysfunction in heart failure.

  18. Improved Adhesion, Growth and Maturation of Vascular Smooth Muscle Cells on Polyethylene Grafted with Bioactive Molecules and Carbon Particles

    Directory of Open Access Journals (Sweden)

    Martina Blazkova

    2009-10-01

    Full Text Available High-density polyethylene (PE foils were modified by an Ar+ plasma discharge and subsequent grafting with biomolecules, namely glycine (Gly, polyethylene glycol (PEG, bovine serum albumin (BSA, colloidal carbon particles (C or BSA and C (BSA + C. As revealed by atomic force microscopy (AFM, goniometry and Rutherford Backscattering Spectroscopy (RBS, the surface chemical structure and surface morphology of PE changed dramatically after plasma treatment. The contact angle decreased for the samples treated by plasma, mainly in relation to the formation of oxygen structures during plasma irradiation. A further decrease in the contact angle was obvious after glycine and PEG grafting. The increase in oxygen concentration after glycine and PEG grafting proved that the two molecules were chemically linked to the plasma-activated surface. Plasma treatment led to ablation of the PE surface layer, thus the surface morphology was changed and the surface roughness was increased. The materials were then seeded with vascular smooth muscle cells (VSMC derived from rat aorta and incubated in a DMEM medium with fetal bovine serum. Generally, the cells adhered and grew better on modified rather than on unmodified PE samples. Immunofluorescence showed that focal adhesion plaques containing talin, vinculin and paxillin were most apparent in cells on PE grafted with PEG or BSA + C, and the fibres containing α-actin, β-actin or SM1 and SM2 myosins were thicker, more numerous and more brightly stained in the cells on all modified PE samples than on pristine PE. An enzyme-linked immunosorbent assay (ELISA revealed increased concentrations of focal adhesion proteins talin and vinculin and also a cytoskeletal protein β-actin in cells on PE modified with BSA + C. A contractile protein α-actin was increased in cells on PE grafted with PEG or Gly. These results showed that PE activated with plasma and subsequently grafted with bioactive molecules and colloidal C

  19. Mitophagy acts as a safeguard mechanism against human vascular smooth muscle cell apoptosis induced by atherogenic lipids.

    Science.gov (United States)

    Swiader, Audrey; Nahapetyan, Hripsime; Faccini, Julien; D'Angelo, Romina; Mucher, Elodie; Elbaz, Meyer; Boya, Patricia; Vindis, Cécile

    2016-05-17

    Mitophagy is a critical cellular process that selectively targets damaged mitochondria for autophagosomal degradation both under baseline conditions and in response to stress preventing oxidative damage and cell death. Recent studies have linked alterations in mitochondria function and reduced autophagy with the development of age-related pathologies. However, the significance of mitochondrial autophagy in vessel wall in response to atherogenic lipid stressors is not known. In the present study, we investigated the role of mitophagy on human vascular smooth muscle cells (VSMC) apoptosis induced by oxidized low-density lipoproteins (LDL). We reported for the first time that the engulfment of defective mitochondria by autophagosomes occurred in human VSMC in response to oxidized LDL. The molecular mechanism mediating mitophagy in human VSMC involved dynamin-related protein 1 (Drp1)-mediated mitochondrial fission, accumulation of PTEN-induced putative kinase 1 (PINK1) and the recruitment of the E3 ubiquitin ligase Parkin to mitochondria. Likewise, we found increased voltage-dependent anion channel 1 (VDAC1) and mitofusin 2 (Mnf2) mitochondrial proteins ubiquitination and LC3 association to mitochondria. Using flow cytometry in the presence of lysosomal inhibitors, we showed that PINK1 and Parkin silencing impaired mitophagy flux and enhanced oxidized LDL-induced VSMC apoptosis. In addition, overexpression of PINK1 and Parkin were protective by limiting cell death. Moreover, reduced Bax levels found in VSMC-overexpressing Parkin indicated cross talk among mitophagy and mitochondrial apoptotic signalling pathways. Altogether these data demonstrate that mitophagy is a safeguard mechanism against human VSMC apoptosis induced by atherogenic stressors and highlight mitophagy as a potential target to stabilize atherosclerotic plaque. PMID:27119505

  20. Experimental Study of 103Pd Stent Affecting Dynamic Equilibrium Between Proliferation and Apoptosis of Vascular Smooth Muscle Cells

    Institute of Scientific and Technical Information of China (English)

    Liu Yingmei; Fu Yuewu; Wei Yulin; Wu Wei

    2006-01-01

    Objectives By observing γ radioactive 103Pd stent affecting the proliferation and apoptosis of vascular smooth muscle cells (VSMCs) to explore the mechanism of radioactive stent preventing in-stent restenosis. Methods Fifty male New Zealand rabbits were randomized into stent group and 103Pd stent group. Control group was set up. The materials were harvested on 3, 7, 14, 28, 56 days after operation and the following investigations were carried out, including pathomorphology, immunohistochemistry, apoptosis (TUNEL) and in situs hybridization studies. Results ①The severity of the stenosis in 103Pd stent group was less than that of stent group.It was the most obvious on 56th day (P < 0.01).②The expression of PCNA of 103Pd stent group was lower than that of stent group on 3 to 28 days. It was the most obvious on 7th day, 16.35%±0.79% vs 24.36%±0.55% (P< 0.01 ). ③TUNEL method showed that the 103Pd stent group had much more apoptosis of VSMCs than that of stent group. The highest rate of apoptosis appeared on day 7, 14.72%±0.53% vs 12.42%±1.13% (P<0.01). ④ By calculating the ratio of PCNA/apoptosis (P:A), a much lower ratio was seen in 103Pd-stent group than that of stent group at 3 to 28 days. There was significant statistic difference between two groups (P<0.05). ⑤For bcl-2/bax ratio, the result in 103Pd-stent group was lower than that of stent group at 3 to 28 days. It had significant statistic difference (P < 0.05). Conclusions γ radioactive stent can inhibit the proliferation and accelerate apoptosis of injured media VSMCs. Also it can decrease the ratio of proliferation to apoptosis and relieve the severity of restenosis.

  1. CaMKIIδ-dependent inhibition of cAMP-response element-binding protein activity in vascular smooth muscle.

    Science.gov (United States)

    Liu, Yongfeng; Sun, Li-Yan; Singer, Diane V; Ginnan, Roman; Singer, Harold A

    2013-11-22

    One transcription factor mediator of Ca(2+)-signals is cAMP response element-binding protein (CREB). CREB expression and/or activity negatively correlates with vascular smooth muscle (VSM) cell proliferation and migration. Multifunctional Ca(2+)/calmodulin-dependent protein kinases, including CaMKII, have been demonstrated to regulate CREB activity through both positive and negative phosphorylation events in vitro, but the function of CaMKII as a proximal regulator of CREB in intact cell systems, including VSM, is not clear. In this study, we used gain- and loss-of-function approaches to determine the function of CaMKIIδ in regulating CREB phosphorylation, localization, and activity in VSM. Overexpression of constitutively active CaMKIIδ specifically increased CREB phosphorylation on Ser(142) and silencing CaMKIIδ expression by siRNA or blocking endogenous CaMKII activity with KN93 abolished thrombin- or ionomycin-induced CREB phosphorylation on Ser(142) without affecting Ser(133) phosphorylation. CREB-Ser(142) phosphorylation correlated with transient nucleocytoplasmic translocation of CREB. Thrombin-induced CREB promoter activity, CREB binding to Sik1 and Rgs2 promoters, and Sik1/Rgs2 transcription were enhanced by a kinase-negative CaMKIIδ2 (K43A) mutant and inhibited by a constitutively active (T287D) mutant. Taken together, these studies establish negative regulation of CREB activity by endogenous CaMKIIδ-dependent CREB-Ser(142) phosphorylation and suggest a potential mechanism for CaMKIIδ/CREB signaling in modulating proliferation and migration in VSM cells. PMID:24106266

  2. Curcumin inhibits cellular cholesterol accumulation by regulating SREBP-1/caveolin-1 signaling pathway in vascular smooth muscle cells

    Institute of Scientific and Technical Information of China (English)

    Hao-yu YUAN; Shuang-yu KUANG; Xing ZHENG; Hong-yan LING; Yun-bo YANG; Peng-ke YAN; Kai LI; Duan-fang LIAO

    2008-01-01

    Aim: To investigate the protective effect and the possible mechanism of curcumin on anti-atherosclerosis. Methods: Morphological changes of atherosclerotic le-sions taken from apoE knockout (apoE-/-) mice were determined by hematoxylin-eosin staining. Intracellular lipid droplets and lipid levels were assayed by oil red O staining and HPLC. The protein expression of caveolin-1 was quantified by West-ern blotting. Translocation and the expression of sterol response element-bind-ing protein-1 (SREBP-1) were indirectly detected by an immunofluorescence analysis. Results: The administration of 20 mg.kg-1.d-1 curcumin to apoE-/1 mice for 4 months induced a 50% reduction of atherosclerotic lesions and yielded a 5-fold increase in the caveolin-1 expression level as compared to the model group. Rat vascular smooth muscle cells (VSMC) pretreated with 50 mg.L-1 ox-lipid den-sity lipoprotein(ox-LDL) for 48 h increased cellular lipid contents, and stimulated SREBP-1 translocation, but decreased the caveolin-1 expression level. Lipid-loaded cells exposed to curcumin at various concentrations (12.5, 25, and 50 μmol.L-1) for different durations (0, 6, 12, 24, and 48 h) significantly diminished the number and area of cellular lipid droplets, total cholesterol, cholesterol ester, and free choles-terol accompanying the elevation of the caveolin-1 expression level (approximately 3-fold); the translocation of SREBP-1 from the cytoplasm to the nucleus was inhibited compared with the models. Lipid-loaded VSMC exposed to N-acetyl-Leu-Leu-norleucinal, a SREBP-1 protease inhibitor, showed increased nuclear trans-location of SREBP-1, reduced caveolin-1 expression level, and upregulated cellu-lar lipid levels. Conclusion: Curcumin inhibits ox-LDL-induced cholesterol accu-mulation in cultured VSMC through increasing the caveolin-1 expression via the inhibition of nuclear translocation of SREBP-1.

  3. Platelet-derived growth factor regulates vascular smooth muscle phenotype via mammalian target of rapamycin complex 1

    Energy Technology Data Exchange (ETDEWEB)

    Ha, Jung Min; Yun, Sung Ji; Kim, Young Whan; Jin, Seo Yeon; Lee, Hye Sun [Medical Research Institute, Department of Pharmacology, Pusan National University School of Medicine, Yangsan (Korea, Republic of); Song, Sang Heon [Department of Internal Medicine, Pusan National University Hospital, Busan (Korea, Republic of); Shin, Hwa Kyoung [Department of Anatomy, Pusan National University School of Korean Medicine, Yangsan (Korea, Republic of); Bae, Sun Sik, E-mail: sunsik@pusan.ac.kr [Medical Research Institute, Department of Pharmacology, Pusan National University School of Medicine, Yangsan (Korea, Republic of)

    2015-08-14

    Mammalian target of rapamycin complex (mTORC) regulates various cellular processes including proliferation, growth, migration and differentiation. In this study, we showed that mTORC1 regulates platelet-derived growth factor (PDGF)-induced phenotypic conversion of vascular smooth muscle cells (VSMCs). Stimulation of contractile VSMCs with PDGF significantly reduced the expression of contractile marker proteins in a time- and dose-dependent manner. In addition, angiotensin II (AngII)-induced contraction of VSMCs was completely blocked by the stimulation of VSMCs with PDGF. PDGF-dependent suppression of VSMC marker gene expression was significantly blocked by inhibition of phosphatidylinositol 3-kinase (PI3K), extracellular signal-regulated kinase (ERK), and mTOR whereas inhibition of p38 MAPK had no effect. In particular, inhibition of mTORC1 by rapamycin or by silencing of Raptor significantly blocked the PDGF-dependent phenotypic change of VSMCs whereas silencing of Rictor had no effect. In addition, loss of AngII-dependent contraction by PDGF was significantly retained by silencing of Raptor. Inhibition of mTORC1 by rapamycin or by silencing of Raptor significantly blocked PDGF-induced proliferation of VSMCs. Taken together, we suggest that mTORC1 plays an essential role in PDGF-dependent phenotypic changes of VSMCs. - Graphical abstract: Regulation of VSMC phenotype by PDGF-dependent activation of mTORC1. - Highlights: • The expression of contractile marker proteins was reduced by PDGF stimulation. • PDGF-dependent phenotypic conversion of VSMCs was blocked by inhibition of mTOR. • PDGF-induced proliferation of VSMCs was attenuated by inhibition of mTORC1. • mTORC1 plays a critical role in PDGF-dependent phenotypic conversion of VSMCs.

  4. A gel-free approach in vascular smooth muscle cell proteome: perspectives for a better insight into activation

    Directory of Open Access Journals (Sweden)

    Tedeschi Lorena

    2010-03-01

    Full Text Available Abstract Background The use of chromatography coupled with mass spectrometry (MS analysis is a powerful approach to identify proteins, owing to its capacity to fractionate molecules according to different chemical features. The first protein expression map of vascular smooth muscle cells (VSMC was published in 2001 and since then other papers have been produced. The most detailed two-dimensional polyacrylamide gel electrophoresis (2D-PAGE map was presented by Mayr et al who identified 235 proteins, corresponding to the 154 most abundant unique proteins in mouse aortic VSMC. A chromatographic approach aimed at fractionating the VSMC proteome has never been used before. Results This paper describes a strategy for the study of the VSMC proteome. Our approach was based on pre-fractionation with ion exchange chromatography coupled with matrix assisted laser desorption-time of flight mass spectrometry analysis assisted by a liquid chromatography (LC-MALDI-TOF/TOF. Ion exchange chromatography resulted in a good strategy designed to simplify the complexity of the cellular extract and to identify a large number of proteins. Selectivity based on the ion-exchange chemical features was adequate if evaluated on the basis of protein pI. The LC-MALDI approach proved to be highly reproducible and sensitive since we were able to identify up to 815 proteins with a concentration dynamic range of 7 orders of magnitude. Conclusions In our opinion, the large number of identified proteins and the promising quantitative reproducibility made this approach a powerful method to analyze complex protein mixtures in a high throughput way and to obtain statistical data for the discovery of key factors involved in VSMC activation and to analyze a label-free differential protein expression.

  5. 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.

  6. Toxicology and pharmacology of some euthenium compounds: vascular smooth muscle relaxation by nitrosyl derivatives of ruthenium and iridium

    Energy Technology Data Exchange (ETDEWEB)

    Kruszyna, H.; Kruszyna, R.; Hurst, J.; Smith, R.P.

    1980-07-01

    A series of compounds were synthesized from ruthenium trichloride, and their ip LD50s were determined in mice: pentamminenitrosylruthenium(II) chloride, 8.9; chloronitrobis(2,2'-dipyridyl)ruthenium(II), 55; dichlorobis(2,2'-dipyridyl)ruthenium(II) 63; ruthenium trichloride, 108; and potassium pentachloronitrosylruthenate(II), 127 mg/kg. The two bis-bipyridyl complexes produced death in convulsions within minutes, whereas the remaining compounds resulted in long, debilitating courses with death occuring in 4 to 7 d. When given in massive overdoses, however, the compounds with inorganic ligands also produced rapid convulsive death in mice, and when given iv to anesthetized cats, they produced respiratory arrest. The major toxic effects of all the complexes appeared to be due to the metal and not to its associated ligands. Only complexes having a nitrosyl ligand specifically relaxed vascular smooth muscle. Potassium pentabromoiridate(III) also relaxed rabbit aortic strips that had been contracted by adrenergic argonists, but potassium pentachloroiridate(III) did not. None of the complexes was as active as nitroprusside in relaxing aortic strips or in decreasing arterial blood pressure in cats. No compound tested was as potent as cisplatin in antitumor activity. The pentamminenitrosylruthenium(II) complex also relaxed guinea pig ileum and frog rectus abdominum when these isolated muscles had been contracted by acetylcholine. It appears that these organoruthenium compounds may produce death in central respiratory arrest, as do the inorganic complexes when given iv or ip in massive overdoses. In minimllylethal doses, the complexes with inorganic ligands may affect a variety of contractile tissues, perhaps by a general mechanism involving Ca. These complexes are apt to be generally cytotoxic as well.

  7. Benefits of Synchrotron Microangiography for Dynamic Studies of Smooth Muscle and Endothelial Roles in the Pathophysiology of Vascular Disease

    International Nuclear Information System (INIS)

    Changes in endothelial and smooth muscle function compromise organ perfusion in the chronic disease states of diabetes, atherosclerosis and hypertension. Moreover, vascular dysfunction increases the likelihood of lethal acute events such as myocardial infarction and stroke, which are now leading causes of adult mortality. Many circulating and local tissue factors in these disease states contribute to impaired vasomotor regulation of the arterial vessels, leading to spasm, chronic constriction and eventually vessel remodelling. X-ray contrast absorption imaging allows assessment of vessel lumen diameter and the factors contributing to steady-state vessel calibre, however, conventional clinical devices (>200 μm resolution) are not adequate to detect microvessels or accurately assess function in real time. Using synchrotron imaging we are now able to detect small vessel calibres (∼30 μm) and quantify regional differences in calibre even under conditions of high heart rate (>500 bpm). Herein we describe recent experiments that were conducted at the Japanese Synchrotron, SPring-8 using anaesthetised Sprague-Dawley rats and C57Bl/6 mice and a synchrotron radiation contrast angiography (single narrow energy bandwidth) approach based on selective arterial injection of iodine contrast agents. Application of this approach to imaging of the heart and other vasculatures are described. Our studies show that within-animal comparisons of 3-4 branching orders of arterial vessels are possible using small bolus contrast injections and appropriate contrast washout times (15-30 min) in many organ systems. Determination of relative calibre changes before and after any treatment allows us to evaluate the contributions of different endogenous factors and ligand-receptor pathways in the maintenance of vasomotor tone. Finally, we will present our findings relating to novel therapies to prevent endothelial dysfunction in heart failure.

  8. 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.

  9. miR-155-dependent regulation of mammalian sterile 20-like kinase 2 (MST2) coordinates inflammation, oxidative stress and proliferation in vascular smooth muscle cells.

    Science.gov (United States)

    Yang, Zhan; Zheng, Bin; Zhang, Yu; He, Ming; Zhang, Xin-hua; Ma, Dong; Zhang, Ruo-nan; Wu, Xiao-li; Wen, Jin-kun

    2015-07-01

    In response to vascular injury, inflammation, oxidative stress, and cell proliferation often occur simultaneously in vascular tissues. We previously observed that microRNA-155 (miR-155), which is implicated in proliferation and inflammation is involved in neointimal hyperplasia; however, the molecular mechanisms by which it regulates these processes remain largely unknown. In this study, we observed that vascular smooth muscle cell (VSMC) proliferation and neointimal formation in wire-injured femoral arteries were reduced by the loss of miR-155 and increased by the gain of miR-155. The proliferative effect of miR-155 was also observed in cultured VSMCs. Notably, expression of the miR-155-target protein mammalian sterile 20-like kinase 2 (MST2) was increased in the injured arteries of miR-155-/- mice. miR-155 directly repressed MST2 and thus activated the extracellular signal-regulated kinase (ERK) pathway by promoting an interaction between RAF proto-oncogene serine/threonine-protein kinase (Raf-1) and mitogen-activated protein kinase kinase (MEK) and stimulating inflammatory and oxidative stress responses; together, these effects lead to VSMC proliferation and vascular remodeling. Our data reveal that MST2 mediates miR-155-promoted inflammatory and oxidative stress responses by altering the interaction of MEK with Raf-1 and MST2 in response to vascular injury. Therefore, suppression of endogenous miR-155 might be a novel therapeutic strategy for vascular injury and remodeling. PMID:25892184

  10. Plasmenylethanolamine is the major storage depot for arachidonic acid in rabbit vascular smooth muscle and is rapidly hydrolyzed after angiotensin II stimulation

    International Nuclear Information System (INIS)

    The present study demonstrates that rabbit aortic intimal smooth muscle cells contain the majority of their endogenous arachidonic acid mass in plasmenylethanolamine molecular species. To demonstrate the potential significance of these plasmenylethanolamines as substrates for the smooth muscle cell phospholipases that are activated during agonist stimulation, aortic rings were prelabeled with [3H]arachidonic acid and stimulated with angiotensin II. Although the specific activities of the choline and inositol glycerophospholipid pools were similar after the labeling interval, ethanolamine glycerophospholipids had a specific activity of only 20% of the specific activity of choline and inositol glycerophospholipids. Despite the marked disparity in the specific activities of these three phospholipid classes, angiotensin II stimulation resulted in similar fractional losses (35-41%) of [3H]arachidonic acid from vascular smooth muscle choline, ethanolamine, and inositol glycerophospholipid classes. Reverse-phase HPLC demonstrated that >60% of the [3H]arachidonic acid released from ethanolamine glycerophospholipids after angiotensin II stimulation originated from plasmenylethanolamine molecular species. Taken together, the results demonstrate that the major phospholipid storage depot for arachidonic acid in vascular smooth muscle cells are plasmenylethanolamine molecular species which are important substrates for the phospholipase(s) that are activated during agonist stimulation

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

    Energy Technology Data Exchange (ETDEWEB)

    Sahni, Abha [Aab Cardiovascular Research Institute, Department of Medicine, University of Rochester School of Medicine and Dentistry, Rochester, NY 14642 (United States); Department of Pathology, University of Texas Medical Branch, Galveston, TX 77555 (United States); Wang, Nadan [Aab Cardiovascular Research Institute, Department of Medicine, University of Rochester School of Medicine and Dentistry, Rochester, NY 14642 (United States); Center for Translational Medicine, Department of Medicine, Thomas Jefferson University, Philadelphia, PA 19107 (United States); Alexis, Jeffrey, E-mail: jeffrey_alexis@urmc.rochester.edu [Aab Cardiovascular Research Institute, Department of Medicine, University of Rochester School of Medicine and Dentistry, Rochester, NY 14642 (United States)

    2013-02-15

    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.

  12. 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)

    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

  13. OVER-EXPRESSION OF EXTRACELLULAR SIGNAL-REGULATED KINASE IN VASCULAR SMOOTH MUSCLE CELL OF HYPERTENSIVE RATS

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    Objective To investigate whether extracellular signal-regulated kinase (ERK1/2) was involved in changes of vascular smooth muscle cell (VSMC) under hypertension. Methods Two-kidney one clip Wistar hypertensive rats (WHR) were sacrificed and their right kidneys were harvested 4 weeks after surgery. The spontaneously hypertensive rats (SHR) were divided into 4, 8, and 16 weeks old groups (SHR4w, SHR8w, and SHR16w), respectively. The control group were sham operated age-matched Wistar rats. Immunohistochemical technique and Western blotting were applied to study ERK1/2 protein expression in VSMC of the renal vascular trees in WHR, SHR, and control rats. Results Blood pressure in two-kidney one clip WHR obviously increased at one week after surgery, and reached to 198.00 ± 33.00 mm Hg at the end of experiment, significantly higher than that in the control rats ( P < 0. 01 ). Blood pressure in SHR4w ( 108.00 ± 11.25 mm Hg) was similar to that in the controls. However, it rose to 122.25 ± 21.75 mm Hg in SHR8w, and even up to 201.75 ± 18.00 mm Hg in SHR16w, which were significantly higher than that of both the SHR4w and the controls ( P < 0. 01 ). The rate and degree of glomerular fibrosis in WHR were significantly higher than controls (P < 0. 05 ). Hyaline degeneration of the afferent arterioles was found in WHR. In contrast, either fibrosis of glomerulus or hyaline degeneration of the arterioles or protein casts was not observed in SHR4w, SHR8w,and SHR16w. Immunohistochemical staining results showed expression of ERK1 was similar to that of ERK2. The positive rates of ERK2 staining in VSMC of afferent arterioles, interlobular, interlobar, and arcuate arteries in two-kidney one clip WHR were significantly higher (7. 09% ± 1.75%, 14. 57% ± 4. 58%, 29.44% ± 7. 35%, and 13.63% ±3.85%, respectively) than that of the controls( P < 0. 01 ). The positive rates of ERK2 staining in VSMC at afferent arterioles, interlobular, interlobar, and arcuate arteries in SHR

  14. Sodium tanshinone IIA silate inhibits high glucose-induced vascular smooth muscle cell proliferation and migration through activation of AMP-activated protein kinase.

    Directory of Open Access Journals (Sweden)

    Wen-yu Wu

    Full Text Available The proliferation of vascular smooth muscle cells may perform a crucial role in the pathogenesis of diabetic vascular disease. AMPK additionally exerts several salutary effects on vascular function and improves vascular abnormalities. The current study sought to determine whether sodium tanshinone IIA silate (STS has an inhibitory effect on vascular smooth muscle cell (VSMC proliferation and migration under high glucose conditions mimicking diabetes without dyslipidemia, and establish the underlying mechanism. In this study, STS promoted the phosphorylation of AMP-activated protein kinase (AMPK at T172 in VSMCs. VSMC proliferation was enhanced under high glucose (25 mM glucose, HG versus normal glucose conditions (5.5 mM glucose, NG, and this increase was inhibited significantly by STS treatment. We utilized western blotting analysis to evaluate the effects of STS on cell-cycle regulatory proteins and found that STS increased the expression of p53 and the Cdk inhibitor, p21, subsequent decreased the expression of cell cycle-associated protein, cyclin D1. We further observed that STS arrested cell cycle progression at the G0/G1 phase. Additionally, expression and enzymatic activity of MMP-2, translocation of NF-κB, as well as VSMC migration were suppressed in the presence of STS. Notably, Compound C (CC, a specific inhibitor of AMPK, as well as AMPK siRNA blocked STS-mediated inhibition of VSMC proliferation and migration. We further evaluated its potential for activating AMPK in aortas in animal models of type 2 diabetes and found that Oral administration of STS for 10 days resulted in activation of AMPK in aortas from ob/ob or db/db mice. In conclusion, STS inhibits high glucose-induced VSMC proliferation and migration, possibly through AMPK activation. The growth suppression effect may be attributable to activation of AMPK-p53-p21 signaling, and the inhibitory effect on migration to the AMPK/NF-κB signaling axis.

  15. 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)

    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

  16. 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.

  17. Capsaicin from chili (Capsicum spp. inhibits vascular smooth muscle cell proliferation [v1; ref status: indexed, http://f1000r.es/4yk

    Directory of Open Access Journals (Sweden)

    Rongxia Liu

    2015-01-01

    Full Text Available Accelerated vascular smooth muscle cell (VSMC proliferation is implied in cardiovascular disease and significantly contributes to vessel lumen reduction following surgical interventions such as percutaneous transluminal coronary angioplasty or bypass surgery. Therefore, identification and characterization of compounds and mechanisms able to counteract VSMC proliferation is of potential therapeutic relevance. This work reveals the anti-proliferative effect of the natural product capsaicin from Capsicum spp. by quantification of metabolic activity and DNA synthesis in activated VSMC. The observed in vitro activity profile of capsaicin warrants further research on its mechanism of action and potential for therapeutic application.

  18. Curcumin inhibits LPS-induced inflammation in rat vascular smooth muscle cells in vitro via ROS-relative TLR4-MAPK/NF-κB pathways

    OpenAIRE

    Meng, Zhe; Yan, Chao; Deng, Qian; Gao, Deng-Feng; Niu, Xiao-lin

    2013-01-01

    Aim: To investigate whether curcumin (Cur) suppressed lipopolysaccharide (LPS)-induced inflammation in vascular smooth muscle cells (VSMCs) of rats, and to determine its molecular mechanisms. Methods: Primary rat VSMCs were treated with LPS (1 μg/L) and Cur (5, 10, or 30 μmol/L) for 24 h. The levels of MCP-1, TNF-α, and iNOS were measured using ELISA and real-time RT-PCR. NO level was analyzed with the Griess reaction. Western-blotting was used to detect the activation of TLR4, MAPKs, IκBα, N...

  19. Regulation of Collagen Type I in Vascular Smooth Muscle Cells by Competition between Nkx2.5 and δEF1/ZEB1

    OpenAIRE

    Ponticos, Markella; Partridge, Terrence; BLACK, CAROL M.; Abraham, David J.; Bou-Gharios, George

    2004-01-01

    A major component of the vessel wall of large arteries and veins is the extracellular matrix (ECM), which consists of collagens, elastin, and proteoglycans. Collagen type I is one of the most abundant of the ECM proteins. We have previously shown that the pro-collagen type I alpha 2 gene contains an enhancer which confers tissue-specific expression in the majority of collagen-producing cells, including blood vessels. In this paper, we delineate a specific vascular smooth muscle cell (vSMC) el...

  20. Osteoprotegerin inhibits calcification of vascular smooth muscle cell via down regulation of the Notch1-RBP-Jκ/Msx2 signaling pathway.

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    Shaoqiong Zhou

    Full Text Available OBJECTIVE: Vascular calcification is a common pathobiological process which occurs among the elder population and in patients with diabetes and chronic kidney disease. Osteoprotegerin, a secreted glycoprotein that regulates bone mass, has recently emerged as an important regulator of the development of vascular calcification. However, the mechanism is not fully understood. The purpose of this study is to explore novel signaling mechanisms of osteoprotegerin in the osteoblastic differentiation in rat aortic vascular smooth muscle cells (VSMCs. METHODS AND RESULTS: VSMCs were isolated from thoracic aorta of Sprague Dawley rats. Osteoblastic differentiation of VSMCs was induced by an osteogenic medium. We confirmed by Von Kossa staining and direct cellular calcium measurement that mineralization was significantly increased in VSMCs cultured in osteogenic medium; consistent with an enhanced alkaline phosphatase activity. This osteoblastic differentiation in VSMCs was significantly reduced by the addition of osteoprotegerin in a dose responsive manner. Moreover, we identified, by real-time qPCR and western blotting, that expression of Notch1 and RBP-Jκ were significantly up-regulated in VSMCs cultured in osteogenic medium at both the mRNA and protein levels, these effects were dose-dependently abolished by the treatment of osteoprotegerin. Furthermore, we identified that Msx2, a downstream target of the Notch1/RBP-Jκ signaling, was markedly down-regulated by the treatment of osteoprotegerin. CONCLUSION: Osteoprotegerin inhibits vascular calcification through the down regulation of the Notch1-RBP-Jκ signaling pathway.

