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Sample records for soluble vascular cell

  1. Soluble Vascular Cell Adhesion Molecule-1 (VCAM-1 as a Biomarker in the Mouse Model of Experimental Autoimmune Myocarditis (EAM.

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    U Grabmaier

    Full Text Available Vascular cell adhesion molecule-1 (VCAM-1 is strongly upregulated in hearts of mice with coxsackie virus-induced as well as in patients with viral infection-triggered dilated cardiomyopathy. Nevertheless, the role of its soluble form as a biomarker in inflammatory heart diseases remains unclear. Therefore, we investigated whether plasma levels of soluble VCAM-1 (sVCAM-1 directly correlated with disease activity and progression of cardiac dysfunction in the mouse model of experimental autoimmune myocarditis (EAM. EAM was induced by immunization of BALB/c mice with heart-specific myosin-alpha heavy chain peptide together with complete Freund`s adjuvant. ELISA revealed strong expression of cardiac VCAM-1 (cVCAM-1 throughout the course of EAM in immunized mice compared to control animals. Furthermore, sVCAM-1 was elevated in the plasma of immunized compared to control mice at acute and chronic stages of the disease. sVCAM-1 did not correlate with the degree of acute cardiac inflammation analyzed by histology or cardiac cytokine expression investigated by ELISA. Nevertheless, heart to body weight ratio correlated significantly with sVCAM-1 at chronic stages of EAM. Cardiac systolic dysfunction studied with positron emission tomography indicated a weak relationship with sVCAM-1 at the chronic stage of the disease. Our data provide evidence that plasma levels of sVCAM-1 are elevated throughout all stages of the disease but showed no strong correlation with the severity of EAM.

  2. Study of serum soluble vascular cell adhesion molecule-1 levels in type 2 diabetic patients with diabetic retinopathy

    International Nuclear Information System (INIS)

    Li Fangdu; Chu Qiaomei

    2002-01-01

    To study the change and the correlation of serum soluble vascular cell adhesion molecule-1 (sV-CAM-1) levels with diabetic retinopathy in type 2 diabetic patients, serum sVCAM-1 levels were measured in duplicate by ELISA in 85 type 2 diabetic patients; fundus examination was performed by an ophthalmologist using ophthalmoscope or fundus fluorescein angiography, and the findings were graded as: no signs of diabetic retinopathy (NDR), background diabetic retinopathy (BDR) and proliferative diabetic retinopathy (PDR). Serum sVCAM-1 levels were significantly higher in the PDR and BDR groups than those in the control and NDR groups respectively (P<0.01). NDR group showed significantly increased serum sVCAM-levels compared with control group (P<0.01). In contrast, serum sVCAM-1 levels were not related to the presence of blood glucose, serum insulin levels or known diabetic duration. Authors' results suggest that serum sVCAM-1 might be implicated in the development of the diabetic retinopathy, and could assess the severity of diabetic retinopathy. The measurement of serum sVCAM-1 levels in 2 type diabetic patients may be clinically useful for early diagnosis or treatment of diabetic retinopathy

  3. Alteration of protein expression pattern of vascular endothelial growth factor (VEGF from soluble to cell-associated isoform during tumourigenesis

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    Lertprasertsuke Nirush

    2005-10-01

    Full Text Available Abstract Background Vascular endothelial growth factor (VEGF is a potent mitogen for endothelial cells, and its expression has been correlated with increased tumour angiogenesis. Although numerous publications dealing with the measurement of circulating VEGF for diagnostic and therapeutic monitoring have been published, the relationship between the production of tissue VEGF and its concentration in blood is still unclear. The aims of this study were to determine: 1 The expression pattern of VEGF isoforms at the protein level in colorectal and lung adenocarcinoma in comparison to the pattern in corresponding adjacent normal tissues 2 The relationship between the expression pattern of VEGF and total level of circulating VEGF in the blood to clarify whether the results of measuring circulating VEGF can be used to predict VEGF expression in tumour tissues. Methods Ninety-four tissue samples were obtained from patients, 76 colorectal tumour tissues and 18 lung tumour tissues. VEGF protein expression pattern and total circulating VEGF were examined using western blot and capture ELISA, respectively. Results Three major protein bands were predominately detected in tumour samples with an apparent molecular mass under reducing conditions of 18, 23 and 26 kDa. The 18 kDa VEGF protein was expressed equally in both normal and colorectal tumour tissues and predominately expressed in normal tissues of lung, whereas the 23 and 26 kDa protein was only detected at higher levels in tumour tissues. The 18, 23 and 26 kDa proteins are believed to represent the VEGF121, the VEGF165 and the VEGF189, respectively. There was a significant correlation of the expression of VEGF165 with a smaller tumour size maximum diameter 189 with advanced clinical stage of colorectal tumours. The measurement of total circulating VEGF in serum revealed that cancer patients significantly (p Conclusion Our findings indicate that the expression patterns of VEGF isoforms are altered during

  4. Alteration of protein expression pattern of vascular endothelial growth factor (VEGF) from soluble to cell-associated isoform during tumourigenesis

    International Nuclear Information System (INIS)

    Cressey, Ratchada; Wattananupong, Onusa; Lertprasertsuke, Nirush; Vinitketkumnuen, Usanee

    2005-01-01

    Vascular endothelial growth factor (VEGF) is a potent mitogen for endothelial cells, and its expression has been correlated with increased tumour angiogenesis. Although numerous publications dealing with the measurement of circulating VEGF for diagnostic and therapeutic monitoring have been published, the relationship between the production of tissue VEGF and its concentration in blood is still unclear. The aims of this study were to determine: 1) The expression pattern of VEGF isoforms at the protein level in colorectal and lung adenocarcinoma in comparison to the pattern in corresponding adjacent normal tissues 2) The relationship between the expression pattern of VEGF and total level of circulating VEGF in the blood to clarify whether the results of measuring circulating VEGF can be used to predict VEGF expression in tumour tissues. Ninety-four tissue samples were obtained from patients, 76 colorectal tumour tissues and 18 lung tumour tissues. VEGF protein expression pattern and total circulating VEGF were examined using western blot and capture ELISA, respectively. Three major protein bands were predominately detected in tumour samples with an apparent molecular mass under reducing conditions of 18, 23 and 26 kDa. The 18 kDa VEGF protein was expressed equally in both normal and colorectal tumour tissues and predominately expressed in normal tissues of lung, whereas the 23 and 26 kDa protein was only detected at higher levels in tumour tissues. The 18, 23 and 26 kDa proteins are believed to represent the VEGF 121 , the VEGF 165 and the VEGF 189 , respectively. There was a significant correlation of the expression of VEGF 165 with a smaller tumour size maximum diameter <5 cm (p < 0.05), and there was a significant correlation of VEGF 189 with advanced clinical stage of colorectal tumours. The measurement of total circulating VEGF in serum revealed that cancer patients significantly (p < 0.001) possessed a higher level of circulating VEGF (1081 ± 652 pg/ml in

  5. Circulating levels of high-sensitivity C-reactive protein and soluble markers of vascular endothelial cell activation in growth hormone-deficient adolescents.

    Science.gov (United States)

    Lanes, Roberto; Marcano, Henry; Villaroel, Omar; Gunczler, Peter; Morillo, Edgar; Paoli, Mariela; Perez, Marvelys; Maulino, Nora; Palacios, Anselmo

    2008-01-01

    Significant endothelial dysfunction as determined by lower flow-mediated vasodilation of the brachial artery was recently reported by us in growth hormone-deficient (GHD) adolescents. The circulating concentrations of markers of vascular endothelial cell and platelet activation and their relationship to inflammatory markers have not been previously evaluated in this group of patients. To assess the relationship between circulating levels of high-sensitivity C-reactive protein (CRP) and soluble markers of vascular endothelial cell activation in GHD adolescents. Twenty-eight GHD children on GH treatment with a chronological age of 15.7 +/- 2.6 years and 16 untreated GHD adolescents with a chronological age of 16.6 +/- 3.3 years were studied. Concentrations of CRP, as an inflammatory marker, were measured in all patients and the association between CRP and the fasting soluble markers of vascular endothelial cell activation intercellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1), E-selectin and P-selectin levels was evaluated. Sixteen healthy adolescents with a mean chronological age of 15.1 +/- 2.2 years served as controls. CRP and P-selectin levels were significantly higher in untreated GHD adolescents than in treated GHD subjects or in healthy controls (p < 0.02), while VCAM-1 concentrations were increased in both untreated and treated GHD adolescents when compared to controls (p < 0.007). E-selectin and ICAM-1 levels were similar in all three groups. CRP was found to be associated with BMI (r: 0.62; p < 0.001), P-selectin (r: 0.43; p < 0.01), E-selectin (r: 0.27; p < 0.03), ICAM-1 (r: 0.23; p < 0.05) and VCAM-1 (r: 0.40; p < 0.001) concentrations in untreated GHD adolescents and with P-selectin (r: 0.88; p < 0.001) and E-selectin (r: 0.29; p < 0.01) in treated GHD subjects. A weak inverse association was observed in a subgroup of patients between brachial artery endothelium-dependent dilation and P-selectin (r: -0.56; p < 0.07). Low

  6. Soluble Vascular Cell Adhesion Molecule-1 (sVCAM-1) Is Elevated in Bronchoalveolar Lavage Fluid of Patients with Acute Respiratory Distress Syndrome.

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    Attia, Engi F; Jolley, Sarah E; Crothers, Kristina; Schnapp, Lynn M; Liles, W Conrad

    2016-01-01

    Pulmonary vascular endothelial activation has been implicated in acute respiratory distress syndrome (ARDS), yet little is known about the presence and role of endothelial activation markers in the alveolar space in ARDS. We hypothesized that endothelial activation biomarkers would be differentially expressed in bronchoalveolar lavage fluid from patients with ARDS compared with healthy volunteers, and that biomarker concentrations would be associated with ARDS severity. We performed a cross-sectional analysis of data from 26 intubated patients with ARDS undergoing evaluation for clinically suspected ventilator-associated pneumonia and five healthy volunteers. Patients underwent bronchoalveolar lavage a median of five days after intubation. Healthy volunteers also underwent bronchoalveolar lavage. Endothelial activation biomarkers (soluble vascular cell adhesion molecule-1 [sVCAM-1], soluble endothelial selectin [sESEL], angiopoietin-1 [Ang-1] and angiopoietin-2 [Ang-2]) were measured in bronchoalveolar lavage fluid. Clinically suspected ventilator-associated pneumonia was confirmed with microbiologic culture data. Patients with ARDS had significantly higher median sVCAM-1 concentrations in the bronchoalveolar lavage fluid compared with healthy volunteers (985 vs 119 pg/mL, p = 0.03). Additionally, there was a trend toward greater bronchoalveolar lavage fluid sVCAM-1 concentrations among patients with moderate/severe compared to mild ARDS (1395 vs 209 pg/mL, p = 0.06). We did not detect significant differences in bronchoalveolar lavage fluid levels of sESEL, Ang-1 or Ang-2 between patients with ARDS and healthy volunteers. Median bronchoalveolar lavage fluid biomarker levels did not differ between patients with and without microbiologically-confirmed ventilator-associated pneumonia. sVCAM-1 concentrations were significantly higher in the bronchoalveolar lavage fluid of patients with ARDS compared to healthy controls, and tended to be higher in moderate/severe ARDS

  7. Soluble vascular endothelial growth factor in various blood transfusion components

    DEFF Research Database (Denmark)

    Nielsen, Hans Jørgen; Werther, K; Mynster, T

    1999-01-01

    BACKGROUND: Blood transfusion may reduce survival after curative surgery for solid tumors. This may be related to extracellular content of cancer growth factors present in transfusion components. Vascular endothelial growth factor (VEGF) is a potent stimulator of angiogenesis in solid tumors....... The potential content of VEGF in various blood components for transfusion was evaluated. STUDY DESIGN AND METHODS: Soluble VEGF (sVEGF, isotype 165) was determined by an enzyme-linked immunosorbent assay (EIA) in serum and plasma samples and in lysed cells from healthy volunteers. Subsequently, total content......-reduced PRP. The sVEGF accumulated significantly in WB, SAGM blood, and BCP pools, depending on the storage time. CONCLUSION: The sVEGF (isotype 165) appears to be present in various blood transfusion components, depending on storage time....

  8. Soluble interleukin 6 receptor (sIL-6R) mediates colonic tumor cell adherence to the vascular endothelium: a mechanism for metastatic initiation?

    LENUS (Irish Health Repository)

    Dowdall, J F

    2012-02-03

    The mechanisms by which surgery increases metastatic proliferation remain poorly characterized, although endotoxin and immunocytes play a role. Recent evidence suggests that endothelial adherence of tumor cells may be important in the formation of metastases. Soluble receptors of interleukin-6 (sIL-6R) shed by activated neutrophils exert IL-6 effects on endothelial cells, which are unresponsive under normal circumstances. This study examined the hypothesis that sIL-6R released by surgical stress increases tumor cell adherence to the endothelium. Neutrophils (PMN) were stimulated with lipopolysaccharide, C-reactive protein (CRP), and tumor necrosis factor-alpha. Soluble IL-6R release was measured by enzyme-linked immunosorbent assay. Colonic tumor cells transfected with green fluorescent protein and endothelial cells were exposed to sIL-6R, and tumor cell adherence and transmigration were measured by fluorescence microscopy. Basal release of sIL-6R from PMN was 44.7 +\\/- 8.2 pg\\/ml at 60 min. This was significantly increased by endotoxin and CRP (131 +\\/- 16.8 and 84.1 +\\/- 5.3, respectively; both P < 0.05). However, tumor necrosis factor-alpha did not significantly alter sIL-6R release. Endothelial and tumor cell exposure to sIL-6R increased tumor cell adherence by 71.3% within 2 h but did not significantly increase transmigration, even at 6 h. Mediators of surgical stress induce neutrophil release of a soluble receptor for IL-6 that enhances colon cancer cell endothelial adherence. Since adherence to the endothelium is now considered to be a key event in metastatic genesis, these findings have important implications for colon cancer treatment strategies.

  9. Insulin-Regulated Increase of Soluble Vascular Adhesion Protein-1 in Diabetes

    OpenAIRE

    Salmi, Marko; Stolen, Craig; Jousilahti, Pekka; Yegutkin, Gennady G.; Tapanainen, Päivi; Janatuinen, Tuula; Knip, Mikael; Jalkanen, Sirpa; Salomaa, Veikko

    2002-01-01

    Vascular adhesion protein-1 (VAP-1) is one of the molecules on the endothelial cell membrane, which may guide inflammatory cells into atherosclerotic lesions. This dual function molecule may also contribute to the pathogenesis of atherosclerosis and other vasculopathies via its enzymatic activity that oxidizes primary amines to produce their corresponding aldehydes, hydrogen peroxide, and ammonium. Because VAP-1 also exists in a soluble form, we analyzed its potential usefulness as a biomarke...

  10. Enhanced Prevalence of Plasmatic Soluble MHC Class I Chain-Related Molecule in Vascular Pregnancy Diseases

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    Jean Baptiste Haumonte

    2014-01-01

    Full Text Available The major histocompatibility complex class I related chain (MIC is a stress-inducible protein modulating the function of immune natural killer (NK cells, a major leukocyte subset involved in proper trophoblast invasion and spiral artery remodeling. Aim of the study was to evaluate whether upregulation of soluble MIC (sMIC may reflect immune disorders associated to vascular pregnancy diseases (VPD. sMIC was more frequently detected in the plasma of women with a diagnostic of VPD (32% than in normal term-matched pregnancies (1.6%, P<0.0001, with highest prevalence in intrauterine fetal death (IUDF, 44% and vascular intrauterine growth restriction (IUGR, 39%. sMIC levels were higher in preeclampsia (PE than in IUFD (P<0.01 and vascular IUGR (P<0.05. sMIC detection was associated with bilateral early diastolic uterine notches (P=0.037, thrombocytopenia (P=0.03, and high proteinuria (P=0.03 in PE and with the vascular etiology of IUGR (P=0.0038. Incubation of sMIC-positive PE plasma resulted in downregulation of NKG2D expression and NK cell-mediated IFN-γ production in vitro. Our work thus suggests that detection of sMIC molecule in maternal plasma may constitute a hallmark of altered maternal immune functions that contributes to vascular disorders that complicate pregnancy, notably by impairing NK-cell mediated production of IFN-γ, an essential cytokine favoring vascular modeling.

  11. Cerebrospinal fluid and plasma concentration of soluble intercellular adhesion molecule1, vascular cell adhesion molecule1 and endothelial leukocyte adhesion molecule in patients with acute ischemic b

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    Selaković Vesna M.

    2003-01-01

    Full Text Available Background. Leukocyte migration into the ischemic area is a complex process controlled by adhesion molecules (AM in leukocytes and endothelium, by migratory capacity of leukocytes and the presence of hemotaxic agents in the tissue. In this research it was supposed that in the blood and cerebrospinal fluid (CSF of patients in the acute phase of ischemic brain disease (IBD there were relevant changes in the concentration of soluble AM (sICAM-1 sVCAM-1 and sE-selectin, that could have been the indicators of the intensity of damaging processes in central nervous system (CNS. Methods. The study included 45 IBD patients, 15 with transient ischemic attack (TIA 15 with reversible ischemic attack (RIA, and 15 with brain infarction (BI of both sexes, mean age 66±7. Control group consisted of 15 patients with radicular lesions of discal origin, subjected to diagnostic radiculography without the signs of interruption in the passage of CSF. Changes of selected biochemical parameters were determined in all patients in frame 72 hours since the occurence of an ischemic episode. Concentrations of soluble AM were determined in plasma and CSF by ELISA. Total number of leukocytes (TNL in peripheral blood was determined by hematological analyzer. Results. The results showed that during the first 72 hrs of IBD significant increases occured in TNL and that the increase was progressive compared to the severeness of the disease. Significant increase of soluble AM concentration was shown in plasma of IBD patients. The increase was highest in BI somewhat lower in RIA and the lowest in TIA patients compared to the control. In CSF concentrations of sICAM-1, sVCAM-1 and sE-selectin demonstrated similar increasing trend as in plasma. Conclusion. TNL, as well as the soluble AM concentrations in plasma and CSF, were increased during the acute IBD phase and progressive in relation to the severeness of the disease, so that they might have been the indicators of CNS inflammatory

  12. Growth hormone increases vascular cell adhesion molecule 1 expression

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    Hansen, Troels Krarup; Fisker, Sanne; Dall, Rolf

    2004-01-01

    We investigated the impact of GH administration on endothelial adhesion molecules, vascular cell adhesion molecule-1 (VCAM-1) and E-selectin, in vivo and in vitro. Soluble VCAM-1, E-selectin, and C-reactive protein concentrations were measured before and after treatment in 25 healthy subjects...

  13. Vascular inflammatory cells in hypertension

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    David G. Harrison

    2012-05-01

    Full Text Available Hypertension is a common disorder with uncertain etiology. In the last several years, it has become evident that components of both the innate and adaptive immune system play an essential role in hypertension. Macrophages and T cells accumulate in the perivascular fat, the heart and the kidney of hypertensive patients and in animals with experimental hypertension. Various immunosuppressive agents lower blood pressure and prevent end-organ damage. Mice lacking lymphocytes are protected against hypertension, and adoptive transfer of T cells, but not B cells in the animals restores their blood pressure response to stimuli such as angiotensin II or high salt. Recent studies have shown that mice lacking macrophages have blunted hypertension in response to angiotensin II and that genetic deletion of macrophages markedly reduces experimental hypertension. Dendritic cells have also been implicated in this disease. Many hypertensive stimuli have triggering effects on the central nervous system and signals arising from the circumventricular organ seem to promote inflammation. Studies have suggested that central signals activate macrophages and T cells, which home to the kidney and vasculature and release cytokines, including IL-6 and IL-17, which in turn cause renal and vascular dysfunction and lead to blood pressure elevation. These recent discoveries provide a new understanding of hypertension and provide novel therapeutic opportunities for treatment of this serious disease.

  14. Bacterial antigen induced release of soluble vascular endothelial growth factor (VEGF) and VEGFR1 before and after surgery

    DEFF Research Database (Denmark)

    Svendsen, Mads N; Lykke, J; Werther, Kim

    2005-01-01

    OBJECTIVE: The influence of surgery on release of soluble vascular endothelial growth factor (sVEGF) and the soluble inhibitory receptor (sVEGFR1) is unknown. The effect of major and minor surgery on variations in sVEGF and sVEGFR1 concentrations in vivo was studied, and on bacterial antigen...... concentrations in plasma changed during surgery. In vitro stimulation of blood samples with bacteria-derived antigens resulted in a significant increase in sVEGF (p Bacterial antigen-induced release of sVEGF correlated...... significantly with neutrophil cell counts (0.53 Bacterial antigen-induced sVEGFR1 release did not correlate with cell counts. CONCLUSIONS: Plasma sVEGF and sVEGFR1 concentrations did not change during surgery. In vitro bacterial stimulation led to increased release of sVEGF, which...

  15. Identification of soluble CD146 as a regulator of trophoblast migration: potential role in placental vascular development.

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    Kaspi, Elise; Guillet, Benjamin; Piercecchi-Marti, Marie-Dominique; Alfaidy, Nadia; Bretelle, Florence; Bertaud-Foucault, Alexandrine; Stalin, Jimmy; Rambeloson, Laka; Lacroix, Odile; Blot-Chabaud, Marcel; Dignat-George, Françoise; Bardin, Nathalie

    2013-04-01

    Both vasculogenesis and angiogenesis occur during normal placental vascular development. Additionally, the placenta undergoes a process of vascular mimicry (pseudo-vasculogenesis) where the placental extravillous trophoblast (EVT) that invade the spiral arteries convert from an epithelial to an endothelial phenotype during normal pregnancy. As soluble CD146 (sCD146) constitutes a new physiological factor with angiogenic properties, we hypothesized that it could be involved in the regulation of placental vascular development by acting on EVT. Using placental villous explants, we demonstrated that sCD146 inhibits EVT outgrowth. Consistently, we showed that sCD146 inhibits the ability of EVT cells (HTR8/SVneo) to migrate, invade and form tubes in Matrigel, without affecting their proliferation or apoptosis. The involvement of sCD146 in human pregnancy was investigated by evaluation of sCD146 levels in 50 pregnant women. We observed physiological down-regulation of sCD146 throughout pregnancy. These results prompted us to investigate the effect of prolonged sCD146 administration in a rat model of pregnancy. Repeated systemic sCD146 injections after coupling caused a significant decrease of pregnancy rate and number of embryos. Histological studies performed on placenta evidenced a reduced migration of glycogen cells (analogous to EVT in rat) in sCD146-treated rats. We propose that in human, sCD146 could represent both an attractive biomarker of placental vascular development and a therapeutic target in pregnancy complications associated with pathological angiogenesis.

  16. Soluble vascular endothelial growth factor receptor-3 suppresses lymphangiogenesis and lymphatic metastasis in bladder cancer

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    Kim Wun-Jae

    2011-04-01

    Full Text Available Abstract Background Most bladder cancer patients experience lymphatic metastasis in the course of disease progression, yet the relationship between lymphangiogenesis and lymphatic metastasis is not well known. The aim of this study is to elucidate underlying mechanisms of how expanded lymphatic vessels and tumor microenvironment interacts each other and to find effective therapeutic options to inhibit lymphatic metastasis. Results The orthotopic urinary bladder cancer (OUBC model was generated by intravesical injection of MBT-2 cell lines. We investigated the angiogenesis, lymphangiogenesis, and CD11b+/CD68+ tumor-associated macrophages (TAM by using immunofluorescence staining. OUBC displayed a profound lymphangiogenesis and massive infiltration of TAM in primary tumor and lymphatic metastasis in lymph nodes. TAM flocked near lymphatic vessels and express higher levels of VEGF-C/D than CD11b- cells. Because VEGFR-3 was highly expressed in lymphatic vascular endothelial cells, TAM could assist lymphangiogenesis by paracrine manner in bladder tumor. VEGFR-3 expressing adenovirus was administered to block VEGF-C/D signaling pathway and clodronate liposome was used to deplete TAM. The blockade of VEGF-C/D with soluble VEGF receptor-3 markedly inhibited lymphangiogenesis and lymphatic metastasis in OUBC. In addition, the depletion of TAM with clodronate liposome exerted similar effects on OUBC. Conclusion VEGF-C/D are the main factors of lymphangiogenesis and lymphatic metastasis in bladder cancer. Moreover, TAM plays an important role in these processes by producing VEGF-C/D. The inhibition of lymphangiogenesis could provide another therapeutic target to inhibit lymphatic metastasis and recurrence in patients with invasive bladder cancer.