  1. Vascular smooth muscle cells for use in vascular tissue engineering obtained by endothelial-to-mesenchymal transdifferentiation (EnMT) on collagen matrices

    NARCIS (Netherlands)

    Krenning, Guido; Moonen, Jan-Renier A. J.; van Luyn, Marja J. A.; Harmsen, Martin C.

    2008-01-01

    The discovery of the endothelial progenitor cell (EPC) has led to an intensive research effort into progenitor cell-based tissue engineering of (small-diameter) blood vessels. Herein, EPC are differentiated to vascular endothelial cells and serve as the inner lining of bioartificial vessels. As yet,

  2. Angiotensin II modulates interleukin-1{beta}-induced inflammatory gene expression in vascular smooth muscle cells via interfering with ERK-NF-{kappa}B crosstalk

    Energy Technology Data Exchange (ETDEWEB)

    Xu, Shanqin [Vascular Biology Unit, Whitaker Cardiovascular Institute, Boston University School of Medicine, Boston, MA (United States); Zhi, Hui [Cardiovascular Division, Department of Medicine, Brigham and Women' s Hospital, Harvard Medical School, Boston, MA (United States); Hou, Xiuyun [Vascular Biology Unit, Whitaker Cardiovascular Institute, Boston University School of Medicine, Boston, MA (United States); Jiang, Bingbing, E-mail: bjiang1@rics.bwh.harvard.edu [Vascular Biology Unit, Whitaker Cardiovascular Institute, Boston University School of Medicine, Boston, MA (United States); Cardiovascular Division, Department of Medicine, Brigham and Women' s Hospital, Harvard Medical School, Boston, MA (United States)

    2011-07-08

    Highlights: {yields} We examine how angiotensin II modulates ERK-NF-{kappa}B crosstalk and gene expression. {yields} Angiotensin II suppresses IL-1{beta}-induced prolonged ERK and NF-{kappa}B activation. {yields} ERK-RSK1 signaling is required for IL-1{beta}-induced prolonged NF-{kappa}B activation. {yields} Angiotensin II modulates NF-{kappa}B responsive genes via regulating ERK-NF-{kappa}B crosstalk. {yields} ERK-NF-{kappa}B 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-1{beta}-induced prolonged phosphorylation of extracellular signal-regulated kinase (ERK) and ribosomal S6 kinase (RSK)-1, and nuclear translocation of nuclear factor (NF)-{kappa}B, 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-1{beta}-induced iNOS expression without influencing VCAM-1 expression. In vivo experiments showed that interleukin-1{beta}, iNOS, and VCAM-1 expression were detectable in the aortic arches of both wild-type and apolipoprotein E-deficient (ApoE{sup -/-}) mice. VCAM-1 and iNOS expression were higher in ApoE{sup -/-} than in wild type mouse aortic arches. Angiotensin II infusion (3.2 mg/kg/day, for 6 days, via subcutaneous osmotic pump) in ApoE{sup -/-} 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

  3. Neuropeptide Y stimulates DNA synthesis in human vascular smooth muscle cells through neuropeptide Y Y1 receptors

    DEFF Research Database (Denmark)

    Nilsson, T; Edvinsson, L

    2000-01-01

    We investigated the mitogenic effect, measured as [3H]thymidine incorporation, of neuropeptide Y (NPY) on smooth muscle cells (SMCs) from human subcutaneous arteries (diameter: 0.4 mm). NPY stimulated DNA synthesis in a concentration-dependent manner, Emax 32 +/- 5% relative to control. The effec...

  4. 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...

  5. Ubiquitin carboxyl terminal hydrolase L1 negatively regulates TNF{alpha}-mediated vascular smooth muscle cell proliferation via suppressing ERK activation

    Energy Technology Data Exchange (ETDEWEB)

    Ichikawa, Tomonaga; Li, Jinqing; Dong, Xiaoyu; Potts, Jay D. [Department of Cell Biology and Anatomy, University of South Carolina School of Medicine, Columbia, SC 29208 (United States); Tang, Dong-Qi [Department of Pathology, Immunology, and Laboratory Medicine, University of Florida College of Medicine, Gainesville, FL 32610-0275 (United States); Li, Dong-Sheng, E-mail: dsli@yymc.edu.cn [Hubei Key Laboratory of Embryonic Stem Cell Research, Tai He Hospital, Yunyang Medical College, 32 S. Renmin Rd., Shiyan, Hubei 442000 (China); 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)

    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 (TNF{alpha})-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 TNF{alpha}-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. TNF{alpha} 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.

  6. Oleic Acid Increases Synthesis and Secretion of VEGF in Rat Vascular Smooth Muscle Cells: Role of Oxidative Stress and Impairment in Obesity

    Directory of Open Access Journals (Sweden)

    Mariella Trovati

    2013-09-01

    Full Text Available Obesity is characterized by poor collateral vessel formation, a process involving vascular endothelial growth factor (VEGF action on vascular smooth muscle cells (VSMC. Free fatty acids are involved in the pathogenesis of obesity vascular complications, and we have aimed to clarify whether oleic acid (OA enhances VEGF synthesis/secretion in VSMC, and whether this effect is impaired in obesity. In cultured aortic VSMC from lean and obese Zucker rats (LZR and OZR, respectively we measured the influence of OA on VEGF-A synthesis/secretion, signaling molecules and reactive oxygen species (ROS. In VSMC from LZR we found the following: (a OA increases VEGF-A synthesis/secretion by a mechanism blunted by inhibitors of Akt, mTOR, ERK-1/2, PKC-beta, NADPH-oxidase and mitochondrial electron transport chain complex; (b OA activates the above mentioned signaling pathways and increases ROS; (c OA-induced activation of PKC-beta enhances oxidative stress, which activates signaling pathways responsible for the increased VEGF synthesis/secretion. In VSMC from OZR, which present enhanced baseline oxidative stress, the above mentioned actions of OA on VEGF-A, signaling pathways and ROS are impaired: this impairment is reproduced in VSMC from LZR by incubation with hydrogen peroxide. Thus, in OZR chronically elevated oxidative stress causes a resistance to the action on VEGF that OA exerts in LZR by increasing ROS.

  7. Association between the Hypomethylation of Osteopontin and Integrin β3 Promoters and Vascular Smooth Muscle Cell Phenotype Switching in Great Saphenous Varicose Veins

    Directory of Open Access Journals (Sweden)

    Han Jiang

    2014-10-01

    Full Text Available Lower extremity varicose veins are a common condition in vascular surgery and proliferation of vascular smooth muscle cells (VSMCs in the intima is a significant pathological feature of varicosity. However, the pathogenesis of varicose veins is not fully understood. Osteopontin (OPN could promote the migration and adhesion of VSMCs through the cell surface receptor integrin β3 and the cooperation of OPN and integrin β3 is involved in many vascular diseases. However, the role of OPN and integrin β3 in varicosity remains unclear. In the current study, we found that the methylation levels in the promoter regions of OPN and integrin β3 genes in the VSMCs of varicose veins are reduced and the protein expression of OPN and integrin β3 are increased, compared with normal veins. Furthermore, it was observed that VSMCs in the neointima of varicose veins were transformed into the synthetic phenotype. Collectively, hypomethylation of the promoter regions for OPN and integrin β3 genes may increase the expression of these genes in varicosity, which is closely related to VSMC phenotype switching. Hypomethylation of the promoter regions for OPN and integrin β3 genes may be a key factor in the pathogenesis of varicosity.

  8. Association between the hypomethylation of osteopontin and integrin β3 promoters and vascular smooth muscle cell phenotype switching in great saphenous varicose veins.

    Science.gov (United States)

    Jiang, Han; Lun, Yu; Wu, Xiaoyu; Xia, Qian; Zhang, Xiaoyu; Xin, Shijie; Zhang, Jian

    2014-10-17

    Lower extremity varicose veins are a common condition in vascular surgery and proliferation of vascular smooth muscle cells (VSMCs) in the intima is a significant pathological feature of varicosity. However, the pathogenesis of varicose veins is not fully understood. Osteopontin (OPN) could promote the migration and adhesion of VSMCs through the cell surface receptor integrin β3 and the cooperation of OPN and integrin β3 is involved in many vascular diseases. However, the role of OPN and integrin β3 in varicosity remains unclear. In the current study, we found that the methylation levels in the promoter regions of OPN and integrin β3 genes in the VSMCs of varicose veins are reduced and the protein expression of OPN and integrin β3 are increased, compared with normal veins. Furthermore, it was observed that VSMCs in the neointima of varicose veins were transformed into the synthetic phenotype. Collectively, hypomethylation of the promoter regions for OPN and integrin β3 genes may increase the expression of these genes in varicosity, which is closely related to VSMC phenotype switching. Hypomethylation of the promoter regions for OPN and integrin β3 genes may be a key factor in the pathogenesis of varicosity.

  9. Dynamic Culturing of Smooth Muscle Cells in Tubular Poly(Trimethylene Carbonate) Scaffolds for Vascular Tissue Engineering

    NARCIS (Netherlands)

    Song, Yan; Wennink, Jos W.H.; Kamphuis, Marloes M.J.; Sterk, Lotus M.T.; Vermes, Istvan; Poot, André A.; Feijen, Jan; Grijpma, Dirk W.

    2011-01-01

    Porous, tubular, flexible, and elastic poly(trimethylene carbonate) (PTMC) scaffolds (length 8 cm and inner diameter 3 mm) for vascular tissue engineering were prepared by means of a dip-coating and particulate leaching procedure. Using NaCl as porogen, scaffolds with an average pore size of 110 μm

  10. C-TYPE NATRIURETIC PEPTIDE INHIBITS UPREGULATION OF αl-ADRENOCEPTOR AND INOSITOL 1,4,5-TRISPHOSPHATE RECEPTOR IN RAT VASCULAR SMOOTH MUSCLE AFTER VASCULAR ENDOTHELIAL INJURY

    Institute of Scientific and Technical Information of China (English)

    王晓红; 杨军; 佟利家; 苏静怡; 唐朝枢; 刘乃奎

    2000-01-01

    Objective. In a model of balloon injury of rat aortic endothelium, the effects of C-type natriuretic peptide(CNP) on al-adrenoreceptar and inositol 1,4,5-triphosphate (IP3) receptor were studied. Methods. Aortic injuries were produced by vascular endothelium-denudation, αl- adrenoreceptor in smooth muscle sarcolemma and IP3 receptor in smooth muscle sarcoplasmic reticulum in the rat aorta were assayed by radioactive analysis method. Results. It was found that neoinfma was formed and the coraents of DNA, collagen and elastin of each intimamedia were significantly increased in 7 days and 21 days after balloon injury of rat aorta, α1-adrenoreceptor in smooth muscle sarcolemma and IP3 receptor in sarcoplasmic reticulum were also upwodated. Results also showed that the administration of CNP i.p significantly decreased the contents of DNA, collagen and elaslin of each iraima-media, and inhibited the up-regulation of α1-adrenoreceptor and IP3 receptor. Conelusion. The inhibition of the up-regulation of α1-adrenoreceptor and IP3 receptor by CNP might be one of the mechanisms of its suppressive action on intimal proliferation.

  11. 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.

  12. Poly(ADP-ribose polymerase 1 is indispensable for transforming growth factor-β Induced Smad3 activation in vascular smooth muscle cell.

    Directory of Open Access Journals (Sweden)

    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.

  13. 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.

  14. 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

  15. Hydrogen sulfide releasing aspirin, ACS14, attenuates high glucose-induced increased methylglyoxal and oxidative stress in cultured vascular smooth muscle cells.

    Science.gov (United States)

    Huang, Qian; Sparatore, Anna; Del Soldato, Piero; Wu, Lingyun; Desai, Kaushik

    2014-01-01

    Hydrogen sulfide is a gasotransmitter with vasodilatory and anti-inflammatory properties. Aspirin is an irreversible cyclooxygenase inhibitor anti-inflammatory drug. ACS14 is a novel synthetic hydrogen sulfide releasing aspirin which inhibits cyclooxygenase and has antioxidant effects. Methylglyoxal is a chemically active metabolite of glucose and fructose, and a major precursor of advanced glycation end products formation. Methylglyoxal is harmful when produced in excess. Plasma methylglyoxal levels are significantly elevated in diabetic patients. Our aim was to investigate the effects of ACS14 on methylglyoxal levels in cultured rat aortic vascular smooth muscle cells. We used cultured rat aortic vascular smooth muscle cells for the study. Methylglyoxal was measured by HPLC after derivatization, and nitrite+nitrate with an assay kit. Western blotting was used to determine NADPH oxidase 4 (NOX4) and inducible nitric oxide synthase (iNOS) protein expression. Dicholorofluorescein assay was used to measure oxidative stress. ACS14 significantly attenuated elevation of intracellular methylglyoxal levels caused by incubating cultured vascular smooth muscle cells with methylglyoxal (30 µM) and high glucose (25 mM). ACS14, but not aspirin, caused a significant attenuation of increase in nitrite+nitrate levels caused by methylglyoxal or high glucose. ACS14, aspirin, and sodium hydrogen sulfide (NaHS, a hydrogen sulfide donor), all attenuated the increase in oxidative stress caused by methylglyoxal and high glucose in cultured cells. ACS14 prevented the increase in NOX4 expression caused by incubating the cultured VSMCs with MG (30 µM). ACS14, aspirin and NaHS attenuated the increase in iNOS expression caused by high glucose (25 mM). In conclusion, ACS14 has the novel ability to attenuate an increase in methylglyoxal levels which in turn can reduce oxidative stress, decrease the formation of advanced glycation end products and prevent many of the known deleterious effects

  16. Hydrogen sulfide releasing aspirin, ACS14, attenuates high glucose-induced increased methylglyoxal and oxidative stress in cultured vascular smooth muscle cells.

    Directory of Open Access Journals (Sweden)

    Qian Huang

    Full Text Available Hydrogen sulfide is a gasotransmitter with vasodilatory and anti-inflammatory properties. Aspirin is an irreversible cyclooxygenase inhibitor anti-inflammatory drug. ACS14 is a novel synthetic hydrogen sulfide releasing aspirin which inhibits cyclooxygenase and has antioxidant effects. Methylglyoxal is a chemically active metabolite of glucose and fructose, and a major precursor of advanced glycation end products formation. Methylglyoxal is harmful when produced in excess. Plasma methylglyoxal levels are significantly elevated in diabetic patients. Our aim was to investigate the effects of ACS14 on methylglyoxal levels in cultured rat aortic vascular smooth muscle cells. We used cultured rat aortic vascular smooth muscle cells for the study. Methylglyoxal was measured by HPLC after derivatization, and nitrite+nitrate with an assay kit. Western blotting was used to determine NADPH oxidase 4 (NOX4 and inducible nitric oxide synthase (iNOS protein expression. Dicholorofluorescein assay was used to measure oxidative stress. ACS14 significantly attenuated elevation of intracellular methylglyoxal levels caused by incubating cultured vascular smooth muscle cells with methylglyoxal (30 µM and high glucose (25 mM. ACS14, but not aspirin, caused a significant attenuation of increase in nitrite+nitrate levels caused by methylglyoxal or high glucose. ACS14, aspirin, and sodium hydrogen sulfide (NaHS, a hydrogen sulfide donor, all attenuated the increase in oxidative stress caused by methylglyoxal and high glucose in cultured cells. ACS14 prevented the increase in NOX4 expression caused by incubating the cultured VSMCs with MG (30 µM. ACS14, aspirin and NaHS attenuated the increase in iNOS expression caused by high glucose (25 mM. In conclusion, ACS14 has the novel ability to attenuate an increase in methylglyoxal levels which in turn can reduce oxidative stress, decrease the formation of advanced glycation end products and prevent many of the known

  17. Inhibition of Phosphate-Induced Vascular Smooth Muscle Cell Osteo-/Chondrogenic Signaling and Calcification by Bafilomycin A1 and Methylamine

    Directory of Open Access Journals (Sweden)

    Ioana Alesutan

    2015-09-01

    Full Text Available Background/Aims: Excessive phosphate concentrations trigger vascular calcification, an active process promoted by osteoinduction of vascular smooth muscle cells (VSMCs with increased expression and activity of transcription factor RUNX2 (Core-binding factor α1, CBFA1, alkaline phosphatase (ALPL, TGFß1, transcription factor NFAT5, and NFAT5-sensitive transcription factor SOX9. The osteoinductive signaling and vascular calcification of hyperphosphatemic klotho-hypomorphic mice could be reversed by treatment with NH4Cl, effects involving decrease of TGFß1 and inhibition of NFAT5-dependent osteoinductive signaling. Known effects of NH4Cl include alkalinization of acidic cellular compartments. The present study explored whether osteo-/chondrogenic signaling could be influenced by alkalinization of acidic cellular compartments following inhibition of the vacuolar H+ ATPase with bafilomycin A1 or following dissipation of the pH gradient across the membranes of acidic cellular compartments with methylamine. Methods: Primary human aortic smooth muscle cells (HAoSMCs were treated with high phosphate to trigger osteo-/chondrogenic signaling and calcification in the absence or presence of bafilomycin A1 or methylamine. Calcium content was determined using a QuantiChrom Calcium assay, ALP activity by a colorimetric assay and transcript levels by quantitative RT-PCR. Results: High phosphate increased significantly the calcium deposition, CBFA1 and ALPL mRNA expression as well as alkaline phosphatase activity in HAoSMCs, all effects ameliorated by both, bafilomycin A1 and methylamine. High phosphate further significantly up-regulated the mRNA levels of TGFB1, NFAT5 and SOX9, effects significantly blunted by additional treatment with bafilomycin A1 or methylamine. Treatment of HAoSMCs with human TGFß1 protein or high phosphate up-regulated NFAT5, SOX9, CBFA1 and ALPL mRNA expression to similarly high levels which could not be further increased by combined

  18. No influence of OPG and its ligands, RANKL and TRAIL, on proliferation and regulation of the calcification process in primary human vascular smooth muscle cells

    DEFF Research Database (Denmark)

    Olesen, Malene; Skov, Vibe; Mechta, Mie;

    2012-01-01

    The aim of this study was to examine the effects of the OPG-RANKL-TRAIL system on proliferation, regulation of calcification-associated genes and calcification of human vascular smooth muscle cells (HVSMCs). Small interfering (si)RNA-mediated knockdown of OPG was followed by treatment of HVSMCs...... with recombinant RANKL or TRAIL. Regulation of a calcification-associated gene set was assayed by pathway analysis of microarray results. The lack of OPG in HVSMCs or treatment with RANKL or TRAIL did not affect proliferation of HVSMCs. In addition, OPG, RANKL or TRAIL did not modify the regulation...... of a calcification-associated gene set. Finally, in the long term calcification assay, we found that cells isolated from seven different human donors showed a great variability in the response to RANKL and insulin. However, overall RANKL and/or insulin did not affect the development of calcification of HVSMCs...

  19. Synthesis and biological evaluation of quinoxaline-5,8-diones that inhibit vascular smooth muscle cell proliferation.

    Science.gov (United States)

    Chung, Hwa-Jin; Jung, Ok-Jai; Chae, Mi Jin; Hong, Sung-Yu; Chung, Kwang-Hoe; Lee, Sang Kook; Ryu, Chung-Kyu

    2005-07-15

    A series of 6-arylamino-2,3-bis(pyridin-2-yl)-7-chloro-quinoxaline-5,8-diones were synthesized and evaluated for their inhibitory activity on the rat aortic smooth muscle cell (RAoSMC) proliferation. The quinoxaline-5,8-diones exhibited a potent antiproliferative activity. Further mechanistic study revealed that the inhibitory effect of one representative quinoxaline-5,8-dione on SMC proliferation was mediated by modulation of the extracellular signal-regulated kinase 1/2 signaling pathway in the RAoSMCs.

  20. 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.

  1. Exendin-4 Prevents Vascular Smooth Muscle Cell Proliferation and Migration by Angiotensin II via the Inhibition of ERK1/2 and JNK Signaling Pathways.

    Directory of Open Access Journals (Sweden)

    Kosuke Nagayama

    Full Text Available Angiotensin II (Ang II is a main pathophysiological culprit peptide for hypertension and atherosclerosis by causing vascular smooth muscle cell (VSMC proliferation and migration. Exendin-4, a glucagon-like peptide-1 (GLP-1 receptor agonist, is currently used for the treatment of type-2 diabetes, and is believed to have beneficial effects for cardiovascular diseases. However, the vascular protective mechanisms of GLP-1 receptor agonists remain largely unexplained. In the present study, we examined the effect of exendin-4 on Ang II-induced proliferation and migration of cultured rat aortic smooth muscle cells (RASMC. The major findings of the present study are as follows: (1 Ang II caused a phenotypic switch of RASMC from contractile type to synthetic proliferative type cells; (2 Ang II caused concentration-dependent RASMC proliferation, which was significantly inhibited by the pretreatment with exendin-4; (3 Ang II caused concentration-dependent RASMC migration, which was effectively inhibited by the pretreatment with exendin-4; (4 exendin-4 inhibited Ang II-induced phosphorylation of ERK1/2 and JNK in a pre-incubation time-dependent manner; and (5 U0126 (an ERK1/2 kinase inhibitor and SP600125 (a JNK inhibitor also inhibited both RASMC proliferation and migration induced by Ang II stimulation. These results suggest that exendin-4 prevented Ang II-induced VSMC proliferation and migration through the inhibition of ERK1/2 and JNK phosphorylation caused by Ang II stimulation. This indicates that GLP-1 receptor agonists should be considered for use in the treatment of cardiovascular diseases in addition to their current use in the treatment of diabetes mellitus.

  2. 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.

  3. Wild-type LRP6 inhibits, whereas atherosclerosis-linked LRP6R611C increases PDGF-dependent vascular smooth muscle cell proliferation

    Science.gov (United States)

    Keramati, Ali R.; Singh, Rajvir; Lin, Aiping; Faramarzi, Saeed; Ye, Zhi-jia; Mane, Shrikant; Tellides, George; Lifton, Richard P.; Mani, Arya

    2011-01-01

    Vascular smooth muscle cell (VSMC) proliferation is an important event in atherosclerosis and other vasculopathies. PDGF signaling is a key mediator of SMC proliferation, but the mechanisms that control its activity remain unclear. We previously identified a mutation in LDL receptor-related protein 6 (LRP6), LRP6R611C, that causes early atherosclerosis. Examination of human atherosclerotic coronary arteries showed markedly increased expression of LRP6 and colocalization with PDGF receptor β (PDGFR-β). Further investigation showed that wild-type LRP6 inhibits but LRP6R611C promotes VSMC proliferation in response to PDGF. We found that wild-type LRP6 forms a complex with PDGFR-β and enhances its lysosomal degradation, functions that are severely impaired in LRP6R611C. Further, we observed that wild-type and mutant LRP6 regulate cell-cycle activity by triggering differential effects on PDGF-dependent pathways. These findings implicate LRP6 as a critical modulator of PDGF-dependent regulation of cell cycle in smooth muscle and indicate that loss of this function contributes to development of early atherosclerosis in humans. PMID:21245321

  4. Pharmacological evidence that potentiation of plasmalemmal Ca(2+)-extrusion is functionally coupled to inhibition of SR Ca(2+)-ATPases in vascular smooth muscle cells.

    Science.gov (United States)

    Zhang, Wen-Bo; Kwan, Chiu-Yin

    2016-04-01

    Cyclopiazonic acid (CPA), a specific inhibitor of sarcoplasmic reticulum (SR) Ca(2+)-ATPases, causes slowly developing and subsequently diminishing characteristic contractions in vascular smooth muscle, and the second application of CPA has incompletely repeatable effects, depending on the vessel type. The objective of the present study was to examine the mechanisms underlying the significant decrease of CPA-induced contractions upon the second application. A pharmacological intervention of Ca(2+) extrusion process as a strategy was performed to modulate vasoconstrictor effects of CPA in rat aortic ring preparations. CPA-induced contractions, expressed as percentages of the contractions induced by KCl (80 mM), were significantly decreased from 44.1 ± 5.7 to 7.6 ± 1.8 % (P CPA-induced contractions were sustained and completely repeatable in Na(+)-free and low Na(+) medium. Furthermore, we found that the contractions were completely repeatable in the presence of 2',4'-dichlorobenzamil, an inhibitor of the forward mode of Na(+)/Ca(2+) exchangers, but not of KBR7943, an inhibitor of the reverse mode of Na(+)/Ca(2+) exchangers. Our findings indicate that CPA by inducing a transient rise in cytosolic Ca(2+) level causes a long-lasting upregulation of plasma membrane (PM) Ca(2+) extruders and thus leads to a diminished contraction upon its second application in blood vessels. This suggests that there is a functional coupling between PM Ca(2+) extruders and SR Ca(2+)-ATPases in rat aortic smooth muscle cells. PMID:26842648

  5. Composition of connective tissues and morphometry of vascular smooth muscle in arterial wall of DOCA-salt hypertensive rats - In relation with arterial remodeling.

    Science.gov (United States)

    Hayashi, Kozaburo; Shimizu, Emiko

    2016-05-01

    Hypertension (HT) was induced in Wistar rats aged 16 and 48 weeks by a deoxycortico-sterone acetate (DOCA)-salt procedure. Common carotid arteries were resected 16 weeks after, and their histological specimens were selectively stained for observations of collagen, elastin, and vascular smooth muscle (VSM) cells. Then, the fractions of collagen and elastin and their radial distributions, and the size and number of VSM cells were determined with an image analyzer. These results were compared with the results from age-matched, non-treated, normotensive (NT) animals and also with those from our previous biomechanical studies. In both age groups, there were no significant differences in the fractions of collagen and elastin, and the ratio of collagen to elastin content between HT and NT arteries. These results correspond well with our previous biomechanical results, which showed no significant difference in wall elasticity between HT and NT vessels. Moreover, in the innermost layer out of 4 layers bordered with thick elastic lamellae, the fraction of collagen was significantly greater in HT arteries than in NT ones, which is attributable to HT-related stress concentration in the layer. VSM cells were significantly hypertrophied and their content was increased by HT, although their total number in the media remained unchanged. The increased size and content of cells correspond to the enhancement of vascular tone and contractility in HT arteries.

  6. Atorvastatin Protects Vascular Smooth Muscle Cells From TGF-β1-Stimulated Calcification by Inducing Autophagy via Suppression of the β-Catenin Pathway

    Directory of Open Access Journals (Sweden)

    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.

  7. Andrographolide, a Novel NF-κB Inhibitor, Induces Vascular Smooth Muscle Cell Apoptosis via a Ceramide-p47phox-ROS Signaling Cascade

    Directory of Open Access Journals (Sweden)

    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.

  8. Composition of connective tissues and morphometry of vascular smooth muscle in arterial wall of DOCA-salt hypertensive rats - In relation with arterial remodeling.

    Science.gov (United States)

    Hayashi, Kozaburo; Shimizu, Emiko

    2016-05-01

    Hypertension (HT) was induced in Wistar rats aged 16 and 48 weeks by a deoxycortico-sterone acetate (DOCA)-salt procedure. Common carotid arteries were resected 16 weeks after, and their histological specimens were selectively stained for observations of collagen, elastin, and vascular smooth muscle (VSM) cells. Then, the fractions of collagen and elastin and their radial distributions, and the size and number of VSM cells were determined with an image analyzer. These results were compared with the results from age-matched, non-treated, normotensive (NT) animals and also with those from our previous biomechanical studies. In both age groups, there were no significant differences in the fractions of collagen and elastin, and the ratio of collagen to elastin content between HT and NT arteries. These results correspond well with our previous biomechanical results, which showed no significant difference in wall elasticity between HT and NT vessels. Moreover, in the innermost layer out of 4 layers bordered with thick elastic lamellae, the fraction of collagen was significantly greater in HT arteries than in NT ones, which is attributable to HT-related stress concentration in the layer. VSM cells were significantly hypertrophied and their content was increased by HT, although their total number in the media remained unchanged. The increased size and content of cells correspond to the enhancement of vascular tone and contractility in HT arteries. PMID:26987272

  9. Pravastatin inhibits fibrinogen- and FDP-induced inflammatory response via reducing the production of IL-6, TNF-α and iNOS in vascular smooth muscle cells.

    Science.gov (United States)

    Lu, Peipei; Liu, Juntian; Pang, Xiaoming

    2015-10-01

    Atherosclerosis is a chronic inflammatory response of the arterial wall to pro‑atherosclerotic factors. As an inflammatory marker, fibrinogen directly participates in the pathogenesis of atherosclerosis. Our previous study demonstrated that fibrinogen and fibrin degradation products (FDP) produce a pro‑inflammatory effect on vascular smooth muscle cells (VSMCs) through inducing the production of interleukin‑6 (IL‑6), tumor necrosis factor‑α (TNF‑α) and inducible nitric oxide synthase (iNOS). In the present study, the effects of pravastatin on fibrinogen‑ and FDP‑induced expression of IL‑6, TNF‑α and iNOS were observed in VSMCs. The results showed that pravastatin dose‑dependently inhibited fibrinogen‑ and FDP‑stimulated expression of IL‑6, TNF‑α and iNOS in VSMCs at the mRNA and protein level. The maximal inhibition of protein expression of IL‑6, TNF‑α and iNOS was 46.9, 42.7 and 49.2% in fibrinogen‑stimulated VSMCs, and 50.2, 49.8 and 53.6% in FDP‑stimulated VSMCs, respectively. This suggests that pravastatin has the ability to relieve vascular inflammation via inhibiting the generation of IL‑6, TNF‑α and iNOS. The results of the present study may aid in further explaining the beneficial effects of pravastatin on atherosclerosis and related cardiovascular diseases. In addition, they suggest that application of pravastatin may be beneficial for prevention of atherosclerosis formation in hyperfibrinogenemia.

  10. Ceramide mediates Ox-LDL-induced human vascular smooth muscle cell calcification via p38 mitogen-activated protein kinase signaling.