  17. Mitochondria, endothelial cell function, and vascular diseases.

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    Tang, Xiaoqiang; Luo, Yu-Xuan; Chen, Hou-Zao; Liu, De-Pei

    2014-01-01

    Mitochondria are perhaps the most sophisticated and dynamic responsive sensing systems in eukaryotic cells. The role of mitochondria goes beyond their capacity to create molecular fuel and includes the generation of reactive oxygen species, the regulation of calcium, and the activation of cell death. In endothelial cells, mitochondria have a profound impact on cellular function under both healthy and diseased conditions. In this review, we summarize the basic functions of mitochondria in endothelial cells and discuss the roles of mitochondria in endothelial dysfunction and vascular diseases, including atherosclerosis, diabetic vascular dysfunction, pulmonary artery hypertension, and hypertension. Finally, the potential therapeutic strategies to improve mitochondrial function in endothelial cells and vascular diseases are also discussed, with a focus on mitochondrial-targeted antioxidants and calorie restriction.

  18. Mitochondria, Endothelial Cell Function and Vascular Diseases

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    Xiaoqiang eTang

    2014-05-01

    Full Text Available Mitochondria are perhaps the most sophisticated and dynamic responsive sensing systems in eukaryotic cells. The role of mitochondria goes beyond their capacity to create molecular fuel and includes the generation of reactive oxygen species, the regulation of calcium, and the activation of cell death. In endothelial cells, mitochondria have a profound impact on cellular function under both healthy and diseased conditions. In this review, we summarize the basic functions of mitochondria in endothelial cells and discuss the roles of mitochondria in endothelial dysfunction and vascular diseases, including atherosclerosis, diabetic vascular dysfunction, pulmonary artery hypertension and hypertension. Finally, the potential therapeutic strategies to improve mitochondrial function in endothelial cells and vascular diseases are also discussed, with a focus on mitochondrial-targeted antioxidants and calorie restriction.

  19. Soluble CD44 and vascular endothelial growth factor levels in patients with acute primary angle closure.

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    Chen, Shida; Huang, Wenbin; Wang, Jiawei; Zhang, Jing; Wang, Wei; Zhou, Minwen; Gao, Xinbo; Zhang, Xiulan

    2015-06-01

    Acute elevation of intraocular pressure (IOP) in acute primary angle closure (APAC) can cause huge damage to the variable cells in the eye; however, the mechanisms that connect the two processes still remain unclear. In this study, we aim to evaluate the levels of soluble CD44 (sCD44) and vascular endothelial growth factor (VEGF) in the aqueous humour of acute primary angle closure patients. This study included 24 eyes of 24 APAC patients (11 eyes with current APAC and 13 eyes with previous APAC) and 15 eyes of 15 cataract subjects. Clinical data were acquired, and aqueous humour was collected. The levels of sCD44 and VEGF in the aqueous humour were determined by ELISA and magnetic bead immunoassay technique. The concentrations of the sCD44 and VEGF in the current APAC were 9.9 ± 8.8 ng/ml and 2440.2 ± 2107.1 ng/ml, respectively, which were significantly higher when compared to the previous APAC group (p = 0.001) and cataract (p APAC and with cataract. Higher IOP was associated with higher concentration of sCD44 (Rho = 0.617, p = 0.001). The concentration of the VEGF in aqueous humour of APAC patients was closely related to the sCD44 levels (Rho = 0.752, p APAC, the level of sCD44 and VEGF increased significantly in the aqueous humour. The damage due to high IOP may therefore be mediated through the release of sCD44. © 2014 Acta Ophthalmologica Scandinavica Foundation. Published by John Wiley & Sons Ltd.

  20. Production of soluble Neprilysin by endothelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Kuruppu, Sanjaya, E-mail: Sanjaya.Kuruppu@monash.edu [Department of Biochemistry and Molecular Biology, Building 77, Monash University, Wellington Rd, Clayton, Vic 3800 (Australia); Rajapakse, Niwanthi W. [Department of Physiology, Building 13F, Monash University, Wellington Rd, Clayton, Vic 3800 (Australia); Minond, Dmitriy [Torrey Pines Institute for Molecular Studies, 11350 SW Village Parkway, Port Saint Lucie, FL 34987 (United States); Smith, A. Ian [Department of Biochemistry and Molecular Biology, Building 77, Monash University, Wellington Rd, Clayton, Vic 3800 (Australia)

    2014-04-04

    Highlights: • A soluble full-length form of Neprilysin exists in media of endothelial cells. • Exosomal release is the key mechanism for the production of soluble Neprilysin. • Inhibition of ADAM-17 by specific inhibitors reduce Neprilysin release. • Exosome mediated release of Neprilysin is dependent on ADAM-17 activity. - Abstract: A non-membrane bound form of Neprilysin (NEP) with catalytic activity has the potential to cleave substrates throughout the circulation, thus leading to systemic effects of NEP. We used the endothelial cell line Ea.hy926 to identify the possible role of exosomes and A Disintegrin and Metalloprotease 17 (ADAM-17) in the production of non-membrane bound NEP. Using a bradykinin based quenched fluorescent substrate (40 μM) assay, we determined the activity of recombinant human NEP (rhNEP; 12 ng), and NEP in the media of endothelial cells (10% v/v; after 24 h incubation with cells) to be 9.35 ± 0.70 and 6.54 ± 0.41 μmols of substrate cleaved over 3 h, respectively. The presence of NEP in the media was also confirmed by Western blotting. At present there are no commercially available inhibitors specific for ADAM-17. We therefore synthesised two inhibitors TPI2155-14 and TPI2155-17, specific for ADAM-17 with IC{sub 50} values of 5.36 and 4.32 μM, respectively. Treatment of cells with TPI2155-14 (15 μM) and TPI2155-17 (4.3 μM) resulted in a significant decrease in NEP activity in media (62.37 ± 1.43 and 38.30 ± 4.70, respectively as a % of control; P < 0.0001), implicating a possible role for ADAM-17 in NEP release. However, centrifuging media (100,000g for 1 h at 4 °C) removed all NEP activity from the supernatant indicating the likely role of exosomes in the release of NEP. Our data therefore indicated for the first time that NEP is released from endothelial cells via exosomes, and that this process is dependent on ADAM-17.

  1. [DEVELOPMENT OF CELL SHEET ENGINEERING TECHNOLOGY IN ENGINEERING VASCULARIZED TISSUE].

    Science.gov (United States)

    Chen, Jia; Ma, Dongyang; Ren, Liling

    2015-03-01

    To review the development of cell sheet engineering technology in engineering vascularized tissue. The literature about cell sheet engineering technology and engineering vascularized tissue was reviewed, analyzed, and summarized. Although there are many methods to engineer vascularized tissue, cell sheet engineering technology provides a promising potential to develop a vascularized tissue. Recently, cell sheet engineering technology has become a hot topic in engineering vascularized tissue. Co-culturing endothelial cells on a cell sheet, endothelial cells are able to form three-dimensional prevascularized networks and microvascular cavities in the cell sheet, which facilitate the formation of functional vascular networks in the transplanted tissue. Cell sheet engineering technology is a promising strategy to engineer vascularized tissue, which is still being studied to explore more potential.

  2. Mononuclear Cells and Vascular Repair in HHT

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    Calinda eDingenouts

    2015-03-01

    Full Text Available Hereditary hemorrhagic telangiectasia (HHT or Rendu-Osler-Weber disease is a rare genetic vascular disorder known for its endothelial dysplasia causing arteriovenous malformations and severe bleedings. HHT-1 and HHT-2 are the most prevalent variants and are caused by heterozygous mutations in endoglin and ALK1, respectively. An undervalued aspect of the disease is that HHT patients experience persistent inflammation. Although endothelial and mural cells have been the main research focus trying to unravel the mechanism behind the disease, wound healing is a process with a delicate balance between inflammatory and vascular cells. Inflammatory cells are part of the mononuclear cells (MNCs fraction, and can, next to eliciting an immune response, also have angiogenic potential. This biphasic effect of MNC can hold a promising mechanism to further elucidate treatment strategies for HHT patients. Before MNC are able to contribute to repair, they need to home to and retain in ischemic and damaged tissue. Directed migration (homing of mononuclear cells following tissue damage is regulated by the stromal cell derived factor 1 (SDF1. MNCs that express the C-X-C chemokine receptor 4 (CXCR4 migrate towards the tightly regulated gradient of SDF1. This directed migration of monocytes and lymphocytes can be inhibited by dipeptidyl peptidase 4 (DPP4. Interestingly, MNC of HHT patients express elevated levels of DPP4 and show impaired homing towards damaged tissue. Impaired homing capacity of the MNCs might therefore contribute to the impaired angiogenesis and tissue repair observed in HHT patients. This review summarizes recent studies regarding the role of MNCs in the etiology of HHT and vascular repair, and evaluates the efficacy of DPP4 inhibition in tissue integrity and repair.

  3. Suppression of lymphangiogenesis by soluble vascular endothelial growth factor receptor-2 in a mouse lung cancer model.

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    Maehana, Shotaro; Nakamura, Masaki; Ogawa, Fumihiro; Imai, Rimika; Murakami, Rei; Kojima, Fumiaki; Majima, Masataka; Kitasato, Hidero

    2016-12-01

    The vascular endothelial growth factor (VEGF) family has a key role in the formation of blood vessels and lymphatics. Among the members of this family, VEGF-C is one of the most important factors involved in lymphangiogenesis via binding with two receptors (vascular endothelial growth factor receptor-2 and -3: VEGFR-2 and VEGFR-3). Soluble VEGFR-2 (sVEGFR-2) has a role in maintaining the alymphatic state of the cornea associated with binding to VEGF-C, and selectively inhibits lymphangiogenesis but not angiogenesis. In this study, we introduced sVEGFR-2 into lung cancer cells and evaluated the influence on tumor progression and on genes regulating lymphatic formation and metastasis in vivo. A retroviral vector was used to introduce the sVEGFR-2 gene into Lewis lung carcinoma cells (LLC), which were designated as LLC-sVEGFR-2 cells. Proteins secreted into the culture supernatant by these cells were detected by western blotting using specific antibodies. To examine lymphangiogenesis by primary lung cancer in vivo, LLC-sVEGFR-2 cells were subcutaneously injected into C57BL/6 mice. At 14days after injection, immunohistochemistry was performed using an antibody directed against lymphatic vessel endothelial hyaluronan receptor 1 (LYVE-1), a marker of lymphatics. Expression of mRNA for VEGFR-2, VEGFR-3 and matrix metalloproteinases (MMPs) was also determined by real-time PCR. Furthermore, LLC-sVEGFR-2 cells were directly inoculated into the left lung in C57BL/6 mice and the number of micro-metastases in pulmonary lymph nodes was determined. Introduction of sVEGFR-2 into LLC cells resulted in secretion of sVEGFR-2 protein into the culture supernatant. There were fewer LYVE-1 positive lymphatics after inoculation of LLC-sVEGFR-2 into mice compared with the control group. In addition, VEGFR-2, VEGFR-3, and MMPs gene expression was suppressed in the primary tumors of the LLC-sVEGFR-2 group compared with the control group. Furthermore, there were fewer micro-metastases in the

  4. Cell based therapeutic approach in vascular surgery: application and review

    Directory of Open Access Journals (Sweden)

    Rocca Aldo

    2017-10-01

    Full Text Available Multipotent stem cells - such as mesenchymal stem/stromal cells and stem cells derived from different sources like vascular wall are intensely studied to try to rapidly translate their discovered features from bench to bedside. Vascular wall resident stem cells recruitment, differentiation, survival, proliferation, growth factor production, and signaling pathways transduced were analyzed. We studied biological properties of vascular resident stem cells and explored the relationship from several factors as Matrix Metalloproteinases (MMPs and regulations of biological, translational and clinical features of these cells. In this review we described a translational and clinical approach to Adult Vascular Wall Resident Multipotent Vascular Stem Cells (VW-SCs and reported their involvement in alternative clinical approach as cells based therapy in vascular disease like arterial aneurysms or peripheral arterial obstructive disease.

  5. Influence of soluble or matrix-bound isoforms of vascular endothelial growth factor-A on tumor response to vascular-targeted strategies.

    Science.gov (United States)

    Akerman, Simon; Fisher, Matthew; Daniel, Rachel A; Lefley, Diane; Reyes-Aldasoro, Constantino C; Lunt, Sarah Jane; Harris, Sheila; Bjorndahl, Meit; Williams, Leigh J; Evans, Helen; Barber, Paul R; Prise, Vivien E; Vojnovic, Borivoj; Kanthou, Chryso; Tozer, Gillian M

    2013-12-01

    Antiangiogenic therapy based on blocking the actions of vascular endothelial growth factor-A (VEGF) can lead to "normalization" of blood vessels in both animal and human tumors. Differential expression of VEGF isoforms affects tumor vascular maturity, which could influence the normalization process and response to subsequent treatment. Fibrosarcoma cells expressing only VEGF120 or VEGF188 isoforms were implanted either subcutaneously (s.c.) or in dorsal skin-fold "window" chambers in SCID mice. VEGF120 was associated with vascular fragility and hemorrhage. Tumor-bearing mice were treated with repeat doses of SU5416, an indolinone receptor tyrosine kinase inhibitor with activity against VEGFR-2 and proven preclinical ability to induce tumor vascular normalization. SU5416 reduced vascularization in s.c. implants of both VEGF120 and VEGF188 tumors. However, in the window chamber, SU5416 treatment increased red cell velocity in VEGF120 (representing vascular normalization) but not VEGF188 tumors. SU5416 treatment had no effect on growth or necrosis levels in either tumor type but tended to counteract the increase in interstitial fluid pressure seen with growth of VEGF120 tumors. SU5416 pretreatment resulted in the normally fragile blood vessels in VEGF120-expressing tumors becoming resistant to the vascular damaging effects of the tubulin-binding vascular disrupting agent (VDA), combretastatin A4 3-O-phosphate (CA4P). Thus, vascular normalization induced by antiangiogenic treatment can reduce the efficacy of subsequent VDA treatment. Expression of VEGF120 made tumors particularly susceptible to vascular normalization by SU5416, which in turn made them resistant to CA4P. Therefore, VEGF isoform expression may be useful for predicting response to both antiangiogenic and vascular-disrupting therapy. Copyright © 2013 UICC.

  6. Soluble RAGE-modulating drugs: state-of-the-art and future perspectives for targeting vascular inflammation.

    Science.gov (United States)

    Lanati, Niccolò; Emanuele, Enzo; Brondino, Natascia; Geroldi, Diego

    2010-01-01

    The expression of the Receptor for Advanced Glycation Endproducts (RAGE) is upregulated at sites of vascular inflammation and plays a crucial role in vessel homeostasis. Soluble RAGE (sRAGE), a truncated soluble form of the receptor, acts as a decoy and prevents the inflammatory response mediated by RAGE activation. sRAGE has recently emerged as a biomarker in several RAGE-mediated vascular disorders, including coronary artery disease, hypertension, diabetic vasculopathy and Kawasaki disease. Given the pivotal role played by RAGE and sRAGE in numerous vascular disorders, there is a growing need to understand how drugs can modulate the RAGE axis in different disease conditions. In this regard, there is evidence to suggest that traditional cardiovascular drugs (statins, thiazolidinediones, ACE-inhibitors, AT-1 receptor antagonists) as well as nutraceuticals (grape seed proanthocyanidin extract) could modulate RAGE expression and circulating sRAGE levels in cardiovascular disease states characterized by enhanced RAGE activation. Additionally, the production of genetically engineered sRAGE may hold promise for targeting the activation of RAGE by proinflammatory ligands in the setting of vascular inflammation. The present review considers current vascular drugs as modulators of the RAGE axis, and highlights future directions in the context of RAGE-directed therapy in cardiovascular disease.

  7. DMPD: Lipoprotein trafficking in vascular cells. Molecular Trojan horses and cellularsaboteurs. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 9287290 Lipoprotein trafficking in vascular cells. Molecular Trojan horses and cell...ml) Show Lipoprotein trafficking in vascular cells. Molecular Trojan horses and cellularsaboteurs. PubmedID ...9287290 Title Lipoprotein trafficking in vascular cells. Molecular Trojan horses

  8. Chorein Sensitivity of Actin Polymerization, Cell Shape and Mechanical Stiffness of Vascular Endothelial Cells

    Directory of Open Access Journals (Sweden)

    Ioana Alesutan

    2013-09-01

    Full Text Available Background/Aims: Endothelial cell stiffness plays a key role in endothelium-dependent control of vascular tone and arterial blood pressure. Actin polymerization and distribution of microfilaments is essential for mechanical cell stiffness. Chorein, a protein encoded by the VPS13A gene, defective in chorea-acanthocytosis (ChAc, is involved in neuronal cell survival as well as cortical actin polymerization of erythrocytes and blood platelets. Chorein is expressed in a wide variety of further cells, yet nothing is known about the impact of chorein on cells other than neurons, erythrocytes and platelets. The present study explored whether chorein is expressed in human umbilical vein endothelial cells (HUVECs and addressed the putative role of chorein in the regulation of cytoskeletal architecture, stiffness and survival of those cells. Methods: In HUVECs with or without silencing of the VPS13A gene, VPS13A mRNA expression was determined utilizing quantitative RT-PCR, cytoskeletal organization visualized by confocal microscopy, G/F actin ratio and phosphorylation status of focal adhesion kinase quantified by western blotting, cell death determined by flow cytometry, mechanical properties studied by atomic force microscopy (AFM and cell morphology analysed by scanning ion conductance microscopy (SICM. Results: VPS13A mRNA expression was detectable in HUVECs. Silencing of the VPS13A gene attenuated the filamentous actin network, decreased the ratio of soluble G-actin over filamentous F-actin, reduced cell stiffness and changed cell morphology as compared to HUVECs silenced with negative control siRNA. These effects were paralleled by a significant decrease in FAK phosphorylation following VPS13A silencing. Moreover, silencing of the VPS13A gene increased caspase 3 activity and induced necrosis in HUVECs. Conclusions: Chorein is a novel regulator of cytoskeletal architecture, cell shape, mechanical stiffness and survival of vascular endothelial cells.

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

    International Nuclear Information System (INIS)

    Davies, P.F.

    1986-01-01

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

  10. Soluble adenylyl cyclase in vascular endothelium: gene expression control of epithelial sodium channel-α, Na+/K+-ATPase-α/β, and mineralocorticoid receptor.

    Science.gov (United States)

    Schmitz, Boris; Nedele, Johanna; Guske, Katrin; Maase, Martina; Lenders, Malte; Schelleckes, Michael; Kusche-Vihrog, Kristina; Brand, Stefan-Martin; Brand, Eva

    2014-04-01

    The Ca(2+)- and bicarbonate-activated soluble adenylyl cyclase (sAC) has been identified recently as an important mediator of aldosterone signaling in the kidney. Nuclear sAC has been reported to stimulate cAMP response element-binding protein 1 phosphorylation via protein kinase A, suggesting an alternative cAMP pathway in the nucleus. In this study, we analyzed the sAC as a potential modulator of endothelial stiffness in the vascular endothelium. We determined the contribution of sAC to cAMP response element-mediated transcriptional activation in vascular endothelial cells and kidney collecting duct cells. Inhibition of sAC by the specific inhibitor KH7 significantly reduced cAMP response element-mediated promoter activity and affected cAMP response element-binding protein 1 phosphorylation. Furthermore, KH7 and anti-sAC small interfering RNA significantly decreased mRNA and protein levels of epithelial sodium channel-α and Na(+)/K(+)-ATPase-α. Using atomic force microscopy, a nano-technique that measures stiffness and deformability of living cells, we detected significant endothelial cell softening after sAC inhibition. Our results suggest that the sAC is a regulator of gene expression involved in aldosterone signaling and an important regulator of endothelial stiffness. Additional studies are warranted to investigate the protective action of sAC inhibitors in humans for potential clinical use.

  11. Soluble and cell-associated haemagglutinins of Helicobacter (Campylobacter) pylori.

    Science.gov (United States)

    Robinson, J; Goodwin, C S; Cooper, M; Burke, V; Mee, B J

    1990-12-01

    Some plate-grown strains of Helicobacter (Campylobacter) pylori that were harvested into phosphate-buffered saline and left for 1 h released soluble haemagglutinins. These caused high-titre agglutination of human and guinea-pig erythrocytes, whereas chicken, sheep and bovine erythrocytes were agglutinated at various titres. Six of 10 strains which had been subcultured repeatedly did not possess soluble haemagglutinins. Slide agglutination of bacterial suspensions demarcated the strains into two groups; Group 1 gave strong agglutination with most types of erythrocyte, Group 2 did not. By microtitration assay, all Group-1 strains but only two Group-2 strains produced a soluble haemagglutinin. Cell-associated haemagglutinins were found by microtitration assay in all strains of H. pylori, but higher titres were found within Group-1 strains. The supernates of broth-grown, shaken cultures also showed the presence of soluble haemagglutinins, with higher titres for recently isolated strains. Pre-treatment of human erythrocytes with neuraminidase from Arthrobacter ureafaciens and Clostridium perfringens abolished haemagglutination by the soluble, but not by the cell-associated haemagglutinin. The soluble haemagglutinin was inhibited by sialoproteins containing predominantly the N-acetylneuraminyl (2-3) galactopyranosyl [NeuAc(2-3)Gal] structure, fetuin, glycophorin and bovine N-acetylneuraminyl-lactose (NeuAc-Lac). Transferrin and human NeuAc-Lac, which contain predominantly the N-acetylneuraminyl (2-6) galactopyranosyl [NeuAc(2-6)Gal] structure were not inhibitory. However, bovine submaxillary mucin (BSM) was strongly inhibitory; it contains several structures with sialic acid linked 2-6 to oligosaccharides. These results suggest that the soluble haemagglutinin recognises a NeuAc(2-3)Gal structure, but has high affinity for another, as yet undetermined, sialic acid-containing structure.

  12. Temperature-dependent rate models of vascular cambium cell mortality

    Science.gov (United States)

    Matthew B. Dickinson; Edward A. Johnson

    2004-01-01

    We use two rate-process models to describe cell mortality at elevated temperatures as a means of understanding vascular cambium cell death during surface fires. In the models, cell death is caused by irreversible damage to cellular molecules that occurs at rates that increase exponentially with temperature. The models differ in whether cells show cumulative effects of...

  13. Soluble Triggering Receptor Expressed on Myeloid Cells-1 as a ...

    African Journals Online (AJOL)

    Soluble Triggering Receptor Expressed on Myeloid Cells-1 as a marker to differentiate septic from aseptic meningitis in children: comparison with procalcitonin and ... Procalcitonin (PCT) was suggested by many researchers as a sensitive marker for early diagnosis of septic meningitis but with varying discriminative power.

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

    Science.gov (United States)

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

  15. Cell proliferation along vascular islands during microvascular network growth

    Directory of Open Access Journals (Sweden)

    Kelly-Goss Molly R

    2012-06-01

    Full Text Available Abstract Background Observations in our laboratory provide evidence of vascular islands, defined as disconnected endothelial cell segments, in the adult microcirculation. The objective of this study was to determine if vascular islands are involved in angiogenesis during microvascular network growth. Results Mesenteric tissues, which allow visualization of entire microvascular networks at a single cell level, were harvested from unstimulated adult male Wistar rats and Wistar rats 3 and 10 days post angiogenesis stimulation by mast cell degranulation with compound 48/80. Tissues were immunolabeled for PECAM and BRDU. Identification of vessel lumens via injection of FITC-dextran confirmed that endothelial cell segments were disconnected from nearby patent networks. Stimulated networks displayed increases in vascular area, length density, and capillary sprouting. On day 3, the percentage of islands with at least one BRDU-positive cell increased compared to the unstimulated level and was equal to the percentage of capillary sprouts with at least one BRDU-positive cell. At day 10, the number of vascular islands per vascular area dramatically decreased compared to unstimulated and day 3 levels. Conclusions These results show that vascular islands have the ability to proliferate and suggest that they are able to incorporate into the microcirculation during the initial stages of microvascular network growth.

  16. Molecular mechanisms for vascular development and secondary cell wall formation

    Directory of Open Access Journals (Sweden)

    Jung Hyun eYang

    2016-03-01

    Full Text Available Vascular tissues are important for transporting water and nutrients throughout the plant and as physical support of upright growth. The primary constituents of vascular tissues, xylem and phloem, are derived from the meristematic vascular procambium and cambium. Xylem cells develop secondary cell walls that form the largest part of plant lignocellulosic biomass that serve as a renewable feedstock for biofuel production. For the last decade, research on vascular development and secondary cell wall biosynthesis has seen rapid progress due to the importance of these processes to plant biology and to the biofuel industry. Plant hormones, transcriptional regulators and peptide signaling regulate procambium/cambium proliferation, vascular patterning, and xylem differentiation. Transcriptional regulatory pathways play a pivot role in secondary cell wall biosynthesis. Although most of these discoveries are derived from research in Arabidopsis, many genes have shown conserved functions in biofuel feedstock species. Here, we review the recent advances in our understanding of vascular development and secondary cell wall formation and discuss potential biotechnological uses.

  17. Scaffold vascularization in vivo driven by primary human osteoblasts in concert with host inflammatory cells.

    Science.gov (United States)

    Ghanaati, Shahram; Unger, Ronald E; Webber, Matthew J; Barbeck, Mike; Orth, Carina; Kirkpatrick, Jenny A; Booms, Patrick; Motta, Antonella; Migliaresi, Claudio; Sader, Robert A; Kirkpatrick, C James

    2011-11-01

    Successful cell-based tissue engineering requires a rapid and thorough vascularization in order to ensure long-term implant survival and tissue integration. The vascularization of a scaffold is a complex process, and is modulated by the presence of transplanted cells, exogenous and endogenous signaling proteins, and the host tissue reaction, among other influencing factors. This paper presents evidence for the significance of pre-seeded osteoblasts for the in vivo vascularization of a biodegradable scaffold. Human osteoblasts, cultured on silk fibroin micronets in vitro, migrated throughout the interconnected pores of the scaffold and produced extensive bone matrix. When these constructs were implanted in SCID mice, a rapid and thorough vascularization of the scaffold by the host blood capillaries occurred. This profound response was not seen for the silk fibroin scaffold alone. Moreover, when the pre-cultivation time of human osteoblasts was reduced from 14 days to only 24 h, the significant effect these cells exerted on vascularization rate in vivo was still detectable. From these studies, we conclude that matrix and soluble factors produced by osteoblasts can serve to instruct host endothelial cells to migrate, proliferate, and initiate the process of scaffold vascularization. This finding represents a potential paradigm shift for the field of tissue engineering, especially in bone, as traditional strategies to enhance scaffold vascularization have focused on endovascular cells and regarded osteoblasts primarily as cell targets for mineralization. In addition, the migration of host macrophages and multinucleated giant cells into the scaffold was also found to influence the vascularization of the biomaterial. Therefore, the robust effect on scaffold vascularization seen by pre-culturing with osteoblasts appears to occur in concert with the pro-angiogenic stimuli arising from host immune cells. Copyright © 2011 Elsevier Ltd. All rights reserved.

  18. Proteolytic cleavage of vascular adhesion protein-1 induced by vascular endothelial growth factor in retinal capillary endothelial cells.

    Science.gov (United States)

    Yoshida, Shiho; Murata, Miyuki; Noda, Kousuke; Matsuda, Takashi; Saito, Michiyuki; Saito, Wataru; Kanda, Atsuhiro; Ishida, Susumu

    2018-02-01

    To investigate the mechanism of soluble vascular adhesion protein-1 (sVAP-1) accumulation induced by vascular endothelial growth factor (VEGF) in the vitreous of patients with diabetic retinopathy (DR). Experimental. Protein levels of sVAP-1 and N epsilon-(hexanoyl)lysine (HEL), an oxidative stress marker, in the vitreous samples from patients with proliferative diabetic retinopathy (PDR) with or without intravitreal bevacizumab (IVB) injection were determined by ELISA. The effect of VEGF on both mRNA expression of Vap-1 and secretion of sVAP-1 in rat retinal capillary endothelial cells (TR-iBRB2) was analyzed by real-time PCR and western blotting, respectively. In addition, the impact of VEGF on production and activation ratios of matrix metalloproteinase (MMP)-2 and MMP-9 was examined by gelatin zymography. Hydrogen peroxide production and reactive oxygen species (ROS) levels were assessed in the supernatants of TR-iBRB2 cells treated with VEGF. IVB injection decreased vitreous levels of sVAP-1 and HEL in patients with PDR. VEGF stimulation released sVAP-1 protein from TR-iBRB2 cells as a consequence of membrane-anchored VAP-1 shedding by MMP-2 and MMP-9. In addition, VEGF increased hydrogen peroxide generation and ROS augmentation through spermine oxidation by sVAP-1 as semicarbazide-sensitive amine oxidase (SSAO) in the supernatant of cultured endothelial cells. The current data demonstrate that proangiogenic factor VEGF induces sVAP-1 release from retinal capillary endothelial cells and facilitates hydrogen peroxide generation via enzymatic property of sVAP-1, followed by the increase of oxidative stress, one of the crucial factors in the pathogenesis of DR.

  19. Interplay between coagulation and vascular inflammation in sickle cell disease

    Science.gov (United States)

    Sparkenbaugh, Erica; Pawlinski, Rafal

    2013-01-01

    Sickle cell disease is the most common inherited hematologic disorder that leads to the irreversible damage of multiple organs. Although sickling of red blood cells and vaso-occlusion are central to the pathophysiology of sickle cell disease the importance of hemolytic anemia and vasculopathy has been recently recognized. Hypercoagulation state is another prominent feature of sickle cell disease and is mediated by activation of both intrinsic and extrinsic coagulation pathways. Growing evidence demonstrates that coagulation may not only contribute to the thrombotic complications, but also to vascular inflammation associated with this disease. This article summarizes the role of vascular inflammation and coagulation activation, discusses potential mechanisms responsible for activation of coagulation and reviews recent data demonstrating the crosstalk between coagulation and vascular inflammation in sickle cell disease. PMID:23593937

  20. Perspectives in inflammation, neoplasia, and vascular cell biology

    Energy Technology Data Exchange (ETDEWEB)

    Edgington, T.S.; Ross, R.; Silverstein, S.C.

    1987-01-01

    This book contains 25 selections. Some of the titles are: Characterization of cDNAs for the Human Interleukin-2 Receptor; Regulation of the Epidermal Growth Factor Receptor by Phosphorylation; Endothelial Cell Proteases and Cellular Invasion; Structure and Chromosomal Localization of the Human Lymphotoxin Gene; and Vascular Endothelial Cells in Cell-Mediated Immunity: Adoptive Transfer with In Vitro Conditioned Cells is Genetically Restricted at the Endothelial Cell Barrier.

  1. Endothelial cell migration and invasiveness are induced by a soluble factor produced by murine endothelioma cells transformed by polyoma virus middle T oncogene.

    Science.gov (United States)

    Taraboletti, G; Belotti, D; Dejana, E; Mantovani, A; Giavazzi, R

    1993-08-15

    Polyoma virus middle T-transformed murine endothelioma cell lines provide a useful model for studying vascular lesions such as hemangiomas, hemangiosarcomas, and Kaposi's sarcoma and tumor-associated angiogenesis. In vivo they produce fast-growing, hemorrhaging, cavernous blood-filled hemangiomas, mainly formed by recruited host endothelial cells, suggesting an angiogenesis-like process underlying the lesion. The molecular mechanism(s) responsible for the recruitment of host endothelial cells by endothelioma cells has not yet been identified. We found that five different cultured endothelioma cell lines produced a soluble factor, named endothelioma-derived motility factor (EDMF) that stimulates chemotaxis (motility induced by a gradient of soluble attractant), haptotaxis (motility in response to substrate-bound attractant), and chemoinvasion (migration through a layer of reconstituted basement membrane, Matrigel) of normal human, bovine, and murine endothelial cells. The inhibitory effect of actinomycin D and of enzymatic treatment on its activity proved that EDMF is a protein. EDMF binds to heparin, since its activity was inhibited by heparin, and it was retained on a heparin-Sepharose column. Its molecular weight, as assessed by Sephacryl S-200 gel filtration, ranges from 40,000-65,000. Although in many aspects EDMF is similar to vascular permeability factor-vascular endothelial growth factor, this was not detected in endothelioma cell supernatants, as assessed by enzyme-linked immunosorbent assay, thus indicating that EDMF might be related to, but is not identical with, vascular permeability factor. Our findings support the notion that recruitment of host endothelial cells by endothelioma cells in vivo might be mediated by a still unidentified, soluble factor that stimulates and directs endothelial cell migration.

  2. Mast cells promote lung vascular remodelling in pulmonary hypertension.

    Science.gov (United States)

    Hoffmann, J; Yin, J; Kukucka, M; Yin, N; Saarikko, I; Sterner-Kock, A; Fujii, H; Leong-Poi, H; Kuppe, H; Schermuly, R T; Kuebler, W M

    2011-06-01

    Left heart disease (LHD) frequently causes lung vascular remodelling and pulmonary hypertension (PH). Yet pharmacological treatment for PH in LHD is lacking and its pathophysiological basis remains obscure. We aimed to identify candidate mechanisms of PH in LHD and to test their relevance and therapeutic potential. In rats, LHD was induced by supracoronary aortic banding. Whole genome microarray analyses were performed, candidate genes were confirmed by RT-PCR and Western blots and functional relevance was tested in vivo by genetic and pharmacological strategies. In lungs of LHD rats, mast cell activation was the most prominently upregulated gene ontology cluster. Mast cell gene upregulation was confirmed at RNA and protein levels and remodelled vessels showed perivascular mast cell accumulations. In LHD rats treated with the mast cell stabiliser ketotifen, or in mast cell deficient Ws/Ws rats, PH and vascular remodelling were largely attenuated. Both strategies also reduced PH and vascular remodelling in monocrotaline-induced pulmonary arterial hypertension, suggesting that the role of mast cells extends to non-cardiogenic PH. In PH of different aetiologies, mast cells accumulate around pulmonary blood vessels and contribute to vascular remodelling and PH. Mast cells and mast cell-derived mediators may present promising targets for the treatment of PH.

  3. 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.

  4. Soluble vascular endothelial growth factor levels in patients with primary colorectal carcinoma. The Danish RANX05 Colorectal Cancer Study Group

    DEFF Research Database (Denmark)

    Werther, K; Christensen, Ib Jarle; Brünner, N

    2000-01-01

    INTRODUCTION: Angiogenesis is decisive in tumour progression and metastasis. Vascular endothelial growth factor (VEGF) is a potent angiogenic factor, and increased VEGF levels in patients with carcinomas may facilitate growth of both primary and secondary tumours. METHODS: Soluble (s) VEGF levels...... were determined in serum from 91 volunteer healthy blood donors and from 614 patients scheduled to undergo resection for primary colorectal cancer. None of the patients received pre- and/or post-operative chemo- and/or radiotherapy. The results of sVEGF were analysed with respect to Dukes>> stage...... disease, who had comparable values. Patients with the primary tumour localized in the colon had significantly (Pprimary tumour localized in the rectum. By classifying the patients into two groups, based on the upper limit of the 95(th)percentile of s...

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

    International Nuclear Information System (INIS)

    Kameyama, Kohji; Aida, Takeo; Asano, Goro

    1982-01-01

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

  6. TLR accessory molecule RP105 (CD180 is involved in post-interventional vascular remodeling and soluble RP105 modulates neointima formation.

    Directory of Open Access Journals (Sweden)

    Jacco C Karper

    Full Text Available BACKGROUND: RP105 (CD180 is TLR4 homologue lacking the intracellular TLR4 signaling domain and acts a TLR accessory molecule and physiological inhibitor of TLR4-signaling. The role of RP105 in vascular remodeling, in particular post-interventional remodeling is unknown. METHODS AND RESULTS: TLR4 and RP105 are expressed on vascular smooth muscle cells (VSMC as well as in the media of murine femoral artery segments as detected by qPCR and immunohistochemistry. Furthermore, the response to the TLR4 ligand LPS was stronger in VSMC from RP105(-/- mice resulting in a higher proliferation rate. In RP105(-/- mice femoral artery cuff placement resulted in an increase in neointima formation as compared to WT mice (4982 ± 974 µm(2 vs.1947 ± 278 µm(2,p = 0.0014. Local LPS application augmented neointima formation in both groups, but in RP105(-/- mice this effect was more pronounced (10316±1243 µm(2 vs.4208 ± 555 µm(2,p = 0.0002, suggesting a functional role for RP105. For additional functional studies, the extracellular domain of murine RP105 was expressed with or without its adaptor protein MD1 and purified. SEC-MALSanalysis showed a functional 2∶2 homodimer formation of the RP105-MD1 complex. This protein complex was able to block the TLR4 response in whole blood ex-vivo. In vivo gene transfer of plasmid vectors encoding the extracellular part of RP105 and its adaptor protein MD1 were performed to initiate a stable endogenous soluble protein production. Expression of soluble RP105-MD1 resulted in a significant reduction in neointima formation in hypercholesterolemic mice (2500 ± 573 vs.6581 ± 1894 µm(2,p<0.05, whereas expression of the single factors RP105 or MD1 had no effect. CONCLUSION: RP105 is a potent inhibitor of post-interventional neointima formation.

  7. Soluble CD163 levels in children with sickle cell disease

    DEFF Research Database (Denmark)

    Moller, Holger Jon; Nielsen, Marianne Jensby; Bartram, Jack

    2011-01-01

    Sickle cell disease (SCD) is characterized by vasculopathy, which has been causally linked to intravascular haemolysis and high levels of free plasma haemoglobin. Soluble CD163 (sCD163) is implicated in the clearance of free plasma haemoglobin and high plasma concentrations have been linked...... to arterial disease. We therefore investigated the value of sCD163 as a biomarker in children with SCD, and also measured haptoglobin levels in this population. We measured sCD163 in 25 control children with no haemoglobinopathy, 41 with sickle cell anaemia (HbSS) in the steady state, 27 with HbSS taking...

  8. Bone Marrow Vascular Niche: Home for Hematopoietic Stem Cells

    Directory of Open Access Journals (Sweden)

    Ningning He

    2014-01-01

    Full Text Available Though discovered later than osteoblastic niche, vascular niche has been regarded as an alternative indispensable niche operating regulation on hematopoietic stem cells (HSCs. As significant progresses gained on this type niche, it is gradually clear that the main work of vascular niche is undertaking to support hematopoiesis. However, compared to what have been defined in the mechanisms through which the osteoblastic niche regulates hematopoiesis, we know less in vascular niche. In this review, based on research data hitherto we will focus on component foundation and various functions of vascular niche that guarantee the normal hematopoiesis process within bone marrow microenvironments. And the possible pathways raised by various research results through which this environment undergoes its function will be discussed as well.

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

    Directory of Open Access Journals (Sweden)

    Jie Su

    2015-01-01

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

  10. Vascular niches for disseminated tumour cells in bone

    Directory of Open Access Journals (Sweden)

    Anjali P. Kusumbe

    2016-09-01

    Full Text Available The vasculature of the skeletal system regulates osteogenesis and hematopoiesis, in addition to its primary function as a transportation network. Recent studies suggest that the vasculature in bone regulates multiple steps involved in the metastatic cascade. Matrix and growth factor abundant vascular microenvironments in bone not only provide a fertile soil for the metastatic growth but also support the dormancy of Disseminated Tumour Cells (DTCs. Interestingly, vasculature also seems to direct the reactivation of dormant DTCs. Targeting such early steps of bone metastasis by directing therapies against vascular niches can lead to the development of effective therapeutic strategies that delay or even prevent the metastatic relapse. However, this would require a detailed understanding of the regulatory mechanisms that govern the interaction between endothelial cells and DTCs in the early stages of bone metastasis. This review aims to highlight the importance of vascular niches and outline their newly identified roles during bone metastasis.

  11. Stem cells: a promising source for vascular regenerative medicine.

    Science.gov (United States)

    Rammal, Hassan; Harmouch, Chaza; Lataillade, Jean-Jacques; Laurent-Maquin, Dominique; Labrude, Pierre; Menu, Patrick; Kerdjoudj, Halima

    2014-12-15

    The rising and diversity of many human vascular diseases pose urgent needs for the development of novel therapeutics. Stem cell therapy represents a challenge in the medicine of the twenty-first century, an area where tissue engineering and regenerative medicine gather to provide promising treatments for a wide variety of diseases. Indeed, with their extensive regeneration potential and functional multilineage differentiation capacity, stem cells are now highlighted as promising cell sources for regenerative medicine. Their multilineage differentiation involves environmental factors such as biochemical, extracellular matrix coating, oxygen tension, and mechanical forces. In this review, we will focus on human stem cell sources and their applications in vascular regeneration. We will also discuss the different strategies used for their differentiation into both mature and functional smooth muscle and endothelial cells.

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

    African Journals Online (AJOL)

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

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

    Science.gov (United States)

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

    2017-06-01

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

  14. Cerebro vascular accident in sickle cell disease

    International Nuclear Information System (INIS)

    Alam, M.; Lodhi, M.A.; Khan, D.

    2003-01-01

    Sickle cell disease (SCD) is a common inherited hemoglobin disorder characterized by the presence of sickle shaped erythrocytes in the blood. It can cause stroke in around 10% of children. Repeated blood transfusions are often used in an attempt to dilute blood thus reducing the risk of vaso-occlusion and stroke. We report a case of an 11 years old girl, known patient of sickle cell disease, who did not follow regular blood transfusion protocol and as a result presented with recurrent stroke. (author)

  15. Mesoderm/mesenchyme homeobox gene l promotes vascular smooth muscle cell phenotypic modulation and vascular remodeling.

    Science.gov (United States)

    Wu, Bing; Zhang, Lei; Zhu, Yun-He; Zhang, You-En; Zheng, Fei; Yang, Jian-Ye; Guo, Ling-Yun; Li, Xing-Yuan; Wang, Lu; Tang, Jun-Ming; Chen, Shi-You; Wang, Jia-Ning

    2018-01-15

    To investigate the role of mesoderm/mesenchyme homeobox gene l (Meox1) in vascular smooth muscle cells (SMCs) phenotypic modulation during vascular remodeling. By using immunostaining, Western blot, and histological analyses, we found that Meox1 was up-regulated in PDGF-BB-treated SMCs in vitro and balloon injury-induced arterial SMCs in vivo. Meox1 knockdown by shRNA restored the expression of contractile SMCs phenotype markers including smooth muscle α-actin (α-SMA) and calponin. In contrast, overexpression of Moex1 inhibited α-SMA and calponin expressions while inducing the expressions of synthetic SMCs phenotype markers such as matrix gla protein, osteopontin, and proliferating cell nuclear antigen. Mechanistically, Meox1 mediated the SMCs phenotypic modulation through FAK-ERK1/2 signaling, which appears to induce autophagy in SMCs. In vivo, knockdown of Meox1 attenuated injury-induced neointima formation and promoted SMCs contractile proteins expressions. Meox1 knockdown also reduced the number of proliferating SMCs, suggesting that Meox1 was important for SMCs proliferation in vivo. Moreover, knockdown of Meox1 attenuated ERK1/2 signaling and autophagy markers expressions, suggesting that Meox1 may promote SMCs phenotypic modulation via ERK1/2 signaling-autophagy in vivo. Our data indicated that Meox1 promotes SMCs phenotypic modulation and injury-induced vascular remodeling by regulating the FAK-ERK1/2-autophagy signaling cascade. Thus, targeting Meox1 may be an attractive approach for treating proliferating vascular diseases. Copyright © 2017. Published by Elsevier B.V.

  16. Inhibition of soluble epoxide hydrolase augments astrocyte release of vascular endothelial growth factor and neuronal recovery after oxygen-glucose deprivation.

    Science.gov (United States)

    Zhang, Yue; Hong, Gina; Lee, Kin Sing Stephen; Hammock, Bruce D; Gebremedhin, Debebe; Harder, David R; Koehler, Raymond C; Sapirstein, Adam

    2017-03-01

    Epoxyeicosatrienoic acids (EETs) are synthesized in astrocytes, and inhibitors of soluble epoxide hydrolase (sEH), which hydrolyzes EETs, reduce infarct volume in ischemic stroke. Astrocytes can release protective neurotrophic factors, such as vascular endothelial growth factor (VEGF). We found that addition of sEH inhibitors to rat cultured astrocytes immediately after oxygen-glucose deprivation (OGD) markedly increased VEGF concentration in the medium 48 h later and the effect was blocked by an EET antagonist. The sEH inhibitors increased EET concentrations to levels capable of increasing VEGF. When the sEH inhibitors were removed from the medium at 48 h, the increase in VEGF persisted for an additional 48 h. Neurons exposed to OGD and subsequently to astrocyte medium previously conditioned with OGD plus sEH inhibitors showed increased phosphorylation of their VEGF receptor-2, less TUNEL staining, and increased phosphorylation of Akt, which was blocked by a VEGF receptor-2 antagonist. Our findings indicate that sEH inhibitors, applied to cultured astrocytes after an ischemia-like insult, can increase VEGF secretion. The released VEGF then enhances Akt-enabled cell survival signaling in neurons through activation of VEGF receptor-2 leading to less neuronal cell death. These results suggest a new strategy by which astrocytes can be leveraged to support neuroprotection. © 2016 International Society for Neurochemistry.

  17. Hypergravity Effects on Dendritic Cells and Vascular Wall Interactions

    Science.gov (United States)

    Bellik, L.; Parenti, A.; Ledda, F.; Basile, V.; Romano, G.; Fusi, F.; Monici, M.

    2009-01-01

    Dendritic cells (DCs), the most potent antigen-presenting cells inducing specific immune responses, are involved in the pathogenesis of atherosclerosis. In this inflammatory disease, DCs increase in number, being particularly abundant in the shoulder regions of plaques. Since the exposure to altered gravitational conditions results in a significant impairment of the immune function, the aim of this study was to investigate the effects of hypergravity on both the function of DCs and their interactions with the vascular wall cells. Monocytes from peripheral blood mononuclear cells of healthy volunteers were sorted by CD14+ magnetic beads selection, cultured for 6 days in medium supplemented with GM-CSF and IL-4, followed by a further maturation stimulus. DC phenotype, assessed by flow cytometry, showed a high expression of the specific DC markers CD80, CD86, HLA-DR and CD83. The DCs obtained were then exposed to hypergravitational stimuli and their phenotype, cytoskeleton, ability to activate lymphocytes and interaction with vascular wall cells were investigated. The findings showed that the exposure to hypergravity conditions resulted in a significant impairment of DC cytoskeletal organization, without affecting the expression of DC markers. Moreover, an increase in DC adhesion to human vascular smooth muscle cells and in their ability to activate lymphocytes was observed.

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

    Science.gov (United States)

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

    2016-01-01

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

  19. Water Soluble Fluorescent Carbon Nanodots from Biosource for Cells Imaging

    Directory of Open Access Journals (Sweden)

    Kumud Malika Tripathi

    2017-01-01

    Full Text Available Carbon nanodots (CNDs derived from a green precursor, kidney beans, was synthesized with high yield via a facile pyrolysis technique. The CND material was easily modified through simple oxidative treatment with nitric acid, leading to a high density “self-passivated” water soluble form (wsCNDs. The synthesized wsCNDs have been extensively characterized by using various microscopic and spectroscopic techniques and were crystalline in nature. The highly carboxylated wsCNDs possessed tunable-photoluminescence emission behavior throughout the visible region of the spectrum, demonstrating their application for multicolor cellular imaging of HeLa cells. The tunable-photoluminescence properties of “self-passivated” wsCNDs make them a promising candidate as a probe in biological cell-imaging applications.

  20. Soluble Factors on Stage to Direct Mesenchymal Stem Cells Fate

    Directory of Open Access Journals (Sweden)

    Cristina Sobacchi

    2017-05-01

    Full Text Available Mesenchymal stem cells (MSCs are multipotent stromal cells that are identified by in vitro plastic adherence, colony-forming capacity, expression of a panel of surface molecules, and ability to differentiate at least toward osteogenic, adipogenic, and chondrogenic lineages. They also produce trophic factors with immunomodulatory, proangiogenic, and antiapoptotic functions influencing the behavior of neighboring cells. On the other hand, a reciprocal regulation takes place; in fact, MSCs can be isolated from several tissues, and depending on the original microenvironment and the range of stimuli received from there, they can display differences in their essential characteristics. Here, we focus mainly on the bone tissue and how soluble factors, such as growth factors, cytokines, and hormones, present in this microenvironment can orchestrate bone marrow-derived MSCs fate. We also briefly describe the alteration of MSCs behavior in pathological settings such as hematological cancer, bone metastasis, and bone marrow failure syndromes. Overall, the possibility to modulate MSCs plasticity makes them an attractive tool for diverse applications of tissue regeneration in cell therapy. Therefore, the comprehensive understanding of the microenvironment characteristics and components better suited to obtain a specific MSCs response can be extremely useful for clinical use.