    Directory of Open Access Journals (Sweden)

    Lizhen Liao

    Full Text Available Vascular calcification is associated with significant cardiovascular morbidity and mortality, and has been demonstrated as an actively regulated process resembling bone formation. Oxidized low density lipoprotein (Ox-LDL has been identified as a regulatory factor involved in calcification of vascular smooth muscle cells (VSMCs. Additionally, over-expression of recombinant human neutral sphingomyelinase (N-SMase has been shown to stimulate VSMC apoptosis, which plays an important role in the progression of vascular calcification. The aim of this study is to investigate whether ceramide regulates Ox-LDL-induced calcification of VSMCs via activation of p38 mitogen-activated protein kinase (MAPK pathway. Ox-LDL increased the activity of N-SMase and the level of ceramide in cultured VSMCs. Calcification and the osteogenic transcription factor, Msx2 mRNA expression were reduced by N-SMase inhibitor, GW4869 in the presence of Ox-LDL. Usage of GW4869 inhibited Ox-LDL-induced apoptosis in VSMCs, an effect which was reversed by C2-ceramide. Additionally, C2-ceramide treatment accelerated VSMC calcification, with a concomitant increase in ALP activity. Furthermore, C2-ceramide treatment enhanced Ox-LDL-induced VSMC calcification. Addition of caspase inhibitor, ZVAD-fmk attenuated Ox-LDL-induced calcification. Both Ox-LDL and C2-ceramide treatment increased the phosphorylation of p38 MAPK. Inhibition of p38 MAPK by SB203580 attenuated Ox-LDL-induced calcification of VSMCs. These data suggest that Ox-LDL activates N-SMase-ceramide signaling pathway, and stimulates phosphorylation of p38 MAPK, leading to apoptosis in VSMCs, which initiates VSMC calcification.

  11. 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

  12. Effect of vascular endothelial growth factor and its receptor KDR on human airway smooth muscle cells proliferation

    Institute of Scientific and Technical Information of China (English)

    ZOU hui; XU Yong-jian; ZHANG Zhen-xiang

    2005-01-01

    @@ Airway remodeling with inflammatory cell infiltration, epithelial shedding, basement membrane thickening and increased mass of airway smooth muscle (ASM) is an important determinant of bronchial obstruction and hyperresponsiveness in asthma.1,2 Increased ASM mass is by far the most important abnormality responsible for excessive airway narrowing and compliance of the airway wall in asthma.1-3 ASM growth and proliferation in asthma is a complex phenomenon of which the underlying mechanisms are difficult to investigate in vivo. The increased amount of ASM in asthmatics is an indication of abnormal cell proliferation and growth, but little is known regarding the molecular mechanisms and factors that regulate ASM cell proliferation and growth in asthma.

  13. Cigarette smoke extracts promote vascular smooth muscle cell proliferation and enhances contractile responses in the vasculature and airway

    DEFF Research Database (Denmark)

    Xu, Cang-Bao; Lei, Ying; Chen, Qingwen;

    2010-01-01

    Cigarette smoke exposure is a strong risk factor for cardiovascular and respiratory diseases. However, the knowledge about how cigarette smoke induces damage to vasculature and airway is limited. The present study was designed to examine the effects of cigarette smoke particles extracted by heptane...... (heptane-soluble smoke particles, HSP), by water (water-soluble smoke particles, WSP) and by DMSO (DMSO-soluble smoke particles, DSP), which represent lipophilic, hydrophilic and ambiphoteric constituents from the cigarette smoke, respectively. Human aortic smooth muscle cell (HASMC) proliferation...... responses to sarafotoxin 6c, U46619 or bradykinin in rat mesenteric artery and/or in bronchi. ERK1/2 is activated by HSP and DSP in HASMCs and inhibition of ERK1/2 abrogated the smoke extracts-induced HASMC proliferation, while blockage of nicotinic receptors had no effects, suggesting that the toxic...

  14. Nuclear Receptor Nurr1 Is Expressed In and Is Associated With Human Restenosis and Inhibits Vascular Lesion Formation In Mice Involving Inhibition of Smooth Muscle Cell Proliferation and Inflammation

    NARCIS (Netherlands)

    P.I. Bonta; T.W.H. Pols; C.M. van Tiel; M. Vos; E.K. Arkenbout; J. Rohlena; K.T. Koch; M.P.M. de Maat; M.W.T. Tanck; R.J. de Winter; H. Pannekoek; E.A.L. Biessen; I. Bot; C.J.M. de Vries

    2010-01-01

    Background-Restenosis is the major drawback of percutaneous coronary interventions involving excessive activation and proliferation of vascular smooth muscle cells (SMCs). The nuclear receptor Nurr1 is an early response gene known mainly for its critical role in the development of dopamine neurons.

  15. Regulation of the sodium/potassium/chloride cotransporter by calcium and cyclic AMP in cultured vascular smooth muscle cells

    Energy Technology Data Exchange (ETDEWEB)

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

    1987-05-01

    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 /sup 86/Rb influx or efflux. Five uM furosemide or 0.2 uM bumetanide inhibited influx by 50%. Furosemide-inhibitable /sup 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 /sup 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 /sup 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.

  16. Co-culture of vascular endothelial cells and smooth muscle cells by hyaluronic acid micro-pattern on titanium surface

    Science.gov (United States)

    Li, Jingan; Li, Guicai; Zhang, Kun; Liao, Yuzhen; Yang, Ping; Maitz, Manfred F.; Huang, Nan

    2013-05-01

    Micro-patterning as an effective bio-modification technique is increasingly used in the development of biomaterials with superior mechanical and biological properties. However, as of now, little is known about the simultaneous regulation of endothelial cells (EC) and smooth muscle cells (SMC) by cardiovascular implants. In this study, a co-culture system of EC and SMC was built on titanium surface by the high molecular weight hyaluronic acid (HMW-HA) micro-pattern. Firstly, the micro-pattern sample with a geometry of 25 μm wide HMW-HA ridges, and 25 μm alkali-activated Ti grooves was prepared by microtransfer molding (μTM) for regulating SMC morphology. Secondly, hyaluronidase was used to decompose high molecular weight hyaluronic acid into low molecular weight hyaluronic acid which could promote EC adhesion. Finally, the morphology of the adherent EC was elongated by the SMC micro-pattern. The surface morphology of the patterned Ti was imaged by SEM. The existence of high molecular weight hyaluronic acid on the modified Ti surface was demonstrated by FTIR. The SMC micro-pattern and EC/SMC co-culture system were characterized by immunofluorescence microscopy. The nitric oxide release test and cell retention calculation were used to evaluate EC function on inhibiting hyperplasia and cell shedding, respectively. The results indicate that EC in EC/SMC co-culture system displayed a higher NO release and cell retention compared with EC cultured alone. It can be suggested that the EC/SMC co-culture system possessed superiority to EC cultured alone in inhibiting hyperplasia and cell shedding at least in a short time of 24 h.

  17. 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.

  18. 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

  19. 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)

    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

  20. Up and Down Expression of Androgen Receptor,Estrogen Receptor beta and Platelet Derived Growth Factor beta by Testosterone in Aortic Vascular Smooth Muscle Tissues

    Institute of Scientific and Technical Information of China (English)

    Wu Saizhu; Lv Hongsong; Zhou Kexiang; Sun Fei; Ma Rui; Zheng Hua; Wei Heming; Rong Zhiyi

    2004-01-01

    Objectives To investigate the effects of testosterone enanthate(TE) on serum lipids and lipoproteins metabolism and the expression of androgen receptor ( AR), estrogen receptor beta ( ER -β) and platelet derived growth factor beta (PDGFR-β ) in aortic vascular smooth muscle tissues(VSMTs). Methods Forty aged male rats were randomly divided into 4 groups, group A (placebo group),group B (2.5 mg/kg intramuscular injection of TE once a week ), group C (5.0 mg/kg intramuscular injection of TE once a week ), group D ( 10.0 mg,/kg intramuscular injection of TE once a week). All animals were fed freely during 16 - week treatment periods. The expression of AR , ER - βand PDGFR - β were studied by Western bolt. Results Average serum LDL - C was lower in group D than that in group A ( p < 0.01 ).Compared with the other groups, average serum TC was also lower in group D ( p < 0.05). AR expression in aortic vascular smooth muscle tissues could be regulated by TE: 99.50 ± 21.74, 125.38 ± 28.68 and 101.98 ±15.42 for TE concentrations at 2.5 mg/kg, 5.0 mg/kgand 10.0 mg/kg, respectively , the expression of ER -β could be regulated by TE: 92.34 ± 18.68, 47.72 ±18.12, 82.13 ±23.50, and the expression of PDGFR -β could be regulated as well by TE: 219.70 ± 45.59,50.16 ± 9.72, 125.36 ± 15.74 ( Data for AR , ER - βand PDGFR - β protein band intensity were expressed with x ± s, with control group taken as 100).Conclusions This study indicates that androgens have significant effects on serum lipids and lipoprotein metabolism. Testosterone enanthate at 5.0 mg/kg can stimulate the expression of AR, but inhibite the expression of PDGFR. Testosterone enanthate at the concentrations of 5.0 mg/kg and 10.0 mg/kg can inhibite the expression of ER - β.

  1. Advanced oxidation protein products induce monocyte chemoattractant protein-1 expression via p38 mitogen-activated protein kinase activation in rat vascular smooth muscle cells

    Institute of Scientific and Technical Information of China (English)

    PENG Kan-fu; WU Xiong-fei; ZHAO Hong-wen; SUN Yan

    2006-01-01

    Background Advanced oxidation protein products (AOPPs) are new uremic toxins reported by Witko-Sarsat in 1996, which are associated with the pathogenesis of atherosclerosis. However, the mechanisms by which AOPPs enhance atherosclerosis have not been fully understood. Monocyte chemoattractant protein-1 (MCP-1) is a chemokine which stimulates migration of monocytes and plays a critical role in the development of atherosclerosis. In this study, we investigated the effect of AOPPs on MCP-1 expression in cultured vascular smooth muscle cells (VSMCs).Methods VSMCs were cultured and then co-incubated with AOPP (200 μ mol/L, 400 μ mol/L) for different times with or without pretreatment with specific p38 mitogen-activated protein kinase (MAPK) inhibitor SB203580. RT-PCR and Western blott were used to detect MCP-1 mRNA and protein expression at different time points after AOPP stimulation in rat smooth muscle cells. Western blot was used to detect the expression of phosphorylated p38 MAPK.Results Treatment of VSMC with AOPPs resulted in a significant increase of the expression of MCP- 1 mRNA and protein in time- and dose-dependent manner, and could activated p38 MAPK. Pretreatment of VSMCs with SB203580 resulted in a dose-dependent inhibition of AOPPs-induced MCP-1 mRNA and protein expression.Conclusions AOPPs can stimulate MCP-1 expression via p38 MAPK in VSMCs. This suggests that AOPPs might contribute to the formation of atherosclerosis through this proinflammatory effect.

  2. Thrombin induced connective tissue growth factor expression in rat vascular smooth muscle cells via the PAR-1/JNK/AP-1 pathway

    Institute of Scientific and Technical Information of China (English)

    Wen-chin KO; Bing-chang CHEN; Ming-jen HSU; Chia-ti TSAI; Chuang-ye HONG; Chien-huang LIN

    2012-01-01

    To investigate the signaling pathways involved in thrombin-induced connective tissue growth factor (CTGF) expression in rat vascular smooth muscle cells (VSMCs).Methods:Experiments were preformed on primary rat aortic smooth muscle cells (RASMCs) and a rat VSMC line (A10).CTGF protein levels were measured using Western blotting.Luciferase reporter genes and dominant negative mutants (DNs) were used to investigate the signaling pathways mediating the induction of CTGF expression by thrombin.Results:Thrombin (0.3-3.0 U/mL) caused a concentration- and time-dependent increase in CTGF expression in both RASMCs and A10 cells.Pretreating A10 cells with the protease-activated receptor 1 (PAR-1) antagonist SCH79797 (0.1 μmol/L) significantly blocked thrombin-induced CTGF expression,while the PAR-4 antagonist tcY-NH2 (30 μmol/L) had no effect.The PAR-1 agonist SFLLRN-NH2 (300 μmol/L) induced CTGF expression,while the PAR-4 agonist GYPGQV-NH2 (300 μmol/L) had no effect.Thrombin (1 U/mL) caused time-dependent phosphorylation of c-Jun N-terminal kinase (JNK).Pretreating with the JNK inhibitor SP600125 (3-30 μmol/L) or transfection with DNs of JNK1/2 significantly attenuated thrombin-induced CTGF expression.Thrombin (0.3-3.0 U/mL) increased activator protein-1 (AP-1)-Iuciferase activity,which was inhibited by the JNK inhibitor SP600125.The AP-1 inhibitor curcumin (1-10 μmol/L) concentration-dependently attenuated thrombin-induced CTGF expression.Conclusion:Thrombin acts on PAR-1 to activate the JNK signaling pathway,which in turn initiates AP-1 activation and ultimately induces CTGF expression in VSMCs.

  3. Blockade of L-type calcium channel in myocardium and calcium-induced contractions of vascular smooth muscle by by CPU 86017

    Institute of Scientific and Technical Information of China (English)

    De-zai DAI; Hui-juan HU; Jing ZHAO; Xue-mei HAO; Dong-mei YANG; Pei-ai ZHOU; Cai-hong WU

    2004-01-01

    AIM: To assess the blockade by CPU 86017 on the L-type calcium channels in the myocardium and on the Ca2+related contractions of vascular smooth muscle. METHODS: The whole-cell patch-clamp was applied to investigate the blocking effect of CPU 86017 on the L-type calcium current in isolated guinea pig myocytes and contractions by KC1 or phenylephrine (Phe) of the isolated rat tail arteries were measured. RESULTS: Suppression of the L-type current of the isolated myocytes by CPU 86017 was moderate, in time- and concentration-dependent manner and with no influence on the activation and inactivation curves. The IC50 was 11.5 μmol/L. Suppressive effect of CPU 86017 on vaso-contractions induced by KC1 100 mmol/L, phenylephrine I μmol/Lin KH solution (phase 1),Ca2+ free KH solution ( phase 2), and by addition of CaCI2 into Ca2+-free KH solution (phase 3) were observed. The IC50 to suppress vaso-contractions by calcium entry via the receptor operated channel (ROC) and Voltage-dependent channel (VDC) was 0.324 μmol/L and 16.3 μmol/L, respectively. The relative potency of CPU 86017 to suppress vascular tone by Ca2+ entry through ROC and VDC is 1/187 of prazosin and 1/37 of verapamil, respectively.CONCLUSION: The blocking effects of CPU 86017 on the L-type calcium channel of myocardium and vessel are moderate and non-selective. CPU 86017 is approximately 50 times more potent in inhibiting ROC than VDC.

  4. 6A3-5/Osa2 is an Early Activated Gene Implicated in the Control of Vascular Smooth Muscle Cell Functions

    Directory of Open Access Journals (Sweden)

    Gwenaele Garin

    2006-01-01

    Full Text Available Vascular smooth muscle cells (VSMC growth plays a key role in the pathophysiology of vascular diseases. However, the molecular mechanisms controlling gene transcription in VSMC remain poorly understood. We previously identified, by differential display, a new gene (6A3-5 overexpressed in proliferating rat VSMC. In this study, we have cloned the full-length cDNA by screening a rat foetal brain cDNA library and investigated its functions. The 6A3-5 protein shows 4 putative conserved functional motifs: a DNA binding domain called ARID (AT-rich interaction domain, two recently described motifs (Osa Homology Domain, and a nuclear localization signal. The deduced protein sequence was observed to be 85% identical to the recently described human Osa2 gene. Immunolabelling, using an anti-6A3-5/Osa2 monoclonal antibody, showed a nuclear localization of the 6A3-5/Osa2 protein. In addition, PDGF upregulated 6A3-5/Osa2 expression at both the transcript and protein levels in a dose and time-dependent fashion. The pattern of upregulation by PDGF was reminiscent of the early responsive gene c-fos. The PDGF-induced upregulation of 6A3-5/Osa2 and proliferation of VSMC were significantly inhibited in a dose and sequence-dependent fashion by an antisense, but not by sense, scrambled or mismatched oligonucleotides directed against 6A3-5/Osa2. In VSMC of aortas derived from hypertensive (LH rats, 6A3-5/Osa2 is overexpressed as compared to that in normotensive (LL rats. The 6A3-5/Osa2-gene expression is downregulated by an ACE inhibitor and upregulated by exogenous AngiotensinII in LH rats. In summary, these results indicate that 6A3-5/Osa2 is an early activated gene that belongs to a new family of proteins involved in the control of VSMC growth.

  5. Alpha1a-Adrenoceptor Genetic Variant Triggers Vascular Smooth Muscle Cell Hyperproliferation and Agonist Induced Hypertrophy via EGFR Transactivation Pathway.

    Directory of Open Access Journals (Sweden)

    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.

  6. Acetylsalicylic acid regulates overexpressed small GTPase RhoA in vascular smooth muscle cells through prevention of new synthesis and enhancement of protein degradation.

    Science.gov (United States)

    Li, Dong-Bo; Fu, Zhi-Xuan; Ruan, Shu-Qin; Hu, Shen-Jiang; Li, Xia

    2012-04-01

    RhoA has been shown to play a major role in vascular processes and acetylsalicylic acid (aspirin) is known to exert a cytoprotective effect via multiple mechanisms. In the present study, we aimed at investigating the effect of aspirin on RhoA expression under a stress state in rat VSMCs (vascular smooth muscle cells) and the underlying mechanisms. The expression of iNOS (inducible nitric oxide synthase) and iNOS activity as well as NO concentration was significantly promoted by LPS (lipopolysaccharide) accompanying the elevation of RhoA expression, which was blocked by the addition of the iNOS inhibitor L-NIL [L-N6-(1-iminoethyl)lysine dihydrochloride]. Aspirin (30 μM) significantly attenuated the elevation of RhoA, while indomethacin and salicylate had no similar effect. The sGC (soluble guanylate cyclase) inhibitor ODQ (1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one) showed the same effect as aspirin in down-regulating RhoA but was reversed by the addition of the cGMP analogue 8-Br-PET-cGMP (β-phenyl-1,N2-ethano-8-bromoguanosine 3',5'-cyclic monophosphorothioate). 8-Br-PET-cGMP solely enhanced the RhoA expression that was abrogated by preincubation with aspirin. Degradation analysis indicated that aspirin enhanced the protein degradation rate of RhoA and GDP-bound RhoA seemed to be more susceptible to aspirin-enhanced degradation compared with the GTP-bound form. Our results indicate that aspirin attenuates the LPS-induced overexpression of RhoA both by inhibiting new synthesis and accelerating protein degradation, which may help elucidate the multiple beneficial effects of aspirin.

  7. Inhibition of Vascular Smooth Muscle Growth via Signaling Crosstalk between AMP-Activated Protein Kinase and cAMP-Dependent Protein Kinase

    Directory of Open Access Journals (Sweden)

    Joshua Daniel Stone

    2012-10-01

    Full Text Available Abnormal vascular smooth muscle (VSM growth is central in the pathophysiology of vascular disease yet fully effective therapies to curb this growth are lacking. Recent findings from our lab and others support growth control of VSM by adenosine monophosphate (AMP-based approaches including the metabolic sensor AMP-activated protein kinase (AMPK and cAMP-dependent protein kinase (PKA. Molecular crosstalk between AMPK and PKA has been previously suggested, yet the extent to which this occurs and its biological significance in VSM remains unclear. Considering their common AMP backbone and similar signaling characteristics, we hypothesized that crosstalk exists between AMPK and PKA in the regulation of VSM growth. Using rat primary VSM cells, the AMPK agonist AICAR increased AMPK activity and phosphorylation of the catalytic Thr172 site on AMPK. Interestingly, AICAR also phosphorylated a suspected PKA-inhibitory Ser485 site on AMPK, and these cumulative events were reversed by the PKA inhibitor PKI suggesting possible PKA-mediated regulation of AMPK. AICAR also increased PKA activity in a reversible fashion. The cAMP stimulator forskolin increased PKA activity and completely ameliorated Ser/Thr protein phosphatase-2C activity, suggesting a potential mechanism of AMPK modulation by PKA since inhibition of PKA by PKI reduced AMPK activity. Functionally, AMPK inhibited serum-stimulated cell cycle progression and cellular proliferation; however, PKA failed to do so. Moreover, AMPK and PKA reduced PDGF-β-stimulated VSM cell migration. Collectively, these results show that AMPK is capable of reducing VSM growth in both anti-proliferative and anti-migratory fashions. Furthermore, these data suggest that AMPK may be modulated by PKA and that positive feedback may exist between these two systems. These findings reveal a discrete nexus between AMPK and PKA in VSM and provide basis for metabolically-directed targets in reducing pathologic VSM growth.

  8. Anti-proliferative actions of 2-decylamino-5,8-dimethoxy-1,4-naphthoquinone in vascular smooth muscle cells

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Jung-Jin [Department of Pharmacology, Chungnam National University College of Pharmacy, Daejeon 305-764 (Korea, Republic of); Institute of Drug Research and Development, Chungnam National University, Daejeon 305-764 (Korea, Republic of); Zhang, Wei-Yun; Yi, Hyoseok; Kim, Yohan; Kim, In-Su [Department of Pharmacology, Chungnam National University College of Pharmacy, Daejeon 305-764 (Korea, Republic of); Shen, Gui-Nan; Song, Gyu-Yong [Department of Medicinal Chemistry, Chungnam National University College of Pharmacy, Daejeon 305-764 (Korea, Republic of); Myung, Chang-Seon, E-mail: cm8r@cnu.ac.kr [Department of Pharmacology, Chungnam National University College of Pharmacy, Daejeon 305-764 (Korea, Republic of); Institute of Drug Research and Development, Chungnam National University, Daejeon 305-764 (Korea, Republic of)

    2011-07-22

    Highlights: {yields} 2-Decylamino-DMNQ inhibited PDGF-BB-induced VSMC proliferation in a dose-dependent manner with no apparent cytotoxicity. {yields} 2-Decylamino-DMNQ inhibited PDGF-BB-induced phosphorylation of Erk1/2 and PLC{gamma}1. {yields} 2-Decylamino-DMNQ arrested a G{sub 0}/G{sub 1} cell cycle progression in association with pRb phosphorylation and PCNA expression. {yields} Both U0126, an Erk inhibitor, and U73122, a PLC{gamma} inhibitor, arrested a G{sub 0}/G{sub 1} phase of the cell cycle. -- Abstract: Naphthoquinone derivatives have been reported to possess various pharmacological activities, such as antiplatelet, anticancer, antifungal, and antiviral properties. In this study, we investigated the effects of a newly-synthesized naphthoquinone derivative, 2-decylamino-5,8-dimethoxy-1,4-naphthoquinone (2-decylamino-DMNQ), on VSMC proliferation and examined the molecular basis of the underlying mechanism. In a dose-dependent manner, 2-decylamino-DMNQ inhibited PDGF-stimulated VSMC proliferation with no apparent cytotoxic effect. While 2-decylamino-DMNQ did not affect PDGF-R{beta} or Akt, it did inhibit the phosphorylation of Erk1/2 and PLC{gamma}1 induced by PDGF. Moreover, 2-decylamino-DMNQ suppressed DNA synthesis through the arrest of cell cycle progression at the G{sub 0}/G{sub 1} phase, including the suppression of pRb phosphorylation and a decrease in PCNA expression, which was related to the downregulation of cell cycle regulatory factors, such as cyclin D1/E and CDK 2/4. It was demonstrated that both U0126, an Erk1/2 inhibitor, and U73122, a PLC{gamma} inhibitor, increased the proportion of cells in the G{sub 0}/G{sub 1} phase of the cell cycle. Thus, these results suggest that 2-decylamino DMNQ has an inhibitory effect on PDGF-induced VSMC proliferation and the mechanism of this action is through cell cycle arrest at the G{sub 0}/G{sub 1} phase. This may be a useful tool for studying interventions for vascular restenosis in coronary

  9. SMOOTH MUSCLE STEM CELLS

    Science.gov (United States)

    Vascular smooth muscle cells (SMCs) originate from multiple types of progenitor cells. In the embryo, the most well-studied SMC progenitor is the cardiac neural crest stem cell. Smooth muscle differentiation in the neural crest lineage is controlled by a combination of cell intrinsic factors, includ...

  10. Relaxation of rabbit corpus cavernosum smooth muscle and aortic vascular endothelium induced by new nitric oxide donor substances of the nitrosyl-ruthenium complex

    Directory of Open Access Journals (Sweden)

    Joao B. G. Cerqueira

    2008-10-01

    Full Text Available INTRODUCTION: Endothelial dysfunction characterized by endogenous nitric oxide (NO deficiency made 56% of patients affected with erectile dysfunction decline treatment with PDE-5 inhibitors. New forms of treatment are currently being developed for this group of patients. MATERIALS AND METHODS: The study compared the effect of sodium nitroprusside (SNP and two substances of the nitrosyl-ruthenium complex, cis-[Ru(bpy2(SO3(NO]PF-6-9 ("FONO1” and trans-[Ru(NH34(caffeine(NO]C13 ("LLNO1” on relaxation of rabbit corpus cavernosum smooth muscle and aortic vascular endothelium. The samples were immersed in isolated baths and precontracted with 0.1 µM phenylephrine (PE and the corresponding relaxation concentration/response curves were plotted. In order to investigate the relaxation mechanisms involved, 100 µM ODQ (a soluble guanylate cyclase-specific inhibitor, 3 µM or 10 µM oxyhemoglobin (an extracellular NO scavenger or 1 mM L-cysteine (a nitrosyl anion-specific scavenger was added to the samples. RESULTS: All the NO donors tested produced a significant level of relaxation in the vascular endothelium. In corpus cavernosum samples, FONO1 produced no significant effect, but LLNO1 and SNP induced dose-dependent relaxation with comparable potency (pEC50 = 6.14 ± 0.08 and 6.4 ± 0.14, respectively and maximum effect (Emax = 82% vs. 100%, respectively. All NO donors were found to activate soluble guanylate cyclase, since the addition of the corresponding inhibitor (100 µM ODQ completely neutralized the relaxation effect observed. The addition of oxyhemoglobin reduced the relaxation effect, but did not inhibit it completely. In aortic vascular endothelium 3 µM oxyhemoglobin decreased the relaxation effect by 26% on the average, while 10 µM oxyhemoglobin reduced it by over 52%. The addition of 100 µM L-cysteine produced no significant inhibiting effect. CONCLUSIONS: These results suggest that LLNO1 and FONO1 are potent vasodilators. LLNO1 was

  11. Effects of serotonin on expression of the LDL receptor family member LR11 and 7-ketocholesterol-induced apoptosis in human vascular smooth muscle cells

    Energy Technology Data Exchange (ETDEWEB)

    Nagayama, Daiji; Ishihara, Noriko [Center of Diabetes, Endocrinology and Metabolism, Toho University, Sakura Medical Center, 564-1, Shimoshizu, Sakura-City, Chiba 285-8741 (Japan); Bujo, Hideaki [Department of Clinical Laboratory Medicine, Toho University, Sakura Medical Center, 564-1, Shimoshizu, Sakura-City, Chiba 285-8741 (Japan); Shirai, Kohji [Department of Vascular Function, Toho University, Sakura Medical Center, 564-1, Shimoshizu, Sakura-City, Chiba 285-8741 (Japan); Tatsuno, Ichiro, E-mail: ichiro.tatsuno@med.toho-u.ac.jp [Center of Diabetes, Endocrinology and Metabolism, Toho University, Sakura Medical Center, 564-1, Shimoshizu, Sakura-City, Chiba 285-8741 (Japan)

    2014-04-18

    Highlights: • The dedifferentiation of VSMCs in arterial intima is involved in atherosclerosis. • 5-HT showed proliferative effect on VSMCs which was abolished by sarpogrelate. • 5-HT enhanced expression of LR11 mRNA in VSMCs which was abolished by sarpogrelate. • 5-HT suppressed 7KCHO-induced apoptosis of VSMCs via caspase-3/7-dependent pathway. • The mechanisms explain the 5-HT-induced remodeling of arterial structure. - Abstract: Serotonin (5-HT) is a known mitogen for vascular smooth muscle cells (VSMCs). The dedifferentiation and proliferation/apoptosis of VSMCs in the arterial intima represent one of the atherosclerotic changes. LR11, a member of low-density lipoprotein receptor family, may contribute to the proliferation of VSMCs in neointimal hyperplasia. We conducted an in vitro study to investigate whether 5-HT is involved in LR11 expression in human VSMCs and apoptosis of VSMCs induced by 7-ketocholesterol (7KCHO), an oxysterol that destabilizes plaque. 5-HT enhanced the proliferation of VSMCs, and this effect was abolished by sarpogrelate, a selective 5-HT2A receptor antagonist. Sarpogrelate also inhibited the 5-HT-enhanced LR11 mRNA expression in VSMCs. Furthermore, 5-HT suppressed the 7KCHO-induced apoptosis of VSMCs via caspase-3/7-dependent pathway. These findings provide new insights on the changes in the differentiation stage of VSMCs mediated by 5-HT.

  12. Alteration of Mevalonate Pathway in Proliferated Vascular Smooth Muscle from Diabetic Mice: Possible Role in High-Glucose-Induced Atherogenic Process

    Directory of Open Access Journals (Sweden)

    Guo-Ping Chen

    2015-01-01

    Full Text Available The proliferation of vascular smooth muscle cells (VSMCs is one of the main features of atherosclerosis induced by high glucose. Mevalonate pathway is an important metabolic pathway that plays a key role in multiple cellular processes. The aim of this study was to define whether the enzyme expression in mevalonate pathway is changed in proliferated VSMCs during atherogenic process in diabetic mice. Diabetes was induced in BALB/c mice with streptozotocin (STZ, 50 mg/kg/day for 5 days. Induction of diabetes with STZ was associated with an increase of lesion area and media thickness after 8 and 16 weeks of diabetes. In aorta, there were overexpressions of some enzymes, including 3-hydroxy-3-methylglutaryl-coenzyme A reductase (HMGR, farnesyl pyrophosphate synthase (FPPS, geranylgeranyl pyrophosphate synthase (GGPPS, farnesyltransferase (FNT, and geranylgeranyltransferase-1 (GGT-1, and unchanged expression of squalene synthase (SQS and phosphor-3-hydroxy-3-methylglutaryl-coenzyme A reductase (P-HMGR in 8 and 16 weeks of diabetes. In vitro, VSMCs were cultured and treated with different glucose concentrations for 48 h. High glucose (22.2 mM induced VSMC proliferation and upregulation of HMGR, FPPS, GGPPS, FNT, and GGT-1 but did not change the expressions of SQS and P-HMGR. In conclusion, altered expression of several key enzymes in the mevalonate pathway may play a potential pathophysiological role in atherogenic process of diabetes macrovascular complication.