  1. Soluble endothelial cell-selective adhesion molecule and incident cardiovascular events in a multiethnic population.

    Science.gov (United States)

    Ren, Hao-Yu; Khera, Amit; de Lemos, James A; Ayers, Colby R; Rohatgi, Anand

    2017-09-01

    Cell adhesion molecules are key regulators of atherosclerotic plaque development, but circulating levels of soluble fragments, such as intercellular adhesion molecule (sICAM-1) and vascular cell adhesion molecule (sVCAM-1), have yielded conflicting associations with atherosclerotic cardiovascular disease (ASCVD). Endothelial cell-selective adhesion molecule (ESAM) is expressed exclusively in platelets and endothelial cells, and soluble ESAM (sESAM) levels have been associated with prevalent subclinical atherosclerosis. We therefore hypothesized that sESAM would be associated with incident ASCVD. sESAM, sICAM-1, and sVCAM-1 were measured in 2,442 participants without CVD in the Dallas Heart Study, a probability-based population sample aged 30-65 years enrolled between 2000 and 2002. ASCVD was defined as first myocardial infarction, stroke, coronary revascularization, or CV death. A total of 162 ASCVD events were analyzed over 10.4 years. Increasing sESAM was associated with ASCVD, independent of risk factors (HR Q4 vs Q1: 2.7, 95% CI 1.6-4.6). Serial adjustment for renal function, sICAM-1, VCAM-1, and prevalent coronary calcium did not attenuate these associations. Continuous ESAM demonstrated similar findings (HR 1.31, 95% CI 1.2-1.4). Addition of sESAM to traditional risk factors improved discrimination and reclassification (delta c-index: P = .009; integrated-discrimination-improvement index P = .001; net reclassification index = 0.42, 95% CI 0.15-0.68). Neither sICAM-1 nor sVCAM-1 was independently associated with ASCVD. sESAM but not sICAM-1 or sVCAM-1 levels are associated with incident ASCVD. Further studies are warranted to investigate the role of sESAM in ASCVD. Copyright © 2017 Elsevier Inc. All rights reserved.

  2. Molecular Characterization of Human Breast Tumor Vascular Cells

    OpenAIRE

    Bhati, Rajendra; Patterson, Cam; Livasy, Chad A.; Fan, Cheng; Ketelsen, David; Hu, Zhiyuan; Reynolds, Evangeline; Tanner, Catherine; Moore, Dominic T.; Gabrielli, Franco; Perou, Charles M.; Klauber-DeMore, Nancy

    2008-01-01

    A detailed understanding of the assortment of genes that are expressed in breast tumor vessels is needed to facilitate the development of novel, molecularly targeted anti-angiogenic agents for breast cancer therapies. Rapid immunohistochemistry using factor VIII-related antibodies was performed on sections of frozen human luminal-A breast tumors (n = 5) and normal breast (n = 5), followed by laser capture microdissection of vascular cells. RNA was extracted and amplified, and fluorescently la...

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

    DEFF Research Database (Denmark)

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

    2017-01-01

    PSCs into specialized cell types such as cardiomyocytes, endothelial cells, and vascular smooth muscle cells (VSMCs) may lead to a better understanding of developmental cardiovascular processes and potentiate progress of safe autologous regenerative therapies in pathological conditions. In this review, we summarize......Cardiovascular diseases remain the leading cause of death worldwide and current treatment strategies have limited effect of disease progression. It would be desirable to have better models to study developmental and pathological processes and model vascular diseases in laboratory settings...

  4. Molecular characterization of human breast tumor vascular cells.

    Science.gov (United States)

    Bhati, Rajendra; Patterson, Cam; Livasy, Chad A; Fan, Cheng; Ketelsen, David; Hu, Zhiyuan; Reynolds, Evangeline; Tanner, Catherine; Moore, Dominic T; Gabrielli, Franco; Perou, Charles M; Klauber-DeMore, Nancy

    2008-05-01

    A detailed understanding of the assortment of genes that are expressed in breast tumor vessels is needed to facilitate the development of novel, molecularly targeted anti-angiogenic agents for breast cancer therapies. Rapid immunohistochemistry using factor VIII-related antibodies was performed on sections of frozen human luminal-A breast tumors (n = 5) and normal breast (n = 5), followed by laser capture microdissection of vascular cells. RNA was extracted and amplified, and fluorescently labeled cDNA was synthesized and hybridized to 44,000-element long-oligonucleotide DNA microarrays. Statistical analysis of microarray was used to compare differences in gene expression between tumor and normal vascular cells, and Expression Analysis Systematic Explorer was used to determine enrichment of gene ontology categories. Protein expression of select genes was confirmed using immunohistochemistry. Of the 1176 genes that were differentially expressed between tumor and normal vascular cells, 55 had a greater than fourfold increase in expression level. The extracellular matrix gene ontology category was increased while the ribosome gene ontology category was decreased. Fibroblast activation protein, secreted frizzled-related protein 2, Janus kinase 3, and neutral sphingomyelinase 2 proteins localized to breast tumor endothelium as assessed by immunohistochemistry, showing significantly greater staining compared with normal tissue. These tumor endothelial marker proteins also exhibited increased expression in breast tumor vessels compared with that in normal tissues. Therefore, these genetic markers may serve as potential targets for the development of angiogenesis inhibitors.

  5. Water Soluble Polymers as Proton Exchange Membranes for Fuel Cells

    Directory of Open Access Journals (Sweden)

    Bing-Joe Hwang

    2012-03-01

    Full Text Available The relentless increase in the demand for useable power from energy-hungry economies continues to drive energy-material related research. Fuel cells, as a future potential power source that provide clean-at-the-point-of-use power offer many advantages such as high efficiency, high energy density, quiet operation, and environmental friendliness. Critical to the operation of the fuel cell is the proton exchange membrane (polymer electrolyte membrane responsible for internal proton transport from the anode to the cathode. PEMs have the following requirements: high protonic conductivity, low electronic conductivity, impermeability to fuel gas or liquid, good mechanical toughness in both the dry and hydrated states, and high oxidative and hydrolytic stability in the actual fuel cell environment. Water soluble polymers represent an immensely diverse class of polymers. In this comprehensive review the initial focus is on those members of this group that have attracted publication interest, principally: chitosan, poly (ethylene glycol, poly (vinyl alcohol, poly (vinylpyrrolidone, poly (2-acrylamido-2-methyl-1-propanesulfonic acid and poly (styrene sulfonic acid. The paper then considers in detail the relationship of structure to functionality in the context of polymer blends and polymer based networks together with the effects of membrane crosslinking on IPN and semi IPN architectures. This is followed by a review of pore-filling and other impregnation approaches. Throughout the paper detailed numerical results are given for comparison to today’s state-of-the-art Nafion® based materials.

  6. Nrf2/Keap1 system regulates vascular smooth muscle cell apoptosis for vascular homeostasis: role in neointimal formation after vascular injury.

    Science.gov (United States)

    Ashino, Takashi; Yamamoto, Masayuki; Numazawa, Satoshi

    2016-05-20

    Abnormal increases in vascular smooth muscle cells (VSMCs) in the intimal region after a vascular injury is a key event in developing neointimal hyperplasia. To maintain vascular function, proliferation and apoptosis of VSMCs is tightly controlled during vascular remodeling. NF-E2-related factor 2 (Nrf2)/Kelch-like ECH-associated protein 1 (Keap1) system, a key component of the oxidative stress response that acts in maintaining homeostasis, plays an important role in neointimal hyperplasia after a vascular injury; however, the role of Nrf2/Keap1 in VSMC apoptosis has not been clarified. Here we report that 14 days after arterial injury in mice, TUNEL-positive VSMCs are detected in both the neointimal and medial layers. These layers contain cells expressing high levels of Nrf2 but low Keap1 expression. In VSMCs, Keap1 depletion induces features of apoptosis, such as positive TUNEL staining and annexin V binding. These changes are associated with an increased expression of nuclear Nrf2. Simultaneous Nrf2 depletion inhibits Keap1 depletion-induced apoptosis. At 14 days after the vascular injury, Nrf2-deficient mice demonstrated fewer TUNEL-positive cells and increased neointimal formation in the neointimal and medial areas. The results suggest that the Nrf2/Keap1 system regulates VSMC apoptosis during neointimal formation, thereby inhibiting neointimal hyperplasia after a vascular injury.

  7. Salt-induced Na+/K+-ATPase-α/β expression involves soluble adenylyl cyclase in endothelial cells.

    Science.gov (United States)

    Mewes, Mirja; Nedele, Johanna; Schelleckes, Katrin; Bondareva, Olga; Lenders, Malte; Kusche-Vihrog, Kristina; Schnittler, Hans-Joachim; Brand, Stefan-Martin; Schmitz, Boris; Brand, Eva

    2017-10-01

    High dietary salt intake may lead to vascular stiffness, which predicts cardiovascular diseases such as heart failure, and myocardial and cerebral infarctions as well as renal impairment. The vascular endothelium is a primary target for deleterious salt effects leading to dysfunction and endothelial stiffness. We hypothesize that the Ca 2+ - and bicarbonate-activated soluble adenylyl cyclase (sAC) contributes to Na + /K + -ATPase expression regulation in vascular endothelial cells and is an important regulator of endothelial stiffness. In vitro stimulation of vascular endothelial cells with high sodium (150 mM Na + )-induced Na + /K + -ATPase-α and Na + /K + -ATPase-β protein expression determined by western blot. Promoter analyses revealed increased cAMP response element (CRE)-mediated Na + /K + -ATPase-α transcriptional activity under high sodium concentrations. Inhibition of sAC by the specific inhibitor KH7 or siRNA reduced the sodium effects. Flame photometry revealed increased intracellular sodium concentrations in response to high sodium stimulations, which were paralleled by elevated ATP levels. Using atomic force microscopy, a nano-technique that measures cellular stiffness and deformability, we detected significant endothelial stiffening under increased sodium concentrations, which was prevented by inhibition of sAC using KH7 and Na + /K + -ATPase using ouabain. Furthermore, analysis of primary aortic endothelial cells in an in vitro aging model revealed an impaired Na + /K + -ATPase-α sodium response and elevated intracellular sodium levels with cellular aging. We conclude that sAC mediates sodium-induced Na + /K + -ATPase expression in vascular endothelium and is an important regulator of endothelial stiffness. The reactivity of Na + /K + -ATPase-α expression regulation in response to high sodium seems to be impaired in aging endothelial cells and might be a component of endothelial dysfunction.

  8. VUV modification promotes endothelial cell proliferation on PTFE vascular grafts

    Science.gov (United States)

    Cezeaux, J. L.; Romoser, C. E.; Benson, R. S.; Buck, C. K.; Sackman, J. E.

    1998-05-01

    Small diameter (⩽6 mm ID ) synthetic vascular grafts, used as lower-limb vessel replacements in patients without suitable autologous saphenous veins, have a failure rate of 53% after 4 yr. Graft failure is due to thrombosis and intimal hyperplasia, an increase in smooth muscle cells in the lumen of the vessel which leads to progressive closing and ultimate occlusion of the vessel. In an effort to increase patency rates of synthetic grafts, investigators have seeded vascular grafts with endothelial cells prior to implantation in an attempt to control both thrombosis and smooth muscle proliferation. This technique has been successful for the development of an endothelial monolayer in animal trials, but has met with limited success in humans. The hydrophobicity, low surface energy, and weak electrical charge of expanded polytetrafluoroethylene (ePTFE) provides conditions which are not optimal for endothelial cell attachment. The purpose of this study is to evaluate the effect of vacuum ultraviolet (VUV) modification of ePTFE on endothelial cell adhesion and proliferation. Pieces of ePTFE graft material were exposed to 10, 20 or 40 W VUV radiation for 10, 20 or 40 min using a UV excimer lamp. Prior to cell adhesion and proliferation experiments, the grafts pieces were autoclaved and cut into pledgets. Half of the pledgets were precoated with fibronectin ( 20 μg/ml). Cell adhesion was measured by seeding 3H-thymidine labeled human umbilical vein endothelial cells (HUVEC) onto the pledgets for 60 min. The pledgets were then washed and the remaining radioactivity assayed using scintillation counting. For the cell proliferation experiments, pledgets were seeded with unlabeled HUVEC which were allowed to adhere to the graft material for 18 h. The cells were then exposed to 3H-thymidine ( 1 μCi/ml) for approximately 48 h and then washed to remove any unincorporated 3H-thymidine. Incorporation of 3H-thymidine was measured using scintillation counting. Four replicate

  9. Effects of infection with recombinant adenovirus on human vascular endothelial and smooth muscle cells

    NARCIS (Netherlands)

    Quax, P.H.A.; Lamfers, M.L.M.; Grimbergen, J.M.; Teeling, J.; Hoeben, R.C.; Nieuw Amerongen, G.P. van; Hinsbergh, V.W.M. van

    1996-01-01

    The plasminogen activation (PA) system is involved in vascular remodelling. Modulating its activity in vascular cells might be a way to interfere in processes such as angiogenesis and restenosis. Adenoviral vectors have become a favourable tool for direct gene transfer into vascular cells. In the

  10. Modeling human endothelial cell transformation in vascular neoplasias.

    Science.gov (United States)

    Wen, Victoria W; MacKenzie, Karen L

    2013-09-01

    Endothelial cell (EC)-derived neoplasias range from benign hemangioma to aggressive metastatic angiosarcoma, which responds poorly to current treatments and has a very high mortality rate. The development of treatments that are more effective for these disorders will be expedited by insight into the processes that promote abnormal proliferation and malignant transformation of human ECs. The study of primary endothelial malignancy has been limited by the rarity of the disease; however, there is potential for carefully characterized EC lines and animal models to play a central role in the discovery, development and testing of molecular targeted therapies for vascular neoplasias. This review describes molecular alterations that have been identified in EC-derived neoplasias, as well as the processes that underpin the immortalization and tumorigenic conversion of ECs. Human EC lines, established through the introduction of defined genetic elements or by culture of primary tumor tissue, are catalogued and discussed in relation to their relevance as models of vascular neoplasia.

  11. Laminaria japonica Polysaccharide Inhibits Vascular Calcification via Preventing Osteoblastic Differentiation of Vascular Smooth Muscle Cells.

    Science.gov (United States)

    Li, Xue-Ying; Li, Qiang-Ming; Fang, Qing; Zha, Xue-Qiang; Pan, Li-Hua; Luo, Jian-Ping

    2018-02-28

    This study aimed to investigate the effect and underlying mechanism of a purified Laminaria japonica polysaccharide (LJP61A) on preventing vascular calcification (VC). In the adenine-induced chronic renal failure (CRF) mice VC model and the β-glycerophosphate (β-GP)-induced vascular smooth muscle cells (VSMC) calcification model, LJP61A was found to significantly inhibit VC phenotypes as determined by biochemical analysis and von Kossa, alizarin red, and immunohistochemical staining. Meanwhile, LJP61A remarkably up-regulated the mRNA levels of VSMC related markers and down-regulated the mRNA levels of sodium-dependent phosphate cotransporter Pit-1. In addition, LJP61A could significantly decrease the protein levels of core-binding factor-1, osteocalcin, bone morphogenetic protein 2, and receptor activator for nuclear factor-κB ligand, and it can increase the protein levels of osteoprotegerin and matrix gla protein. These results indicated that LJP61A ameliorated VC both in vivo and in vitro via preventing osteoblastic differentiation of VSMC, suggesting LJP61A might be a potential therapeutic agent for VC in CRF patients.

  12. Characterization of vascular endothelial progenitor cells from chicken bone marrow

    Directory of Open Access Journals (Sweden)

    Bai Chunyu

    2012-05-01

    Full Text Available Abstract Background Endothelial progenitor cells (EPC are a type of stem cell used in the treatment of atherosclerosis, vascular injury and regeneration. At present, most of the EPCs studied are from human and mouse, whereas the study of poultry-derived EPCs has rarely been reported. In the present study, chicken bone marrow-derived EPCs were isolated and studied at the cellular level using immunofluorescence and RT-PCR. Results We found that the majority of chicken EPCs were spindle shaped. The growth-curves of chicken EPCs at passages (P 1, -5 and -9 were typically “S”-shaped. The viability of chicken EPCs, before and after cryopreservation was 92.2% and 81.1%, respectively. Thus, cryopreservation had no obvious effects on the viability of chicken EPCs. Dil-ac-LDL and FITC-UAE-1 uptake assays and immunofluorescent detection of the cell surface markers CD34, CD133, VEGFR-2 confirmed that the cells obtained in vitro were EPCs. Observation of endothelial-specific Weibel-Palade bodies using transmission electron microscopy further confirmed that the cells were of endothelial lineage. In addition, chicken EPCs differentiated into endothelial cells and smooth muscle cells upon induction with VEGF and PDGF-BB, respectively, suggesting that the chicken EPCs retained multipotency in vitro. Conclusions These results suggest that chicken EPCs not only have strong self-renewal capacity, but also the potential to differentiate into endothelial and smooth muscle cells. This research provides theoretical basis and experimental evidence for potential therapeutic application of endothelial progenitor cells in the treatment of atherosclerosis, vascular injury and diabetic complications.

  13. From here to there, progenitor cells and stem cells are everywhere in lung vascular remodeling

    Directory of Open Access Journals (Sweden)

    Rebecca L. Heise

    2016-08-01

    Full Text Available The field of stem cell biology, cell therapy and regenerative medicine has expanded almost exponentially in the last decade. Clinical trials are evaluating the potential therapeutic use of stem cells in many adult and pediatric lung diseases with vascular component, such as bronchopulmonary dysplasia (BPD, chronic obstructive pulmonary disease (COPD, idiopathic pulmonary fibrosis (IPF or pulmonary arterial hypertension (PAH. Extensive research activity is exploring lung resident and circulating progenitor cells and their contribution to vascular complications of chronic lung diseases, and researchers hope to use resident or circulating stem/progenitor cells to treat chronic lung diseases and their vascular complications. It is becoming more and more clear that progress in mechanobiology will help to understand the various influences of physical forces and extracellular matrix composition on the phenotype and features of the progenitor cells and stem cells. The current review provides an overview of current concepts in the field.

  14. Association of postmenopausal hormone replacement therapy with carotid atherosclerosis and soluble thrombomodulin: the vascular aging (EVA) study. Etude du Vieillissement Artériel.

    Science.gov (United States)

    Petit, Laure; van Oort, Floor V A; Le Gal, Grégoire; Mennen, Louise I; Alhenc-Gelas, Martine; Touboul, Pierre-Jean; Zureik, Mahmoud; Scarabin, Pierre-Yves

    2002-02-15

    Hormone replacement therapy (HRT) may reduce atherosclerosis among postmenopausal women, partly by reducing vascular endothelium damage. We have tested this hypothesis by evaluating the association of HRT with firstly, carotid intima media thickness (IMT) and plaques, and secondly, with endothelial cell damage, indicated by soluble thrombomodulin (sTM). Then, we tested the association between the two markers of atherosclerosis and the levels of sTM. Among 747 postmenopausal women included into the EVA study, we compared 154 HRT users (including 80% transdermal treatment) with 593 never users. Carotid IMT and plaques were measured with B-mode ultrasonography and sTM with ELISA. At least one plaque was detected among 13.6% of HRT users and 27.3% of never users. After adjustment for confounding factors, the odds ratio for the presence of plaque was 0.45 (95% confidence interval, 0.25-0.78, P=0.005) in HRT users in comparison with nonusers. HRT users had a slightly lower crude mean IMT than nonusers, but the difference was not significant. sTM was positively associated with mean IMT (P for trend=0.001) but not with plaques. Finally, estrogen users had a lower sTM level than nonusers (difference 0.14 ng/ml, P=0.03). As HRT was associated with sTM and plaques, but not with IMT, while sTM was only associated with IMT, our hypothesis was not confirmed. This suggests that the possible beneficial effects of HRT on atherosclerosis may not go through the endothelial cell damage assessed by plasma thrombomodulin.

  15. Vascular Wall-Resident Multipotent Stem Cells of Mesenchymal Nature within the Process of Vascular Remodeling: Cellular Basis, Clinical Relevance, and Implications for Stem Cell Therapy

    Directory of Open Access Journals (Sweden)

    Diana Klein

    2016-01-01

    Full Text Available Until some years ago, the bone marrow and the endothelial cell compartment lining the vessel lumen (subendothelial space were thought to be the only sources providing vascular progenitor cells. Now, the vessel wall, in particular, the vascular adventitia, has been established as a niche for different types of stem and progenitor cells with the capacity to differentiate into both vascular and nonvascular cells. Herein, vascular wall-resident multipotent stem cells of mesenchymal nature (VW-MPSCs have gained importance because of their large range of differentiation in combination with their distribution throughout the postnatal organism which is related to their existence in the adventitial niche, respectively. In general, mesenchymal stem cells, also designated as mesenchymal stromal cells (MSCs, contribute to the maintenance of organ integrity by their ability to replace defunct cells or secrete cytokines locally and thus support repair and healing processes of the affected tissues. This review will focus on the central role of VW-MPSCs within vascular reconstructing processes (vascular remodeling which are absolute prerequisite to preserve the sensitive relationship between resilience and stability of the vessel wall. Further, a particular advantage for the therapeutic application of VW-MPSCs for improving vascular function or preventing vascular damage will be discussed.

  16. Hypoxic contraction of cultured pulmonary vascular smooth muscle cells

    International Nuclear Information System (INIS)

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

    1990-01-01

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

  17. Resveratrol inhibits vascular smooth muscle cell proliferation and induces apoptosis.

    Science.gov (United States)

    Poussier, Bertrand; Cordova, Alfredo C; Becquemin, Jean-Pierre; Sumpio, Bauer E

    2005-12-01

    In France, despite a high intake of dietary cholesterol and saturated fat, the cardiovascular death rate is one of the lowest among developed countries. This "French paradox" has been postulated to be related to the high red wine intake in France. The aim of this study was to determine the effects of resveratrol, a major polyphenol component of red wine, on vascular smooth muscle cell (SMC) proliferation in vitro. SMCs were exposed to 10(-6) to 10(-4) M resveratrol and cell proliferation was assessed by cell counting. Cell cycle analysis was done by treating cells with propidium iodide followed by flow-activated cell sorting. Apoptosis was determined by terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick-end labeling staining. We demonstrate that resveratrol inhibited bovine aortic SMC proliferation in a dose-dependent manner. The lowest concentration of resveratrol resulting in a significant decrease in SMC proliferation compared with control was 10(-5) M. By flow cytometry, we observed a block in the G1-S phase of the SMC cycle. Resveratrol treatment also resulted in a dose-dependent apoptosis of SMCs but had no effects on SMC morphology. The results indicated that vascular SMC proliferation could be inhibited by resveratrol through a block on G1-S phase and by an increase in apoptosis. It supports the conjecture that red wine consumption may have a beneficial effect on cardiovascular mortality. Our results suggest that resveratrol inhibits, in a dose-dependent manner, smooth muscle cell proliferation, which may help to partially explain a beneficial effect of wine drinking. This inhibition is related to an early block in the cell cycle and also to a dose-dependent apoptotic effect. The present study demonstrates that resveratrol not only is an indirect marker of a healthy life style and alimentation but may also be directly responsible for the French paradox.

  18. 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.

  19. Decreased soluble cell adhesion molecules after tirofiban infusion in patients with unstable angina pectoris

    Directory of Open Access Journals (Sweden)

    Aliyev Emil

    2004-04-01

    Full Text Available Abstract Aim The inflammatory response, initiated by neutrophil and monocyte adhesion to endothelial cells, is important in the pathogenesis of acute coronary syndromes. Platelets play an important role in inflammatory process by interacting with monocytes and neutrophils. In this study, we investigated the effect of tirofiban on the levels of cell adhesion molecules (soluble intercellular adhesion molecule-1, sICAM-1, and vascular cell adhesion molecule-1, sVCAM-1 in patients with unstable angina pectoris (AP. Methods Thirty-five patients with unstable AP (Group I, ten patients with stable AP (Group II and ten subjects who had angiographycally normal coronary arteries (Group III were included the study. Group I was divided into two subgroups for the specific treatment regimens: Group IA (n = 15 received tirofiban and Group IB (n = 20 did not. Blood samples for investigating the cell adhesion molecules were drawn at zero time (baseline; 0 h in all patients and at 72 h in Group I. Results The baseline levels of sICAM-1 and sVCAM-1 were higher in Group I than in Groups II and III. They were higher in Group IA than in Group IB. However, the sICAM-1 and sVCAM-1 levels decreased significantly in Group IA after tirofiban infusion. In contrast, these levels remained unchanged or were increased above the baseline value in Group IB at 72 h. Conclusion The levels of cell adhesion molecules in patients with unstable AP decreased significantly after tirofiban infusion. Inhibition of platelet function by specific glycoprotein IIb/IIIa antagonists may decrease platelet-mediated inflammation and the ischemic end-point.