  13. Effects of serotonin on expression of the LDL receptor family member LR11 and 7-ketocholesterol-induced apoptosis in human vascular smooth muscle cells

    International Nuclear Information System (INIS)

    Highlights: • The dedifferentiation of VSMCs in arterial intima is involved in atherosclerosis. • 5-HT showed proliferative effect on VSMCs which was abolished by sarpogrelate. • 5-HT enhanced expression of LR11 mRNA in VSMCs which was abolished by sarpogrelate. • 5-HT suppressed 7KCHO-induced apoptosis of VSMCs via caspase-3/7-dependent pathway. • The mechanisms explain the 5-HT-induced remodeling of arterial structure. - Abstract: Serotonin (5-HT) is a known mitogen for vascular smooth muscle cells (VSMCs). The dedifferentiation and proliferation/apoptosis of VSMCs in the arterial intima represent one of the atherosclerotic changes. LR11, a member of low-density lipoprotein receptor family, may contribute to the proliferation of VSMCs in neointimal hyperplasia. We conducted an in vitro study to investigate whether 5-HT is involved in LR11 expression in human VSMCs and apoptosis of VSMCs induced by 7-ketocholesterol (7KCHO), an oxysterol that destabilizes plaque. 5-HT enhanced the proliferation of VSMCs, and this effect was abolished by sarpogrelate, a selective 5-HT2A receptor antagonist. Sarpogrelate also inhibited the 5-HT-enhanced LR11 mRNA expression in VSMCs. Furthermore, 5-HT suppressed the 7KCHO-induced apoptosis of VSMCs via caspase-3/7-dependent pathway. These findings provide new insights on the changes in the differentiation stage of VSMCs mediated by 5-HT

  14. 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.

  15. 2D and 3D collagen and fibrin biopolymers promote specific ECM and integrin gene expression by vascular smooth muscle cells

    Science.gov (United States)

    HONG, HELEN; STEGEMANN, JAN P.

    2009-01-01

    Collagen Type I and fibrin are polymeric proteins commonly used in the field of regenerative medicine as the foundational matrix of engineered tissues. We examined the response of vascular smooth muscle cells (VSMC) to both two-dimensional (2D) substrates as well as three-dimensional (3D) matrices of these biopolymers. Pure collagen Type I, pure fibrin and composite matrices consisting of 1:1 mixtures of collagen and fibrin were studied. Relative gene expression of three ECM molecules (collagen Type I and III, and tropoelastin) and three integrin subunits (integrins α1, β1 and β3) was determined over 7 days in culture using quantitative RT-PCR. Expression of all of these marker genes was up-regulated in 3D matrices, relative to 2D substrates. Tropoelastin, integrin α1 and integrin β1 were highest in collagen matrices, while collagen III and integrin β3 expression were highest in pure fibrin, and collagen I expression was highest in the collagen-fibrin composite materials. Both the compositional and temporal expression patterns of these specific ECM-related genes were suggestive of a wound healing response. These results illuminate the short-term responses of VSMC to 2D and 3D biopolymer matrices, and have relevance to tissue engineering and cardiovascular biology. PMID:18854122

  16. Heat shock protein 60 stimulates the migration of vascular smooth muscle cells via Toll-like receptor 4 and ERK MAPK activation

    Science.gov (United States)

    Zhao, Ying; Zhang, Chenxu; Wei, Xuge; Li, Pei; Cui, Ying; Qin, Yuanhua; Wei, Xiaoqing; Jin, Minli; Kohama, Kazuhiro; Gao, Ying

    2015-01-01

    Accumulating evidence indicates that heat shock protein (HSP) 60 is strongly associated with the pathology of atherosclerosis (AS). However, the precise mechanisms by which HSP60 promotes atherosclerosis remain unclear. In the present study, we found that HSP60 mRNA and protein expression levels in the thoracic aorta are enhanced not only in a mouse model of AS but also in high-fat diet (HFD) mice. HSP60 expression and secretion was activated by platelet-derived growth factor-BB (PDGF-BB) and interleukin (IL)-8 in both human umbilical vein endothelial cells (HUVECs) and vascular smooth muscle cells (VSMCs). HSP60 was found to induce VSMC migration, and exposure to HSP60 activated ERK MAPK signaling. U0126, an inhibitor of ERK, reduced VSMC migration. The HSP60-stimulated VSMCs were found to express TLR4 mRNA but not TLR2 mRNA. Knockdown of TLR4 by siRNA reduced HSP60-induced VSMC migration and HSP60-induced ERK activation. Finally, HSP60 induced IL-8 secretion in VSMCs. Together these results suggest that HSP60 is involved in the stimulation of VSMC migration, via TLR4 and ERK MAPK activation. Meanwhile, activation of HSP60 is one of the most powerful methods of sending a ‘danger signal’ to the immune system to generate IL-8, which assists in the management of an infection or disease. PMID:26477505

  17. Angiotensin Ⅱ stimulates phosphorylation of 4E-binding protein 1 and p70 S6 kinase in cultured vascular smooth muscle cells

    Institute of Scientific and Technical Information of China (English)

    Na LI; Ke-gui WU; Xiang-yu WANG; Liang-di XIE; Chang-sheng XU; Hua-jun WANG

    2004-01-01

    AIM: To examine the regulatory effects of angiotensin Ⅱ (Ang Ⅱ) on the phosphorylation of 4E-binding protein 1 (4E-BP1) and p70 S6 kinase in cultured vascular smooth muscle cells (VSMC), and the contribution of phosphatidylinositol 3-kinase (PI3K)/protein kinase B (PKB) signaling pathway in this process. METHODS: VSMC obtained from rat thoracic aortas were cultured. The phosphorylation of 4E-BP1 and p70 S6 kinase was detected by immunoblotting. RESULTS: Ang Ⅱ significantly increased the phosphorylation of 4E-BP1 and p70 S6 kinase,with the peaks occurring at, respectively, 10 min and 30 min, after stimulation with Ang Ⅱ. The stimulatory effect of Ang Ⅱ on 4E-BP1 and p70 S6 kinase phosphorylation was abrogated by Ang Ⅱ type 1 receptor (AT1 receptor)antagonist losartan, and suppressed by PI3K inhibitor LY294002 in a concentration-dependent manner.CONCLUSION: Ang Ⅱ treatment of VSMC induces the phosphorylation of 4E-BP1 and p70 S6 kinase via AT1 receptor, and PI3K signaling pathway is involved in this process.

  18. Mechanical trapping of the nucleus on micropillared surfaces inhibits the proliferation of vascular smooth muscle cells but not cervical cancer HeLa cells.

    Science.gov (United States)

    Nagayama, Kazuaki; Hamaji, Yumi; Sato, Yuji; Matsumoto, Takeo

    2015-07-16

    The interaction between cells and the extracellular matrix on a topographically patterned surface can result in changes in cell shape and many cellular functions. In the present study, we demonstrated the mechanical deformation and trapping of the intracellular nucleus using polydimethylsiloxane (PDMS)-based microfabricated substrates with an array of micropillars. We investigated the differential effects of nuclear deformation on the proliferation of healthy vascular smooth muscle cells (SMCs) and cervical cancer HeLa cells. Both types of cell spread normally in the space between micropillars and completely invaded the extracellular microstructures, including parts of their cytoplasm and their nuclei. We found that the proliferation of SMCs but not HeLa cells was dramatically inhibited by cultivation on the micropillar substrates, even though remarkable deformation of nuclei was observed in both types of cells. Mechanical testing with an atomic force microscope and a detailed image analysis with confocal microscopy revealed that SMC nuclei had a thicker nuclear lamina and greater expression of lamin A/C than those of HeLa cells, which consequently increased the elastic modulus of the SMC nuclei and their nuclear mechanical resistance against extracellular microstructures. These results indicate that the inhibition of cell proliferation resulted from deformation of the mature lamin structures, which might be exposed to higher internal stress during nuclear deformation. This nuclear stress-induced inhibition of cell proliferation occurred rarely in cancer cells with deformable nuclei. PMID:26054426

  19. Surface modification of poly-L-lactic acid films by electrostatic self-assembly to promote vascular smooth muscle cells growth

    Institute of Scientific and Technical Information of China (English)

    XU Li; HU Kun; JIAO Yanpeng; CUI Fuzhai; AI Hongbin

    2007-01-01

    The intention of this study was to surface modify the poly-L-1actic acid(PLLA)film and evaluate the effects of the surfaces on the growth of vascular smooth muscle eells (VSMCs)in vitro.Collagen and hyaluronic acid(HA) were utilized as polycation and polyanion in this study.Layerby-1ayer(LBL)self-assembly technique was used to lead to the formation of multilayer moleculer on the poly-L-lactic acid(PLLA)film surfaces.Collagen/HA layers was overcasted coating on the PLLA surface after the activation layers by poly-(ethyleneimine)(PEI).The structure and morphology of the multilayer molecular were examined by attenuated total reflectance Fourier transform infrared (ATR-FTIR) spectrophotometer and atomic force microscope (AFM),respectively.The ATR-FTIR analysis illuminated the presence of collagen on the PLLA surface.The AFM results showed the multilayer appeared on PLLA surface.The VSMCs were adopted to evaluate me cyto-compatibility of the modified PLLA films.It was found that the viability of VSMCs on the modified PLLA films were greater than that on original PLLA films and tissue culture plastic after ten days culture (p<0.05).Scanning electron microscopy(SEM) and laser scanning confocal microscopy (LSCM) data also confirmed the homogeneous results.These data suggest that collagen/HA coat Can be successfully adopted in the surface modification of PLLA film through LBL technique,and also can enhance its eell compatibility.

  20. Expression of Inositol 1,4, 5-trisphosphate Receptor mRNA in Myocardium of Spontaneous Hypertension Rats and Cultured Vascular Smooth Muscle Cells of Rats

    Institute of Scientific and Technical Information of China (English)

    刘乃丰; 张寄南; 耿茜; 杨笛; 董莉; 马文珠

    2002-01-01

    Objective To investigate expression of inositol 1,4,5-trisphosphate receptor ( IP3R) mRNA on sacroplasmic reticular in myocardium of spontaneous hypertension rats ( SHRs) and cultured vascular smooth muscle cells (VSMC) of rats and effects of perindopril and urapidil on them. Methods SHRs were orally given perindopril (1. 0 mg@ kg-1 @ d-1) or urapidil (15 mg@kg-1 @ d-1) for 24 weeks, respectively. Expression of IP3R mRNA was examined by semi-quantitatwe reverse transcription polymers chain reaction ( RT-PCR ) using three oligonuclotide primers for each subtype of IP3R with β-actin as internal label. Results All subtypes of IP3R were expressed in myocardium of SHR, WKY and cultured VSMC. Expression of IPsR mRNA in left ventricle of SHR was markedly enhanced. Urapidil could down-regulate expression of IP3R- I and IP3R- iⅢ , perindopril slightly increased expression of IP3R- Ⅱ and decreased expression of IP3R- I and IP3R- Ⅲ in myocardium of SHR. Conclusion Our results suggest that expression of IP3R mRNA in cardiovascular system could be regulated by urapidil and perindopril.

  1. Inhibitory effect of puerarin on vascular smooth muscle cells proliferation induced by oxidised low-density lipoprotein via suppressing ERK 1/2 phosphorylation and PCNA expression.

    Science.gov (United States)

    Hu, Yanwu; Liu, Kai; Bo, Sun; Yan, Mengtong; Zhang, Yang; Miao, Chunsheng; Ren, Liqun

    2016-02-01

    Puerarin, an isoflavonoid isolated from the traditional Chinese herbal medicine Pueraria lobata (Wild.) Ohwi, has been shown to process antioxidant, anti-inflammatory, anti-cancer, anti-hypercholesterolemic, and anti-hyperglycemic activities in vivo and in vitro. The aim of the present study was to investigate the antiproliferative effects and the possible mechanisms of puerarin in vascular smooth muscle cells (VSMCs) stimulated with oxidised low-density lipoprotein (ox-LDL). VSMCs were cultured and pretreated with different concentrations of puerarin (0, 1, 10, 50 µM) before stimulated by ox-LDL (50 µg/mL). Cell proliferation was evaluated by MTT assay. Flow cytometry was used to study the influence of puerarin on cell cycle. Proliferating cell nuclear antigen (PCNA) expression and phosphorylation levels of extracellular signal-regulated kinase (ERK) 1/2 were detected by western blotting analysis. The results indicated that puerarin significantly inhibited VSMCs proliferation induced by ox-LDL and phosphorylation of ERK 1/2. Furthermore, puerarin also blocked the ox-LDL-induced cell-cycle progression at G1/S-interphase and down-regulated the expression of PCNA of VSMCs. The results suggest puerarin inhibits ox-LDL-induced proliferation of VSMCs by suppressing ERK 1/2 phosphorylation and PCNA expression.

  2. Phorbol ester promotes a sustained down-regulation of endothelin receptors and cellular responses to endothelin in human vascular smooth muscle cells.

    Science.gov (United States)

    Resink, T J; Scott-Burden, T; Weber, E; Bühler, F R

    1990-02-14

    The effect of phorbol ester pretreatment of human vascular smooth muscle cells (hVSMC) was studied with respect to regulation of endothelin (ET)-receptor binding and cellular responses to ET. The capacity of hVSMC to bind ET was decreased (by approximately 50% at maximum) after phorbol exposure, and this reductive effect was both rapid (t 1/2 approximately 10 min.) and sustained (for up to 24 hrs. of chronic phorbol exposure). Phorbol pretreatment inhibited both inositol phosphate and diacylclycerol production responses of hVSMC to ET in a manner that was time-dependent and sustained. Phorbol pretreatment also produced a persistent reduction in the ability of ET to release isotopically-labelled arachidonic and/or its metabolites from hVSMC, but importantly ionomycin-stimulated release was similarly negatively affected. Furthermore, ET-induced accumulation of the phospholipase A2/phospholipase B-derived inositol phospholipid metabolite, glycerophosphoinositol, was not different between control and phorbol-treated hVMSC. The mechanism whereby phorbol exerts differential, but notably sustained inhibitory effects on ET-promoted signal transduction pathways are thus complex and illustrative of the selectivity of protein kinase C in regulating cellular responses. PMID:2154974

  3. Selective biological response of human pulmonary microvascular endothelial cells and human pulmonary artery smooth muscle cells on cold-plasma-modified polyester vascular prostheses

    International Nuclear Information System (INIS)

    The aim of this work was to improve the hemocompatibility and the selectivity according to cells of non-woven poly(ethylene terephthalate) (PET) membranes. Non-woven PET membranes were modified by a combined plasma-chemical process. The surface of these materials was pre-activated by cold-plasma treatment and poly(acrylic acid) (PAA) was grafted by the in situ free radical polymerization of acrylic acid (AA). The extent of this reaction and the number of carboxylic groups incorporated were evaluated by colorimetric titration using toluidine blue O. All samples were characterized by SEM, AFM and thermogravimetric analysis, and the mechanical properties of the PAA grafted sample were determined. A selective cell response was observed when human pulmonary artery smooth muscle cells (HPASMC) or human pulmonary micro vascular endothelial cells (HPMEC) were seeded on the modified surfaces. HPASMC proliferation decreased about 60%, while HPMEC proliferation was just reduced about 10%. PAA grafted samples did not present hemolytic activity and the platelet adhesion decreased about 28% on PAA grafted surfaces.

  4. Enhanced Levels of microRNA-125b in Vascular Smooth Muscle Cells of Diabetic db/db Mice Lead to Increased Inflammatory Gene Expression by Targeting the Histone Methyltransferase Suv39h1

    OpenAIRE

    Villeneuve, Louisa M.; KATO, MITSUO; Reddy, Marpadga A.; Wang, Mei; Lanting, Linda; Natarajan, Rama

    2010-01-01

    OBJECTIVE Diabetes remains a major risk factor for vascular complications that seem to persist even after achieving glycemic control, possibly due to “metabolic memory.” Using cultured vascular smooth muscle cells (MVSMC) from type 2 diabetic db/db mice, we recently showed that decreased promoter occupancy of the chromatin histone H3 lysine-9 methyltransferase Suv39h1 and the associated repressive epigenetic mark histone H3 lysine-9 trimethylation (H3K9me3) play key roles in sustained inflamm...

  5. C-TYPE NATRIURETIC PEPTIDE INHIBITS UPR EGULATION OF α1-ADRENOCEPTOR AND INOSITO L 1,4,5-TRISPHOSPHATE RECEPTOR IN RAT VASCULAR SMOOTH MUSCLE AFTER VASCULAR ENDOTHELIAL INJURY

    Institute of Scientific and Technical Information of China (English)

    2000-01-01

    Objective.In a model o f balloon injury of rat aortic endotheli um, the effects of C-type natriuretic pe ptide(CNP) on α1-adrenoreceptor and ino sitol 1,4,5-triphosphate (IP3) receptor were studied Methods. Aortic injuri es were produced by vascular endothelium -denudation.α1- adrenoreceptor in smoot h muscle sarcolemma and IP3 receptor in smooth muscle sarcoplasmic reticulum in the rat aorta were assayed by radioactiv e analysis method.Results. It was found that neointima was formed and the conten ts of DNA, collagen and elastin of each int ima-media were significantly increased i n 7 days and 21 days after balloon injury of rat aorta. α1-adrenoreceptor in smo oth muscle sarcolemma and IP3 receptor in sarcoplasmic reticulum were also upre gul ated. Results also showed that the admin i stration of CNP i.p significantly decrea sed the contents of DNA, collagen and el as tin of each intima-media, and inhibited the up-regulation of α1-adrenoreceptor and IP3 receptor.Conclusion. The inhibition of the up-regulation of α 1-adrenoreceptor and IP3 receptor by C NP might be one of the mechanisms of its suppressive action on intimal proliferation.

  6. Coexpression of voltage-dependent calcium channels Cav1.2, 2.1a, and 2.1b in vascular myocytes

    DEFF Research Database (Denmark)

    Andreasen, Ditte; Friis, Ulla Glenert; Uhrenholt, Torben Rene;

    2006-01-01

    Voltage-dependent Ca2+ channels Cav1.2 (L type) and Cav2.1 (P/Q type) are expressed in vascular smooth muscle cells (VSMCs) and are important for the contraction of renal resistance vessels. In the present study we examined whether native renal VSMCs coexpress L-, P-, and Q-type Ca2+ currents. The...... microscopy revealed expression of both channels in all of the smooth muscle cells. Whole-cell patch clamp on single preglomerular VSMCs from mice showed L-, P-, and Q-type currents. Blockade of the L-type currents by calciseptine (20 nmol/L) inhibited 35.6+/-3.9% of the voltage-dependent Ca2+ current, and...... blocking P-type currents (omega-agatoxin IVA 10 nmol/L) led to 20.2+/-3.0% inhibition, whereas 300 nmol/L of omega agatoxin IVA (blocking P/Q-type) inhibited 45.0+/-7.3%. In rat aortic smooth muscle cells (A7r5), blockade of L-type channels resulted in 28.5+/-6.1% inhibition, simultaneous blockade of L...

  7. Localization of adenylyl cyclase isoforms and G protein-coupled receptors in vascular smooth muscle cells: expression in caveolin-rich and noncaveolin domains.

    Science.gov (United States)

    Ostrom, Rennolds S; Liu, Xiaoqiu; Head, Brian P; Gregorian, Caroline; Seasholtz, Tammy M; Insel, Paul A

    2002-11-01

    A number of different agonists activate G protein-coupled receptors to stimulate adenylyl cyclase (AC), increase cAMP formation, and promote relaxation in vascular smooth muscle. To more fully understand this stimulation of AC, we assessed the expression, regulation, and compartmentation of AC isoforms in rat aortic smooth muscle cells (RASMC). Reverse transcription-polymerase chain reaction detected expression of AC3, AC5, and AC6 mRNA, whereas immunoblot analysis indicated expression of AC3 and AC5/6 protein primarily in caveolin-rich membrane (cav) fractions relative to noncaveolin (noncav) fractions. Beta(1)-adrenergic receptors (AR), beta(2)AR, and G(s) were detected in both cav and noncav fractions, whereas the prostanoid receptors EP(2)R and EP(4)R were excluded from cav fractions. We used an adenoviral construct to increase AC6 expression. Overexpressed AC6 localized only in noncav fractions. Two-fold overexpression of AC6 caused enhancement of forskolin-, isoproterenol- and prostaglandin E(2)-stimulated cAMP formation but no changes in basal levels of cAMP. At higher levels of AC6 overexpression, basal and adenosine receptor-stimulated cAMP levels were increased. Stimulation of cAMP levels by agents that increase Ca(2+) in native cells was consistent with the expression of AC3, but overexpression of AC6, which is inhibited by Ca(2+), blunted the Ca(2+)-stimulable cAMP response. These data indicate that: 1) RASMC express multiple AC isoforms that localize in both caveolin-rich and noncaveolin domains, 2) expression of AC6 in non-caveolin-rich membranes can increase basal levels of cAMP and response to several stimulatory agonists, and 3) Ca(2+)-mediated regulation of cAMP formation depends upon expression of different AC isoforms in RASMC. Compartmentation of GPCRs and AC is different in cardiomyocytes than in RASMC, indicating that targeting of these components to caveolin-rich membranes can be cell-specific. Moreover, our results imply that the

  8. Up-regulation of Fas Ligand Expression by Sirtuin 1 in both Flow-restricted Vessels and Serum-stimulated Vascular Smooth Muscle Cells

    Institute of Scientific and Technical Information of China (English)

    Li Li; Peng Gao; Hou-zao Chen; Zhu-qin Zhang; Ting-ting Xu; Yu-yan Jia; Hui-na Zhang; Guan-hua Du; De-pei Liu

    2013-01-01

    Objective To study the role of sirtuin 1 (SIRT1) in Fas ligand (FasL) expression regulation duringvascular lesion formation and to elucidate the potential mechanisms.Methods SIRT1 and FasL protein levels were detected byWestern blotting in either mouse arteries extract or the whole rat aortic vascular smooth muscle cell (VSMC) lysate. Smooth muscle cell (SMC)-specific human SIRT1 transgenic (Tg) C57BL/6 mice and their littermate wild-type (WT) controlsunderwent complete carotid artery ligation (ligation groups) or the ligation-excluded operation (sham groups). The carotid arteries were collected 1 day after operation. Reverse transcription-polymerase chain reaction was performed to detect the mRNA levels of SIRT1 and FasL. Luciferase reporter assays wereperformed todetect the effect of WT-SIRT1, a dominant-negative form of SIRT1 (SIRT1H363Y), andGATA-6 on the promoter activity of FasL. Flow cytometry assay was applied to measure the hypodiploidDNA content of VSMC so as to monitor cellular apoptosis.Results SIRT1 was expressed in both rat aortic VSMCs and mouse arteries. Forced SIRT1 expressionincreased FasL expression both in injured mouse carotid arteries 1 day after ligation (P<0.001) and VSMCs treated with serum (P<0.05 at the transcriptional level, P<0.001 at the protein level). No notable apoptosis was observed. Furthermore, transcription factor GATA-6 increased the promoter activity of FasL (P<0.001).The induction of FasL promoter activity by GATA-6 was enhanced by WT-SIRT1 (P<0.001), whileSIRT1H363Y significantly relieved the enhancing effect of WT-SIRT1 on GATA-6 (P<0.001).ConclusionsOverexpression of SIRT1 up-regulates FasL expression in both flow-restricted mousecarotid arteries and serum-stimulated VSMCs. The transcription factor GATA-6 participates in thetranscriptional regulation of FasL expression by SIRT1.

  9. The P2Y2 receptor mediates uptake of matrix-retained and aggregated low density lipoprotein in primary vascular smooth muscle cells

    Science.gov (United States)

    Dissmore, Tixieanna; Seye, Cheikh I.; Medeiros, Denis M.; Weisman, Gary A.; Bardford, Barry; Mamedova, Laman

    2016-01-01

    Background and aims The internalization of aggregated low-density lipoproteins (agLDL) mediated by low-density lipoprotein receptor related protein (LRP1) may involve the actin cytoskeleton in ways that differ from the endocytosis of soluble LDL by the LDL receptor (LDLR). This study aims to define novel mechanisms of agLDL uptake through modulation of the actin cytoskeleton, to identify molecular targets involved in foam cell formation in vascular smooth muscle cells (VSMCs). The critical observation that formed the basis for these studies is that under pathophysiological conditions, nucleotide release from blood-derived and vascular cells activates SMC P2Y2 receptors (P2Y2Rs) leading to rearrangement of the actin cytoskeleton and cell motility. Therefore, we tested the hypothesis that P2Y2R activation mediates agLDL uptake by VSMCs. Methods Primary VSMCs were isolated from aortas of wild type (WT) C57BL/6 and.P2Y2R−/− mice to investigate whether P2Y2R activation modulates LRP1 expression. Cells were transiently transfected with cDNA encoding a hemagglutinin-tagged (HA-tagged) WT P2Y2R, or a mutant P2Y2R that unlike the WT P2Y2R does not bind the cytoskeletal actin-binding protein filamin-A (FLN-A). Results P2Y2R activation significantly increased agLDL uptake, and LRP1 mRNA expression decreased in P2Y2R−/− VSMCs versus WT. SMCs, expressing P2Y2R defective in FLN-A binding, exhibit 3-fold lower LDLR expression levels than SMCs expressing WT P2Y2R, while cells transfected with WT P2Y2R show greater agLDL uptake in both WT and P2Y2R−/− VSMCs versus cells transfected with the mutant P2Y2R. Conclusions Together, these results show that both LRP1 and LDLR expression and agLDL uptake are regulated by P2Y2R in VSMCs, and that agLDL uptake due to P2Y2R activation is dependent upon cytoskeletal reorganization mediated by P2Y2R binding to FLN-A. PMID:27522265

  10. 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.

  11. Secondary Amines Containing One Aromatic Nitro Group: Preparation, Nitrosation, Sustained Nitric Oxide Release, and the Synergistic Effects of Released Nitric Oxide and an Arginase Inhibitor on Vascular Smooth Muscle Cell Proliferation

    OpenAIRE

    Curtis, Brandon; Payne, Thomas J.; Ash, David E.; Mohanty, Dillip K.

    2013-01-01

    Atherosclerosis, a leading cause of death worldwide, is associated with the excessive proliferation of vascular smooth muscle cells. Nitrogen monoxide, more commonly known as nitric oxide, inhibits this uncontrolled proliferation. Herein we report the preparation of two families of nitric oxide donors; beginning with the syntheses of secondary amine precursors, obtained through the reaction between two equivalents of various monoamines with 2,4 or 2,6-difluoronitrobenzene. The purified second...

  12. Dynamic expression of alpha 1 beta 1 and alpha 2 beta 1 integrin receptors by human vascular smooth muscle cells. Alpha 2 beta 1 integrin is required for chemotaxis across type I collagen-coated membranes.

    OpenAIRE

    Skinner, M P; Raines, E W; Ross, R.

    1994-01-01

    Vascular smooth muscle cells (SMCs) in the media of normal arteries express alpha 1 beta 1 integrin with no detectable alpha 2 beta 1 as determined by immunocytochemistry. In contrast, immunoprecipitation of integrins expressed by human SMCs cultured from medial explants shows strong expression of alpha 2 beta 1 and no expression of alpha 1 beta 1. The apparent reciprocal expression of these two collagen and laminin receptors was confirmed by flow cytometric analysis of fluorescent labeled ce...

  13. Ligand selectivity of 105 kDa and 130 kDa lipoprotein-binding proteins in vascular-smooth-muscle-cell membranes is unique.

    Science.gov (United States)

    Bochkov, V N; Tkachuk, V A; Philippova, M P; Stambolsky, D V; Bühler, F R; Resink, T J

    1996-07-01

    Using ligand blotting techniques, with low-density lipoprotein (LDL) as ligand, we have previously described the existence of atypical lipoprotein-binding proteins (105 kDa and 130 kDa) in membranes from human aortic medical tissue. The present study demonstrates that these proteins are also present in membranes from cultured human (aortic and mesenteric) and rat (aortic) vascular smooth-muscle cells (VSMCs). To assess the relationship of 105 and 130 kDa lipoprotein-binding proteins to known lipoprotein receptors, ligand binding specificity was studied. We tested effects of substances known to antagonize ligand binding to either the LDL [apolipoprotein B,E (apo B,E)] receptor (dextran sulphate, heparin, pentosan polysulphate, protamine, spermine, histone), the scavenger receptor (dextran sulphate, fucoidin), the very-low-density-lipoprotein (VLDL) receptor [receptor-associated protein (RAP)], or LDL receptor-related protein (RAP, alpha 2-macroglobulin, lipoprotein lipase, exotoxin-A). None of these substances, with the exception of dextran sulphate, influenced binding of LDL to either 105 or 130 kDa proteins. Sodium oleate or oleic acid, known stimuli for the lipoprotein binding activity of the lipolysis-stimulated receptor, were also without effect. LDL binding to 105 and 130 kDa proteins was inhibited by anti-LDL (apo B) antibodies. LDL and VLDL bound to 105 and 130 kDa proteins with similar affinities (approximately 50 micrograms/ml). The unique ligand selectivity of 105 and 130 kDa proteins supports the existence of a novel lipoprotein-binding protein that is distinct from all other currently identified LDL receptor family members. The similar ligand selectivity of 105 and 130 kDa proteins suggests that they may represent variant forms of an atypical lipoprotein-binding protein. PMID:8694779

  14. Lithium chloride inhibits vascular smooth muscle cell proliferation and migration and alleviates injury-induced neointimal hyperplasia via induction of PGC-1α.

    Directory of Open Access Journals (Sweden)

    Zhuyao Wang

    Full Text Available The proliferation and migration of vascular smooth muscle cells (VSMCs contributes importantly to the development of in-stent restenosis. Lithium has recently been shown to have beneficial effects on the cardiovascular system, but its actions in VSMCs and the direct molecular target responsible for its action remains unknown. On the other hand, PGC-1α is a transcriptional coactivator which negatively regulates the pathological activation of VSMCs. Therefore, the purpose of the present study is to determine if lithium chloride (LiCl retards VSMC proliferation and migration and if PGC-1α mediates the effects of lithium on VSMCs. We found that pretreatment of LiCl increased PGC-1α protein expression and nuclear translocation in a dose-dependent manner. MTT and EdU incorporation assays indicated that LiCl inhibited serum-induced VSMC proliferation. Similarly, deceleration of VSMC migration was confirmed by wound healing and transwell assays. LiCl also suppressed ROS generation and cell cycle progression. At the molecular level, LiCl reduced the protein expression levels or phosphorylation of key regulators involved in the cell cycle re-entry, adhesion, inflammation and motility. In addition, in vivo administration of LiCl alleviated the pathophysiological changes in balloon injury-induced neointima hyperplasia. More importantly, knockdown of PGC-1α by siRNA significantly attenuated the beneficial effects of LiCl on VSMCs both in vitro and in vivo. Taken together, our results suggest that LiCl has great potentials in the prevention and treatment of cardiovascular diseases related to VSMC abnormal proliferation and migration. In addition, PGC-1α may serve as a promising drug target to regulate cardiovascular physiological homeostasis.

  15. 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.

  16. 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.

  17. Primary culture of differentiated phenotype vascular smooth muscle cells%分化型血管平滑肌细胞的原代培养

    Institute of Scientific and Technical Information of China (English)

    牛建平; 周志斌; 史树海; 彭瑞强

    2008-01-01

    目的 建立大鼠主动脉分化表型血管平滑肌细胞(vascular smooth muscle cell,VSMC)原代培养,为研究VSMC表型转化的分子机制提供体外研究模型.方法 无菌条件下分离大鼠胸主动脉中膜,组织贴壁法在Ⅳ型胶原蛋白包被培养瓶内培养大鼠胸主动脉VSMC,消化、过滤、离心,收集从组织块分离的单个细胞后,接种于含0.2%小牛血清、胰岛素样生长因子Ⅰ、DMEM培养液的层粘连蛋白包被的培养板上,培养1天后将培养液更换为不合血清及生长因子的DMEM.结果 根据细胞形态观察、免疫组织化学鉴定、兴奋剂刺激引发的收缩反应、分化型VSMC标志基因的检测,证实为分化型VSMC,分化状态可持续1周以上.结论 在组织贴壁法基础上,改变培养环境,可使VSMC较长时间保持于分化状态,是研究VSMC表型转化的分子机制良好的体外研究模型.

  18. 17β-Estradiol inhibits TNF-α-induced proliferation and migration of vascular smooth muscle cells via suppression of TRAIL.

    Science.gov (United States)

    Li, Hengchang; Cheng, Yang; Simoncini, Tommaso; Xu, Shiyuan

    2016-07-01

    Atherosclerosis is an inflammatory disease and involves migration of vascular smooth muscle cells (VSMCs). Estrogen inhibits VSMCs migration, while the underlying mechanism remains to be revealed. Recent years, there is emerging evidence showing that TNF-related apoptosis-inducing ligand (TRAIL) increases proliferation and migration of VSMCs. In this study, we investigated the regulatory effect of estrogen on TRAIL expression in VSMCs. TNF-α greatly enhanced TRAIL protein expression and stimulated VSMCs proliferation and migration. This effect was partially inhibited by the addition of TRAIL neutralizing antibody, suggesting that TRAIL is important in TNF-α-induced migration. 17β-estradiol (E2) inhibited TRAIL expression under TNF-α stimulation in a time- and concentration-dependent manner. This effect was was mimicked by ERα agonist 4',4″,4‴-(4-propyl-[1H]-pyrazole-1,3,5-triyl) trisphenol (PPT), but not ERβ agonist 2,3-bis-(4-hydroxyphenyl)-propionitrile (DPN), indicating that ERα is involved in this action. TNF-α led to nuclear factor kappa B (NF-κB) p65 phosphorylation and the inhibitor pyrrolidine dithiocarbama (PDTC) inhibited TRAIL expression, suggesting that NF-κB signaling is crucial for TARIL production. E2 suppressed p65 phosphorylation in VSMCs and the overexpression of p65 subunit reversed the inhibitory effect of E2 on TRAIL expression and cell proliferation and migration. Taken together, our results indicate that E2 inhibits VSMCs proliferation and migration by downregulation of TRAIL expression via suppression of NF-κB pathway.

  19. Impact of losartan and angiotensin II on the expression of matrix metalloproteinase-9 and tissue inhibitor of metalloproteinase-1 in rat vascular smooth muscle cells.

    Science.gov (United States)

    Guo, Yan-Song; Wu, Zong-Gui; Yang, Jun-Ke; Chen, Xin-Jing

    2015-03-01

    The present study aimed to investigate the impact of losartan and angiotensin II (AngII) on the expression of matrix metalloproteinase-9 (MMP-9) and tissue inhibitor of metalloproteinase-1 (TIMP-1), secreted by rat vascular smooth muscle cells (VSMCs). Rat VSMCs were isolated and cultured in different concentrations of AngII and losartan for 24 h and western blot analysis and quantitative polymerase chain reaction were performed to observe the subsequent impact on the gene and protein expression of MMP-9 and TIMP-1. AngII was shown to promote the protein and gene expression of MMP-9 in VSMCs in a concentration-dependent manner. No effect was observed on the expression of TIMP-1, therefore, an increase in the MMP-9/TIMP-1 ratio was observed. Losartan was shown to be able to inhibit MMP-9 protein and gene expression in a concentration-dependent manner, whilst promoting an increase in TIMP-1 expression, thus decreasing the ratio of MMP-9/TIMP-1. The combined action of losartan and AngII resulted in the same directional changes in MMP-9 and TIMP-1 expression as observed for losartan alone. The comparison of AngII, losartan and the combinatory effect on the expression of MMP-9 and TIMP-1 in VSMCs indicated that losartan inhibited the effects of AngII, therefore reducing the MMP-9/TIMP-1 ratio, which may contribute to the molecular mechanism of losartan in preventing atherosclerosis. In atherosclerosis, the development of the extracellular matrix of plaque is closely correlated with the evolution of AS. The balance between MMPs and TIMPs is important in maintaining the dynamic equilibrium between the ECM, and the renin-angiotensin-aldosterone system, which is involved in the pathologenesis of AS, and in which AngII has a central role.

  20. Hypertension secondary to early-stage kidney disease: the pathogenetic role of altered cytosolic calcium (Ca2+) homeostasis of vascular smooth muscle cells.

    Science.gov (United States)

    Schiffl, H; Fricke, H; Sitter, T

    1993-05-01

    We have examined cardiovascular pressor responsiveness to infused norepinephrine (NE) as related to endogenous plasma NE and plasma renin and to platelet free cytosolic (Ca2+) in 36 patients with early-stage kidney disease and 27 matched normal subjects. The 27 hypertensive patients and the normal subjects did not differ in blood volume, plasma renin, and NE; however, the hypertensive patients had a higher exchangeable body sodium content. Basal plasma NE levels, the relationship between plasma NE measured during NE infusion and the corresponding NE infusion rate, as well as the total plasma clearance for NE did also not differ significantly between the two study groups. In contrast, the threshold or pressor doses of infused NE significantly decreased in the patients with kidney disease. Antihypertensive pharmacotherapy with (Ca2+) channel blockers and/or loop diuretics normalized blood pressure and cardiovascular NE hyperresponsiveness and reduced blood volume, exchangeable body sodium, and platelet free cytosolic (Ca2+). In contrast, experimental digitalisation as a model for in vivo sodium/potassium adenosine triphosphatase inhibition augmented NE responsiveness and raised platelet free cytosolic (Ca2+). Incubation of platelets from normal subjects with plasma ultrafiltrate from hypertensive patients gave evidence for an endogenous factor capable to raise free cytosolic (Ca2+) and to act synergistically with digoxin. Hypertension secondary to early-stage kidney disease is related to an impairment of sodium excretion leading to an expansion of blood volume and exchangeable body sodium. This may result in increased secretion of endogenous factors, leading to alterations of cytosolic (Ca2+) homeostasis of vascular smooth muscle cells followed by elevated peripheral resistance and thus blood pressure. PMID:8494019

  1. Effect of Nuclear Factor-kappa B on Vascular Endothelial Growth Factor mRNA Expression of Human Pulmonary Artery Smooth Muscle Cells in Hypoxia

    Institute of Scientific and Technical Information of China (English)

    张焕萍; 徐永健; 张珍祥; 许淑云; 倪望; 陈士新

    2004-01-01

    Summary: In order to investigate the effect of nuclear factor-kappa B (NF-κB) on vascular endothelial growth factor (VEGF) mRNA expression of human pulmonary artery smooth muscle cells (HPASMCs) in hypoxia, the cultured HPASMCs in vitro were stimulated with pyrrolidine dithiocarbamate (PDTC), an inhibitor of NF-κB. The NF-κB p65 nuclei positive expression was detected by immunocytochemical technique. The IκBa protein expression was measured by Western blot.RT-PCR was used to detect the VEGF mRNA expression of HPASMCs. The results showed that no significant change was observed in the NF-κB p65 nuclei positive expression of cultured HPASMCs during 6 h-24 h in normoxia, but the levels of NF-κB p65 nuclei positive expression of cultured HPASMCs were significantly increased in hypoxia groups as compared with those in all normoxia groups (P<0.05). The IκBα protein expression of cultured HPASMCs showed no significant change during 6 h-24 h in normoxia, but significantly decreased in hypoxia as comapred with that in normoxia groups (P<0.05). PDTC (1 to 100 μmol/L) could inhibit the VEGF mRNA expression of HPASMCs in a concentration-dependent manner in hypoxia. In conclusion, NF-κB can be partly translocation activated from cytoplasm into nuclei in the cultured HPASMCs under hypoxia. The inhibition of NF-κB activation can decrease the VEGF mRNA expression. h is suggested that the activation of NF-κB is involved in the VEGFmRNA expression of HPASMCs under hypoxia.

  2. Polydatin inhibits the oxidative stress-induced proliferation of vascular smooth muscle cells by activating the eNOS/SIRT1 pathway.

    Science.gov (United States)

    Ma, Yi; Gong, Xun; Mo, Yingli; Wu, Saizhu

    2016-06-01

    Oxidative stress-mediated proliferation of vascular smooth muscle cells (VSMCs) contributes to plaque formation and the progression of atherosclerosis. Polydatin is a derivative of resveratrol, and is widely present in certain herbal medications used for the treatment of cardiovascular diseases. In the present study, we examined whether polydatin was capable of attenuating VSMC proliferation induced by oxidative stress as well as the potential involvement of the endothelial nitric oxide synthetase (eNOS)/SIRT1 pathway. Briefly, VSMCs were exposed to H2O2 for 24 h in the absence or presence of polydatin (10-100 µM) prior to performing a cell proliferation assay. In mechanistic studies, the cells were incubated with the silent information regulator 1 (SIRT1) inhibitor, EX527, or the eNOS inhibitor, L-NAME, prior to polydatin treatment. The results showed that polydatin inhibited VSMC proliferation and the level of reactive oxygen species, increased the expression of Kip1/p27, SIRT1 and eNOS, whereas the expression of cyclin B1, Cdk1 and c-myc was decreased. The number of cells in the G2/M phase was increased. Pre-treatment with L-NAME attenuated the inhibitory effects of polydatin on cell proliferation, inhibited the expression of SIRT1 and the phosphorylation of eNOS. Pre-treatment with EX527 also attenuated the inhibitory effects of polydatin on cell proliferation, but failed to reduce the activation of eNOS and the production of nitric oxide. Taken together, these findings suggest that, polydatin inhibited the oxidative stress-induced proliferation of VMSCs by activating the eNOS/SIRT1 pathway. PMID:27081912

  3. Arginase inhibition reduces interleukin-1β-stimulated vascular smooth muscle cell proliferation by increasing nitric oxide synthase-dependent nitric oxide production

    Energy Technology Data Exchange (ETDEWEB)

    Yoon, Jeongyeon; Ryoo, Sungwoo, E-mail: ryoosw08@kangwon.ac.kr

    2013-06-07

    Highlights: •Arginase inhibition suppressed proliferation of IL-1β-stimulated VSMCs in dose-dependent manner. •NO production from IL-1β-induced iNOS expression was augmented by arginase inhibition, reducing VSMC proliferation. •Incubation with cGMP analogues abolished IL-1β-dependent proliferation of VSMCs. -- Abstract: We investigated whether arginase inhibition suppressed interleukin (IL)-1β-stimulated proliferation in vascular smooth muscle cells (VSMCs) and the possible mechanisms involved. IL-1β stimulation increased VSMC proliferation, while the arginase inhibitor BEC and transfection of the antisense (AS) oligonucleotide against arginase I decreased VSMC proliferation and was associated with increased protein content of the cell cycle regulator p21Waf1/Cip1. IL-1β incubation induced inducible nitric oxide synthase (iNOS) mRNA expression and protein levels in a dose-dependent manner, but did not affect arginase I and II expression. Consistent with this data, IL-1β stimulation resulted in increase in NO production that was significantly augmented by arginase inhibition. The specific iNOS inhibitor 1400W abolished IL-1β-mediated NO production and further accentuated IL-1β-stimulated cell proliferation. Incubation with NO donors GSNO and DETA/NO in the presence of IL-1β abolished VSMCs proliferation and increased p21Waf1/Cip1 protein content. Furthermore, incubation with the cGMP analogue 8-Br-cGMP prevented IL-1β-induced VSMCs proliferation. In conclusion, arginase inhibition augmented iNOS-dependent NO production that resulted in suppression of IL-1β-induced VSMCs proliferation in a cGMP-dependent manner.

  4. The inhibitory effect of dexamethasone on platelet-derived growth factor-induced vascular smooth muscle cell migration through up-regulating PGC-1{alpha} expression

    Energy Technology Data Exchange (ETDEWEB)

    Xu, Wei [Jiangsu Diabetes Center, State Key Laboratory of Pharmaceutical Biotechnology, School of Life Sciences, Nanjing University, 22 Hankou Road, Nanjing 210093 (China); Department of cardiology, the Second Affiliated Hospital of Harbin Medical University, 246 Xuefu Road, Harbin 150081 (China); Guo, Ting; Zhang, Yan; Jiang, Xiaohong; Zhang, Yongxian [Jiangsu Diabetes Center, State Key Laboratory of Pharmaceutical Biotechnology, School of Life Sciences, Nanjing University, 22 Hankou Road, Nanjing 210093 (China); Zen, Ke, E-mail: kzen@nju.edu.cn [Jiangsu Diabetes Center, State Key Laboratory of Pharmaceutical Biotechnology, School of Life Sciences, Nanjing University, 22 Hankou Road, Nanjing 210093 (China); Yu, Bo, E-mail: yubodr@163.com [Department of cardiology, the Second Affiliated Hospital of Harbin Medical University, 246 Xuefu Road, Harbin 150081 (China); Zhang, Chen-Yu, E-mail: cyzhang@nju.edu.cn [Jiangsu Diabetes Center, State Key Laboratory of Pharmaceutical Biotechnology, School of Life Sciences, Nanjing University, 22 Hankou Road, Nanjing 210093 (China)

    2011-05-01

    Dexamethasone has been shown to inhibit vascular smooth muscle cell (VSMC) migration, which is required for preventing restenosis. However, the mechanism underlying effect of dexamethasone remains unknown. We have previously demonstrated that peroxisome proliferator-activated receptor gamma (PPAR{gamma}) coactivator-1 alpha (PGC-1{alpha}) can inhibit VSMC migration and proliferation. Here, we investigated the role of PGC-1{alpha} in dexamethasone-reduced VSMC migration and explored the possible mechanism. We first examined PGC-1{alpha} expression in cultured rat aortic VSMCs. The results revealed that incubation of VSMCs with dexamethasone could significantly elevate PGC-1{alpha} mRNA expression. In contrast, platelet-derived growth factor (PDGF) decreased PGC-1{alpha} expression while stimulating VSMC migration. Mechanistic study showed that suppression of PGC-1{alpha} by small interfering RNA strongly abrogated the inhibitory effect of dexamethasone on VSMC migration, whereas overexpression of PGC-1{alpha} had the opposite effect. Furthermore, an analysis of MAPK signal pathways showed that dexamethasone inhibited ERK and p38 MAPK phosphorylation in VSMCs. Overexpression of PGC-1{alpha} decreased both basal and PDGF-induced p38 MAPK phosphorylation, but it had no effect on ERK phosphorylation. Finally, inhibition of PPAR{gamma} activation by a PPAR{gamma} antagonist GW9662 abolished the suppressive effects of PGC-1{alpha} on p38 MAPK phosphorylation and VSMC migration. These effects of PGC-1{alpha} were enhanced by a PPAR{gamma} agonist troglitazone. Collectively, our data indicated for the first time that one of the anti-migrated mechanisms of dexamethasone is due to the induction of PGC-1{alpha} expression. PGC-1{alpha} suppresses PDGF-induced VSMC migration through PPAR{gamma} coactivation and, consequently, p38 MAPK inhibition.

  5. The inhibitory effect of dexamethasone on platelet-derived growth factor-induced vascular smooth muscle cell migration through up-regulating PGC-1α expression

    International Nuclear Information System (INIS)

    Dexamethasone has been shown to inhibit vascular smooth muscle cell (VSMC) migration, which is required for preventing restenosis. However, the mechanism underlying effect of dexamethasone remains unknown. We have previously demonstrated that peroxisome proliferator-activated receptor gamma (PPARγ) coactivator-1 alpha (PGC-1α) can inhibit VSMC migration and proliferation. Here, we investigated the role of PGC-1α in dexamethasone-reduced VSMC migration and explored the possible mechanism. We first examined PGC-1α expression in cultured rat aortic VSMCs. The results revealed that incubation of VSMCs with dexamethasone could significantly elevate PGC-1α mRNA expression. In contrast, platelet-derived growth factor (PDGF) decreased PGC-1α expression while stimulating VSMC migration. Mechanistic study showed that suppression of PGC-1α by small interfering RNA strongly abrogated the inhibitory effect of dexamethasone on VSMC migration, whereas overexpression of PGC-1α had the opposite effect. Furthermore, an analysis of MAPK signal pathways showed that dexamethasone inhibited ERK and p38 MAPK phosphorylation in VSMCs. Overexpression of PGC-1α decreased both basal and PDGF-induced p38 MAPK phosphorylation, but it had no effect on ERK phosphorylation. Finally, inhibition of PPARγ activation by a PPARγ antagonist GW9662 abolished the suppressive effects of PGC-1α on p38 MAPK phosphorylation and VSMC migration. These effects of PGC-1α were enhanced by a PPARγ agonist troglitazone. Collectively, our data indicated for the first time that one of the anti-migrated mechanisms of dexamethasone is due to the induction of PGC-1α expression. PGC-1α suppresses PDGF-induced VSMC migration through PPARγ coactivation and, consequently, p38 MAPK inhibition.

  6. Iroquois homeobox transcription factor (Irx5) promotes G1/S-phase transition in vascular smooth muscle cells by CDK2-dependent activation.

    Science.gov (United States)

    Liu, Dong; Pattabiraman, Vaishnavi; Bacanamwo, Methode; Anderson, Leonard M

    2016-08-01

    The Iroquois homeobox (Irx5) gene is essential in embryonic development and cardiac electrophysiology. Although recent studies have reported that IRX5 protein is involved in regulation of the cell cycle and apoptosis in prostate cancer cells, little is known about the role of IRX5 in the adult vasculature. Here we report novel observations on the role of IRX5 in adult vascular smooth muscle cells (VSMCs) during proliferation in vitro and in vivo. Comparative studies using primary human endothelial cells, VSMCs, and intact carotid arteries to determine relative expression of Irx5 in the peripheral vasculature demonstrate significantly higher expression in VSMCs. Sprague-Dawley rat carotid arteries were subjected to balloon catherization, and the presence of IRX5 was examined by immunohistochemistry after 2 wk. Results indicate markedly elevated IRX5 signal at 14 days compared with uninjured controls. Total RNA was isolated from injured and uninjured arteries, and Irx5 expression was measured by RT-PCR. Results demonstrate a significant increase in Irx5 expression at 3-14 days postinjury compared with controls. Irx5 genetic gain- and loss-of-function studies using thymidine and 5-bromo-2'-deoxyuridine incorporation assays resulted in modulation of DNA synthesis in primary rat aortic VSMCs. Quantitative RT-PCR results revealed modulation of cyclin-dependent kinase inhibitor 1B (p27(kip1)), E2F transcription factor 1 (E2f1), and proliferating cell nuclear antigen (Pcna) expression in Irx5-transduced VSMCs compared with controls. Subsequently, apoptosis was observed and confirmed by morphological observation, caspase-3 cleavage, and enzymatic activation compared with control conditions. Taken together, these results indicate that Irx5 plays an important role in VSMC G1/S-phase cell cycle checkpoint control and apoptosis. PMID:27170637

  7. Identification and characterization of a novel angiotensin binding site in cultured vascular smooth muscle cells that is specific for the hexapeptide (3-8) fragment of angiotensin II, angiotensin IV.

    Science.gov (United States)

    Hall, K L; Hanesworth, J M; Ball, A E; Felgenhauer, G P; Hosick, H L; Harding, J W

    1993-03-19

    This study demonstrates the existence of a previously unrecognized class of angiotensin binding sites on vascular smooth muscle that exhibit high affinity and specificity for the hexapeptide (3-8) fragment of angiotensin II (AngIV). Binding of [125I]AngIV is saturable, reversible and describes a pharmacologic profile that is distinct and separate from the classic AT1 or AT2 angiotensin receptors. Saturation binding studies utilizing cultured vascular smooth muscle cells obtained from bovine aorta (BVSM) revealed that [125I]AngIV bound to a single high affinity site with an associated Hill coefficient of 0.99 +/- 0.003, exhibiting a KD = 1.85 +/- 0.45 nM and a corresponding Bmax = 960 +/- 100 fmol mg-1 protein. Competition binding curves in BVSM demonstrated the following rank order effectiveness: AngIV > AngII(3-7) > AngIII > Sar1,Ile8 AngII > AngII > AngII(1-7) > AngII(4-8), DuP 753, PD123177. The presence of the non-hydrolyzable GTP analog GTP gamma S, had no effect on [125I]AngIV binding affinity in BVSM. The presence of this novel angiotensin binding site on smooth muscle in high concentration suggests the possibility that this system may play an important, yet unrecognized role in vascular control.

  8. TGF-beta Inhibits Ang II-Induced MAPK p44/42 Signaling in Vascular Smooth Muscle Cells by Ang II Type 1 Receptor Downregulation

    NARCIS (Netherlands)

    Meijering, Bernadet D. M.; van der Wouden, Els A.; Pelgrom, Vincent; Henning, Robert H.; Sharma, Kumar; Deelman, Leo E.

    2009-01-01

    Vascular changes in diabetes are characterized by reduced vasoconstriction and vascular remodeling. Previously, we demonstrated that TGF-beta 1 impairs Ang II-induced contraction through reduced calcium mobilization. However, the effect of TGF-beta 1 on Ang II-induced vascular remodeling is unknown.

  9. TGF-beta inhibits Ang II-induced MAPK p44/42 signaling in vascular smooth muscle cells by Ang II type 1 receptor downregulation.

    NARCIS (Netherlands)

    Meijering, B.D.; Wouden, E.A. van der; Pelgrom, V.; Henning, R.H.; Sharma, K.; Deelman, L.E.

    2009-01-01

    Vascular changes in diabetes are characterized by reduced vasoconstriction and vascular remodeling. Previously, we demonstrated that TGF-beta1 impairs Ang II-induced contraction through reduced calcium mobilization. However, the effect of TGF-beta1 on Ang II-induced vascular remodeling is unknown. T

  10. 心脉龙注射液对家兔离体血管平滑肌的影响%Effects of sinmailong Injection on Isolated Vascular Smooth Muscle in Rabbit

    Institute of Scientific and Technical Information of China (English)

    彭芳; 瞿培红; 等

    2001-01-01

    目的:研究心脉龙注射液升压作用与血管平滑肌钙通道之间的关系。方法:观察心脉龙注射液对不同处理的离体血管平滑肌的影响。结果:心脉龙注射液能兴奋家兔离体降主动脉及肺动脉,该药引起上述动脉条收缩 50%效应的浓度分别是 1.99、 4.67mg.ml- 1,分别在 3× 10- 5~ 6× 10- 3、 10- 3~ 6× 10- 3g.ml- 1范围呈量效关系。 50μ mol维拉帕米能完全取消心脉龙注射液兴奋降主动脉、肺动脉的作用。该药对两种血管条在无钙及含钙 Krebs液中分别引起无兴奋及收缩反应。结论:心脉龙注射液收缩血管平滑肌与细胞外钙内流有密切关系。%Objective: To study the relationship between Xinmailong injection raising pressure and calcium channel in vascular smooth muscle.Methods: To observe the effects of Xinmailong injection on isolated vascular smooth muscle by various handles.Results: Xinmailong injection could constrict isolated thoracic artery and pulmonary artery in rabbits, and the concentrations causing the median constrict were 1.99、 4.67 mg.ml 1respectively.Xinmailong injection could appear concentration- effect relationship on thoracic artery and pulmonary artery in scope of 3× 10- 5~ 6× 10- 3、 10- 3~ 6× 10- 3g.ml- 1respectively.50 umol verapamil could entirely conceal the effects of Xinmailong injection exciting the two vascular smooth. Xinmailong injection could cause no excitg and contracting with or without calcium Krebs solution in the two vascular smooth.Conclusion:There is close relationship between Xinmailong injection on contracting vascular smooth muscle and the calcium influx.

  11. The retardation of vasculopathy induced by attenuation of insulin resistance in the corpulent JCR:LA-cp rat is reflected by decreased vascular smooth muscle cell proliferation in vivo.

    Science.gov (United States)

    Absher, P M; Schneider, D J; Baldor, L C; Russell, J C; Sobel, B E

    1999-04-01

    Proliferation in vivo of vascular smooth muscle cells occurs early in the course of atherosclerosis. Cultured smooth muscle cells (SMCs) explanted from aortas of JCR:LA-cp corpulent rats known to exhibit metabolic derangements and insulin resistance typical of type II diabetes early in life and to develop atherosclerosis later in life exhibit increased proliferation compared with SMCs from lean, normal rats. Vascular smooth muscle proliferation in vitro was found to be positively and significantly correlated with plasma insulin levels in vivo. Proliferation of aortic SMCs from JCR:LA-cp cp/cp corpulent rats cultured in vitro exhibited increased proliferation in the presence of exogenous insulin. Exercise and diet, selected as interventions designed to ameliorate the insulin resistance and hyperinsulinemia in the JCR:LA-cp cp/cp rat, effectively lowered blood insulin levels and decreased subsequent proliferation in vitro of aortic SMCs explanted from these animals. The results indicate that assessment of proliferation of vascular smooth muscle cells ex vivo may provide insight into the presence and severity of atherogenicity in association with insulin resistance in diverse species under diverse circumstances. Accordingly, with appropriate controls, it may be possible to use SMC proliferation ex vivo as a marker of the extent to which an intervention such as administration of insulin sensitizers to experimental animals and human subjects results in a change in behavior of vessel wall elements potentially indicative of amelioration of atherogenicity and detectable as judged from reduced proliferative rates of the cells ex vivo when they have been harvested from vessels exposed to a milieu in which insulin resistance has been attenuated.

  12. Differential baseline expression and angiotensin II stimulation of leukemia-associated RhoGEF in vascular smooth muscle cells of spontaneously hypertensive rats

    Directory of Open Access Journals (Sweden)

    Chiu WC

    2012-12-01

    Full Text Available Wei-Chiao Chiu,1 Jyh-ming Juang,1,2 Shen-nan Chang,2 Cho-kai Wu,2 Chia-ti Tsai,2 Chuen-den Tseng,2 Yung-zu Tseng,1,2 Ming-Jai Su,3 Fu-tien Chiang1,2,41Graduate Institute of Physiology, National Taiwan University College of Medicine, 2Division of Cardiology, Department of Internal Medicine, National Taiwan University Hospital and National Taiwan University College of Medicine, 3Graduate Institute of Pharmacology, National Taiwan University College of Medicine, 4Department of Laboratory Medicine, National Taiwan University College of Medicine, Taipei, Taiwan, Republic of ChinaPurpose: Studies to explore angiotensin II (Ang II and its downstream signaling pathways via Rho guanine nucleotide exchange factors (RhoGEFs and RhoA signaling are crucial to understanding the mechanisms of smooth muscle contraction leading to hypertension. This study aimed to investigate the Ang II–induced expression of RhoGEFs in vascular smooth muscle cells (VSMCs of spontaneously hypertensive rats (SHRs and to identify the possible regulator associated with hypertension.Methods: Cultured VSMCs of the aorta from SHRs and Wistar-Kyoto (WKY rats were treated with or without Ang II or Ang II plus Ang II type 2 receptor antagonists. The expression levels of RhoGEF messenger RNA (mRNA and protein were determined. To evaluate the changes of aortic ring contractile force in response to Ang II, a nonviral carrier system was adopted to deliver the leukemia-associated RhoGEF (LARG small interfering RNA via nanoparticles into aortic rings.Results: The baseline mRNA levels of three RhoGEFs in cultured VSMCs of WKY rats did not increase with age, but they were significantly higher in 12-week-old SHRs than in 5-week-old SHRs. Expression levels of LARG mRNA were higher in SHRs than in age-matched WKY rats. The baseline LAGR protein of 12-week-old SHRs was about four times higher than that of WKY rats of the same age. After Ang II stimulation, LAGR protein expression was significantly

  13. Angiotensin II-induced Akt activation through the epidermal growth factor receptor in vascular smooth muscle cells is mediated by phospholipid metabolites derived by activation of phospholipase D.