  20. 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.

  1. The cytotoxicity and genotoxicity of soluble and particulate cobalt in human lung fibroblast cells

    Energy Technology Data Exchange (ETDEWEB)

    Smith, Leah J.; Holmes, Amie L. [Wise Laboratory of Environmental and Genetic Toxicology, University of Southern Maine, 96 Falmouth St., P.O. Box 9300, Portland, ME 04101-9300 (United States); Maine Center for Environmental Toxicology and Health, University of Southern Maine, 96 Falmouth St., P.O. Box 9300, Portland, ME 04101-9300 (United States); Department of Applied Medical Science, University of Southern Maine, 96 Falmouth St., P.O. Box 9300, Portland, ME 04101-9300 (United States); Kandpal, Sanjeev Kumar; Mason, Michael D. [Department of Chemical and Biological Engineering, University of Maine, Orono, ME (United States); Zheng, Tongzhang [Department of Environmental Health Sciences, Yale School of Public Health, New Haven, CT (United States); Wise, John Pierce, E-mail: John.Wise@usm.maine.edu [Wise Laboratory of Environmental and Genetic Toxicology, University of Southern Maine, 96 Falmouth St., P.O. Box 9300, Portland, ME 04101-9300 (United States); Maine Center for Environmental Toxicology and Health, University of Southern Maine, 96 Falmouth St., P.O. Box 9300, Portland, ME 04101-9300 (United States); Department of Applied Medical Science, University of Southern Maine, 96 Falmouth St., P.O. Box 9300, Portland, ME 04101-9300 (United States)

    2014-08-01

    Cobalt exposure is increasing as cobalt demand rises worldwide due to its use in enhancing rechargeable battery efficiency, super-alloys, and magnetic products. Cobalt is considered a possible human carcinogen with the lung being a primary target. However, few studies have considered cobalt-induced toxicity in human lung cells. Therefore, in this study, we sought to determine the cytotoxicity and genotoxicity of particulate and soluble cobalt in human lung cells. Cobalt oxide and cobalt chloride were used as representative particulate and soluble cobalt compounds, respectively. Exposure to both particulate and soluble cobalt induced a concentration-dependent increase in cytotoxicity, genotoxicity, and intracellular cobalt ion levels. Based on intracellular cobalt ion levels, we found that soluble cobalt was more cytotoxic than particulate cobalt while particulate and soluble cobalt induced similar levels of genotoxicity. However, soluble cobalt induced cell cycle arrest indicated by the lack of metaphases at much lower intracellular cobalt concentrations compared to cobalt oxide. Accordingly, we investigated the role of particle internalization in cobalt oxide-induced toxicity and found that particle-cell contact was necessary to induce cytotoxicity and genotoxicity after cobalt exposure. These data indicate that cobalt compounds are cytotoxic and genotoxic to human lung fibroblasts, and solubility plays a key role in cobalt-induced lung toxicity. - Highlights: • Particulate and soluble cobalt are cytotoxic and genotoxic to human lung cells. • Soluble cobalt induces more cytotoxicity compared to particulate cobalt. • Soluble and particulate cobalt induce similar levels of genotoxicity. • Particle-cell contact is required for particulate cobalt-induced toxicity.

  2. Uncommon vascular tumor of the ovary. Primary ovarian epithelioid hemangioendothelioma or vascular sarcomatous transformation in ovarian germ cell tumor?

    Science.gov (United States)

    Illueca, Carmen; Machado, Isidro; García, Ana; Covisa, Amparo; Morales, Javier; Cruz, Julia; Traves, Victor; Almenar, Sergio

    2011-12-01

    Epithelioid hemangioendothelioma (EHE) is an unusual vascular tumor, which usually occurs in the soft tissue, liver, breast, lung and bone. We submit a case of EHE, a tumor never before reported in the ovary. A 20-year-old woman was admitted with a medical history of unilateral ovarian tumor. The right ovary was totally removed and histologically, the tumor was composed of epithelioid cells with eosinophilic cytoplasm and prominent intracytoplasmic vacuoles associated with myxohyaline matrix. No morphologic evidence of germ cell tumor was observed. Immunohistochemically, the tumor cells were positive for CD31 and CD34. However, all germ cell tumor markers were negative. The final diagnosis was EHE of the ovarian gland and sarcomatous transformation in ovarian germ cell tumor was excluded after extensive histopathological and immunohistochemical study. EHE is an uncommon vascular tumor, which is rarely seen in female genital tract and this is the first report of EHE in ovarian gland. Final diagnosis depends on histopathological and immunohistochemical features.

  3. Lipid-soluble smoke particles upregulate vascular smooth muscle ETB receptors via activation of mitogen-activating protein kinases and NF-kappaB pathways

    DEFF Research Database (Denmark)

    Xu, C.B.; Zheng, J.P.; Zhang, W.

    2008-01-01

    Cigarette smoke is a strong risk factor for cardiovascular disease. However, the underlying molecular mechanisms that lead to cigarette smoke-associated cardiovascular disease remain elusive. With functional and molecular methods, we demonstrate for the first time that lipid-soluble cigarette smoke...... particles (dimethylsulfoxide-soluble cigarette smoke particles; DSP) increased the expression of endothelin type B (ET(B)) receptors in arterial smooth muscle cells. The increased ET(B) receptors in arterial smooth muscle cells was documented as enhanced contractility (sensitive myograph technique...

  4. Soluble interleukin 2 receptors are released from activated human lymphoid cells in vitro

    International Nuclear Information System (INIS)

    Rubin, L.A.; Kurman, C.C.; Fritz, M.E.; Biddison, W.E.; Boutin, B.; Yarchoan, R.; Nelson, D.L.

    1985-01-01

    With the use of an enzyme-linked immunoabsorbent assay to measure soluble human interleukin 2 receptors (IL 2R), certain human T cell leukemia virus I (HTLV I)-positive T cell lines were found to spontaneously release large quantities of IL 2R into culture supernatants. This was not found with HTLV I-negative and IL 2 independent T cell lines, and only one of seven B cell-derived lines examined produced small amounts of IL 2R. In addition to this constitutive production of soluble IL 2R by certain cell lines, normal human peripheral blood mononuclear cells (PBMC) could be induced to release soluble IL 2R by plant lectins, the murine monoclonal antibody OKT3, tetanus toxoid, and allogeneic cells. Such activated cells also expressed cellular IL 2R measurable in detergent solubilized cell extracts. The generation of cellular and supernatant IL 2R was: dependent on cellular activation, rapid, radioresistant (3000 rad), and inhibited by cycloheximide treatment. NaDodSO 4 -polyacrylamide gel electrophoresis analysis of soluble IL 2R demonstrated molecules of apparent Mr = 35,000 to 40,000, and 45,000 to 50,000, respectively, somewhat smaller than the mature surface receptor on these cells. The release of soluble IL 2R appears to be a characteristic marker of T lymphocyte activation and might serve an immunoregulatory function during both normal and abnormal cell growth and differentiation

  5. Thrombomodulin Induces a Quiescent Phenotype and Inhibits Migration in Vascular Smooth Muscle Cells In Vitro.

    Science.gov (United States)

    Bass, Heather M; Beard, Richard S; Cha, Byeong J; Yuan, Sarah Y; Nelson, Peter R

    2016-01-01

    Loss of critical endothelial cell function and subsequent vascular smooth muscle cell (VSMC) migration is central to the pathology of injury-induced neointimal hyperplasia and recurrent stenosis. Thrombomodulin (TM), well known for its function as an endothelial surface anticoagulant, may have an unknown direct effect on VSMC physiology that would be lost after injury. Here, we examined a novel effect of TM on VSMC by testing the hypothesis that direct application of TM induces favorable changes to the morphology of VSMC and inhibits their migration. Primary human VSMC were harvested using the explant technique and used in early passage (1-4) for all experiments. Laser-scanning confocal fluorescent imaging was performed to assess the effect of soluble TM on VSMC morphology. In vitro, migration of VSMC was measured using: (1) a 4-hr modified Boyden chemotaxis assay and (2) a 24-hr electric cell-substrate impedance sensing injury migration assay. Migration experiments were conducted with VSMC exposed to increasing doses of soluble recombinant TM. Recombinant thrombin served as a positive control and serum-free media as a negative control for all experimentation. Data were analyzed using a Student's t-test or repeated measures analysis of variance where appropriate (α < 0.05). VSMC exposed to TM clearly demonstrated a quiescent morphology with organized stress fibers consistent with a quiescent, differentiated, contractile phenotype; whereas, thrombin stimulation led to an activated, dedifferentiated, synthetic phenotype. VSMC demonstrated a low, baseline level of migration in unstimulated serum-free conditions. Thrombin significantly stimulated VSMC migration as expected. TM, independent of thrombin, significantly inhibited baseline VSMC migration in a dose-response fashion. The maximal inhibition was observed at (5 μg/mL) with 70% reduction (56 ± 1.7 vs. 18 ± 3.5 cells/5 high-power fields, P = 0.0005). TM has a direct effect on VSMC resulting in a quiescent

  6. Construction of three-dimensional vascularized cardiac tissue with cell sheet engineering.

    Science.gov (United States)

    Sakaguchi, Katsuhisa; Shimizu, Tatsuya; Okano, Teruo

    2015-05-10

    Construction of three-dimensional (3D) tissues with pre-isolated cells is a promising achievement for novel medicine and drug-discovery research. Our laboratory constructs 3D tissues with an innovative and unique method for layering multiple cell sheets. Cell sheets maintain a high-efficiently regenerating function, because of the higher cell density and higher transplantation efficiency, compared to other cell-delivery methods. Cell sheets have already been applied in clinical applications for regenerative medicine in treating patients with various diseases. Therefore, in our search to develop a more efficient treatment with cell sheets, we are constructing 3D tissues by layering cell sheets. Native animal tissues and organs have an abundance of capillaries to supply oxygen and nutrients, and to remove waste molecules. In our investigation of vascularized cardiac cell sheets, we have found that endothelial cells within cell sheets spontaneously form blood vessel networks as in vivo capillaries. To construct even thicker 3D tissues by layering multiple cell sheets, it is critical to have a medium or blood flow within the vascular networks of the cell sheets. Therefore, to perfuse medium or blood in the cell sheet vascular network to maintain the viability of all cells, we developed two types of vascular beds; (1) a femoral muscle-based vascular bed, and (2) a synthetic collagen gel-based vascular bed. Both vascular beds successfully provide the critical flow of culture medium, which allows 12-layer cell sheets to survive. Such bioreactor systems, when combined with cell sheet engineering techniques, have produced functional vascularized 3D tissues. Here we explain and discuss the various processes to obtain vascular networks by properly connecting cell sheets and the engineering of 3D tissues. Copyright © 2014 Elsevier B.V. All rights reserved.

  7. Soluble suppressor supernatants elaborated by concanavalin A-activated human mononuclear cells. Characterization of a soluble suppressor of B cell immunoglobulin production

    International Nuclear Information System (INIS)

    Fleisher, T.A.; Greene, W.C.; Blaese, R.M.; Waldmann, T.A.

    1981-01-01

    Human peripheral blood mononuclear cells (PBMC) activated with the mitogenic lectin concanavalin A (Con A) elaborate a soluble immune suppressor supernatant (SISS) that contains at least 2 distinct suppressor factors. One of these, SISS-B, inhibits polyclonal B cell immunoglobulin production, whereas the other, SISS-T, suppresses T cell proliferation to both mitogens and antigens. The latter mediator is discussed in the companion paper. Characteristics of the human soluble suppressor of B cell immunoglobulin production (SISS-B) include: 1) inhibition by a noncytotoxic mechanism, 2) loss of activity in the presence of the monosaccharide L-rhamnose, 3) appearance within 8 to 16 hr after the addition of Con A, 4) elaboration by cells irradiated with 500 or 2000 rads, 5) production by highly purified T cells, 6) stability at pH 2.5 but instability at 56/sup o/C, and 7) m.w. of 60 to 80,000. These data indicate that after Con A activation, selected T cells not only become potent suppressor cells, but also generate a soluble saccharide-specific factor(s) that inhibits polyclonal immunoglobulin production by human B cells

  8. Role of glycosaminoglycans (GAGs) in regulation of the immunogenicity of human vascular endothelial cells.

    Science.gov (United States)

    Rix, D A; Douglas, M S; Talbot, D; Dark, J H; Kirby, J A

    1996-04-01

    Heparan sulphate is a common glycosaminoglycan component of proteoglycans present on the luminal surface of vascular endothelium. It has been proposed that an important function of these molecules is the sequestration of a range of proinflammatory and proadhesive cytokines. Such cytokines play a vital role during lymphocyte recruitment from the blood at sites of inflammation. In this study it is shown that the effects of interferon-gamma (IFN-gamma), but not of tumour necrosis factor-alpha (TNF-alpha), are inhibited by treatment with soluble heparin. Specifically, heparin was shown to inhibit the induction of class II MHC antigens and the up-regulation of intercellular adhesion molecule-1 (ICAM-1) produced by treatment of cultured human endothelial cells with IFN-gamma. Furthermore, it was shown that heparin blocked the enhanced adhesion of T lymphocytes to IFN-gamma-treated endothelial cells. Investigation of the inhibitory effects of other GAG molecules demonstrated a requirement for heparin-like structural domains as chondroitin sulphate was unable to inhibit the function of IFN-gamma. These results may explain reported immunosuppressive properties of heparin, and are consistent with the model that heparin may compete with cell surface GAGs to bind IFN-gamma, thereby reducing effective biological activity.

  9. 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.

  10. Nano- and microstructured materials for in vitro studies of the physiology of vascular cells

    Directory of Open Access Journals (Sweden)

    Alexandra M. Greiner

    2016-11-01

    Full Text Available The extracellular environment of vascular cells in vivo is complex in its chemical composition, physical properties, and architecture. Consequently, it has been a great challenge to study vascular cell responses in vitro, either to understand their interaction with their native environment or to investigate their interaction with artificial structures such as implant surfaces. New procedures and techniques from materials science to fabricate bio-scaffolds and surfaces have enabled novel studies of vascular cell responses under well-defined, controllable culture conditions. These advancements are paving the way for a deeper understanding of vascular cell biology and materials–cell interaction. Here, we review previous work focusing on the interaction of vascular smooth muscle cells (SMCs and endothelial cells (ECs with materials having micro- and nanostructured surfaces. We summarize fabrication techniques for surface topographies, materials, geometries, biochemical functionalization, and mechanical properties of such materials. Furthermore, various studies on vascular cell behavior and their biological responses to micro- and nanostructured surfaces are reviewed. Emphasis is given to studies of cell morphology and motility, cell proliferation, the cytoskeleton and cell-matrix adhesions, and signal transduction pathways of vascular cells. We finalize with a short outlook on potential interesting future studies.

  11. Expression of glycoprotein VI in vascular endothelial cells.

    Science.gov (United States)

    Sun, Bing; Tao, Lian; Lin, Shihua; Calingasan, Noel Y; Li, Jess; Tandon, Narendra N; Yoshitake, Masuhiro; Kambayashi, Jun-ichi

    2003-06-01

    Glycoprotein (GP) VI, a collagen receptor, plays a important role in collagen-mediated platelet aggregation and adhesion. To date, GPVI expression has been found only in platelets and megakaryocytes. In the present studies, we have demonstrated that GPVI was also expressed in cultured human umbilical vein endothelial cells (HUVEC) at both transcript and protein levels. Using a GPVI-specific probe, a approximately 6-kb band was detected in HUVEC as well as in platelets and megakaryoblastic cell lines by Northern blotting. Using polyclonal antibodies raised against platelet GPVI peptides, the same size band (57 kDa) was labeled with convulxin (CVX) after immuo-precipitation in both HUVEC and platelet lysates. In addition, a approximately 70-kDa band was also labeled in HUVEC. Surface expression of GPVI in HUVEC was confirmed by flow cytometry with GPVI-specific IgG or by direct labeling with FITC-conjugated CVX. Since HUVEC lack FcRgamma chain that forms complex with GPVI in platelets for signaling process, the function of GPVI in vascular endothelial cells remains to be determined.

  12. Effect of New Water-Soluble Dendritic Phthalocyanines on Human Colorectal and Liver Cancer Cell Lines

    Directory of Open Access Journals (Sweden)

    Ebru YABAŞ

    2017-08-01

    Full Text Available Human hepatocellular carcinoma (HepG2 cells and colorectal adenocarcinoma (DLD-1 cells were treated with the synthesized water soluble phthalocyanine derivatives to understand the effect of the compounds both on colorectal and liver cancer cells. The compounds inhibited cell proliferation and displayed cytotoxic effect on these cancer cell lines however; the effect of the compounds on healthy control fibroblast cell line was comparatively lower. The compounds can be employed for cancer treatment as anticancer agents.

  13. Nicotine promotes vascular endothelial growth factor secretion by human trophoblast cells under hypoxic conditions and improves the proliferation and tube formation capacity of human umbilical endothelial cells.

    Science.gov (United States)

    Zhao, Hongbo; Wu, Lanxiang; Wang, Yahui; Zhou, Jiayi; Li, Ruixia; Zhou, Jiabing; Wang, Zehua; Xu, Congjian

    2017-04-01

    Pre-eclampsia, characterized as defective uteroplacental vascularization, remains the major cause of maternal and fetal mortality and morbidity. Previous epidemiological studies demonstrated that cigarette smoking reduced the risk of pre-eclampsia. However, the molecular mechanism remains elusive. In the present study, it is demonstrated that a low dose of nicotine decreased soluble vascular endothelial growth factor receptor 1 (sFlt1) secretion in human trophoblast cells under hypoxic conditions. Nicotine was then observed to promote vascular endothelial growth factor (VEGF) secretion by reducing sFlt1 secretion and increasing VEGF mRNA transcription. Further data showed that nicotine enhanced hypoxia-mediated hypoxia-inducible factor-1α (HIF-1α) expression and HIF-1α small interfering RNA abrogated nicotine-induced VEGF secretion, indicating that HIF-1α may be responsible for nicotine-mediated VEGF transcription under hypoxic conditions. Moreover, conditioned medium from human trophoblast cells treated with nicotine under hypoxic conditions promoted the proliferation and tube formation capacity of human umbilical endothelial cells (HUVEC) by promoting VEGF secretion. These findings indicate that nicotine may promote VEGF secretion in human trophoblast cells under hypoxic conditions by reducing sFlt1 secretion and up-regulating VEGF transcription and improve the proliferation and tube formation of HUVEC cells, which may contribute to elucidate the protective effect of cigarette smoking against pre-eclampsia. Copyright © 2017 Reproductive Healthcare Ltd. Published by Elsevier Ltd. All rights reserved.

  14. Soluble Factors Secreted by T Cells Promote β-Cell Proliferation

    Science.gov (United States)

    Dirice, Ercument; Kahraman, Sevim; Jiang, Wenyu; El Ouaamari, Abdelfattah; De Jesus, Dario F.; Teo, Adrian K.K.; Hu, Jiang; Kawamori, Dan; Gaglia, Jason L.; Mathis, Diane; Kulkarni, Rohit N.

    2014-01-01

    Type 1 diabetes is characterized by infiltration of pancreatic islets with immune cells, leading to insulin deficiency. Although infiltrating immune cells are traditionally considered to negatively impact β-cells by promoting their death, their contribution to proliferation is not fully understood. Here we report that islets exhibiting insulitis also manifested proliferation of β-cells that positively correlated with the extent of lymphocyte infiltration. Adoptive transfer of diabetogenic CD4+ and CD8+ T cells, but not B cells, selectively promoted β-cell proliferation in vivo independent from the effects of blood glucose or circulating insulin or by modulating apoptosis. Complementary to our in vivo approach, coculture of diabetogenic CD4+ and CD8+ T cells with NOD.RAG1−/− islets in an in vitro transwell system led to a dose-dependent secretion of candidate cytokines/chemokines (interleukin-2 [IL-2], IL-6, IL-10, MIP-1α, and RANTES) that together enhanced β-cell proliferation. These data suggest that soluble factors secreted from T cells are potential therapeutic candidates to enhance β-cell proliferation in efforts to prevent and/or delay the onset of type 1 diabetes. PMID:24089508

  15. Energy consumption and water-soluble protein release by cell wall disruption of Nannochloropsis gaditana

    NARCIS (Netherlands)

    Safi, C.; Cabas Rodriguez, L.; Mulder, W.J.; Engelen-Smit, N.; Spekking, W.; Broek, van den L.A.M.; Olivieri, G.; Sijtsma, L.

    2017-01-01

    Several cell disruption methods were tested on Nannochloropsis gaditana, to evaluate their efficiency in terms of cell disintegration, energy input and release of soluble proteins. High-pressure homogenization (HPH) and bead milling were the most efficient with >95% cell disintegration, ±50%

  16. Matrix Metalloproteinases: Inflammatory Regulators of Cell Behaviors in Vascular Formation and Remodeling

    Directory of Open Access Journals (Sweden)

    Qishan Chen

    2013-01-01

    Full Text Available Abnormal angiogenesis and vascular remodeling contribute to pathogenesis of a number of disorders such as tumor, arthritis, atherosclerosis, restenosis, hypertension, and neurodegeneration. During angiogenesis and vascular remodeling, behaviors of stem/progenitor cells, endothelial cells (ECs, and vascular smooth muscle cells (VSMCs and its interaction with extracellular matrix (ECM play a critical role in the processes. Matrix metalloproteinases (MMPs, well-known inflammatory mediators are a family of zinc-dependent proteolytic enzymes that degrade various components of ECM and non-ECM molecules mediating tissue remodeling in both physiological and pathological processes. MMPs including MMP-1, MMP-2, MMP-3, MMP-7, MMP-8, MMP-9, MMP-12, and MT1-MMP, are stimulated and activated by various stimuli in vascular tissues. Once activated, MMPs degrade ECM proteins or other related signal molecules to promote recruitment of stem/progenitor cells and facilitate migration and invasion of ECs and VSMCs. Moreover, vascular cell proliferation and apoptosis can also be regulated by MMPs via proteolytically cleaving and modulating bioactive molecules and relevant signaling pathways. Regarding the importance of vascular cells in abnormal angiogenesis and vascular remodeling, regulation of vascular cell behaviors through modulating expression and activation of MMPs shows therapeutic potential.

  17. Cell Therapy Applications for Retinal Vascular Diseases: Diabetic Retinopathy and Retinal Vein Occlusion.

    Science.gov (United States)

    Park, Susanna S

    2016-04-01

    Retinal vascular conditions, such as diabetic retinopathy and retinal vein occlusion, remain leading causes of vision loss. No therapy exists to restore vision loss resulting from retinal ischemia and associated retinal degeneration. Tissue regeneration is possible with cell therapy. The goal would be to restore or replace the damaged retinal vasculature and the retinal neurons that are damaged and/or degenerating from the hypoxic insult. Currently, various adult cell therapies have been explored as potential treatment. They include mesenchymal stem cells, vascular precursor cells (i.e., CD34+ cells, hematopoietic cells or endothelial progenitor cells), and adipose stromal cells. Preclinical studies show that all these cells have a paracrine trophic effect on damaged ischemic tissue, leading to tissue preservation. Endothelial progenitor cells and adipose stromal cells integrate into the damaged retinal vascular wall in preclinical models of diabetic retinopathy and ischemia-reperfusion injury. Mesenchymal stem cells do not integrate as readily but appear to have a primary paracrine trophic effect. Early phase clinical trials have been initiated and ongoing using mesenchymal stem cells or autologous bone marrow CD34+ cells injected intravitreally as potential therapy for diabetic retinopathy or retinal vein occlusion. Adipose stromal cells or pluripotent stem cells differentiated into endothelial colony-forming cells have been explored in preclinical studies and show promise as possible therapies for retinal vascular disorders. The relative safety or efficacy of these various cell therapies for treating retinal vascular disorders have yet to be determined.