    Science.gov (United States)

    Li, Fang; Malik, Kafait U

    2005-03-01

    Angiotensin II (Ang II) activates cytosolic Ca(2+)-dependent phospholipase A(2) (cPLA(2)), phospholipase D (PLD), p38 mitogen-activated protein kinase (MAPK), epidermal growth factor receptor (EGFR) and Akt in vascular smooth muscle cells (VSMC). This study was conducted to investigate the relationship between Akt activation by Ang II and other signaling molecules in rat VSMC. Ang II-induced Akt phosphorylation was significantly reduced by the PLD inhibitor 1-butanol, but not by its inactive analog 2-butanol, and by brefeldin A, an inhibitor of the PLD cofactor ADP-ribosylation factor, and in cells infected with retrovirus containing PLD(2) siRNA or transfected with PLD(2) antisense but not control LacZ or sense oligonucleotide. Diacylglycerol kinase inhibitor II diminished Ang II-induced and diC8-phosphatidic acid (PA)-increased Akt phosphorylation, suggesting that PLD-dependent Akt activation is mediated by PA. Ang II-induced EGFR phosphorylation was inhibited by 1-butanol and PLD(2) siRNA and also by cPLA(2) siRNA. In addition, the inhibitor of arachidonic acid (AA) metabolism 5,8,11,14-eicosatetraynoic acid (ETYA) reduced both Ang II- and AA-induced EGFR transactivation. Furthermore, ETYA, cPLA(2) antisense, and cPLA(2) siRNA attenuated Ang II-elicited PLD activation. p38 MAPK inhibitor SB202190 [4-(4-flurophenyl)-2-(4-hydroxyphenyl)-5-(4-pyridyl)1H-imidazole] reduced PLD activity and EGFR and Akt phosphorylation elicited by Ang II. Pyrrolidine-1, a cPLA(2) inhibitor, and cPLA(2) siRNA decreased p38 MAPK activity. These data indicate that Ang II-stimulated Akt activity is mediated by cPLA(2)-dependent, p38 MAPK regulated PLD(2) activation and EGFR transactivation. We propose the following scheme of the sequence of events leading to activation of Akt in VSMC by Ang II: Ang II-->cPLA(2)-->AA-->p38 MAPK-->PLD(2)-->PA-->EGFR-->Akt. PMID:15525798

  14. Angiotensin II-induced Akt activation through the epidermal growth factor receptor in vascular smooth muscle cells is mediated by phospholipid metabolites derived by activation of phospholipase D.

    Science.gov (United States)

    Li, Fang; Malik, Kafait U

    2005-03-01

    Angiotensin II (Ang II) activates cytosolic Ca(2+)-dependent phospholipase A(2) (cPLA(2)), phospholipase D (PLD), p38 mitogen-activated protein kinase (MAPK), epidermal growth factor receptor (EGFR) and Akt in vascular smooth muscle cells (VSMC). This study was conducted to investigate the relationship between Akt activation by Ang II and other signaling molecules in rat VSMC. Ang II-induced Akt phosphorylation was significantly reduced by the PLD inhibitor 1-butanol, but not by its inactive analog 2-butanol, and by brefeldin A, an inhibitor of the PLD cofactor ADP-ribosylation factor, and in cells infected with retrovirus containing PLD(2) siRNA or transfected with PLD(2) antisense but not control LacZ or sense oligonucleotide. Diacylglycerol kinase inhibitor II diminished Ang II-induced and diC8-phosphatidic acid (PA)-increased Akt phosphorylation, suggesting that PLD-dependent Akt activation is mediated by PA. Ang II-induced EGFR phosphorylation was inhibited by 1-butanol and PLD(2) siRNA and also by cPLA(2) siRNA. In addition, the inhibitor of arachidonic acid (AA) metabolism 5,8,11,14-eicosatetraynoic acid (ETYA) reduced both Ang II- and AA-induced EGFR transactivation. Furthermore, ETYA, cPLA(2) antisense, and cPLA(2) siRNA attenuated Ang II-elicited PLD activation. p38 MAPK inhibitor SB202190 [4-(4-flurophenyl)-2-(4-hydroxyphenyl)-5-(4-pyridyl)1H-imidazole] reduced PLD activity and EGFR and Akt phosphorylation elicited by Ang II. Pyrrolidine-1, a cPLA(2) inhibitor, and cPLA(2) siRNA decreased p38 MAPK activity. These data indicate that Ang II-stimulated Akt activity is mediated by cPLA(2)-dependent, p38 MAPK regulated PLD(2) activation and EGFR transactivation. We propose the following scheme of the sequence of events leading to activation of Akt in VSMC by Ang II: Ang II-->cPLA(2)-->AA-->p38 MAPK-->PLD(2)-->PA-->EGFR-->Akt.

  15. Ablation of adenosine monophosphate-activated protein kinaseα1 in vascular smooth muscle cells promotes diet-induced atherosclerotic calcification in vivo

    Institute of Scientific and Technical Information of China (English)

    CAI Zhe-jun; DING Ye; ZHANG Miao; LU Qiu-lun; WU Sheng-nan; ZHU Huai-ping; SONG Ping; ZOU Ming-hui

    2016-01-01

    AIM:Atherosclerotic calcification is highly linked with plaque instability and cardiovascular events .Adenosine monophosphate-activated protein kinase ( AMPK) has been involved in the pathogenesis of various cardiovascular disease .The contributions of AMPKαsubunits to the development of atherosclerotic calcification in vivo remained unknown .We hypothesized that AMPKαsubunits may play a role in the development of atherosclerotic calcification .METHODS: Atherosclerotic calcification was generated by 24-week fed of western diet in ApoE-/-background mice .Calcification was evaluated in aortic roots and innominate arteries of ApoE-/-mice or in mice with dual deficiencies of ApoE and AMPKαsubunits globally ( AMPKα1 and AMPKα2 ) , or vascular smooth muscle cell ( VSMC)-specific or macrophage-specific knockout of AMPKα1 with atherosclerotic calcification pone diet . The mechanism of AMPKα1 in regulating Runx2 was further explored in human aortic VSMC .RESULTS: Ablation of AMPKα1 but not AMPKα2 in ApoE-/-background promoted atherosclerotic calcification with increased Runt -related transcription factor ( Runx2 ) expression in VSMC compared with ApoE-/-mice.Conversely, chronic administration of metformin, which activated AMPK, markedly reduced ath-erosclerotic calcification and Runx2 expression in ApoE-/-mice but had less effects in ApoE-/-/AMPKα1 -/-mice.Furthermore, VSMC-but not macrophage-specific deficiency of AMPKα1 in ApoE-/-background promoted atherosclerotic calcification in vivo com-pared with the controls .AMPKα1 silencing in human aortic VSMC prevented Runx 2 from proteasome degradation to trigger osteoblastic differentiation of VSMC .Conversely , activation of AMPK led to Runx 2 instability by inducing its small ubiquitin-like modifier modifi-cation (SUMOylation).Protein inhibitor of activated STAT-1 (PIAS1), the SUMO E3-ligase of Runx2, was directly phosphorylated by AMPKα1 at serine 510, to enhance its SUMO E3-ligase activity.Ablation of PIAS1

  16. Effects of rosuvastatin on the production and activation of matrix metalloproteinase-2 and migration of cultured rat vascular smooth muscle cells induced by homocysteine

    Institute of Scientific and Technical Information of China (English)

    Ya-fei SHI; Ju-fang CHI; Wei-liang TANG; Fu-kang XU; Long-bin LIU; Zheng JI; Hai-tao LV

    2013-01-01

    Objective:To test the influence of homocysteine on the production and activation of matrix metalloproteinase-2 (MMP-2) and tissue inhibitors of matrix metalloproteinase-2 (TIMP-2) and on cell migration of cultured rat vascular smooth muscle cells (VSMCs).Also,to explore whether rosuvastatin can alter the abnormal secretion and activation of MMP-2 and TIMP-2 and migration of VSMCs induced by homocysteine.Methods:Rat VSMCs were incubated with different concentrations of homocysteine (50-5000 μmol/L).Western blotting and gelatin zymography were used to investigate the expressions and activities of MMP-2 and TIMP-2 in VSMCs in culture medium when induced with homocysteine for 24,48,and 72 h.Transwell chambers were employed to test the migratory ability of VSMCs when incubated with homocysteine for 48 h.Different concentrations of rosuvastatin (10-9-10-5 mol/L) were added when VSMCs were induced with 1000 μmol/L homocysteine.The expressions and activities of MMP-2 and TIMP-2 were examined after incubating for 24,48,and 72 h,and the migration of VSMCs was also examined after incubating for 48 h.Results:Homocysteine (50-1000 μmol/L) increased the production and activation of MMP-2 and expression of TIMP-2 in a dose-dependent manner.However,when incubated with 5000 μmol/L homocysteine,the expression of MMP-2 was up-regulated,but its activity was down-regulated.Increased homocysteine-induced production and activation of MMP-2 were reduced by rosuvastatin in a dose-dependent manner whereas secretion of TIMP-2 was not significantly altered by rosuvastatin.Homocysteine (50-5000 μmol/L) stimulated the migration of VSMCs in a dose-dependent manner,but this effect was eliminated by rosuvastatin.Conclusions:Homocysteine (50-1000 μmol/L) significantly increased the production and activation of MMP-2,the expression of TIMP-2,and the migration of VSMCs in a dose-dependent manner.Additional extracellular rosuvastatin can decrease the excessive expression and activation of MMP-2

  17. Effects of Total Flavonoids of Hippophae Rhamnoides L. on Intracellular Free Calcium in Cultured Vascular Smooth Muscle Cells of Spontaneously Hypertensive Rats and Wistar-Kyoto Rats

    Institute of Scientific and Technical Information of China (English)

    ZHU Fu; CHEN Wei; HUANG Pan-hua; HUANG Bo; HU Chun-yan; JIANG Qing-yuan; LU Zhen-guo; LU Ming; WANG Mei-hua; GONG Min; QIAO Chun-ping

    2005-01-01

    Objective: To explore the effects of total flavonoids of Hippophae rhamnoides L. (TFH),quercetin (Que) and isorhamnetin (Isor) on the intracellular free calcium ([Ca2+ ]i) in vascular smooth muscle cells (VSMC) of spontaneously hypertensive rats (SHR) and Wistar-Kyoto rats (WKY). Methods: Fluo 3-acetoxymethylester(Fluo-3/AM) was used to observe the effects of TFH (100mg/L) and its essential monomers, namely Que (10-4mol/L) and Isor (10-4mol/L) on changes of [Ca2+] i in cultured SHR and WKY VSMC (abbr. to Ca-SHR & Ca-WKY) following exposure to high K+, norepinephrine (NE) and angiotensin Ⅱ (Ang Ⅱ ), and to compare with the effects of verapamil (Ver). Results: (1) TFH, Que and Isor had inhibitory effects on resting Ca-SHR (P<0.05), but had no significant effects on Ca-WKY (P>0.05). (2) High K+ could increase Ca-SHR more significantly than Ca-WKY (P<0.05); TFH, Que and Isor could inhibit the elevation of [Ca2+ ]i induced by high K+-depolarization, with the effects similar to that of Ver, and the effect on Ca-SHR was more significant than that on Ca-WKY ( P<0.05). (3) NE and Ang Ⅱ could increase Ca-SHR more significantly than Ca-WKY ( P<0.05), TFH, Que and Isor had remarkably inhibitory effect on the elevation of Ca-SHR and Ca-WKY induced by NE or Ang Ⅱ. (4) In the absence of extracellular Ca2+ , TFH, Que and Isor also had certain inhibitory effect on Ca-SHR and Ca-WKY induced by NE, and the effect on the former was more significant than that on the latter( P<0.05). Conclusion: TFH, Que and Isor might decrease the levels of [Ca2+ ]i in VSMCs by blocking both voltage-dependent calcium channels (VDC) and receptoroperated calcium channels (ROC) in physiological or pathological state, which may be one of the important mechanisms of their hypotensive and protective effects on target organs in patients with hypertension.

  18. Screening of Active Compounds from Gastrodia elata Blume for Vascular Smooth Muscle Relaxation%云南昭通天麻松弛血管平滑肌活性成分的筛选

    Institute of Scientific and Technical Information of China (English)

    张维明; 杨莲; 李秀芳; 林青; 李国花; 魏文彬

    2011-01-01

    Objective:To study the vascular smooth muscle relaxation effect and explicit the material base of Gastrodia elata Blume. Method:Tension recording for rat isolated aortic artery was used to study the effect of vascular smooth muscle relaxation. Column chromatography and mass spectrography and nuclear magnetic resonance spectrometry were used to isolate and identify the active compounds of G. elata Blume. Result: G. elata Blume.could significantly inhibit the smooth muscle constriction of isolated aortic artery induced by KCI. Five esterified phenolic compounds ( Ⅰ-Ⅴ ) were isolated from G. elata Blume, such as p-hydroxybenz aldehyde ( Ⅰ ); phydroxybenzyl methylether ( Ⅱ ); p-hydroxybenzyl alcohol ( Ⅲ ); 4,4'-dihydroxydiphenyl methane ( Ⅳ ); 4,4'-dihydroxydibenzyl ether( Ⅴ ). The results showed that the vascular smooth muscle relaxation effects were the result of the role of five compounds. Conclusion: The five esterified phenolic compounds ( Ⅰ - Ⅴ ) from G. elata Blume.play a combined role for vascular smooth muscle relaxation.%目的:观察天麻血管平滑肌松弛作用,明确其作用物质基础.方法:采用大鼠离体胸主动脉环灌流实验方法,对天麻血管平滑肌松弛作用进行考察,采用柱色谱法、光谱法(MS,NMR)对其活性成分进行了分离鉴定.结果:天麻能够显著抑制KCl引起的大鼠胸主动脉环收缩,从天麻乙酸乙酯提取物中分离并鉴定了5个酯溶性酚性成分(Ⅰ~Ⅴ):对羟基苯甲醛(Ⅰ)、对羟苄基甲醚(Ⅱ)、对羟基苯甲醇(Ⅲ)、4,4'-二羟基二苯基甲烷(Ⅳ),4,4-二羟基二苄醚(Ⅴ),明确了天麻的血管平滑肌松弛作用是这5个成分共同作用的结果.结论:天麻具有显著的血管平滑肌松弛作用,其作用主要由5个酯溶性酚性成分共同发挥.

  19. Erratum: Hayward IP, Bridle KR, Campbell GR, Underwood PA, Campbell JH (1995) Effect of Extracellular Matrix Proteins on Vascular Smooth Muscle Cell Phenotype. Cell Biology International 19: 839-846. doi: 10.1006/cbir.1995.1019.

    Science.gov (United States)

    2016-02-01

    The above article, published in print in Cell Biology International in September 1995 and online on 2 January 2013 in Wiley Online Library (http://onlinelibrary.wiley.com/doi/10.1006/cbir.1995.1019/abstract), is an exact duplicate of the article 'Effect of Extracellular Matrix Proteins on Vascular Smooth Muscle Cell Phenotype' (Hayward et al., ), which was published in the previous issue of Cell Biology International (Hayward et al., ). The duplicate publication is the result of an administrative error. The publisher apologises for any inconvenience.

  20. Compressive Elasticity of Three-Dimensional Nanofiber Matrix Directs Mesenchymal Stem Cell Differentiation to Vascular Cells with Endothelial or Smooth Muscle Cell Markers

    OpenAIRE

    Wingate, Kathryn; Bonani, Walter; Tan, Yan; Bryant, Stephanie J.; Tan, Wei

    2012-01-01

    The importance of mesenchymal stem cell (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. We utilized electrospinning and photopolymerization techniques to fabricate a 3D PEGdma nanofiber hydrogel matrix with a tunable elasticity for use as a cellular substrate. Compression testing demonstrated that the elastic modulus ...

  1. Poly(ADP-ribose) Polymerase 1 Is Indispensable for Transforming Growth Factor-β Induced Smad3 Activation in Vascular Smooth Muscle Cell

    OpenAIRE

    Dan Huang; Yan Wang; Lin Wang; Fengxiao Zhang; Shan Deng; Rui Wang; Yun Zhang; Kai Huang

    2011-01-01

    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 trans...

  2. 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, Cang-Bao; Zheng, Jian-Pu; Zhang, Wei;

    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...

  3. Effects of tetrandrine on phenotypic modulation of vascular smooth muscle cells and expression of p38 MAPK as well as MKP-1 after intimal injury of rabbit carotid arteries

    Institute of Scientific and Technical Information of China (English)

    Xinping Zhang; Lihong Xiang; Yibai Feng; Yongzhi Deng; Zhuolin Fu; Chunzhi Shi; Xiang Gu

    2006-01-01

    Objective: To study the effects of tetrandrine (Tet) on phenotypic modulation of vascular smooth muscle cells (VSMCs) and expression of p38 mitogen-activated protein kinase (p38MAPK) as well as mitogen-activated protein kinase phosphatase-1(MKP-1) after vascular intimal injury. Methods: HE staining was used to analyze vascular morphology of sham-injured group, injured group and Tet-treated group at day 28. Immunohistochemistry, Western blot and RT-PCR were respectively used to detect the expression change of smooth muscle α-actin (SMα-actin), proliferation cell nuclear antigen (PCNA), p38MAPK and MKP-1 of injured group and Tet neointimal area was significantly increased and the lumen area notably decreased in injured group at day 28. The neointimal proliferation in Tet treated group was less than that in injured group, and the lumen area of Tet group was significantly increased than that of injured group was no difference, and the neointimal proliferation condition was also basically as same as injured group at day 7 after injury. The expression of PCNA and p38MAKP in Tet group was obviously lower than that in injured group, and the expression of MKP-1 in Tet group was obviously higher than that in injured group at days 14 and 28 after injury. The expression of SMa-actin in Tet group was slightly higher than that in injured group at days 14 and 28 after injury. Conclusions: Tet could reduce neointimal proliferation by inhibiting VSMCs phenotypic modulation and p38MAPK signaling transduction pathway as well as its down regulation.

  4. 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...

  5. Adiponectin attenuates angiotensin II-induced vascular smooth muscle cell remodeling through nitric oxide and the RhoA/ROCK pathway.

    Directory of Open Access Journals (Sweden)

    Wared eNour-Eldine

    2016-04-01

    Full Text Available INTRODUCTION: Adiponectin (APN, an adipocytokine, exerts protective effects on cardiac remodeling, while angiotensin II (Ang II induces hypertension and vascular remodeling. The potential protective role of APN on the vasculature during hypertension has not been fully elucidated yet. Here, we evaluate the molecular mechanisms of the protective role of APN in the physiological response of the vascular wall to Ang II.METHODS AND RESULTS: Rat aortic tissues were used to investigate the effect of APN on Ang II-induced vascular remodeling and hypertrophy. We investigated whether nitric oxide (NO, the RhoA/ROCK pathway, actin cytoskeleton remodeling, and reactive oxygen species (ROS mediate the anti-hypertrophic effect of APN. Ang II-induced protein synthesis was attenuated by pre-treatment with APN, NO donor (SNAP, or cGMP. The hypertrophic response to Ang II was associated with a significant increase in RhoA activation and vascular force production, which were prevented by APN and SNAP. NO was also associated with inhibition of Ang II-induced phosphorylation of cofilin. In addition, immunohistochemistry revealed that 24 hr Ang II treatment increased the F- to G-actin ratio, an effect that was inhibited by SNAP. Ang II-induced ROS formation and upregulation of p22phox mRNA expression were inhibited by APN and NO. Both compounds failed to inhibit Nox1 and p47phox expression. CONCLUSIONS: Our results suggest that the anti-hypertrophic effects of APN are due, in part, to NO-dependent inhibition of the RhoA/ROCK pathway and ROS formation.

  6. Expression of c-Myc in Pulmonary Vascular Smooth Muscle Cells in the Development of Pulmonary Vascular Remodeling in Broilers Induced by Low Ambient Temperature%低温诱发肉鸡肺血管重塑过程中肺动脉平滑肌细胞c-Myc蛋白表达的变化

    Institute of Scientific and Technical Information of China (English)

    王建琳; 尹燕博

    2011-01-01

    The study was conducted to assess the expression of c-Myc in pulmonary vascular smooth muscle cells in the development of pulmonary vascular remodeling in broilers induced by low ambient temperature. 120 male broiler chicks (Arbor Acre) of commercial strain were randomly allocated to control group (raised at the temperature of (22 + 1. 5) °C ) and low temperature group (raised at the temperature of (11 + 2) °C) at 15 days old. Six broilers in each group were sampled every week from 15 to 50 days of age and lungs were paraffin-embedded, sectioned. The percentage of relative medial thickness (MT%) and the percentage of relative lumen area (RLA%) ,the indexes of pulmonary vascular remodeling,were examined by computer-image analyzing system. Positive indexes (PI) of c-Myc in pulmonary vascular smooth muscle cells were assessed. The result indicated that pulmonary vascular remodeling were significantly elevated in low temperature group from 36 days of age (P<0. 01 or P<0. 05). PI of c-Myc in pulmonary vascular smooth muscle cells were significantly higher than those in the control group from 29 days of age (P<0. 01). The study demonstrated that c-Myc in pulmonary vascular smooth muscle cells were expressed significantly induced by low ambient temperature and have a pivotal role in the development of pulmonary vascular remodeling.%试验旨在研究环境低温诱发肉鸡肺血管重塑过程中肺小动脉血管平滑肌细胞c-Myc蛋白的表达变化,初步探讨肉鸡肺血管重塑的发生机制.120只雄性AA商品代肉鸡15日龄时随机分为对照组((22±1.5)℃)条件下饲养)和低温组((11±2)℃条件下饲养).15~50日龄,每周每组随机取6只,取肺组织做石蜡切片,Weigert-间苯二酚复红染色,观察并测定m管重塑情况;采用免疫组织化学方法榆测肺动脉血管平滑肌细胞c-Myc蛋白表达,并对其进行半定量化.结果显示:(1)低温组肉鸡肺小动脉结构从36日龄开始较对照

  7. Baicalin inhibits PDGF-BB-stimulated vascular smooth muscle cell proliferation through suppressing PDGFRβ-ERK signaling and increase in p27 accumulation and prevents injury-induced neointimal hyperplasia

    Institute of Scientific and Technical Information of China (English)

    Li-Hua Dong; Jin-Kun Wen; Sui-Bing Miao; Zhenhua Jia; Hai-Juan Hu; Rong-Hua Sun; Yiling Wu; Mei Han

    2011-01-01

    The authors would like to clarify a deficiency in our paper recently published in Cell Research (CR) (2010;20:1252-1262).We did not reference the results in the first part of our paper reporting the effect of baicalin on vascular smooth muscle cells (VSMCs) in vitro which we had previously published in the Chinese language only Chinese Journal of Cell Biology (CJCB) (2010; 32(1):91-96); the overlap includes the re-use of some western blot data from the CJCB paper (including those in upper panels of Figures 2D, 3A and 3C; and ICAM1 and VCAM-1 of Figure 5B).These results suggest that baicalin inhibits PDGF-BB-induced expression of genes related to cell proliferation and migration, and blocks cell cycle progression.

  8. Proofs concerning the existence, in the blood of hypertensive patients, of some serum factors influencing the vascular smooth muscle and the myocardium physiology.

    Science.gov (United States)

    Mocanu, M; Botea, S; Dragomir, C T

    1991-01-01

    Starting from the existence of some autoimmune diseases (i.e. bronchial asthma or miastenia gravis) we asked ourselves if some plasmatic factors do exist, influencing the receptor--mediator relations in cardiovascular system during some illnesses having unknown etiology, as arterial hypertension. For this reason, in this work was tested the hypothesis that, in some chronic cardiovascular diseases would exist factors circulating and affecting the functions of the cellular membranes of the arterial wall, particularly of the smooth muscle cells and myocardial cells. Our results show a significant modification of the calcium fluxes and of some neuromediators uptake at the hypertensive patients.

  9. Effects of advanced glycation end-products on the expression of parathyroid hormone related peptide and vascular calcification on vascular smooth muscle cells%糖基化终产物对人血管平滑肌细胞表达及分泌甲状旁腺激素相关肽和血管钙化的影响

    Institute of Scientific and Technical Information of China (English)

    张琴; 刘乃丰

    2014-01-01

    Objective:To investigate the effects of advanced glycation end-products(AGEs) on expression and secretion of parathyroid hormone related peptide(PTHrP)in vitro cultured human vascular smooth muscle cells (HVSMCs), and explore the related mechanism of PTHrP influencing vascular smooth muscle cells calcification . Methods:HVSMCs were treated with AGE-BSA of indicated concentration or non-glycated BSA for same periods . The calcium contents and activity of alkaline phosphatase of cells were analyzed by microplate reader ;PTHrP levels in the supernatant were detected by the enzyme-linked immunosorbent method .Real-time fluorescent quantitative reverse transcription polymerase chain reaction ( RT-PCR) was performed to detect the expressive of PTHrP , core- binding factor α1(cbfα1) in human and bone morphogenetic protein-2(BMP-2) in vascular smooth muscle cells . Results:AGE-BSA increased calcium deposition and activity of alkaline phosphatase in HVSMCs in dose-independent manners( P<0.05) , but reduced the secretion of PTHrP.Futhermore, the elevated AGE-BSA treatment on HVSMCs significantly enhanced the expression of cbfα1, BMP-2 and PTHrP, compared with the controls( P <0.05 ) .Conclusion: The expression and secretion of PTHrP on human vascular smooth muscle cells can be effected by AGEs , and PTHrP may induce deposition of calcium on vascular smooth muscle cells, thereby contributing to the vascular calcification .However, the related mechanism will be further explored .%目的:观察糖基化终末产物(AGEs)对人血管平滑肌细胞表达及分泌甲状旁腺激素相关肽(PTHrP)功能的影响,探讨PTHrP影响血管平滑肌细胞钙化发生的相关机制。方法:不同浓度的AGE-BSA分别与人血管平滑肌细胞孵育相同时间后,检测各组细胞钙含量及碱性磷酸酶( ALP)活性判断钙化程度;酶联免疫吸附法检测各组细胞PTHrP的分泌量;实时荧光定量逆转录聚合酶链反应检

  10. Secondary amines containing one aromatic nitro group: preparation, nitrosation, sustained nitric oxide release, and the synergistic effects of released nitric oxide and an arginase inhibitor on vascular smooth muscle cell proliferation.

    Science.gov (United States)

    Curtis, Brandon; Payne, Thomas J; Ash, David E; Mohanty, Dillip K

    2013-03-01

    Atherosclerosis, a leading cause of death worldwide, is associated with the excessive proliferation of vascular smooth muscle cells. Nitrogen monoxide, more commonly known as nitric oxide, inhibits this uncontrolled proliferation. Herein we report the preparation of two families of nitric oxide donors; beginning with the syntheses of secondary amine precursors, obtained through the reaction between 2 equiv of various monoamines with 2,4 or 2,6-difluoronitrobenzene. The purified secondary amines were nitrosated then subjected to a Griess reagent test to examine the slow and sustained nitric oxide release rate for each compound in both the absence and presence of reduced glutathione. The release rate profiles of these two isomeric families of NO-donors were strongly dependent on the number of side chain methylene units and the relative orientations of the nitro groups with respect to the N-nitroso moieties. The nitrosated compounds were then added to human aortic smooth muscle cell cultures, individually and in tandem with S-2-amino-6-boronic acid (ABH), a potent arginase inhibitor. Cell viability studies indicated a lack of toxicity of the amine precursors, in addition to anti-proliferative effects exhibited by the nitrosated compounds, which were enhanced in the presence of ABH. PMID:23375096

  11. Relationship Between Pulmonary Artery Smooth Muscle Cells and Mechanism of Hypoxia-induced Pulmonary Vascular Remodeling%肺动脉平滑肌细胞与低氧性肺血管重塑形成机制

    Institute of Scientific and Technical Information of China (English)

    张凌云

    2013-01-01

    低氧条件下肺血管收缩、重塑,继而导致肺血管的持续对抗,其中以中膜增厚为主的肺血管重塑是导致低氧性肺动脉高压持续不可逆性病理改变的重要因素.肺动脉平滑肌细胞是肺动脉中膜的主要构成部分,慢性缺氧条件下由于各种活性介质及细胞生长因子稳态的失衡,肺动脉平滑肌细胞聚集、增殖、肥大及分泌胞外基质;另外,肺动脉平滑肌细胞通过各种信号通路与内膜的内皮细胞及外膜的成纤维细胞相互作用,在低氧性肺血管重塑过程中起着至关重要的作用,本文将对肺动脉平滑肌细胞与低氧性肺血管重塑形成机制的最新研究概况作一综述.%Under conditions of hypoxia generalized vasoconstriction and remodeling of the pulmonary vascular leads to pulmonary vascular persistent resistance. The medial thickening is the main reason of pulmonary vascular remodeling and hypoxic pulmonary artery hypertension, pulmonary artery smooth muscle cells (PASMC) are the principal structure of media, and chronic hypoxia induces the imbalance of vasoactive substances and growth factors. Under this condition, the main medial thickening is believed to be attributable to proliferation, hypertrophy and increased accumulation of PASMC as well as expression of extracellular matrix proteins. Moreover, PASMC has an interaction with endothelial cell of intima and fibroblast of adventitia through multiple signal pathways and plays a crucial role in the development of pulmonary vascular remodeling. The article will make a summary of latest research on PASMC and mechanism of hypoxic pulmonary vascular remodeling.

  12. 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

    ), elevated levels of ET(B) receptor mRNA (quantitative real-time PCR), and protein expressions (immunohistochemistry and Western blotting). Intracellular signaling was studied with Western blotting and phosphoELISA; this revealed that DSP induced extracellular-regulated protein kinases 1 and 2 (ERK1/2), p38......, and nuclear factor-kappaB (NF-kappaB) phosphorylation within 3 h. Blocking ERK1/2, p38, or NF-kappaB activation by their specific inhibitors significantly attenuated the DSP-induced upregulation of ET(B) receptor-mediated contraction and both ET(B) receptor mRNA and protein expression. In addition......, dexamethasone abolished the DSP-induced upregulation of ET(B) receptor-mediated contraction. In conclusion, upregulation of ET(B) receptors by DSP in arterial smooth muscle cells involves activation of mitogen-activating protein kinases (ERK1/2 and p38) and the downstream transcriptional factor NF...

  13. Prevention of vascular smooth muscle cell proliferation and injury-induced neointimal hyperplasia by CREB-mediated p21 induction: An insight from a plant polyphenol.

    Science.gov (United States)

    Sun, Lan; Zhao, Rui; Zhang, Li; Zhang, Weiku; He, GuoRong; Yang, Shengqian; Song, Junke; Du, Guanhua

    2016-03-01

    Cyclic adenosine monophosphate (cAMP)/protein kinase A (PKA)/cAMP response element (CRE)-binding protein (CREB) signaling cascade negatively regulates platelet-derived growth factor BB (PDGF-BB)-induced smooth muscle cell (SMC) proliferation, which is a critical event in the initiation and development of restenosis and atherosclerotic lesions. Salvianolic acid A (SAA) is one of the most abundant polyphenols extracted from salvia. The aim of this study is to investigate whether SAA exerts an action on PDGF-BB-induced proliferation via cAMP/PKA/CREB mechanism. SAA blunts PDGF-BB-induced human umbilical artery smooth muscle cell (hUASMC) proliferation via p21 induction, as evidenced by its increased mRNA and protein expression levels. The SAA-induced upregulation of p21 involves the cAMP/PKA signaling pathway; a cAMP analog mimicked the effects of SAA and a specific cAMP/PKA inhibitor opposed these effects. SAA also activated CREB, including phosphorylation at Ser133, and induced its nuclear translocation. Deletion and mutational analysis of p21 promoters, co-immunoprecipitation, and western blot analysis showed that CRE is essential for SAA-induced p21 protein expression. Transfection of dominant-negative CREB (mutated Ser133) plasmids into hUASMCs attenuated SAA-stimulated p21 expression. SAA upregulated p21 expression and activated CREB in the neointima of balloon-injured arteries in vivo. Our results indicate that SAA promotes p21 expression in SMCs through the cAMP/PKA/CREB signaling cascade in vitro and prevents injury-induced neointimal hyperplasia.

  14. Efficient generation of smooth muscle cells from adipose-derived stromal cells by 3D mechanical stimulation can substitute the use of growth factors in vascular tissue engineering.

    Science.gov (United States)

    Parvizi, Mojtaba; Bolhuis-Versteeg, Lydia A M; Poot, André A; Harmsen, Martin C

    2016-07-01

    Occluding artery disease causes a high demand for bioartificial replacement vessels. We investigated the combined use of biodegradable and creep-free poly (1,3-trimethylene carbonate) (PTMC) with smooth muscle cells (SMC) derived by biochemical or mechanical stimulation of adipose tissue-derived stromal cells (ASC) to engineer bioartificial arteries. Biochemical induction of cultured ASC to SMC was done with TGF-β1 for 7d. Phenotype and function were assessed by qRT-PCR, immunodetection and collagen contraction assays. The influence of mechanical stimulation on non-differentiated and pre-differentiated ASC, loaded in porous tubular PTMC scaffolds, was assessed after culturing under pulsatile flow for 14d. Assays included qRT-PCR, production of extracellular matrix and scanning electron microscopy. ASC adhesion and TGF-β1-driven differentiation to contractile SMC on PTMC did not differ from tissue culture polystyrene controls. Mesenchymal and SMC markers were increased compared to controls. Interestingly, pre-differentiated ASC had only marginal higher contractility than controls. Moreover, in 3D PTMC scaffolds, mechanical stimulation yielded well-aligned ASC-derived SMC which deposited ECM. Under the same conditions, pre-differentiated ASC-derived SMC maintained their SMC phenotype. Our results show that mechanical stimulation can replace TGF-β1 pre-stimulation to generate SMC from ASC and that pre-differentiated ASC keep their SMC phenotype with increased expression of SMC markers. PMID:26989865

  15. Efficient generation of smooth muscle cells from adipose-derived stromal cells by 3D mechanical stimulation can substitute the use of growth factors in vascular tissue engineering.

    Science.gov (United States)

    Parvizi, Mojtaba; Bolhuis-Versteeg, Lydia A M; Poot, André A; Harmsen, Martin C

    2016-07-01

    Occluding artery disease causes a high demand for bioartificial replacement vessels. We investigated the combined use of biodegradable and creep-free poly (1,3-trimethylene carbonate) (PTMC) with smooth muscle cells (SMC) derived by biochemical or mechanical stimulation of adipose tissue-derived stromal cells (ASC) to engineer bioartificial arteries. Biochemical induction of cultured ASC to SMC was done with TGF-β1 for 7d. Phenotype and function were assessed by qRT-PCR, immunodetection and collagen contraction assays. The influence of mechanical stimulation on non-differentiated and pre-differentiated ASC, loaded in porous tubular PTMC scaffolds, was assessed after culturing under pulsatile flow for 14d. Assays included qRT-PCR, production of extracellular matrix and scanning electron microscopy. ASC adhesion and TGF-β1-driven differentiation to contractile SMC on PTMC did not differ from tissue culture polystyrene controls. Mesenchymal and SMC markers were increased compared to controls. Interestingly, pre-differentiated ASC had only marginal higher contractility than controls. Moreover, in 3D PTMC scaffolds, mechanical stimulation yielded well-aligned ASC-derived SMC which deposited ECM. Under the same conditions, pre-differentiated ASC-derived SMC maintained their SMC phenotype. Our results show that mechanical stimulation can replace TGF-β1 pre-stimulation to generate SMC from ASC and that pre-differentiated ASC keep their SMC phenotype with increased expression of SMC markers.

  16. Cigarette smoke extract promotes human vascular smooth muscle cell proliferation and survival through ERK1/2- and NF-κB-dependent pathways

    DEFF Research Database (Denmark)

    Chen, Qing-Wen; Edvinsson, Lars; Xu, Cang-Bao

    2010-01-01

    aortic smooth muscle cell (HASMC) cultures, and to explore the mitogen-activated protein kinase (MAPK)/extracellular signal-regulated protein kinase 1 and 2 (ERK1/2) and nuclear factor-kappaB (NF-κB) signal mechanisms involved. Serum-starved HASMCs were treated with DSPs for up to 48 h. DSPs promoted...... cell proliferation in a concentration-dependent manner from 0.05 to 0.2 μl/ml. Activation of ERK1/2 and NF-κB was seen after exposure to DSPs. This occurred in parallel with the increase in cell population, bromodeoxyuridine incorporation, and cyclinD1/cyclin-dependent kinase 4 expression. Blocking...... phosphorylation of ERK1/2 by MAPK inhibitors U0126 and PD98059, and inhibiting activation of NF-κB by IkappaB (IκB) kinase inhibitors wedelolactone or IMD-0354, abolished the DSP effects. However, either a p38 inhibitor (SB203580) or an inhibitor of lipopolysaccharide (polymyxin B), or nicotinic receptor blockers...

  17. Cigarette smoke extract promotes human vascular smooth muscle cell proliferation and survival through ERK1/2- and NF-κB-dependent pathways

    DEFF Research Database (Denmark)

    Chen, Qing-Wen; Edvinsson, Lars; Xu, Cang-Bao

    2010-01-01

    aortic smooth muscle cell (HASMC) cultures, and to explore the mitogen-activated protein kinase (MAPK)/extracellular signal-regulated protein kinase 1 and 2 (ERK1/2) and nuclear factor-kappaB (NF-¿B) signal mechanisms involved. Serum-starved HASMCs were treated with DSPs for up to 48 h. DSPs promoted...... cell proliferation in a concentration-dependent manner from 0.05 to 0.2 µl/ml. Activation of ERK1/2 and NF-¿B was seen after exposure to DSPs. This occurred in parallel with the increase in cell population, bromodeoxyuridine incorporation, and cyclinD1/cyclin-dependent kinase 4 expression. Blocking...... phosphorylation of ERK1/2 by MAPK inhibitors U0126 and PD98059, and inhibiting activation of NF-¿B by IkappaB (I¿B) kinase inhibitors wedelolactone or IMD-0354, abolished the DSP effects. However, either a p38 inhibitor (SB203580) or an inhibitor of lipopolysaccharide (polymyxin B), or nicotinic receptor blockers...

  18. Smooth manifolds

    CERN Document Server

    Sinha, Rajnikant

    2014-01-01

    This book offers an introduction to the theory of smooth manifolds, helping students to familiarize themselves with the tools they will need for mathematical research on smooth manifolds and differential geometry. The book primarily focuses on topics concerning differential manifolds, tangent spaces, multivariable differential calculus, topological properties of smooth manifolds, embedded submanifolds, Sard’s theorem and Whitney embedding theorem. It is clearly structured, amply illustrated and includes solved examples for all concepts discussed. Several difficult theorems have been broken into many lemmas and notes (equivalent to sub-lemmas) to enhance the readability of the book. Further, once a concept has been introduced, it reoccurs throughout the book to ensure comprehension. Rank theorem, a vital aspect of smooth manifolds theory, occurs in many manifestations, including rank theorem for Euclidean space and global rank theorem. Though primarily intended for graduate students of mathematics, the book ...

  19. Using FM4-64 FX to Lable Transport Vesicles of Rat Vascular Smooth Muscle Cells%利用FM4-64FX标记大鼠血管平滑肌细胞的囊泡运输

    Institute of Scientific and Technical Information of China (English)

    姜隽; 王勇; 李涛; 聂利霞

    2015-01-01

    囊泡运输是大分子物质进入细胞的途径,血管平滑肌细胞(vascular smooth muscle cells,VSMCs)与外界存在频繁的信息和物质交换,该研究通过标识内吞囊泡来研究VSMCs的囊泡运输.体外培养大鼠胸主动脉VSMCs,用血管紧张素Ⅱ(angiotensin Ⅱ,Ang Ⅱ)刺激,加入FM4-64FX短暂孵育后固定.通过免疫组化方法标记VSMCs血管紧张素Ⅱ 1型受体(angiotensin Ⅱ receptor type 1,AT1R),检测内吞囊泡和AR1R转运之间的关系.受到Ang Ⅱ的激活后,VSMC快速形成内吞囊泡,将AT1R转运至胞质;存在血管紧张素受体阻断剂(angiotensin receptor blocker,ARB)时,内吞囊泡数量少,AT1R较少进入胞质.通过FM4-64 FX对胞内囊泡进行标识可以显示VSMCs的大分子物质运输,可观察特定的分子在内吞囊泡上的分布和运输情况.%This study aims to investigate vascular smooth muscle cells (VSMCs) vesicle transport by endocytic vesicles labeling.Rat VSMCs from the thoracic aortic were cultivated in vitro.VSMCs were stimulated with angiotensin Ⅱ (Ang Ⅱ) and incubated with FM4-64 FX shortly.Then VSMCs were fixed with paraformaldehyde and marked with angiotensin Ⅱ type 1 receptor (AT1R) antibody by immunohistochemistry.With Ang Ⅱ stimu-lation,VSMCs rapid formed endocytic vesicles and with it,AT1R was transported into the cytoplasm.Presence of an angiotensin receptor blocker (ARB) inhibited the number of endocytic vesicles formation and less AT1R entered the cytoplasm.Macromolecules transport of VSMCs can be illustrated by labeling the intracellular vesicles with FM4-64 FX dye.With this method,early endocytosis of VSMCs when the external environment changed can be investigated.

  20. Enhanced expression of Gqα and PLC-β1 proteins contributes to vascular smooth muscle cell hypertrophy in SHR: role of endogenous angiotensin II and endothelin-1.

    Science.gov (United States)

    Atef, Mohammed Emehdi; Anand-Srivastava, Madhu B

    2014-07-01

    Vascular Gqα signaling has been shown to contribute to cardiac hypertrophy. In addition, angiotensin II (ANG II) was shown to induce vascular smooth muscle cell (VSMC) hypertrophy through Gqα signaling; however, the studies on the role of Gqα and PLC-β1 proteins in VSMC hypertrophy in animal model are lacking. The present study was therefore undertaken to examine the role of Gqα/PLC-β1 proteins and the signaling pathways in VSMC hypertrophy using spontaneously hypertensive rats (SHR). VSMC from 16-wk-old SHR and not from 12-wk-old SHR exhibited enhanced levels of Gqα/PLC-β1 proteins compared with age-matched Wistar-Kyoto (WKY) rats as determined by Western blotting. However, protein synthesis as determined by [(3)H]leucine incorporation was significantly enhanced in VSMC from both 12- and 16-wk-old SHR compared with VSMC from age-matched WKY rats. Furthermore, the knockdown of Gqα/PLC-β1 in VSMC from 16-wk-old SHR by antisense and small interfering RNA resulted in attenuation of protein synthesis. In addition, the enhanced expression of Gqα/PLC-β1 proteins, enhanced phosphorylation of ERK1/2, and enhanced protein synthesis in VSMC from SHR were attenuated by the ANG II AT1 and endothelin-1 (ET-1) ETA receptor antagonists losartan and BQ123, respectively, but not by the ETB receptor antagonist BQ788. In addition, PD98059 decreased the enhanced expression of Gqα/PLC-β1 and protein synthesis in VSMC from SHR. These results suggest that the enhanced levels of endogenous ANG II and ET-1 through the activation of AT1 and ETA receptors, respectively, and MAP kinase signaling, enhanced the expression of Gqα/PLC-β1 proteins in VSMC from 16-wk-old SHR and result in VSMC hypertrophy.

  1. Inhibition of VCAM-1 expression on mouse vascular smooth muscle cells by lobastin via downregulation of p38, ERK 1/2 and NF-κB signaling pathways.

    Science.gov (United States)

    Lee, Kyoungran; Yim, Joung-Han; Lee, Hong-Kum; Pyo, Suhkneung

    2016-01-01

    Atherosclerosis is a chronic inflammatory disease, the progression of which is associated with the increased expression of cell adhesion molecules on vascular smooth muscle cells (VSMCs). Lobastin is a new pseudodepsidone isolated from Stereocaulon alpinum, Antarctic lichen, which is known to have antioxidant and antibacterial activities. However, the nature of the biological effects of lobastin still remains unclear. In the present study, we examine the effect of lobastin on the expression of vascular cell adhesion molecules (VCAM-1) induced by TNF-α in the cultured mouse VSMC cell line, MOVAS-1. Pretreatment of VSMCs for 2 h with lobastin (0.1-10 μg/ml) concentration-dependently inhibited TNF-α-induced protein expression of VCAM-1. Lobastin also inhibited TNF-α-induced production of intracellular reactive oxygen species (ROS). Lobastin abrogated TNF-α-induced phosphorylation of p38 and ERK 1/2, but not JNK, and also inhibited TNF-α-induced NK-κB activation. In addition, lobastin suppressed TNF-α-induced IκB kinase activation, subsequent degradation of IκBα and nuclear translocation of p65 NF-κB. Our results indicate that lobastin downregulates the TNF-α-mediated induction of VCAM-1 in VSMC by inhibiting the p38, ERK 1/2 and NF-κB signaling pathways and intracellular ROS generation. Thus, lobastin may be an important regulator of inflammation in the atherosclerotic lesion and a novel therapeutic drug for the treatment of atherosclerosis.

  2. Expression of GRP78 and caspase-12 in aortic vascular smooth muscle cells of hypertension rats and effects of enalapril on vascular remodeling%高血压大鼠血管平滑肌细胞GRP78和caspase-12表达的变化及依那普利的干预作用

    Institute of Scientific and Technical Information of China (English)

    郭丽; 赵连友; 刘静; 张志敏; 李雪; 丁璐

    2013-01-01

    AIM: To examine the effect of endoplasmic reticulum stress (ERS) on vascular remodeling induced by hypertension and to observe the effects of enalapril on the expressions of the correlation factors GRP78 and caspase-12 in vascular smooth muscle cells (VSMCs) and their relationship with vascular changes. METHODS: Forty adult male SD rats were randomly divided into sham operation group, sham + Ena group, AAB group and the AAB + Ena group. Four weeks later, MBP was measured through carotid artery incubation, the aortic media thickness was measured by image analyses software and the expressions of GRP78 and caspase-12 in the vascular smooth muscle cells were examined by immunohisto-chemistry. RESULTS: MAP of AAB group rats was significantly higher than those of rats in sham operation group, sham + Ena group and AAB + Ena group (P < 0. 05). The aortic media thickness of AAB group rats was significantly thicker than those of rats in sham operation group, sham + Ena group and the AAB + Ena group (P <0. 05). The expressions of GRP78 and caspase-12 of AAB group rats were significantly higher than those of rats in sham group, sham +Ena group and AAB +Ena group. CONCLUSION: High blood pressure resulting from abdominal aortic bounding causes endoplasmic reticulum stress response of vascular smooth muscle cells. Enalapril may exert some protective effects on VSMCs by inhibiting the endoplasmic reticulum stress via downregulation of the expression of caspase-12.%目的:观察高血压大鼠动脉血管平滑肌细胞(VSMCs)内质网应激(ERS)相关因子葡萄糖调节蛋白78(GRP78)和半胱氨酸门冬氨酸特异性蛋白酶12(caspase-12)表达的变化,并探讨高血压时VSMCs中ERS的分子机制,以及依那普利(enalapril,Ena)对血管重构的影响.方法:将40只健康雄性SD大鼠随机分为4组,即假手术(sham)组、sham+ Ena组,腹主动脉缩窄术(AAB)组及AAB+ Ena组,每组10只.4周后,通过颈动脉插管法测定大鼠血压和利用图像分析系

  3. Icariin inhibits oxidized low-density lipoprotein-induced proliferation of vascular smooth muscle cells by suppressing activation of extracellular signal-regulated kinase 1/2 and expression of proliferating cell nuclear antigen.

    Science.gov (United States)

    Hu, Yanwu; Liu, Kai; Yan, Mengtong; Zhang, Yang; Wang, Yadi; Ren, Liqun

    2016-03-01

    Icariin, a flavonoid isolated from the traditional Chinese herbal medicine Epimedium brevicornum Maxim, has been shown to possess anti-inflammatory, anti‑oxidant and anti-atherosclerotic activities in vivo and in vitro. The aim of the present study was to investigate the effects of icariin on oxidized low‑density lipoprotein (ox-LDL)-induced proliferation of vascular smooth muscle cells (VSMCs) and the possible underlying mechanism. VSMCs were cultured and pre‑treated with various concentrations of icariin (0, 10, 20 or 40 µm) prior to stimulation by ox‑LDL (50 µg/ml). Cell proliferation was evaluated by an MTT assay. Flow cytometry was used to study the influence of icariin on the cell cycle. Proliferating cell nuclear antigen (PCNA) expression and phosphorylation levels of extracellular signal-regulated kinase (ERK)1/2 were detected by western blot analysis. The results indicated that icariin significantly inhibited ox‑LDL‑induced proliferation of VSMCs and phosphorylation of ERK1/2. Furthermore, icariin also blocked the ox‑LDL‑induced cell‑cycle progression at G1/S‑interphase and downregulated the expression of PCNA in VSMCs. In conclusion, the present study indicated for the first time that icariin reduced the amount of ox‑LDL‑induced proliferation of VSMCs through suppression of PCNA expression and inactivation of ERK1/2.

  4. Smooth magnetogenesis

    CERN Document Server

    Campanelli, L

    2016-01-01

    In the Ratra scenario of inflationary magnetogenesis, the kinematic coupling between the photon and the inflaton undergoes a nonanalytical jump at the end of inflation. Using smooth interpolating analytical forms of the coupling function, we show that such unphysical jump does not invalidate the main prediction of the model, which still represents a viable mechanism for explaining cosmic magnetization. Nevertheless, there is a spurious result associated with the nonanaliticity of the coupling, to wit, the prediction that the spectrum of created photons has a power-law decay in the ultraviolet regime. This issue is discussed using both semiclassical approximation and smooth coupling functions.

  5. The Effect of Sulfur Dioxide on The Potassium Channel of Rat Vascular Smooth Muscle Cells%SO2对胸主动脉血管平滑肌细胞钾离子通道的影响

    Institute of Scientific and Technical Information of China (English)

    武文杰; 赵玲; 杜正清

    2011-01-01

    为了探讨二氧化硫(SO2)引起大鼠血管平滑肌的降压机制,采用急性酶分离法分离大鼠单个血管平滑肌细胞,运用全细胞膜片钳技术记录平滑肌细胞外向钾电流(IKv),观察SO2及其衍生物对平滑肌细胞膜钾电流的作用,从离子通道角度研究SO2对血压的影响.结果发现:SO2衍生物可使外向IKv显著增大,10 μmol/L SO2衍生物可使电流-电压曲线(I-V曲线)显著上移,即增大IKv,且呈一定的电压依赖性,并且,SO2衍生物可使IKv增大呈现出剂量-效应关系.当使用5 mmol/L 4-氨基吡啶(4-AP)抑制IKv后,加入10μmol/L SO2衍生物,IKv有一定程度增加.TEA能抑制SO2衍生物对IKv的增大效应.10μmol/L SO2衍生物可使IKv的激活曲线显著向超极化方向移动,但并不影响其斜率因子.说明SO2衍生物作用于血管平滑肌细胞,可引起外向钾电流幅度增大,使钾电流提前激活,这是SO2及其衍生物降压的作用机制之一;TEA、4AP对SO2衍生物引起的血管平滑肌细胞钾电流的增大具有拮抗作用.%To investigate the depressing blood pressure mechanism of the sulfur dioxide (SO2) in vascular smooth muscle cells, we studied the outward potassium current (IKv) with whole-cell configuration patch clamp technique in these cells which were obtained through acute enzyme separation method. The results show as follows: SO2 derivatives significantly increased outward IKv which was in voltage-dependent and dosedependent manner. 10 μmol/L SO2 derivative made I-V curves significantly upward. 10 μmol/L SO2 derivatives evaluated IKv which inhabited by 5mmol/L 4-AP. TEA inhibits which SO2 derivatives induced the augmentation of IKv. Moreover, the sulfur dioxide derivatives at 10 μmol/L caused the activation curve to hyperpolarization without changing the slope coefficient. In conclusion, SO2 derivatives increased outward IKv in vascular smooth muscle cells which advanced the activation of potassium current. This may be one

  6. PDTC, metal chelating compound, induces G1 phase cell cycle arrest in vascular smooth muscle cells through inducing p21Cip1 expression: involvement of p38 mitogen activated protein kinase.

    Science.gov (United States)

    Moon, Sung-Kwon; Jung, Sun-Young; Choi, Yung-Hyun; Lee, Young-Choon; Patterson, Cam; Kim, Cheorl-Ho

    2004-02-01

    Pyrrolidine dithiocarbamate (PDTC), a metal chelating compound, is known to induce cell death in vascular smooth muscle cells (VSMC). However, the molecular mechanism for PDTC-induced VSMC death is not well understood. Addition of PDTC reduced cell growth and DNA synthesis on VSMC in low density conditions. However, in serum depleted medium, PDTC did not affect the cell viability, suggesting that certain factors in serum may mediate the cytotoxic effect of PDTC. Several metal chelators prevented the cell death induced by PDTC. In a serum-deprived condition, addition of exogenous metals, copper, iron, and zinc, restored the cytotoxic effect of PDTC. These data indicate that metals such as copper, iron, and zinc in serum may mediate the cytotoxic effect of PDTC. At low VSMC density in 10% FBS, treatment of PDTC, which induced a cell-cycle block in G1-phase, induced down-regulation of cyclins and CDKs and up-regulation of the CDK inhibitor p21 expression, whereas up-regulation of p27 or p53 by PDTC was not observed. Finally, we determined PDTC-mediated signaling pathway involved in VSMC death. Among relevant pathways, PDTC induced marked activation of p38MAPK and JNK. Expression of dominant negative p38MAPK and SB203580, a p38MAPK specific inhibitor, blocked PDTC-dependent p38MAPK, growth inhibition, and p21 expression. These data demonstrate that the p38MAPK pathway participates in p21 induction, which consequently leads to decrease of cyclin D1/cdk4 and cyclin E/cdk2 complexes and PDTC-dependent VSMC growth inhibition. In conclusion, an understanding of the molecular mechanisms of PDTC in VSMC provides a theoretical basis for clinical approaches using antioxidant therapies in atherosclerosis. PMID:14603533

  7. Altered phosphodiesterase 3-mediated cAMP hydrolysis contributes to a hypermotile phenotype in obese JCR:LA-cp rat aortic vascular smooth muscle cells: implications for diabetes-associated cardiovascular disease.

    Science.gov (United States)

    Netherton, Stuart J; Jimmo, Sandra L; Palmer, Daniel; Tilley, Douglas G; Dunkerley, Heather A; Raymond, Daniel R; Russell, James C; Absher, P Marlene; Sage, E Helene; Vernon, Robert B; Maurice, Donald H

    2002-04-01

    Cardiovascular diseases represent a significant cause of morbidity and mortality in diabetes. Of the many animal models used in the study of non-insulin-dependent (type 2) diabetes, the JCR:LA-cp rat is unique in that it develops insulin resistance in the presence of obesity and manifests both peripheral and coronary vasculopathies. In this animal model, arterial vascular smooth muscle cells (VSMCs) from homozygous obese (cp/cp) rats, but not from age-matched healthy (+/+ or + /cp, collectively defined +/?) littermates, display an " activated" phenotype in vitro and in vivo and have an elevated level of cAMP phosphodiesterase (PDE) activity. In this report, we confirm that cp/cp rat aortic VSMCs have an elevated level of PDE3 activity and show that only particulate PDE3 (PDE3B) activity is elevated. In marked contrast to results obtained in + /? VSMCs, simultaneous activation of adenylyl cyclase and inhibition of PDE3 activity in cp/cp VSMCs synergistically increased cAMP. Although PDE3 inhibition did not potentiate the antimigratory effects of forskolin on +/? VSMCs, PDE3 inhibition did markedly potentiate the forskolin-induced inhibition of migration of cp/cp-derived VSMCs. Although PDE3 activity was elevated in cp/cp rat aortic VSMCs, levels of expression of cytosolic PDE3 (PDE3A) and PDE3B in +/? and cp/cp VSMCs, as well as activation of these enzymes following activation of the cAMP-protein kinase A signaling cascade, were not different. Our data are consistent with an increased role for PDE3 in regulating cAMP-dependent signaling in cp/cp VSMCs and identify PDE3 as a cellular activity potentially responsible for the phenotype of cp/cp VSMCs.

  8. The mineralocorticoid receptor-p38MAPK-NFκB or ERK-Sp1 signal pathways mediate aldosterone-stimulated inflammatory and profibrotic responses in rat vascular smooth muscle cells

    Institute of Scientific and Technical Information of China (English)

    Chuan-jiang ZHU; Qing-qing WANG; Jun-lan ZHOU; Han-zhi LIU; Fang HUA; Hong-zheng YANG; Zhuo-wei HU

    2012-01-01

    Aim:To explore the signalling pathways involved in aldosterone-induced inflammation and fibrosis in rat vascular smooth muscle cells (VSMCs).Methods:Using Western blotting and real-time RT-PCR,we investigated the effects of aldosterone on the expression of cyclooxygenase-2 (Cox-2) and IL-6,two important proinflammatory factors,and TGFβ1,a critical profibrotic factor,in VSMCs.Results:Aldosterone treatment significantly increased the expression of Cox-2 and IL-6 and activation of p38MAPK and NF-κB.The expression of both Cox-2 and IL-6 could be blocked by the mineralocorticoid receptor (MR) antagonist spironolactone and the p38MAPK inhibitor SB203580.Also,the rapid phosphorylation of p38MAPK could be suppressed by SB203580 but not by spironolactone,implicating in nongenomic effects of aldosterone.Similar to SB203580 and spironolactone,the NF-κB inhibitor α-p-tosyl-L-lysine chloromethyl ketone (TLCK) markedly attenuated expression of Cox-2,indicating that MR,p38MAPK and NF-KB are associated with aldosterone-induced inflammatory responses.Furthermore,aldosterone enhanced expression of TGFβ1 in rat VSMCs.This result may be related to activation of the MR/ERK-Sp1 signalling pathway because PD98059,an ERK1/2 inhibitor,significantly blocked the rapid phosphorylation of ERK1/2 and function of Sp1 and led to reduced expression of TGFβ1.Spironolactone was also shown to significantly inhibit TGFβ1 and Sp1 expression but not ERK1/2 phosphorylation.Conclusion:These results suggest that aldosterone-induced inflammatory responses and fibrotic responses may be mediated by the MR/p38MAPK-NF-κB pathways and the MR/ERK-Sp1 pathways in VSMCs,respectively.

  9. Vascular Stem/Progenitor Cell Migration Induced by Smooth Muscle Cell-Derived Chemokine (C-C Motif) Ligand 2 and Chemokine (C-X-C motif) Ligand 1 Contributes to Neointima Formation.

    Science.gov (United States)

    Yu, Baoqi; Wong, Mei Mei; Potter, Claire M F; Simpson, Russell M L; Karamariti, Eirini; Zhang, Zhongyi; Zeng, Lingfang; Warren, Derek; Hu, Yanhua; Wang, Wen; Xu, Qingbo

    2016-09-01

    Recent studies have shown that Sca-1(+) (stem cell antigen-1) stem/progenitor cells within blood vessel walls may contribute to neointima formation, but the mechanism behind their recruitment has not been explored. In this work Sca-1(+) progenitor cells were cultivated from mouse vein graft tissue and found to exhibit increased migration when cocultured with smooth muscle cells (SMCs) or when treated with SMC-derived conditioned medium. This migration was associated with elevated levels of chemokines, CCL2 (chemokine (C-C motif) ligand 2) and CXCL1 (chemokine (C-X-C motif) ligand 1), and their corresponding receptors on Sca-1(+) progenitors, CCR2 (chemokine (C-C motif) receptor 2) and CXCR2 (chemokine (C-X-C motif) receptor 2), which were also upregulated following SMC conditioned medium treatment. Knockdown of either receptor in Sca-1(+) progenitors significantly inhibited cell migration. The GTPases Cdc42 and Rac1 were activated by both CCL2 and CXCL1 stimulation and p38 phosphorylation was increased. However, only Rac1 inhibition significantly reduced migration and p38 phosphorylation. After Sca-1(+) progenitors labeled with green fluorescent protein (GFP) were applied to the adventitial side of wire-injured mouse femoral arteries, a large proportion of GFP-Sca-1(+) -cells were observed in neointimal lesions, and a marked increase in neointimal lesion formation was seen 1 week post-operation. Interestingly, Sca-1(+) progenitor migration from the adventitia to the neointima was abrogated and neointima formation diminished in a wire injury model using CCL2(-/-) mice. These findings suggest vascular stem/progenitor cell migration from the adventitia to the neointima can be induced by SMC release of chemokines which act via CCR2/Rac1/p38 and CXCR2/Rac1/p38 signaling pathways. Stem Cells 2016;34:2368-2380.

  10. Palmitic acid exerts pro-inflammatory effects on vascular smooth muscle cells by inducing the expression of C-reactive protein, inducible nitric oxide synthase and tumor necrosis factor-α.