  18. Vasostatin-2 inhibits cell proliferation and adhesion in vascular smooth muscle cells, which are associated with the progression of atherosclerosis

    International Nuclear Information System (INIS)

    Hou, Jianghong; Xue, Xiaolin; Li, Junnong

    2016-01-01

    Recently, the serum expression level of vasostatin-2 was found to be reduced and is being studied as an important indicator to assess the presence and severity of coronary artery disease; the functional properties of vasostatin-2 and its relationship with the development of atherosclerosis remains unclear. In this study, we attempted to detect the expression of vasostatin-2 and its impact on human vascular smooth muscle cells (VSMCs). Quantitative real-time PCR (qRT-PCR) and western blot were used to assess the expression level of vasostatin-2 in VSMCs between those from atherosclerosis and disease-free donors; we found that vasostatin-2 was significantly down-regulated in atherosclerosis patient tissues and cell lines. In addition, the over-expression of vasostatin-2 apparently inhibits cell proliferation and migration in VSMCs. Gain-of-function in vitro experiments further show that vasostatin-2 over-expression significantly inhibits inflammatory cytokines release in VSMCs. In addition, cell adhesion experimental analysis showed that soluble adhesion molecules (sICAM-1, sVCAM-1) had decreased expression when vasostatin-2 was over-expressed in VSMCs. Therefore, our results indicate that vasostatin-2 is an atherosclerosis-related factor that can inhibit cell proliferation, inflammatory response and cell adhesion in VSMCs. Taken together, our results indicate that vasostatin-2 could serve as a potential diagnostic biomarker and therapeutic option for human atherosclerosis in the near future. - Highlights: • Vasostatin-2 levels were down-regulated in atherosclerosis patient tissues and VSMCs. • Ectopic expression of vasostatin-2 directly affects cell proliferation and migration in vitro. • Ectopic expression of vasostatin-2 protein affects pro-inflammatory cytokines release in VSMCs. • Ectopic expression of vasostatin-2 protein affects cell adhesion in VSMCs.

  19. Vasostatin-2 inhibits cell proliferation and adhesion in vascular smooth muscle cells, which are associated with the progression of atherosclerosis

    Energy Technology Data Exchange (ETDEWEB)

    Hou, Jianghong, E-mail: jianghonghou@163.com [Department of Cardiovascular, Weinan Center Hospital, The Middle of Victory Avenue, Linwei District, Weinan City 714000 (China); Xue, Xiaolin [Department of Cardiovascular, The First Affiliated Hospital, College of Medicine, Xi' an Jiaotong University, Xi' an 710061 (China); Li, Junnong [Department of Cardiovascular, Weinan Center Hospital, The Middle of Victory Avenue, Linwei District, Weinan City 714000 (China)

    2016-01-22

    Recently, the serum expression level of vasostatin-2 was found to be reduced and is being studied as an important indicator to assess the presence and severity of coronary artery disease; the functional properties of vasostatin-2 and its relationship with the development of atherosclerosis remains unclear. In this study, we attempted to detect the expression of vasostatin-2 and its impact on human vascular smooth muscle cells (VSMCs). Quantitative real-time PCR (qRT-PCR) and western blot were used to assess the expression level of vasostatin-2 in VSMCs between those from atherosclerosis and disease-free donors; we found that vasostatin-2 was significantly down-regulated in atherosclerosis patient tissues and cell lines. In addition, the over-expression of vasostatin-2 apparently inhibits cell proliferation and migration in VSMCs. Gain-of-function in vitro experiments further show that vasostatin-2 over-expression significantly inhibits inflammatory cytokines release in VSMCs. In addition, cell adhesion experimental analysis showed that soluble adhesion molecules (sICAM-1, sVCAM-1) had decreased expression when vasostatin-2 was over-expressed in VSMCs. Therefore, our results indicate that vasostatin-2 is an atherosclerosis-related factor that can inhibit cell proliferation, inflammatory response and cell adhesion in VSMCs. Taken together, our results indicate that vasostatin-2 could serve as a potential diagnostic biomarker and therapeutic option for human atherosclerosis in the near future. - Highlights: • Vasostatin-2 levels were down-regulated in atherosclerosis patient tissues and VSMCs. • Ectopic expression of vasostatin-2 directly affects cell proliferation and migration in vitro. • Ectopic expression of vasostatin-2 protein affects pro-inflammatory cytokines release in VSMCs. • Ectopic expression of vasostatin-2 protein affects cell adhesion in VSMCs.

  20. Soluble urokinase plasminogen activator receptor during allogeneic stem cell transplantation

    DEFF Research Database (Denmark)

    Haastrup, E; Andersen, J; Ostrowski, S R

    2011-01-01

    Previous studies have found that soluble urokinase plasminogen activation receptor (suPAR) increases during inflammatory and malignant illness and elevated suPAR levels may be associated with poor clinical outcome. The purpose of this study was to investigate plasma levels of suPAR during...... to pretreatment levels. High suPAR levels at day 0 were associated with increased mortality (P = 0.011). The present study found increased suPAR levels during the conditioning in SCT patients. Further, the data indicated that increased suPAR levels may be associated with increased mortality, suggesting su...

  1. A Fluid Membrane-Based Soluble Ligand Display System for Live CellAssays

    Energy Technology Data Exchange (ETDEWEB)

    Nam, Jwa-Min; Nair, Pradeep N.; Neve, Richard M.; Gray, Joe W.; Groves, Jay T.

    2005-10-14

    Cell communication modulates numerous biological processes including proliferation, apoptosis, motility, invasion and differentiation. Correspondingly, there has been significant interest in the development of surface display strategies for the presentation of signaling molecules to living cells. This effort has primarily focused on naturally surface-bound ligands, such as extracellular matrix components and cell membranes. Soluble ligands (e.g. growth factors and cytokines) play an important role in intercellular communications, and their display in a surface-bound format would be of great utility in the design of array-based live cell assays. Recently, several cell microarray systems that display cDNA, RNAi, or small molecules in a surface array format were proven to be useful in accelerating high-throughput functional genetic studies and screening therapeutic agents. These surface display methods provide a flexible platform for the systematic, combinatorial investigation of genes and small molecules affecting cellular processes and phenotypes of interest. In an analogous sense, it would be an important advance if one could display soluble signaling ligands in a surface assay format that allows for systematic, patterned presentation of soluble ligands to live cells. Such a technique would make it possible to examine cellular phenotypes of interest in a parallel format with soluble signaling ligands as one of the display parameters. Herein we report a ligand-modified fluid supported lipid bilayer (SLB) assay system that can be used to functionally display soluble ligands to cells in situ (Figure 1A). By displaying soluble ligands on a SLB surface, both solution behavior (the ability to become locally enriched by reaction-diffusion processes) and solid behavior (the ability to control the spatial location of the ligands in an open system) could be combined. The method reported herein benefits from the naturally fluid state of the supported membrane, which allows

  2. Vascular Endothelial Growth Factor (VEGF) Bioavailability Regulates Angiogenesis and Intestinal Stem and Progenitor Cell Proliferation during Postnatal Small Intestinal Development.

    Science.gov (United States)

    Schlieve, Christopher R; Mojica, Salvador Garcia; Holoyda, Kathleen A; Hou, Xiaogang; Fowler, Kathryn L; Grikscheit, Tracy C

    2016-01-01

    Vascular endothelial growth factor (VEGF) is a highly conserved, master regulatory molecule required for endothelial cell proliferation, organization, migration and branching morphogenesis. Podocoryne carnea and drosophila, which lack endothelial cells and a vascular system, express VEGF homologs, indicating potential roles beyond angiogenesis and vasculogenesis. The role of VEGF in the development and homeostasis of the postnatal small intestine is unknown. We hypothesized regulating VEGF bioavailability in the postnatal small intestine would exhibit effects beyond the vasculature and influence epithelial cell stem/progenitor populations. VEGF mutant mice were created that overexpressed VEGF in the brush border of epithelium via the villin promotor following doxycycline treatment. To decrease VEGF bioavailability, sFlt-1 mutant mice were generated that overexpressed the soluble VEGF receptor sFlt-1 upon doxycycline administration in the intestinal epithelium. Mice were analyzed after 21 days of doxycycline administration. Increased VEGF expression was confirmed by RT-qPCR and ELISA in the intestine of the VEGF mutants compared to littermates. The VEGF mutant duodenum demonstrated increased angiogenesis and vascular leak as compared to littermate controls. The VEGF mutant duodenum revealed taller villi and increased Ki-67-positive cells in the transit-amplifying zone with reduced Lgr5 expression. The duodenum of sFlt-1 mutants revealed shorter villi and longer crypts with reduced proliferation in the transit-amplifying zone, reduced expression of Dll1, Bmp4 and VE-cadherin, and increased expression of Sox9 and EphB2. Manipulating VEGF bioavailability leads to profound effects on not only the intestinal vasculature, but epithelial stem and progenitor cells in the intestinal crypt. Elucidation of the crosstalk between VEGF signaling in the vasculature, mesenchyme and epithelial stem/progenitor cell populations may direct future cell therapies for intestinal

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

    Directory of Open Access Journals (Sweden)

    Sayo Koike

    2016-09-01

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

  4. Promoting Tropoelastin Expression in Arterial and Venous Vascular Smooth Muscle Cells and Fibroblasts for Vascular Tissue Engineering.

    Science.gov (United States)

    Rothuizen, Tonia C; Kemp, Raymond; Duijs, Jacques M G J; de Boer, Hetty C; Bijkerk, Roel; van der Veer, Eric P; Moroni, Lorenzo; van Zonneveld, Anton Jan; Weiss, Anthony S; Rabelink, Ton J; Rotmans, Joris I

    2016-10-01

    Elastin, critical for its structural and regulatory functions, is a missing link in vascular tissue engineering. Several elastin-inducting compounds have previously been reported, but their relative efficiency in promoting elastogenesis by adult arterial and venous vascular smooth muscle cells (VSMCs) and fibroblasts, four main vascular and elastogenic cells, has not been described. In addition to elasto-inductive substances, microRNA-29a was recently established as a potent post-transcriptional inhibitor of elastogenesis. Here, we explored if stimulating positive regulators or blocking inhibitors of elastogenesis could maximize elastin production. We tested whether the elasto-inducing compounds IGF-1, TGF-β1, and minoxidil could indeed augment elastin production, and whether microRNA-29a antagonism could block elastin production in adult arterial and venous fibroblasts and VSMCs. The effects on elastin, lysyl oxidase, and fibrillin-1 mRNA expression levels and tropoelastin protein were determined. IGF-1 and minoxidil exerted little effect on tropoelastin mRNA expression levels in all cell types, while TGF-β1 predominantly enhanced mRNA tropoelastin levels, but this mRNA increase did not impact tropoelastin protein abundance. In contrast, microRNA29a inhibition resulted in the upregulation of tropoelastin mRNA in all cell types, but most pronounced in venous VSMCs. Importantly, microRNA-29a-antagonism also enhanced lysyl oxidase and fibrillin-1 mRNA expression, and revealed a dose-dependent increase in tropoelastin protein expression in venous VSMCs. Our studies suggest that the elastogenic potential of microRNA-29a inhibition in vascular cells is superior to that of established elastin-stimulating compounds IGF-1, TGF-β1, and minoxidil. Thus, microRNA-29a antagonism could serve as an attractive means of enhancing elastin synthesis in tissue-engineered blood vessels.

  5. The mast cell-B cell axis in lung vascular remodeling and pulmonary hypertension.

    Science.gov (United States)

    Breitling, Siegfried; Hui, Zhang; Zabini, Diana; Hu, Yijie; Hoffmann, Julia; Goldenberg, Neil M; Tabuchi, Arata; Buelow, Roland; Dos Santos, Claudia; Kuebler, Wolfgang M

    2017-05-01

    Over past years, a critical role for the immune system and, in particular, for mast cells in the pathogenesis of pulmonary hypertension (PH) has emerged. However, the way in which mast cells promote PH is still poorly understood. Here, we investigated the mechanisms by which mast cells may contribute to PH, specifically focusing on the interaction between the innate and adaptive immune response and the role of B cells and autoimmunity. Experiments were performed in Sprague-Dawley rats and B cell-deficient JH-KO rats in the monocrotaline, Sugen/hypoxia, and the aortic banding model of PH. Hemodynamics, cell infiltration, IL-6 expression, and vascular remodeling were analyzed. Gene array analyses revealed constituents of immunoglobulins as most prominently regulated mast cell-dependent genes in the lung in experimental PH. IL-6 was shown to link mast cells to B cells, as 1 ) IL-6 was upregulated and colocalized with mast cells and was reduced by mast-cell stabilizers and 2 ) IL-6 or mast cell blockade reduced B cells in lungs of monocrotaline-treated rats. A functional role for B cells in PH was demonstrated in that either blocking B cells by an anti-CD20 antibody or B-cell deficiency in JH-KO rats attenuated right ventricular systolic pressure and vascular remodeling in experimental PH. We here identify a mast cell-B cell axis driven by IL-6 as a critical immune pathway in the pathophysiology of PH. Our results provide novel insights into the role of the immune system in PH, which may be therapeutically exploited by targeted immunotherapy. Copyright © 2017 the American Physiological Society.

  6. Reactive oxygen species and angiotensin II signaling in vascular cells: implications in cardiovascular disease

    Directory of Open Access Journals (Sweden)

    Touyz R.M.

    2004-01-01

    Full Text Available Diseases such as hypertension, atherosclerosis, hyperlipidemia, and diabetes are associated with vascular functional and structural changes including endothelial dysfunction, altered contractility and vascular remodeling. Cellular events underlying these processes involve changes in vascular smooth muscle cell (VSMC growth, apoptosis/anoikis, cell migration, inflammation, and fibrosis. Many factors influence cellular changes, of which angiotensin II (Ang II appears to be amongst the most important. The physiological and pathophysiological actions of Ang II are mediated primarily via the Ang II type 1 receptor. Growing evidence indicates that Ang II induces its pleiotropic vascular effects through NADPH-driven generation of reactive oxygen species (ROS. ROS function as important intracellular and intercellular second messengers to modulate many downstream signaling molecules, such as protein tyrosine phosphatases, protein tyrosine kinases, transcription factors, mitogen-activated protein kinases, and ion channels. Induction of these signaling cascades leads to VSMC growth and migration, regulation of endothelial function, expression of pro-inflammatory mediators, and modification of extracellular matrix. In addition, ROS increase intracellular free Ca2+ concentration ([Ca2+]i, a major determinant of vascular reactivity. ROS influence signaling molecules by altering the intracellular redox state and by oxidative modification of proteins. In physiological conditions, these events play an important role in maintaining vascular function and integrity. Under pathological conditions ROS contribute to vascular dysfunction and remodeling through oxidative damage. The present review focuses on the biology of ROS in Ang II signaling in vascular cells and discusses how oxidative stress contributes to vascular damage in cardiovascular disease.

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

    Energy Technology Data Exchange (ETDEWEB)

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

    2016-05-01

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

  8. Effect of lovastatin on rabbit vascular smooth muscle cells

    International Nuclear Information System (INIS)

    Luan Zhaoxia; Pei Zhuguo

    2003-01-01

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

  9. Evolution of plant conducting cells: perspectives from key regulators of vascular cell differentiation.

    Science.gov (United States)

    Ohtani, Misato; Akiyoshi, Nobuhiro; Takenaka, Yuto; Sano, Ryosuke; Demura, Taku

    2017-01-01

    One crucial problem that plants faced during their evolution, particularly during the transition to growth on land, was how to transport water, nutrients, metabolites, and small signaling molecules within a large, multicellular body. As a solution to this problem, land plants developed specific tissues for conducting molecules, called water-conducting cells (WCCs) and food-conducting cells (FCCs). The well-developed WCCs and FCCs in extant plants are the tracheary elements and sieve elements, respectively, which are found in vascular plants. Recent molecular genetic studies revealed that transcriptional networks regulate the differentiation of tracheary and sieve elements, and that the networks governing WCC differentiation are largely conserved among land plant species. In this review, we discuss the molecular evolution of plant conducting cells. By focusing on the evolution of the key transcription factors that regulate vascular cell differentiation, the NAC transcription factor VASCULAR-RELATED NAC-DOMAIN for WCCs and the MYB-coiled-coil (CC)-type transcription factor ALTERED PHLOEM DEVELOPMENT for sieve elements, we describe how land plants evolved molecular systems to produce the specialized cells that function as WCCs and FCCs. © The Author 2016. Published by Oxford University Press on behalf of the Society for Experimental Biology. All rights reserved. For permissions, please email: journals.permissions@oup.com.

  10. Vascular Cell Induction Culture System Using Arabidopsis Leaves (VISUAL) Reveals the Sequential Differentiation of Sieve Element-Like Cells

    Science.gov (United States)

    Kondo, Yuki; Nurani, Alif Meem; Saito, Chieko; Ichihashi, Yasunori; Saito, Masato; Yamazaki, Kyoko; Mitsuda, Nobutaka; Ohme-Takagi, Masaru; Fukuda, Hiroo

    2016-01-01

    Cell differentiation is a complex process involving multiple steps, from initial cell fate specification to final differentiation. Procambial/cambial cells, which act as vascular stem cells, differentiate into both xylem and phloem cells during vascular development. Recent studies have identified regulatory cascades for xylem differentiation. However, the molecular mechanism underlying phloem differentiation is largely unexplored due to technical challenges. Here, we established an ectopic induction system for phloem differentiation named Vascular Cell Induction Culture System Using Arabidopsis Leaves (VISUAL). Our results verified similarities between VISUAL-induced Arabidopsis thaliana phloem cells and in vivo sieve elements. We performed network analysis using transcriptome data with VISUAL to dissect the processes underlying phloem differentiation, eventually identifying a factor involved in the regulation of the master transcription factor gene APL. Thus, our culture system opens up new avenues not only for genetic studies of phloem differentiation, but also for future investigations of multidirectional differentiation from vascular stem cells. PMID:27194709

  11. Vascular smooth muscle cells remodel collagen matrices by long-distance action and anisotropic interaction

    NARCIS (Netherlands)

    van den Akker, Jeroen; Guvenc Tuna, Bilge; Pistea, Adrian; Sleutel, Arie J. J.; Bakker, Erik N. T. P.; van Bavel, Ed

    2012-01-01

    While matrix remodeling plays a key role in vascular physiology and pathology, the underlying mechanisms have remained incompletely understood. We studied the remodeling of collagen matrices by individual vascular smooth muscle cells (SMCs), clusters and monolayers. In addition, we focused on the

  12. The Cytotoxicity and Genotoxicity of Particulate and Soluble Cobalt in Human Urothelial Cells.

    Science.gov (United States)

    Speer, Rachel M; The, Therry; Xie, Hong; Liou, Louis; Adam, Rosalyn M; Wise, John Pierce

    2017-11-01

    Cobalt use is increasing particularly due to its use as one of the primary metals in cobalt-chromium-molybdenum (CoCrMo) metal-on-metal prosthetics. CoCrMo is a high-strength, wear-resistant alloy with reduced risk for prosthetic loosening and device fracture. More than 500,000 people receive hip implants each year in the USA which puts them at potential risk for exposure to metal ions and particles released by the prosthetic implants. Data show cobalt ions released from prosthetics reach the bloodstream and accumulate in the bladder. As patients with failed hip implants show increased urinary and blood cobalt levels, no studies have considered the effects of cobalt on human urothelial cells. Accordingly, we investigated the cytotoxic and genotoxic effects of particulate and soluble cobalt in urothelial cells. Exposure to both particulate and soluble cobalt resulted in a concentration-dependent increase in cytotoxicity, genotoxicity, and intracellular cobalt ions. Based on intracellular cobalt ion levels, we found, when compared to particulate cobalt, soluble cobalt was more cytotoxic, but induced similar levels of genotoxicity. Interestingly, at similar intracellular cobalt ion concentrations, soluble cobalt induced cell cycle arrest indicated by a lack of metaphases not observed after particulate cobalt treatment. These data indicate that cobalt compounds are cytotoxic and genotoxic to human urothelial cells and solubility may play a key role in cobalt-induced toxicity.

  13. Soluble guanylyl cyclase is involved in PDT-induced injury of crayfish glial cells

    Science.gov (United States)

    Kovaleva, V. D.; Uzdensky, A. B.

    2016-04-01

    Photodynamic therapy (PDT) is a potential tool for selective destruction of malignant brain tumors. However, not only malignant but also healthy neurons and glial cells may be damaged during PDT. Nitric oxide is an important modulator of cell viability and intercellular neuroglial communications. NO have been already shown to participate in PDT-induced injury of neurons and glial cells. As soluble guanylyl cyclase is the only known receptor for NO, we have studied the possible role of soluble guanylyl cyclase in the regulation of survival and death of neurons and surrounding glial cells under photo-oxidative stress induced by photodynamic treatment (PDT). The crayfish stretch receptor consisting of a single identified sensory neuron enveloped by glial cells is a simple but informative model object. It was photosensitized with alumophthalocyanine photosens (10 nM) and irradiated with a laser diode (670 nm, 0.4 W/cm2). Using inhibitory analysis we have shown that during PDT soluble guanylyl cyclase, probably, has proapoptotic and antinecrotic effect on the glial cells of the isolated crayfish stretch receptor. Proapoptotic effect of soluble guanylyl cyclase could be mediated by protein kinase G (PKG). Thus, the involvement of NO/sGC/cGMP/PKG signaling pathway in PDT-induced apoptosis of glial cells was indirectly demonstrated.

  14. Clarification of mural cell coverage of vascular endothelial cells by live imaging of zebrafish.

    Science.gov (United States)

    Ando, Koji; Fukuhara, Shigetomo; Izumi, Nanae; Nakajima, Hiroyuki; Fukui, Hajime; Kelsh, Robert N; Mochizuki, Naoki

    2016-04-15

    Mural cells (MCs) consisting of vascular smooth muscle cells and pericytes cover the endothelial cells (ECs) to regulate vascular stability and homeostasis. Here, we clarified the mechanism by which MCs develop and cover ECs by generating transgenic zebrafish lines that allow live imaging of MCs and by lineage tracing in vivo To cover cranial vessels, MCs derived from either neural crest cells or mesoderm emerged around the preformed EC tubes, proliferated and migrated along EC tubes. During their migration, the MCs moved forward by extending their processes along the inter-EC junctions, suggesting a role for inter-EC junctions as a scaffold for MC migration. In the trunk vasculature, MCs derived from mesoderm covered the ventral side of the dorsal aorta (DA), but not the posterior cardinal vein. Furthermore, the MCs migrating from the DA or emerging around intersegmental vessels (ISVs) preferentially covered arterial ISVs rather than venous ISVs, indicating that MCs mostly cover arteries during vascular development. Thus, live imaging and lineage tracing enabled us to clarify precisely how MCs cover the EC tubes and to identify the origins of MCs. © 2016. Published by The Company of Biologists Ltd.

  15. 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.

  16. MCPIP1 Downregulation in Clear Cell Renal Cell Carcinoma Promotes Vascularization and Metastatic Progression.

    Science.gov (United States)

    Marona, Paulina; Górka, Judyta; Mazurek, Zofia; Wilk, Waclaw; Rys, Janusz; Majka, Marcin; Jura, Jolanta; Miekus, Katarzyna

    2017-09-15

    Clear cell renal cell carcinoma (ccRCC) is the most common type of kidney cancer and it forms highly vascularized tumors. The monocyte endoribonuclease MCPIP1 negatively regulates inflammation by degrading mRNA encoding proinflammatory cytokines, such as IL6, IL1, and IL12. MCPIP1 is also a negative regulator of NFκB and AP1 activity and it influences a broad range of miRNA activities. Here we report that MCPIP1 protein levels are decreased during renal cancer progression. In patient-derived tumors and xenografts established in NOD-SCID or nude mice, low MCPIP1 levels correlated strongly with increased proliferation, tumor outgrowth, and vascularity. MCPIP1 activity regulated secretion of VEGF, IL8, and CXCL12 leading to chemotaxis of microvascular endothelial cells, phosphorylation of VE-cadherin, and increased vascular permeability. Mechanistic investigations showed that MCPIP1 regulated ccRCC cell motility, lung metastasis, and mesenchymal phenotype by regulating key elements in the EMT signaling axis. Overall, our results illuminate how MCPIP1 serves as a key nodal point in coordinating tumor growth, angiogenesis, and metastatic spread in ccRCC. Cancer Res; 77(18); 4905-20. ©2017 AACR . ©2017 American Association for Cancer Research.

  17. Soluble CD54 induces human endothelial cells ex vivo expansion useful for cardiovascular regeneration and tissue engineering application

    KAUST Repository

    Malara, N.M.

    2015-03-01

    Aim: Consistent expansion of primary human endothelial cells in vitro is critical in the development of engineered tissue. A variety of complex culture media and techniques developed from different basal media have been reported with alternate success. Incongruous results are further confounded by donor-to-donor variability and cellular source of derivation. Our results demonstrate how to overcome these limitations using soluble CD54 (sCD54) as additive to conventional culture medium. Methods and results: Isolated primary fragment of different vessel types was expanded in Ham\\'s F12 DMEM, enriched with growth factors, Fetal Calf Serum and conditioned medium of Human Umbilical Vein Endothelial Cells (HUVEC) collected at different passages. Cytokine content of culture media was analyzed in order to identify the soluble factors correlating with better proliferation profile. sCD54 was found to induce the in vitro expansion of human endothelial cells (HECs) independently from the vessels source and even in the absence of HUVEC-conditioned medium. The HECs cultivated in the presence of sCD54 (50 ng/ml), resulted positive for the expression of CD146 and negative for CD45, and lower fibroblast contamination. Cells were capable to proliferate with an S phase of 25%, to produce vascular endothelial growth factor, VEGF, (10 ng/ml) and to give origin to vessel-like tubule in vitro. Conclusion: Our results demonstrate that sCD54 is an essential factor for the in-vitro expansion of HECs without donor and vessel-source variability. Resulting primary cultures can be useful, for tissue engineering in regenerative medicine (e.g. artificial micro tissue generation, coating artificial heart valve etc.) and bio-nanotechnology applications. © 2015 The Authors. Published by Elsevier Ireland Ltd.

  18. Activation of human natural killer cells by the soluble form of cellular prion protein

    Energy Technology Data Exchange (ETDEWEB)

    Seong, Yeon-Jae [Laboratory of Immunology and Infectious Diseases, Graduate School of Medical Science and Engineering, KAIST, Daejeon (Korea, Republic of); Hafis Clinic, Seoul (Korea, Republic of); Sung, Pil Soo; Jang, Young-Soon; Choi, Young Joon [Laboratory of Immunology and Infectious Diseases, Graduate School of Medical Science and Engineering, KAIST, Daejeon (Korea, Republic of); Park, Bum-Chan [Aging Research Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon (Korea, Republic of); Park, Su-Hyung [Laboratory of Translational Immunology and Vaccinology, Graduate School of Medical Science and Engineering, KAIST, Daejeon (Korea, Republic of); Park, Young Woo [Aging Research Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon (Korea, Republic of); Shin, Eui-Cheol, E-mail: ecshin@kaist.ac.kr [Laboratory of Immunology and Infectious Diseases, Graduate School of Medical Science and Engineering, KAIST, Daejeon (Korea, Republic of)

    2015-08-21

    Cellular prion protein (PrP{sup C}) is widely expressed in various cell types, including cells of the immune system. However, the specific roles of PrP{sup C} in the immune system have not been clearly elucidated. In the present study, we investigated the effects of a soluble form of recombinant PrP{sup C} protein on human natural killer (NK) cells. Recombinant soluble PrP{sup C} protein was generated by fusion of human PrP{sup C} with the Fc portion of human IgG{sub 1} (PrP{sup C}-Fc). PrP{sup C}-Fc binds to the surface of human NK cells, particularly to CD56{sup dim} NK cells. PrP{sup C}-Fc induced the production of cytokines and chemokines and the degranulation of granzyme B from NK cells. In addition, PrP{sup C}-Fc facilitated the IL-15-induced proliferation of NK cells. PrP{sup C}-Fc induced phosphorylation of ERK-1/2 and JNK in NK cells, and inhibitors of the ERK or the JNK pathways abrogated PrP{sup C}-Fc-induced cytokine production in NK cells. In conclusion, the soluble form of recombinant PrP{sup C}-Fc protein activates human NK cells via the ERK and JNK signaling pathways. - Highlights: • Recombinant soluble PrP{sup C} (PrP{sup C}-Fc) was generated by fusion of human PrP{sup C} with IgG1 Fc portion. • PrP{sup C}-Fc protein induces the production of cytokines and degranulation from human NK cells. • PrP{sup C}-Fc protein enhances the IL-15-induced proliferation of human NK cells. • PrP{sup C}-Fc protein activates human NK cells via the ERK and JNK signaling pathways.

  19. Activation of human natural killer cells by the soluble form of cellular prion protein

    International Nuclear Information System (INIS)

    Seong, Yeon-Jae; Sung, Pil Soo; Jang, Young-Soon; Choi, Young Joon; Park, Bum-Chan; Park, Su-Hyung; Park, Young Woo; Shin, Eui-Cheol

    2015-01-01

    Cellular prion protein (PrP C ) is widely expressed in various cell types, including cells of the immune system. However, the specific roles of PrP C in the immune system have not been clearly elucidated. In the present study, we investigated the effects of a soluble form of recombinant PrP C protein on human natural killer (NK) cells. Recombinant soluble PrP C protein was generated by fusion of human PrP C with the Fc portion of human IgG 1 (PrP C -Fc). PrP C -Fc binds to the surface of human NK cells, particularly to CD56 dim NK cells. PrP C -Fc induced the production of cytokines and chemokines and the degranulation of granzyme B from NK cells. In addition, PrP C -Fc facilitated the IL-15-induced proliferation of NK cells. PrP C -Fc induced phosphorylation of ERK-1/2 and JNK in NK cells, and inhibitors of the ERK or the JNK pathways abrogated PrP C -Fc-induced cytokine production in NK cells. In conclusion, the soluble form of recombinant PrP C -Fc protein activates human NK cells via the ERK and JNK signaling pathways. - Highlights: • Recombinant soluble PrP C (PrP C -Fc) was generated by fusion of human PrP C with IgG1 Fc portion. • PrP C -Fc protein induces the production of cytokines and degranulation from human NK cells. • PrP C -Fc protein enhances the IL-15-induced proliferation of human NK cells. • PrP C -Fc protein activates human NK cells via the ERK and JNK signaling pathways

  20. Cheiradone: a vascular endothelial cell growth factor receptor antagonist

    Directory of Open Access Journals (Sweden)

    Ahmed Nessar

    2008-01-01

    Full Text Available Abstract Background Angiogenesis, the growth of new blood vessels from the pre-existing vasculature is associated with physiological (for example wound healing and pathological conditions (tumour development. Vascular endothelial growth factor (VEGF, fibroblast growth factor-2 (FGF-2 and epidermal growth factor (EGF are the major angiogenic regulators. We have identified a natural product (cheiradone isolated from a Euphorbia species which inhibited in vivo and in vitro VEGF- stimulated angiogenesis but had no effect on FGF-2 or EGF activity. Two primary cultures, bovine aortic and human dermal endothelial cells were used in in vitro (proliferation, wound healing, invasion in Matrigel and tube formation and in vivo (the chick chorioallantoic membrane models of angiogenesis in the presence of growth factors and cheiradone. In all cases, the concentration of cheiradone which caused 50% inhibition (IC50 was determined. The effect of cheiradone on the binding of growth factors to their receptors was also investigated. Results Cheiradone inhibited all stages of VEGF-induced angiogenesis with IC50 values in the range 5.20–7.50 μM but did not inhibit FGF-2 or EGF-induced angiogenesis. It also inhibited VEGF binding to VEGF receptor-1 and 2 with IC50 values of 2.9 and 0.61 μM respectively. Conclusion Cheiradone inhibited VEGF-induced angiogenesis by binding to VEGF receptors -1 and -2 and may be a useful investigative tool to study the specific contribution of VEGF to angiogenesis and may have therapeutic potential.

  1. Anchored and soluble gangliosides contribute to myelosupportivity of stromal cells

    Energy Technology Data Exchange (ETDEWEB)

    Ziulkoski, Ana L. [Programa de Pos-Graduacao em Ciencias Biologicas: Bioquimica, ICBS, Universidade Federal do Rio Grande do Sul, Porto Alegre, RS (Brazil); Departamento de Bioquimica, ICBS, Universidade Federal do Rio Grande do Sul, Porto Alegre, RS (Brazil); Instituto de Ciencias da Saude, Centro Universitario Feevale, Novo Hamburgo, RS (Brazil); Santos, Aline X.S. dos; Andrade, Claudia M.B.; Trindade, Vera M.T. [Programa de Pos-Graduacao em Ciencias Biologicas: Bioquimica, ICBS, Universidade Federal do Rio Grande do Sul, Porto Alegre, RS (Brazil); Departamento de Bioquimica, ICBS, Universidade Federal do Rio Grande do Sul, Porto Alegre, RS (Brazil); Daniotti, Jose Luis [Departamento de Quimica Biologica, Faculdad de Ciencias Quimicas, Universidad Nacional de Cordoba, Cordoba (Argentina); Borojevic, Radovan [Departamento de Histologia e Embriologia, Universidade Federal do Rio de Janeiro, Rio de Janeiro (Brazil); Guma, Fatima C.R., E-mail: fatima.guma@ufrgs.br [Programa de Pos-Graduacao em Ciencias Biologicas: Bioquimica, ICBS, Universidade Federal do Rio Grande do Sul, Porto Alegre, RS (Brazil); Departamento de Bioquimica, ICBS, Universidade Federal do Rio Grande do Sul, Porto Alegre, RS (Brazil)

    2009-10-09

    Stroma-mediated myelopoiesis depends upon growth factors and an appropriate intercellular microenvironment. Previous studies have demonstrated that gangliosides, produced by hepatic stromal cell types, are required for optimal myelosupportive function. Here, we compared the mielossuportive functions of a bone marrow stroma (S17) and skin fibroblasts (SF) regarding their ganglioside pattern of synthesis and shedding. The survival and proliferation of a myeloid precursor cell (FDC-P1) were used as reporter. Although the ganglioside synthesis of the two stromal cells was similar, their relative content and shedding were distinct. The ganglioside requirement for mielossuportive function was confirmed by the decreased proliferation of FDC-P1 cells in ganglioside synthesis-inhibited cultures and in presence of an antibody to GM3 ganglioside. The distinct mielossuportive activities of the S17 and SF stromata may be related to differences on plasma membrane ganglioside concentrations or to differences on the gangliosides shed and their subsequent uptake by myeloid cells, specially, GM3 ganglioside.

  2. Redox-sensitive transcription factor Nrf2 regulates vascular smooth muscle cell migration and neointimal hyperplasia.

    Science.gov (United States)

    Ashino, Takashi; Yamamoto, Masayuki; Yoshida, Takemi; Numazawa, Satoshi

    2013-04-01

    Reactive oxygen species are important mediators for platelet-derived growth factor (PDGF) signaling in vascular smooth muscle cells, whereas excess reactive oxygen species-induced oxidative stress contributes to the development and progression of vascular diseases, such as atherosclerosis. Activation of the redox-sensitive transcription factor, nuclear factor erythroid 2-related factor 2 (Nrf2), is pivotal in cellular defense against oxidative stress by transcriptional upregulation of antioxidant proteins. This study aimed to elucidate the role of Nrf2 in PDGF-mediated vascular smooth muscle cell migration and neointimal hyperplasia. PDGF promoted nuclear translocation of Nrf2, followed by the induction of target genes, including NAD(P)H:quinone oxidoreductase-1, heme oxygenase-1, and thioredoxin-1. Nrf2 depletion by small interfering RNA enhanced PDGF-promoted Rac1 activation and reactive oxygen species production and persistently phosphorylated downstream extracellular signal-regulated kinase-1/2. Nrf2 depletion enhanced vascular smooth muscle cell migration in response to PDGF and wound scratch. In vivo, Nrf2-deficient mice showed enhanced neointimal hyperplasia in a wire injury model. These findings suggest that the Nrf2 system is important for PDGF-stimulated vascular smooth muscle cell migration by regulating reactive oxygen species elimination, which may contribute to neointimal hyperplasia after vascular injury. Our findings provide insight into the Nrf2 system as a novel therapeutic target for vascular remodeling and atherosclerosis.

  3. Analysis of the soluble cell wall proteome of gymnosperms.

    Science.gov (United States)

    Uzal, Esther Novo; Gómez-Ros, Laura V; Hernández, Jose A; Pedreño, María A; Cuello, Juan; Ros Barceló, Alfonso

    2009-05-15

    We analyzed the cell wall proteome of lignifying suspension cell cultures (SCCs) from four gymnosperms that differ in evolution degree. This analysis showed the presence of "peptide sequence tags" (PSTs) corresponding to glucan endo-1,3-beta-D-glucosidase, xyloglucan-endotrans-glucosylase/hydrolase, chitinases, thaumatin-like proteins and proteins involved in lignin/lignan biosynthesis, such as dirigent-like proteins and peroxidases. Surprisingly, and given the abundance of peroxidases in the cell wall proteome of these gymnosperms, PSTs corresponding to peroxidases were only detected in tryptic fragments of the cell wall proteome of Cycas revoluta. The current lack of knowledge regarding C. revoluta peroxidases led us to purify, characterize and partially sequence the peroxidases responsible for lignin biosynthesis in this species. This yielded three peroxidase-enriched fractions: CrPrx 1, CrPrx 2 and CrPrx 3. Analyses of tryptic peptides of CrPrx 2 (32kDa) and CrPrx 3 (26kDa) suggest that CrPrx 3 arises from CrPrx 2 by protein truncation, and that CrPrx 3 apparently constitutes a post-translational modification of CrPrx 2. That CrPrx 2 and CrPrx 3 are apparently the same enzyme was also deduced from the similarity between the k(cat) shown by both peroxidases for the three monolignols. These results emphasize the analogies between the cell wall proteome of gymnosperms and angiosperms, the complexity of the peroxidase proteome, and the difficulties involved in establishing fine structure-function relationships.

  4. Synergistic actions of hematopoietic and mesenchymal stem/progenitor cells in vascularizing bioengineered tissues.

    Directory of Open Access Journals (Sweden)

    Eduardo K Moioli

    Full Text Available Poor angiogenesis is a major road block for tissue repair. The regeneration of virtually all tissues is limited by angiogenesis, given the diffusion of nutrients, oxygen, and waste products is limited to a few hundred micrometers. We postulated that co-transplantation of hematopoietic and mesenchymal stem/progenitor cells improves angiogenesis of tissue repair and hence the outcome of regeneration. In this study, we tested this hypothesis by using bone as a model whose regeneration is impaired unless it is vascularized. Hematopoietic stem/progenitor cells (HSCs and mesenchymal stem/progenitor cells (MSCs were isolated from each of three healthy human bone marrow samples and reconstituted in a porous scaffold. MSCs were seeded in micropores of 3D calcium phosphate (CP scaffolds, followed by infusion of gel-suspended CD34(+ hematopoietic cells. Co-transplantation of CD34(+ HSCs and CD34(- MSCs in microporous CP scaffolds subcutaneously in the dorsum of immunocompromised mice yielded vascularized tissue. The average vascular number of co-transplanted CD34(+ and MSC scaffolds was substantially greater than MSC transplantation alone. Human osteocalcin was expressed in the micropores of CP scaffolds and was significantly increased upon co-transplantation of MSCs and CD34(+ cells. Human nuclear staining revealed the engraftment of transplanted human cells in vascular endothelium upon co-transplantation of MSCs and CD34(+ cells. Based on additional in vitro results of endothelial differentiation of CD34(+ cells by vascular endothelial growth factor (VEGF, we adsorbed VEGF with co-transplanted CD34(+ and MSCs in the microporous CP scaffolds in vivo, and discovered that vascular number and diameter further increased, likely owing to the promotion of endothelial differentiation of CD34(+ cells by VEGF. Together, co-transplantation of hematopoietic and mesenchymal stem/progenitor cells may improve the regeneration of vascular dependent tissues such as bone

  5. The role of vascular actors in two dimensional dialogue of human bone marrow stromal cell and endothelial cell for inducing self-assembled network.

    Directory of Open Access Journals (Sweden)

    Haiyan Li

    Full Text Available Angiogenesis is very important for vascularized tissue engineering. In this study, we found that a two-dimensional co-culture of human bone marrow stromal cell (HBMSC and human umbical vein endothelial cell (HUVEC is able to stimulate the migration of co-cultured HUVEC and induce self-assembled network formation. During this process, expression of vascular endothelial growth factor (VEGF₁₆₅ was upregulated in co-cultured HBMSC. Meanwhile, VEGF₁₆₅-receptor2 (KDR and urokinase-type plasminogen activator (uPA were upregulated in co-cultured HUVEC. Functional studies show that neutralization of VEGF₁₆₅ blocked the migration and the rearrangement of the cells and downregulated the expression of uPA and its receptor. Blocking of vascular endothelial-cadherin (VE-cad did not affect the migration of co-cultured HUVEC but suppressed the self-assembled network formation. In conclusion, co-cultures upregulated the expression of VEGF₁₆₅ in co-cultured HBMSC; VEGF₁₆₅ then activated uPA in co-cultured HUVEC, which might be responsible for initiating the migration and the self-assembled network formation with the participation of VE-cad. All of these results indicated that only the direct contact of HBMSC and HUVEC and their respective dialogue are sufficient to stimulate secretion of soluble factors and to activate molecules that are critical for self-assembled network formation which show a great application potential for vascularization in tissue engineering.

  6. Soluble triggering receptor expressed on myeloid cells 1: a biomarker for bacterial meningitis

    NARCIS (Netherlands)

    Determann, Rogier M.; Weisfelt, Martijn; de Gans, Jan; van der Ende, Arie; Schultz, Marcus J.; van de Beek, Diederik

    2006-01-01

    OBJECTIVE: To evaluate whether soluble triggering receptor expressed on myeloid cells 1 (sTREM-1) in CSF can serve as a biomarker for the presence of bacterial meningitis and outcome in patients with this disease. DESIGN: Retrospective study of diagnostic accuracy. SETTING AND PATIENTS: CSF was

  7. Role of the Vasa Vasorum and Vascular Resident Stem Cells in Atherosclerosis

    Directory of Open Access Journals (Sweden)

    Jun-ichi Kawabe

    2014-01-01

    Full Text Available Atherosclerosis is considered an “inside-out” response, that begins with the dysfunction of intimal endothelial cells and leads to neointimal plaque formation. The adventitia of large blood vessels has been recognized as an active part of the vessel wall that is involved in the process of atherosclerosis. There are characteristic changes in the adventitial vasa vasorum that are associated with the development of atheromatous plaques. However, whether vasa vasorum plays a causative or merely reactive role in the atherosclerotic process is not completely clear. Recent studies report that the vascular wall contains a number of stem/progenitor cells that may contribute to vascular remodeling. Microvessels serve as the vascular niche that maintains the resident stem/progenitor cells of the tissue. Therefore, the vasa vasorum may contribute to vascular remodeling through not only its conventional function as a blood conducting tube, but also its new conceptual function as a stem cell reservoir. This brief review highlights the recent advances contributing to our understanding of the role of the adventitial vasa vasorum in the atherosclerosis and discusses new concept that involves vascular-resident factors, the vasa vasorum and its associated vascular-resident stem cells, in the atherosclerotic process.

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

    Science.gov (United States)

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

    2016-06-01

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

  9. Tumor cells secrete an angiogenic factor that stimulates basic fibroblast growth factor and urokinase expression in vascular endothelial cells

    NARCIS (Netherlands)

    Peverali, F.A.; Mandriota, S.J.; Ciana, P.; Marelli, R.; Quax, P.; Rifkin, D.B.; Della Valle, G.; Mignatti, P.

    1994-01-01

    Culture medium conditioned by human SK-Hep1 hepatoma cells or mouse S180 sarcoma cells rapidly up-regulates endothelial cell expression of basic fibroblast growth factor (bFGF) and induces formation of capillary-like structures by vascular endothelial cells grown on three-dimensional fibrin gels (in

  10. Isolation and differentiation of stromal vascular cells to beige/brite cells

    DEFF Research Database (Denmark)

    Aune, Ulrike Liisberg; Ruiz, Lauren; Kajimura, Shingo

    2013-01-01

    Brown adipocytes have the ability to uncouple the respiratory chain in mitochondria and dissipate chemical energy as heat. Development of UCP1-positive brown adipocytes in white adipose tissues (so called beige or brite cells) is highly induced by a variety of environmental cues such as chronic...... cold exposure or by PPARγ agonists, therefore, this cell type has potential as a therapeutic target for obesity treatment. Although most immortalized adipocyte lines cannot recapitulate the process of "browning" of white fat in culture, primary adipocytes isolated from stromal vascular fraction...... in subcutaneous white adipose tissue (WAT) provide a reliable cellular system to study the molecular control of beige/brite cell development. Here we describe a protocol for effective isolation of primary preadipocytes and for inducing differentiation to beige/brite cells in culture. The browning effect can...

  11. Conversion of vascular endothelial cells into multipotent stem-like cells

    Science.gov (United States)

    Medici, Damian; Shore, Eileen M.; Lounev, Vitali Y.; Kaplan, Frederick S.; Kalluri, Raghu; Olsen, Bjorn R.

    2011-01-01

    Mesenchymal stem cells can give rise to several cell types, but variations depending on isolation method and tissue source have led to controversies about their usefulness in clinical medicine. Here we show that vascular endothelial cells can transform into multipotent stem-like cells by an ALK2 receptor-dependent mechanism. In lesions from patients with Fibrodysplasia Ossificans Progressiva, a disease where heterotopic ossification occurs as a result of activating ALK2 mutations, or from a mutant ALK2 transgenic mouse model, chondrocytes and osteoblasts express endothelial markers. Tie2-Cre lineage tracing also suggests an endothelial origin of these cells. Expressing mutant ALK2 in endothelial cells, or treatment with the ALK2 ligands TGF-β2 or BMP4, causes endothelial-mesenchymal transition and acquisition of a stem cell-like phenotype. In selective media, these cells differentiate into osteoblasts, chondrocytes, or adipocytes. The process is inhibited by ALK2-specific siRNA. Conversion of endothelial cells to stem-like cells may provide a novel approach to tissue engineering. PMID:21102460

  12. Vascularization mediated by mesenchymal stem cells from bone marrow and adipose tissue: a comparison

    Directory of Open Access Journals (Sweden)

    Karoline Pill

    2015-01-01

    Full Text Available Tissue-engineered constructs are promising to overcome shortage of organ donors and to reconstruct at least parts of injured or diseased tissues or organs. However, oxygen and nutrient supply are limiting factors in many tissues, especially after implantation into the host. Therefore, the development of a vascular system prior to implantation appears crucial. To develop a functional vascular system, different cell types that interact with each other need to be co-cultured to simulate a physiological environment in vitro. This review provides an overview and a comparison of the current knowledge of co-cultures of human endothelial cells (ECs with human adipose tissue-derived stem/stromal cells (ASCs or bone marrow-mesenchymal stem cells (BMSCs in three dimensional (3D hydrogel matrices. Mesenchymal stem cells (MSCs, BMSCs or ASCs, have been shown to enhance vascular tube formation of ECs and to provide a stabilizing function in addition to growth factor delivery and permeability control for ECs. Although phenotypically similar, MSCs from different tissues promote tubulogenesis through distinct mechanisms. In this report, we describe differences and similarities regarding molecular interactions in order to investigate which of these two cell types displays more favorable characteristics to be used in clinical applications. Our comparative study shows that ASCs as well as BMSCs are both promising cell types to induce vascularization with ECs in vitro and consequently are promising candidates to support in vivo vascularization.

  13. Effect of soluble factors derived from oral cancer cells on the production of interferon-γ from peripheral blood mononuclear cells following stimulation with OK-432.

    Science.gov (United States)

    Ohe, Go; Sasai, Akiko; Uchida, Daisuke; Tamatani, Tetsuya; Nagai, Hirokazu; Miyamoto, Youji

    2013-08-01

    The streptococcal antitumor agent OK-432 is commonly used as an immunopotentiator for immunotherapy in various types of malignant tumors including oral cancer. It has been demonstrated that OK-432 elicits an antitumor effect by stimulating immunocompetent cells, thereby inducing multiple cytokines including interferon (IFN)-γ, interleukin (IL)-2 and IL-12. Serum concentrations of IFN-γ in patients with oral cancer were examined 24 h after administration of OK-432. Serum concentrations of IFN-γ in patients with advanced cancer were significantly lower than those in patients with early cancer. These results suggested that some soluble factors produced by cancer cells may inhibit IFN-γ production with OK-432. Thus, in the present study, an in vitro simulation model was established for the immune status of patients with oral cancer by adding conditioned medium (CM) derived from oral cancer cell lines into a culture of peripheral blood mononuclear cells (PBMCs) derived from a healthy volunteer. We investigated whether soluble factors derived from oral cancer cells affected IFN-γ production from PBMCs following stimulation with OK-432. PBMCs stimulated with OK-432 produced a large amount of IFN-γ; however, both IFN-γ production and cytotoxic activity from PBMCs induced by OK-432 were inhibited by the addition of CM in a dose-dependent manner. In order to examine these inhibitory effects against IFN-γ production, the contribution of inhibitory cytokines such as IL-4, IL-6, IL-10, transforming growth factor-β and vascular endothelial growth factor was investigated. However, neutralization of these inhibitory cytokines did not recover IFN-γ production inhibited by CM. These results indicated that unknown molecules may inhibit IFN-γ production from PBMCs following stimulation with OK-432.