    Science.gov (United States)

    Wu, Di; Liu, Juntian; Pang, Xiaoming; Wang, Shuyue; Zhao, Jingjing; Zhang, Xiaolu; Feng, Liuxin

    2014-12-01

    Atherosclerosis is a chronic inflammatory disease in the vessel, and inflammatory cytokines play an important role in the inflammatory process of atherosclerosis. A high level of free fatty acids (FFAs) produced in lipid metabolism disorders are known to participate in the formation of atherosclerosis through multiple bioactivities. As the main saturated fatty acid in FFAs, palmitic acid stimulates the expression of inflammatory cytokines in macrophages. However, it is unclear whether palmitic acid exerts a pro-inflammatory effect on vascular smooth muscle cells (VSMCs). The purpose of the present study was to observe the effect of palmitic acid on the expression of C-reactive protein (CRP), tumor necrosis factor α (TNF-α) and inducible nitric oxide synthase (iNOS) in VSMCs. Rat VSMCs were cultured, and palmitic acid was used as a stimulant for CRP, TNF-α and iNOS expression. mRNA expression was assayed with reverse transcription-polymerase chain reaction, and protein expression was detected with western blot analysis and immunocytochemistry. The results showed that palmitic acid significantly stimulated mRNA and protein expression of CRP, TNF-α and iNOS in VSMCs in time- and concentration-dependent manners, and therefore, palmitic acid is able to exert a pro-inflammatory effect on VSMCs via stimulating CRP, TNF-α and iNOS expression. The findings provide a novel explanation for the direct pro-inflammatory and atherogenic effects of palmitic acid, and for the association with metabolic syndrome, such as type 2 diabetes mellitus, obesity and atherosclerosis. Therefore, the intervention with anti-inflammatory agents may effectively delay the formation and progression of atherosclerosis in patients with metabolic syndrome.

  11. Cyclic strain amplitude dictates the growth response of vascular smooth muscle cells in vitro: role in in-stent restenosis and inhibition with a sirolimus drug-eluting stent.

    Science.gov (United States)

    Colombo, Alberto; Guha, Shaunta; Mackle, Joseph N; Cahill, Paul A; Lally, Caitríona

    2013-08-01

    The putative effects of changes in mean strain and cyclic strain amplitude on vascular smooth muscle cell (vSMC) growth (proliferation and apoptosis) were examined. Subsequently, a quantitative measure of vSMC growth was obtained to determine the prolonged effect of changes in mechanical burden following bare-metal stent (BMS) and sirolimus drug-eluting stent (DES) deployment in vitro. Bovine aortic vSMCs were exposed to prolonged cyclic strain using a Flexercell(TM) Tension system and a novel Sylgard(TM) phantom vessel following stent implantation before the level of vSMC proliferation and apoptosis was assessed by FACS analysis, cell counting, and immunocytochemistry. Physiological cyclic strain (5%) decreased vSMC proliferation and increased apoptosis in a temporal manner. There was no significant difference in cell growth following exposure to varying mean strains with similar amplitude. In contrast, exposure to varying strain amplitudes with similar mean strains resulted in significant differences in cell proliferation and apoptosis. In parallel studies, the level of vSMC proliferation and cell survival was significantly increased within low amplitude, high mean strain regions of a phantom vessel following BMS implantation when compared to regions of higher strain amplitude upstream and downstream of the stent, respectively. Moreover, the level of vSMC growth within the stented region was significantly attenuated following implantation of a sirolimus-coated DES independent of significant changes in cell survival. Cyclic strain amplitude is an important regulator of vSMC growth capacity within a stent and is a target for inhibition using a sirolimus-coated DES.

  12. The present studies of the role of the aortic vascular smooth muscle apoptosis and Hippo-YAP signaling pathway in the development of aortic dissection%主动脉血管平滑肌细胞凋亡和Hippo-YAP信号通路作用于主动脉夹层发病的研究进展

    Institute of Scientific and Technical Information of China (English)

    姜文剑; 兰峰; 张宏家

    2016-01-01

    Aortic dissection is a kind of fatal cardiovascular disease, and the apoptosis of aortic vascular smooth muscle plays an important role in aortic dissection.The new discovered Hippo-YAP signal transduction pathway is significant in regulation of the function of vascular smooth muscle, and it can cause many cardiovascular diseases.This paper aims to review the present studies of the mechanism of the apoptosis of aortic vascular smooth muscle and Hippo-YAP signaling pathway in the pathogenesis of cardiovascular diseases, especially the aortic dissection.%主动脉夹层是一种致命的心血管疾病,主动脉血管平滑肌的凋亡是造成主动脉夹层发生的重要原因.而新近发现的Hippo-YAP信号传导通路在血管平滑肌的功能调节中发挥重要作用,进而造成许多心血管疾病的发生.本文旨在回顾现有的关于主动脉血管平滑肌凋亡和Hippo-YAP信号传导通路在心血管疾病,特别是主动脉夹层发病中作用机制的研究.

  13. Green tea polyphenols inhibit low-density lipoprotein-induced proliferation of rat vascular smooth muscle cells%绿茶多酚抑制LDL诱导的血管平滑肌细胞增殖

    Institute of Scientific and Technical Information of China (English)

    欧阳平; 彭文烈; 赖文岩; 徐安龙

    2004-01-01

    The proliferation and migration of vascular smooth muscle cells (VSMCs) is one of the major mechanisms of intimal thickening in atherosclerosis and post-angioplasty restenosis. Elevated plasma levels of low-density lipoprotein (LDL) have been implicated in the pathogenesis of atherosclerotic vascular diseases. The purpose of this study was to determine the effects of green tea polyphenols on the proliferation and p44/42 mitogen-activated protein kinase (MAPK) activity in rat VSMCs simulated by native LDL. Rat aortic VSMCs were cultured and treated with LDL (100 μg/ml) in the absence or presence of green tea polyphenols, and the cell proliferation was subsequently quantified by non-radioactive MTS/PES assay and the cell cycle analyzed by flow cytometry. The p44/42 MAPK activity was evaluated by immunoblotting using anti-p44/42 phospho-MAPK antibody. Compared with the cells without polyphenol treatment, the proliferation of the VSMCs induced by LDL was dose-dependently inhibited by green tea polyphenols (P<0.05), with more numerous cells in G0/G1 phase (P<0.05) as shown by flow cytometry analysis. LDL significantly enhanced the p44/42 MAPK activity, an effect obviously inhibited by green tea polyphenols (at 100 μg/ml). These results suggest that green tea polyphenols can inhibit high levels of LDL-induced proliferation of phosphorylated p44/42 MAPK expression in rat VSMCs. Green tea polyphenols may, therefore,offer vascular protection by inhibiting VSMC growth in response to hypercholesterolemia.%观察绿茶多酚对低密度脂蛋白(LDL)刺激下离体大鼠胸主动脉血管平滑肌细胞增殖和p44/42MAPK表达的影响.体外培养大鼠胸主动脉血管平滑肌细胞,在LDL(100μg/ml)刺激时,应用不同剂量的绿茶多酚作用24 h后,采用MTS/PES法确定血管平滑肌细胞的增殖状态.应用流式细胞术测定细胞周期,p44/42磷酸化抗MAPK抗体的蛋白免疫印迹法测定MAPK蛋白表达.与未经LDL处理的平滑肌细胞相

  14. Vascular Cures

    Science.gov (United States)

    ... our CEO Board of Directors Scientific Advisory Board History of Vascular Cures Impact Contact Us Vascular Disease What is Vascular Disease? Education and Awareness Vascular Diseases Abdominal Aortic Aneurysm Aortic ...

  15. The Effects of the Mechanical Stretch on the Adhesion and Growth of Vascular Smooth Muscle Cells in vitro%机械拉伸对血管平滑肌细胞粘附及生长的影响

    Institute of Scientific and Technical Information of China (English)

    王红兵; 黄岂平; 卢晓; 秦建; 王远亮; 蔡绍皙

    2001-01-01

    通过自行研制的“四点弯曲梁”实验装置对血管平滑肌细胞(VSMC)加载培养,并结合显微形态观察和 计算机图像处理系统测量细胞铺展投影面积、微管吸吮实验系统检测细胞与表面的粘附力、α-肌动蛋白(actin) 免疫组化试验,了解细胞骨架发育和排列取向、流式细胞仪检测细胞动力学以及细胞生长行为等认识VSMC对 应变刺激的响应.发现VSMC粘附铺展与实验时间正相关,细胞粘附力、铺展面积、单位面积粘附力4 h后实 验组与对照组无显著性差异.VSMC内α-actin发育随加载时间延长呈增加趋势.细胞动力学检测实验组加载24 h后VSMC增殖活动受到抑制.VSMC可能通过调节细胞铺展行为、胞内应力纤维发育等主动机制,实现对机 械拉伸的适应性改建.应变刺激有利于体外培养的VSMC维持收缩表型.%An in vitro model was built for researching the effects of strain on vascular smooth muscle cells (VSMCs). The cultured VSMCs was stretched by four-support-bending-beam system, then the project area of cells was measured by computer-image-processing, the adhesion force was measured by micropipette-aspirating system, the α-actin of VSMCs was distinguished by immunocytochemistry and the dynamic of VSMCs was determined by FCM. The results show that: (1) The adhesion force of VSMCs is positively related to time. The adhesion force of unit area is indistinct after stretched for four hour. (2) The amount of α-actin increases with stretching time. (3) The proliferation of VSMCs is a little inhibited by stretched 24 h. These results suggest that the VSMCs in vitro could adjust their behavious to adapt the tension.

  16. 人参皂甙Rg3对血管平滑肌细胞衰老的影响及机制%Effect of ginsenoside Rg3 on ageing of vascular smooth muscle cells and its mechanism

    Institute of Scientific and Technical Information of China (English)

    鄢梦竹; 李书国

    2015-01-01

    目的 探讨人参皂甙Rg3对D-半乳糖诱导的血管平滑肌细胞(vascular smooth muscle cells,VSMC)衰老的影响及机制.方法 取SD大鼠胸主动脉中层,分离并原代培养VSMC,选取3~6代VSMC,通过D-半乳糖共孵育的方法诱导细胞衰老,β-半乳糖苷酶染色和透射电镜鉴定衰老VSMC;VSMC随机分成对照组、D-半乳糖组、Rg3高浓度组、Rg3低浓度组,Western blot方法检测p16、p21和p53蛋白的表达水平,流式细胞术检测细胞周期.结果 与对照组比较,D-半乳糖组%-半乳糖苷酶阳性细胞明显升高[(58.67±2.52)% vs (4.67±0.58)%,P<0.05].电镜下表现为细胞核膜内折,细胞线粒体肿胀,脂褐素堆积;p16、p21和p53蛋白表达水平明显升高(P<0.05),细胞周期停滞在G0/G1期.与D-半乳糖组比较,Rg3低浓度组和Rg3高浓度组β-半乳糖苷酶阳性细胞明显减少,p16、p21和p53蛋白表达水平明显降低,G0/G1期细胞数明显减少(P<0.05).结论人参皂甙Rg3可抑制VSMC衰老,其机制可能部分通过抑制细胞生长周期p16INK4a/Rb、p53-p21Cip1/Wan信号通路来实现.

  17. The amiodarone derivative 2-methyl-3-(3,5-diiodo-4-carboxymethoxybenzyl)benzofuran (KB130015) opens large-conductance Ca2+-activated K+ channels and relaxes vascular smooth muscle.

    Science.gov (United States)

    Gessner, Guido; Heller, Regine; Hoshi, Toshinori; Heinemann, Stefan H

    2007-01-26

    2-methyl-3-(3,5-diiodo-4-carboxymethoxybenzyl)benzofuran (KB130015) has been developed to retain the antiarrhythmic properties of the parent molecule amiodarone but to eliminate its undesired side effects. In patch-clamp experiments, KB130015 activated large-conductance, Ca2+-activated BK(Ca) channels formed by hSlo1 (alpha) subunits in HEK 293 cells. Channels were reversibly activated by shifting the open-probability/voltage (P(o)/V) relationship by about -60 mV in 3 muM intracellular free Ca2+ ([Ca2+]in). No effect on the single-channel conductance was observed. KB130015-mediated activation of BK(Ca) channels was half-maximal at 20 microM with a Hill coefficient of 2.8. BK(Ca) activation by KB130015 did not require the presence of Ca2+ and still occurred with saturating (100 microM) [Ca2+]in. Effects of the prototypic BK(Ca) activator NS1619 (1,3-dihydro-1-[2-hydroxy-5-(trifluoromethyl)phenyl]-5-(trifluoromethyl)-2H-benzimidazol-2-one) and those of KB130015 were not additive suggesting that both activators may at least partially share a common mechanism of action. KB130015-mediated activation was observed also for BK(Ca) channels from insects and for human BK(Ca) channels with already profoundly left-shifted voltage-dependence. In contrast, human intermediate conductance Ca2+-activated channels were inhibited by KB130015. Using segments of porcine pulmonary arteries, KB130015 induced endothelium-independent vasorelaxation, half-maximal at 43 microM KB130015. Relaxation was inhibited by 1 mM tetraethylammonium, suggesting that KB130015 can activate vascular smooth muscle type BK(Ca) channels under physiological conditions. Interestingly, the shift in the P(o)/V relationship was considerably stronger (-90 mV in 3 microM [Ca2+]in) for BK(Ca) channels containing Slo-beta1 subunits. Thus, KB130015 belongs to a novel class of BK(Ca) channel openers that exert an effect depending on the subunit composition of the channel complex.

  18. Vascular potassium channels in NVC.

    Science.gov (United States)

    Yamada, K

    2016-01-01

    It has long been proposed that the external potassium ion ([K(+)]0) works as a potent vasodilator in the dynamic regulation of local cerebral blood flow. Astrocytes may play a central role for producing K(+) outflow possibly through calcium-activated potassium channels on the end feet, responding to a rise in the intracellular Ca(2+) concentration, which might well reflect local neuronal activity. A mild elevation of [K(+)]0 in the end feet/vascular smooth muscle space could activate Na(+)/K(+)-ATPase concomitant with inwardly rectifying potassium (Kir) channels in vascular smooth muscle cells, leading to a hyperpolarization of vascular smooth muscle and relaxation of smooth muscle actin-positive vessels. Also proposed notion is endothelial calcium-activated potassium channels and/or inwardly rectifying potassium channel-mediated hyperpolarization of vascular smooth muscle. A larger elevation of [K(+)]0, which may occur pathophysiologically in such as spreading depression or stroke, can trigger a depolarization of vascular smooth muscle cells and vasoconstriction instead. PMID:27130411

  19. The NA+/K+-ATPase controls gap junctions via membrane microdomain interactions in rat smooth muscles.

    DEFF Research Database (Denmark)

    Matchkov, Vladimir; Nilsson, Holger; Aalkjær, Christian

    The Na+/K+-ATPase is known to interact with many membrane and cytosolic proteins by organizing various signaling complexes. These interactions were suggested to be important in regulation of various cellular responses. Pumping activity of the Na+/K+-ATPase is suggested to be essential for some...... in rat mesenteric small arteries. Paired cultured rat smooth muscle cells (A7r5) were used as a model for electrical coupling of SMC by measuring membrane capacitance (Cm). PCR, Western blotting and immunohistochemistry were used to identify the membrane transporters. SMCs were uncoupled (evaluated...... in regulation of the intercellular communication. We have here shown that gap junctions between SMCs are regulated through an interaction between the Na+/K+-ATPase and the Na+/Ca2+-exchanger leading to an increase in [Ca2+]i in discrete areas near the plasma membrane. We have also suggested that this Na...

  20. Effect of oleanolic acid on apoptosis of vascular smooth muscle cell induced by hydrogen peroxide%齐墩果酸对过氧化氢诱导血管平滑肌细胞凋亡的影响

    Institute of Scientific and Technical Information of China (English)

    冯健; 何国祥

    2012-01-01

    Objective To investigate the protective effect of oleanolic acid(OA) against vascular smooth muscle cells(VSMCs) apoptosis induced by hydrogen peroxideC H2 O2) and to explore the related mechanism. Methods Rat aortic VSMCs were isolated, cultured and divided into normal control group, H2O2 group, H2O2 + OA group and H2O2 + OA + wortmannin group. Results Compared with normal control group, H2O2 significantly induced VSMCs apoptosis and reduced the protein level of p-Akt(P<0. 05). Compared with H2O2 group,OA attenuated H2O2-induced apoptosis of VSMCs and up-regulated the protein level of p-Akt(P<0, 05). Compared with H2O2+ OA group, pretreatment of VSMCs with wortmannin could increase the apoptosis of VSMCs and reduce the protein level of p-Akt(P<0. 05). Conclusions OA can reduce the apoptosis of VSMCs induced by H2O2,which may be brought about through PI3K/Akt signaling pathway.%目的 研究齐墩果酸(OA)对H2O2诱导血管平滑肌细胞( VSMCs)凋亡的影响.方法 采用贴块法培养大鼠主动脉VSMCs,随机分为对照组、H2O2组、H2O2 +OA组、阻断荆组.采用Hoechst 33342染色和Annexin V/FITC染色检测各组细胞凋亡情况;Western blot检测各组细胞中磷酸化Akt蛋白表达变化.结果 与对照组比较,H2O2组细胞凋亡率明显升高,p-Akt蛋白表达明显降低(P<0.05);与H2O2组比较,H2O2+OA组细胞凋亡率明显降低,p-Akt蛋白表达明显升高(P<0.05);与H2O2 +OA组比较,阻断剂组细胞凋亡率明显升高,p-Akt蛋白表达明显降低(P<0.05).荧光显微镜显示,对照组细胞核呈蓝色,H2O2组细胞核呈致密浓染,H2O2+ OA组细胞核呈蓝染,有少量细胞核致密浓染,阻断刺组细胞核呈致密浓染.结论 OA能减轻H2O2导致的VSMCs凋亡,其机制可能与激活细胞内磷脂酰肌醇-3激酶/Akt信号通路引起下游促生存基因的表达有关.

  1. The role of hypertension-related gene in aortic vascular smooth muscle cells from mice and rats%高血压相关基因(HRG-1)在心血管病发病中的作用

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    目的研究高血压相关基因(HRG-1)在心血管病发病中的作用。 方法用RT-PCR和Northern blot分析HRG-1的表达。用3 H-TdR掺入和组织学方法检测血管平滑肌细胞(VSMC)的增殖。 结果 Northern blot分析显示,HRG-1 mRNA不仅可在VSMC中表达,亦可在大鼠多种组织中表达(心、脑、肺、肾和肝)。此外,在SHR大鼠的心、脑、肾和肝组织的HRG-1基因表达均明显低于正常WKY大鼠。半定量RT-PCR和组织学分析表明,ApoE敲除小鼠和再狭窄动物模型的HRG-1 mRNA表达减少,并且均可见到新生内膜形成。刺激VSMC增殖的ET, AII和IL-1可减少VSMC HRG-1 mRNA的表达。ANF, CGRP和Adm可抑制VSMC增殖,增加HRG-1 mRNA表达。这些作用能被相应的阻断剂或抗体所阻断或抑制。 结论 HRG-1是一种与VSMC增殖相关的基因,它在动脉粥样硬化,再狭窄和高血压等心血管疾病中可能具有重要作用。%Objective To study the role of hypertension-related gene (HRG-1) in cardiovascular disease. Methods The expression of HRG-1 was analyzed with RT-PCR and Northern blotting. Vascular smooth muscle cell (VSMC) proliferation was measured with 3 H-TdR incorporation and was confirmed with histological analysis. Results Northern blot analysis showed that HRG-1 mRNA was expressed not only in VSMC, but also in various rat tissues (heart, brain, lung, kidney, and liver). In addition, the expression of HRG-1 mRNA in heart, brain, kidney and liver of spontaneously hypertensive rat (SHR) was lower than that in the same tissues of Wistar-Kyotorat (WKY). Semi-quantitative RT-PCR and histological analysis showed that the expression of HRG-1 mRNA in ApoE-knockout mice and in animal models of restenosis was decreased and neointimal formation was observed in both models. ET, AII, and IL-1 stimulating VSMC proliferation reduced the expression of HRG-1 mRNA of VSMC. Atrial natriuretic factor (ANF), calcitonin gene-related peptide and

  2. PD98059抑制Peroxynitrite诱导人脑血管平滑肌细胞DDR2的表达%PD98059 inhibits Peroxynitrite-induced discoidin domain receptor-2 expression in human brain vascular smooth muscle cells

    Institute of Scientific and Technical Information of China (English)

    万小娟; 黄凌燕; 张国栋; 李建峰

    2012-01-01

    目的 探讨过氧亚硝酸盐(ONOO-)诱导人脑血管平滑肌细胞(HBVSMCs)盘状结构域受体2(DDR2)表达的机制.方法 体外培养HBVSMCs,用倒置相差显微镜、吖啶橙染色法和MTT比色法对PD98059预处理的细胞进行细胞形态学及细胞生存率检测,采用Western blot及Real-time PCR技术,检测ONOO-作用下HBVSMCs中DDR2表达及施加ERK1/2抑制剂PD98059后DDR2/MMP-9表达的变化.结果 10 μmol/L ONOO-诱导DDR2在蛋白及mRNA水平上呈现高表达(P<0.05),随着ONOO-浓度的增加,DDR2逐渐呈现低表达(P<0.05).PD98059预处理后的HBVSMCs,无明显形态学变化,对其进行生存率检测亦无显著影响(P>0.05).PD98059显著抑制ONOO-诱导的DDR2/MMP-9表达(P<0.05).结论 ERK1/2信号转导途径抑制剂PD98059可以抑制ONOO-诱导的DDR2/MMP-9表达,ONOO-诱导HBVSMCsDDR2的表达机制与ERK1/2信号转导途径有关.%Objective To explore the possible mechanism underlying peroxynitrite (ONOO ) -induced discoidin domain receptor-2 (DDR2) expression in human brain vascular smooth muscle cells (HBVSMCs). Methods HBVSMCs were cultured in vitro in different concentrations of ONOO- , and PD98059, a specific inhibitor of ERK1/2. Morphology and viability of HBVSMCs pretreated with PD98059 were detected by an inverted phase contrast microscope, acri-dine orange staining and MTT assay. Western blot and Real-time PCR were employed to determine expressions of DDR2/ MMP-9. Results 10 μmol/L of ONOO- significantly induced expression of DDR2 at both protein and mRNA levels (P0.05). Pretreatment on HBVSMCs with PD98059 significantly inhibited expressions of DDR2/ MMP-9 in HBVSMCs induced by 10 μmol/L of ONOO- (P < 0.05). Conclusion PD98059 inhibits ONOO- -induced DDR2 expression. The ERK1/2 signal transduction pathway might involve in ONOO -induced DDR2 expression in HBVSMCs.

  3. MicroRNAs in Vascular Biology

    OpenAIRE

    Munekazu Yamakuchi

    2012-01-01

    Vascular inflammation is an important component of the pathophysiology of cardiovascular diseases, such as hypertension, atherosclerosis, and aneurysms. All vascular cells, including endothelial cells (ECs) and vascular smooth muscle cells (VSMCs), and infiltrating cells, such as macrophages, orchestrate a series of pathological events. Despite dramatic improvements in the treatment of atherosclerosis, the molecular basis of vascular inflammation is not well understood. In the last decade, mi...

  4. 匹伐他汀对血管平滑肌细胞金属基质蛋白酶9表达的影响%Pitavastatin inhibits MMP-9 expression in vascular smooth muscle cells of human aorta

    Institute of Scientific and Technical Information of China (English)

    费宇行; 张蓉; 李晶; 胡明; 裘毅钢; 张沂

    2013-01-01

    (TNF-α) in vascular smooth muscle cells (VSMCs). Methods The atherosclerotic specimen from open-operation abdominal a-neurism was examined with routine and immunohistochemical stain. The specimen of control group was get from healthy adults died from accident or patients accepted partial aorta resection with non-vascular disease. The VSMCs were cultured from aortic tunica media of abdominal aneurism. The VSMCs in interventional groups were cultured with pitavastatin in different concentration (1 ng/ml, 10 ng/ml, 100 ng/ml, 500 ng/ml) before TNF-α stimulation. The expression of MMP-9 protein and mRNA were measured. Results It was showed that the distribution of TNF-α, MMP-9 were significantly augmented in AS group than control, and the distribution were centered at the area with inflammation cells. Pitavastatin inhibited the expression of MMP-9 protein and mRNA by a concentration-dependent manner which responsed to TNF-α significantly. The expression of MMP-9 protein was reduced 10.5%, 24.1%, 46.0% and 61.0% and mRNA 9%, 21%, 35% and 46% compared with control groups. The difference were significantly. Conclusion The distribution of MMP-9, TNF-α protein in AS tissue suggested that cy-tokine can influence each other. Pitavastatin preprocess suppressed TNF-α induced MMP-9 expression in VSMCs at both the mRNA and protein levels. These results indicate that pitavastatin can defer the progress of AS and to stabilize atherosclerotic plaque by inhibiting the expression of MMP-9, earlier intervention can provide better effect.

  5. Effects of X-ray radiation on cultured vascular smooth muscle cells%X射线对血管平滑肌细胞作用机制的实验研究

    Institute of Scientific and Technical Information of China (English)

    朱丽红; 申文江; 李平; 张强

    2001-01-01

    Objective  Proliferation of vascular smooth muscle cells (VSMC) represents an important event in vascular lesion formation. The aim of this investigation is to study the effect of X-ray irradiation on the viability of VSMC and the cellular mechanism, depending on which irradiation inhibits VSMC proliferation. Methods The different 8MV doses ranging from 2~20 Gy were delivered to cultured VSMC with 0 Gy serving as control. Cytometry, clonogenic assay and MTT calorimetric assay, eosin exclusion test, flow cytometry and the terminal deoxynucleotidyltransferase-mediated DUTP nick end labeling (TUNEL) technique were to determine the cell amount, proliferation, growth, death and aptosis rates of VSMC. According to these references, the VSMC irradiated repopulation model can be established. Aptosis cells were identified and quantified by the TUNEL and flow cytometry techniques. Results  ①The cell amount of VSMC began to decrease 24 hours after irradiation and to recover up to 72 hours(<0.05).The decrement of VSMC cellularity was proportionally related to the escalated dose effect. ②With MTT assay, the A570 value of VSMC decreased within 48 hours after irradiation and the value began to rise 72 hours later, faster in the 2Gy and 5Gy groups(P<0.05). ③ Eosin stain exclusion revealed the very high VSMC's death rate after irradiation of 10,15 and 20 Gy. The mortalities of VSMC in all groups came down to a lower level(P<0.05) 6 days later. ④ The VSMC survival data from the clonogenic assay were consistent with the mechanism of radiation-induced cell killing. ⑤ DNA hypodiploid peaks were found on the flow cytometry histograms and the TUNEL assay , revealing the presence of VSMC DNA fragmentation. Conclusions  It can be concluded that there are two mechanisms to inhibit VSMC proliferation and even result in death, lethal and/or sublethal cellular injury and apoptosis. Therefore, X-ray irradiation can effectively prevent restenosis after

  6. 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...

  7. Cardiac, Skeletal, and smooth muscle mitochondrial respiration

    DEFF Research Database (Denmark)

    Park, Song-Young; Gifford, Jayson R; Andtbacka, Robert H I;

    2014-01-01

    Unlike cardiac and skeletal muscle, little is known about vascular smooth muscle mitochondrial function. Therefore, this study examined mitochondrial respiratory rates in the smooth muscle of healthy human feed arteries and compared with that of healthy cardiac and skeletal muscle. Cardiac......, skeletal, and smooth muscle was harvested from a total of 22 subjects (53±6 yrs) and mitochondrial respiration assessed in permeabilized fibers. Complex I+II, state 3 respiration, an index of oxidative phosphorylation capacity, fell progressively from cardiac, skeletal, to smooth muscle (54±1; 39±4; 15......±1 pmol•s(-1)•mg (-1), pmitochondrial density, also fell progressively from cardiac, skeletal, to smooth muscle (222±13; 115±2; 48±2 umol•g(-1)•min(-1), p

  8. Roles of kallikrein-kinin system in vascular smooth muscle cell proliferation and migration%激肽释放酶-激肽系统在血管平滑肌细胞增殖和迁移中的作用

    Institute of Scientific and Technical Information of China (English)

    葛良; 刘玲; 杨昉; 张仁良

    2012-01-01

    Vascular smooth muscle cell (VSMC) proliferation and migation play important roles in the occurrence and development of atherosclerosis and restenosis after intravascular stenting.The studies in recent years have shown that kallikrein-kinin system (KKS) is closely correlated with VSMC proliferation and migration in cytokines and transduction pathways.Therefore,investigating the roles of KKS in the VSMC proliferation and migation process has great significance in clinical prevention and treatment of atherosclerosis and restenosis after intravascular stenting.%血管平滑肌细胞(vascular smooth muscle cell,VSMC)增殖和迁移在动脉粥样硬化以及血管内支架成形术后再狭窄的发生和发展中起着重要作用.近年来的研究显示,激肽释放酶-激肽系统(kallikrein-kinin system,KKS)在细胞因子和转导通路方面与VSMC增殖和迁移密切相关.因此,探讨KKS在VSMC增殖和迁移过程中的作用,对于临床防治动脉粥样硬化、血管内支架成形术后再狭窄具有十分重要的意义.

  9. [Vascular parkinsonism].

    Science.gov (United States)

    Marxreiter, F; Winkler, J

    2016-07-01

    Parkinsonism may result from cerebral vascular disorders that feature white matter lesions and small vessel pathology. Vascular Parkinsonism typically presents as lower body Parkinsonism with predominant gait impairment. Urinary incontinence and cognitive decline are additional features of the disease. There is a considerable overlap between vascular Parkinsonism and vascular dementia. We review the clinical characteristics of vascular Parkinsonism and discuss the current treatment approaches, as well as the role of brain imaging for the diagnostic workup. . PMID:27299942

  10. Effect of S-adenosylmethionine on vascular smooth cells proliferation and migration%S-腺苷甲硫氨酸对大鼠血管平滑肌细胞增生和迁移的影响

    Institute of Scientific and Technical Information of China (English)

    金成吉; 王晓梅; 段薇; 李湘; 曲建梅; 张弢; 刘璟瑶

    2012-01-01

    目的 观察S-