  14. Anatomical region-dependent enhancement of 3-dimensional chondrogenic differentiation of human mesenchymal stem cells by soluble meniscus extracellular matrix.

    Science.gov (United States)

    Rothrauff, Benjamin B; Shimomura, Kazunori; Gottardi, Riccardo; Alexander, Peter G; Tuan, Rocky S

    2017-02-01

    Extracellular matrix (ECM) derived from decellularized tissues has been found to promote tissue neogenesis, most likely mediated by specific biochemical and physical signaling motifs that promote tissue-specific differentiation of progenitor cells. Decellularized ECM has been suggested to be efficacious for the repair of tissue injuries. However, decellularized meniscus contains a dense collagenous structure, which impedes cell seeding and infiltration and is not readily applicable for meniscus repair. In addition, the meniscus consists of two distinct anatomical regions that differ in vascularity and cellular phenotype. The purpose of this study was to explore the region-specific bioactivity of solubilized ECM derived from the inner and outer meniscal regions as determined in 2D and 3D cultures of adult mesenchymal stem cells (MSCs). When added as a medium supplement to 2D cultures of MSCs, urea-extracted fractions of the inner (imECM) and outer meniscal ECM (omECM) enhanced cell proliferation while imECM most strongly upregulated fibrochondrogenic differentiation on the basis of gene expression profiles. When added to 3D cultures of MSCs seeded in photocrosslinked methacrylated gelatin (GelMA) hydrogels, both ECM fractions upregulated chondrogenic differentiation as determined by gene expression and protein analyses, as well as elevated sulfated glycosaminoglycan sGAG content, compared to ECM-free controls. The chondrogenic effect at day 21 was most pronounced with imECM supplementation, but equivalent between ECM groups by day 42. Despite increased cartilage matrix, imECM and omECM constructs possessed compressive moduli similar to controls. In conclusion, soluble meniscal ECM may be considered for use as a tissue-specific reagent to enhance chondrogenesis for MSC-based 3D cartilage tissue engineering. The inner region of the knee meniscus is frequently injured and possesses a poor intrinsic healing capacity. Solubilized extracellular matrix (ECM) derived from

  15. Ionizing radiation activates vascular endothelial growth factor-A transcription in human umbilical vein endothelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Hyounji; Kim, Kwang Seok; Jeong, Jae Hoon; Lim, Young Bin [Radiation Cancer Biology Team, Korea Institute of Radiological and Medical Sciences, Seoul (Korea, Republic of)

    2016-12-15

    Vascular endothelial growth factor (VEGF) is an essential paracrine factor for developmental and pathological angiogenesis. VEGF also exerts its effects in an autocrine manner in VEGF-producing cells. For instance, autocrine VEGF signaling occurs in tumor cells and contributes to key aspects of tumorigenesis, such as in the function of cancer stem cells and tumor initiation, which are independent of angiogenesis. In addition to tumors cells, non-transformed cells also express VEGF. For example, a VEGF dependent intracellular autocrine mechanism is crucial for the survival of hematopoietic stem cells and hematopoiesis. Stereotactic body radiation therapy (SBRT) is a novel treatment modality for early primary cancer and oligometastatic disease. SBRT delivers high-dose hypofractionated radiation, such as 20-60 Gy, to tumors in a single fraction or 2-5 fractions. As VEGF is a critical regulator of functional integrity and viability of vascular endothelial cells, we examined whether high-dose irradiation alters VEGF signaling by measuring the expression levels of VEGFA transcript. It is generally believed that endothelial cells do not produce VEGF in response to radiation. In present study, however, we provide the first demonstration of transcriptional regulation of VEGFA in human vascular endothelial cells by IR treatment. Irradiation with doses higher than 10 Gy in a single exposure triggers up-regulation of VEGFA transcription within 2 hours in HUVECs, whereas irradiation with 10 Gy does not alter VEGFA levels. Our data have shown that high-dose irradiation triggers immediate transactivation of VEGFA in human vascular endothelial cells.

  16. Taurine inhibits osteoblastic differentiation of vascular smooth muscle cells via the ERK pathway.

    Science.gov (United States)

    Liao, Xiao-bo; Zhou, Xin-min; Li, Jian-ming; Yang, Jin-fu; Tan, Zhi-ping; Hu, Zhuo-wei; Liu, Wei; Lu, Ying; Yuan, Ling-qing

    2008-05-01

    Vascular calcification develops within atherosclerotic lesions and results from a process similar to osteogenesis. Taurine is a free beta-amino acid and plays an important physiological role in mammals. We have recently demonstrated that vascular smooth muscle cells (VSMCs) express a functional taurine transporter. To evaluate the possible role of taurine in vascular calcification, we assessed its effects on osteoblastic differentiation of VSMCs in vitro. The results showed that taurine inhibited the beta-glycerophosphate-induced osteoblastic differentiation of VSMCs as evidenced by both the decreasing alkaline phosphate (ALP) activity and expression of the core binding factor alpha1 (Cbfalpha1). Taurine also activated the extracellular signal-regulated protein kinase (ERK) pathway. Inhibition of ERK pathway reversed the effect of taurine on ALP activity and Cbfalpha1 expression. These results suggested that taurine inhibited osteoblastic differentiation of vascular cells via the ERK pathway.

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

    Directory of Open Access Journals (Sweden)

    Chang Hyun Byon

    2016-10-01

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

  18. Incorporation of bone marrow cells in pancreatic pseudoislets improves posttransplant vascularization and endocrine function.

    Directory of Open Access Journals (Sweden)

    Christine Wittig

    Full Text Available Failure of revascularization is known to be the major reason for the poor outcome of pancreatic islet transplantation. In this study, we analyzed whether pseudoislets composed of islet cells and bone marrow cells can improve vascularization and function of islet transplants. Pancreatic islets isolated from Syrian golden hamsters were dispersed into single cells for the generation of pseudoislets containing 4×10(3 cells. To create bone marrow cell-enriched pseudoislets 2×10(3 islet cells were co-cultured with 2×10(3 bone marrow cells. Pseudoislets and bone marrow cell-enriched pseudoislets were transplanted syngeneically into skinfold chambers to study graft vascularization by intravital fluorescence microscopy. Native islet transplants served as controls. Bone marrow cell-enriched pseudoislets showed a significantly improved vascularization compared to native islets and pseudoislets. Moreover, bone marrow cell-enriched pseudoislets but not pseudoislets normalized blood glucose levels after transplantation of 1000 islet equivalents under the kidney capsule of streptozotocin-induced diabetic animals, although the bone marrow cell-enriched pseudoislets contained only 50% of islet cells compared to pseudoislets and native islets. Fluorescence microscopy of bone marrow cell-enriched pseudoislets composed of bone marrow cells from GFP-expressing mice showed a distinct fraction of cells expressing both GFP and insulin, indicating a differentiation of bone marrow-derived cells to an insulin-producing cell-type. Thus, enrichment of pseudoislets by bone marrow cells enhances vascularization after transplantation and increases the amount of insulin-producing tissue. Accordingly, bone marrow cell-enriched pseudoislets may represent a novel approach to increase the success rate of islet transplantation.

  19. Podocalyxin regulates murine lung vascular permeability by altering endothelial cell adhesion.

    Directory of Open Access Journals (Sweden)

    Erin J Debruin

    Full Text Available Despite the widespread use of CD34-family sialomucins (CD34, podocalyxin and endoglycan as vascular endothelial cell markers, there is remarkably little known of their vascular function. Podocalyxin (gene name Podxl, in particular, has been difficult to study in adult vasculature as germ-line deletion of podocalyxin in mice leads to kidney malformations and perinatal death. We generated mice that conditionally delete podocalyxin in vascular endothelial cells (Podxl(ΔEC mice to study the homeostatic role of podocalyxin in adult mouse vessels. Although Podxl(ΔEC adult mice are viable, their lungs display increased lung volume and changes to the matrix composition. Intriguingly, this was associated with increased basal and inflammation-induced pulmonary vascular permeability. To further investigate the etiology of these defects, we isolated mouse pulmonary endothelial cells. Podxl(ΔEC endothelial cells display mildly enhanced static adhesion to fibronectin but spread normally when plated on fibronectin-coated transwells. In contrast, Podxl(ΔEC endothelial cells exhibit a severely impaired ability to spread on laminin and, to a lesser extent, collagen I coated transwells. The data suggest that, in endothelial cells, podocalyxin plays a previously unrecognized role in maintaining vascular integrity, likely through orchestrating interactions with extracellular matrix components and basement membranes, and that this influences downstream epithelial architecture.

  20. Cadherins in vascular smooth muscle cell (patho)biology: Quid nos scimus?

    Science.gov (United States)

    Frismantiene, Agne; Philippova, Maria; Erne, Paul; Resink, Therese J

    2018-05-01

    Vascular smooth muscle cells (SMCs) phenotypes span a reversible continuum from quiescent/contractile (differentiated) to proliferative/synthetic (dedifferentiated) enabling them to perform a diversity of functions that are context-dependent and important for vascular tone-diameter homeostasis, vasculogenesis, angiogenesis or vessel reparation after injury. Dysregulated phenotype modulation and failure to maintain/regain the mature differentiated and contractile phenotypic state is pivotal in the development of vascular diseases such as atherosclerosis and restenosis after angioplasty and coronary bypass grafting. Many functions of SMCs such as adhesion, migration, proliferation, contraction, differentiation and apoptosis are regulated by a broad spectrum of cell-cell and cell-matrix adhesion molecules. Cadherins represent a superfamily of cell surface homophilic adhesion molecules with fundamental roles in morphogenetic and differentiation processes during development and in the maintenance of tissue integrity and homeostasis in adults. The cadherins have major inputs on signalling pathways and cytoskeletal assemblies that participate in regulating processes such as cell polarity, migration, proliferation, survival, phenotype and differentiation. Abnormalities in these processes have long been recognized to underlie pathological SMC-driven reparation, but knowledge on the involvement of cadherins is remarkably limited. This article presents a comprehensive review of cadherin family members currently identified on vascular SMCs in relation to their functions, molecular mechanisms of action and relevance for vascular pathology. Copyright © 2018 Elsevier Inc. All rights reserved.

  1. Interaction of L-lysine and soluble elastin with the semicarbazide-sensitive amine oxidase in the context of its vascular-adhesion and tissue maturation functions.

    LENUS (Irish Health Repository)

    Olivieri, Aldo

    2010-04-01

    The copper-containing quinoenzyme semicarbazide-sensitive amine oxidase (EC 1.4.3.21; SSAO) is a multifunctional protein. In some tissues, such as the endothelium, it also acts as vascular-adhesion protein 1 (VAP-1), which is involved in inflammatory responses and in the chemotaxis of leukocytes. Earlier work had suggested that lysine might function as a recognition molecule for SSAO\\/VAP-1. The present work reports the kinetics of the interaction of L-lysine and some of its derivatives with SSAO. Binding was shown to be saturable, time-dependent but reversible and to cause uncompetitive inhibition with respect to the amine substrate. It was also specific, since D-lysine, L-lysine ethyl ester and epsilon-acetyl-L-lysine, for example, did not bind to the enzyme. The lysine-rich protein soluble elastin bound to the enzyme relatively tightly, which may have relevance to the reported roles of SSAO in maintaining the extracellular matrix (ECM) and in the maturation of elastin. Our data show that lysyl residues are not oxidized by SSAO, but they bind tightly to the enzyme in the presence of hydrogen peroxide. This suggests that binding in vivo of SSAO to lysyl residues in physiological targets might be regulated in the presence of H(2)O(2), formed during the oxidation of a physiological SSAO substrate, yet to be identified.

  2. Mechanism of the toxicity induced by natural humic acid on human vascular endothelial cells

    OpenAIRE

    Kihara, Yusuke; Yustiawati; Tanaka, Masato; Gumiri, Sulmin; Ardianor; Hosokawa, Toshiyuki; Tanaka, Shunitz; Saito, Takeshi; Kurasaki, Masaaki

    2012-01-01

    Humic acid (HA), a group of high-molecular weight organic compounds characterized by an ability to bind heavy metals, is normally found in natural water. Although the impairment of vascular endothelial cells in the presence of humic substances has been reported to be involved in some diseases, the mechanisms responsible for this involvement remain unclear. In this study, we examined the cytotoxicity of HA obtained from peatland in Central Kalimantan, Indonesia, to human vascular endothelial c...

  3. 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.

  4. Carbon black nanoparticles and vascular dysfunction in cultured endothelial cells and artery segments

    DEFF Research Database (Denmark)

    Vesterdal, Lise K; Mikkelsen, Lone; Folkmann, Janne K

    2012-01-01

    surface expression of intercellular cell adhesion molecule 1 (ICAM-1) and vascular adhesion molecule 1 (VCAM-1) in HUVECs at 100µg/ml. CB exposure was also associated with increased reactive oxygen species production and damage to the cell membranes in the form of increased lactate dehydrogenase leakage...

  5. 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

    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...

  6. 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.

    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,

  7. Chloroquine inhibits accessory cell presentation of soluble natural and synthetic protein antigens

    DEFF Research Database (Denmark)

    Buus, S; Werdelin, O

    1984-01-01

    We have studied the in vitro effect of the lysosomotrophic agent, chloroquine, on the presentation of soluble protein antigens by guinea pig accessory cells. Chloroquine inhibited the capacity of antigen-pulsed accessory cells to stimulate proliferation in appropriately primed T cells. The effect...... was time- and dose-dependent. A brief treatment solely of the accessory cells with the drug compromised their ability to stimulate primed T cells in a subsequent culture provided the accessory cells were treated with chloroquine before their exposure to the antigen. These results suggest that chloroquine...... acts on an early event in the antigen handling by accessory cells. Chloroquine is a well known inhibitor of lysosomal proteolysis, and it is likely that its effect on antigen presentation is caused by an inhibition of antigen degradation....

  8. Bone marrow derived mesenchymal stem cells inhibit inflammation and preserve vascular endothelial integrity in the lungs after hemorrhagic shock.

    Directory of Open Access Journals (Sweden)

    Shibani Pati

    Full Text Available Hemorrhagic shock (HS and trauma is currently the leading cause of death in young adults worldwide. Morbidity and mortality after HS and trauma is often the result of multi-organ failure such as acute lung injury (ALI and acute respiratory distress syndrome (ARDS, conditions with few therapeutic options. Bone marrow derived mesenchymal stem cells (MSCs are a multipotent stem cell population that has shown therapeutic promise in numerous pre-clinical and clinical models of disease. In this paper, in vitro studies with pulmonary endothelial cells (PECs reveal that conditioned media (CM from MSCs and MSC-PEC co-cultures inhibits PEC permeability by preserving adherens junctions (VE-cadherin and β-catenin. Leukocyte adhesion and adhesion molecule expression (VCAM-1 and ICAM-1 are inhibited in PECs treated with CM from MSC-PEC co-cultures. Further support for the modulatory effects of MSCs on pulmonary endothelial function and inflammation is demonstrated in our in vivo studies on HS in the rat. In a rat "fixed volume" model of mild HS, we show that MSCs administered IV potently inhibit systemic levels of inflammatory cytokines and chemokines in the serum of treated animals. In vivo MSCs also inhibit pulmonary endothelial permeability and lung edema with concurrent preservation of the vascular endothelial barrier proteins: VE-cadherin, Claudin-1, and Occludin-1. Leukocyte infiltrates (CD68 and MPO positive cells are also decreased in lungs with MSC treatment. Taken together, these data suggest that MSCs, acting directly and through soluble factors, are potent stabilizers of the vascular endothelium and inflammation. These data are the first to demonstrate the therapeutic potential of MSCs in HS and have implications for the potential use of MSCs as a cellular therapy in HS-induced lung injury.

  9. Perinatal testosterone exposure potentiates vascular dysfunction by ERβ suppression in endothelial progenitor cells.

    Science.gov (United States)

    Xie, Weiguo; Ren, Mingming; Li, Ling; Zhu, Yin; Chu, Zhigang; Zhu, Zhigang; Ruan, Qiongfang; Lou, Wenting; Zhang, Haimou; Han, Zhen; Huang, Xiaodong; Xiang, Wei; Wang, Tao; Yao, Paul

    2017-01-01

    Recent clinical cohort study shows that testosterone therapy increases cardiovascular diseases in men with low testosterone levels, excessive circulating androgen levels may play a detrimental role in the vascular system, while the potential mechanism and effect of testosterone exposure on the vascular function in offspring is still unknown. Our preliminary results showed that perinatal testosterone exposure in mice induces estrogen receptor β (ERβ) suppression in endothelial progenitor cells (EPCs) in offspring but not mothers, while estradiol (E2) had no effect. Further investigation showed that ERβ suppression is due to perinatal testosterone exposure-induced epigenetic changes with altered DNA methylation on the ERβ promoter. During aging, EPCs with ERβ suppression mobilize to the vascular wall, differentiate into ERβ-suppressed mouse endothelial cells (MECs) with downregulated expression of SOD2 (mitochondrial superoxide dismutase) and ERRα (estrogen-related receptor α). This results in reactive oxygen species (ROS) generation and DNA damage, and the dysfunction of mitochondria and fatty acid metabolism, subsequently potentiating vascular dysfunction. Bone marrow transplantation of EPCs that overexpressed with either ERβ or a SIRT1 single mutant SIRT1-C152(D) that could modulate SIRT1 phosphorylation significantly ameliorated vascular dysfunction, while ERβ knockdown worsened the problem. We conclude that perinatal testosterone exposure potentiates vascular dysfunction through ERβ suppression in EPCs.

  10. Construction of a fucoidan/laminin functional multilayer to direction vascular cell fate and promotion hemocompatibility

    Energy Technology Data Exchange (ETDEWEB)

    Ye, Changrong; Wang, Yan; Su, Hong; Yang, Ping; Huang, Nan [Key Laboratory of Advanced Materials Technology of Ministry of Education, Department of Materials Science and Engineering, Southwest Jiaotong University, Chengdu 610031 (China); Maitz, Manfred F. [Leibniz Institute of Polymer Research Dresden, Max Bergmann Center of Biomaterials Dresden, Dresden 01069 (Germany); Zhao, Anshan [Key Laboratory of Advanced Materials Technology of Ministry of Education, Department of Materials Science and Engineering, Southwest Jiaotong University, Chengdu 610031 (China)

    2016-07-01

    Surface biofunctional modification of cardiovascular stents is a versatile approach to reduce the adverse effects after implantation. In this work, a novel multifunctional coating was fabricated by coimmobilization of the sulfated polysaccharide of brown algae fucoidan and laminin to biomimic the vascular intimal conditions in order to support rapid endothelialization, prevent restenosis and improve hemocompatibility. The surface properties of the coating such as hydrophilicity, bonding density of biomolecules and stability were evaluated and optimized. According to the biocompatibility tests, the fucoidan/laminin multilayer coated surface displayed less platelet adhesion with favorable anticoagulant property. In addition, the fucoidan/laminin complex showed function to selectively regulate vascular cells growth behavior. The proliferation of endothelial cells (ECs) on the fucoidan/laminin biofunctional coating was significantly promoted. For the smooth muscle cells (SMCs), inhibitory effects on cell adhesion and proliferation were observed. In conclusion, the fucoidan/laminin biofunctional coating was successfully fabricated with desirable anticoagulant and endothelialization properties which show a promising application in the vascular devices such as vascular stents or grafts surface modification. - Highlights: • Construction of fucoidan/laminin functional multilayer to biomimic the basement membrane of vascular • The fucoidan/laminin complex demonstrates anti-coagulation property. • The fucoidan/laminin complex can selectively regulate EC and SMC growth behavior to prevent restenosis.

  11. Quantification of stromal vascular cell mechanics with a linear cell monolayer rheometer

    Energy Technology Data Exchange (ETDEWEB)

    Elkins, Claire M., E-mail: cma9@stanford.edu; Fuller, Gerald G. [Department of Chemical Engineering, Stanford University, Stanford, California 94305 (United States); Shen, Wen-Jun; Khor, Victor K.; Kraemer, Fredric B. [Division of Endocrinology, Gerontology and Metabolism, Stanford University, Stanford, California 94305 and Veterans Affairs Palo Alto Health Care System, Palo Alto, California 94304 (United States)

    2015-01-15

    Over the past few decades researchers have developed a variety of methods for measuring the mechanical properties of whole cells, including traction force microscopy, atomic force microscopy (AFM), and single-cell tensile testing. Though each of these techniques provides insight into cell mechanics, most also involve some nonideal conditions for acquiring live cell data, such as probing only one portion of a cell at a time, or placing the cell in a nonrepresentative geometry during testing. In the present work, we describe the development of a linear cell monolayer rheometer (LCMR) and its application to measure the mechanics of a live, confluent monolayer of stromal vascular cells. In the LCMR, a monolayer of cells is contacted on both top and bottom by two collagen-coated plates and allowed to adhere. The top plate then shears the monolayer by stepping forward to induce a predetermined step strain, while a force transducer attached to the top plate collects stress information. The stress and strain data are then used to determine the maximum relaxation modulus recorded after step-strain, G{sub r}{sup 0}, referred to as the zero-time relaxation modulus of the cell monolayer. The present study validates the ability of the LCMR to quantify cell mechanics by measuring the change in G{sub r}{sup 0} of a confluent cell monolayer upon the selective inhibition of three major cytoskeletal components (actin microfilaments, vimentin intermediate filaments, and microtubules). The LCMR results indicate that both actin- and vimentin-deficient cells had ∼50% lower G{sub r}{sup 0} values than wild-type, whereas tubulin deficiency resulted in ∼100% higher G{sub r}{sup 0} values. These findings constitute the first use of a cell monolayer rheometer to quantitatively distinguish the roles of different cytoskeletal elements in maintaining cell stiffness and structure. Significantly, they are consistent with results obtained using single-cell mechanical testing methods

  12. The effect of ionizing radiation on the filamentous actin of vascular endothelial cell

    International Nuclear Information System (INIS)

    Yao Xiaowu; Chen Shisheng; Yang Lihe; Lin Juelong; Yang Haiwei

    2006-01-01

    Objective: To observe the ionizing radiation effect on filamentous actin of vascular endothelial cell and explore its mechanism. Methods: The vascular endothelial cells were irradiated with 0, 2, 4, 6, 8, 10 and 12 Gy 60 Co γ-rays. The cytoskeleton was observed with CLSM at 6 hs after the irradiation and the cytoskeleton protein F-actin detected with flow cytometry after 12 and 24 hs. Results: The damage to cytoskeletons increased with the radiation dose. The cytoskeleton protein F-actin was significantly decreased at 12 hs after the irradiation, and then recovered after 24 hs. Conclusion: Ionizing radiation caused vascular endothelial cell injury by damaging the cytoskeleton and depolymerizating the F-actin. (authors)

  13. 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.

  14. The cytotoxicity and genotoxicity of particulate and soluble hexavalent chromium in leatherback sea turtle lung cells.

    Science.gov (United States)

    Speer, Rachel M; Wise, Catherine F; Young, Jamie L; Aboueissa, AbouEl-Makarim; Martin Bras, Mark; Barandiaran, Mike; Bermúdez, Erick; Márquez-D'Acunti, Lirio; Wise, John Pierce

    2018-05-01

    Hexavalent chromium [Cr(VI)] is a marine pollution of concern as recent studies show it has a global distribution, with some regions showing high Cr concentrations in marine animal tissue, and it is extensively used. Leatherback sea turtles (Dermochelys coriacea) are an endangered marine species that may experience prolonged exposures to environmental contaminants including Cr(VI). Human activities have led to global Cr(VI) contamination of the marine environment. While Cr(VI) has been identified as a known human carcinogen, the health effects in marine species are poorly understood. In this study, we assessed the cytotoxic and genotoxic effects of particulate and soluble Cr(VI) in leatherback sea turtle lung cells. Both particulate and soluble Cr(VI) induced a concentration-dependent increase in cytotoxicity. Next, using a chromosome aberration assay, we assessed the genotoxic effects of Cr(VI) in leatherback sea turtle lung cells. Particulate and soluble Cr(VI) induced a concentration-dependent increase in clastogenicity in leatherback sea turtle lung cells. These data indicate that Cr(VI) may be a health concern for leatherback sea turtles and other long-lived marine species. Additionally, these data provide foundational support to use leatherback sea turtles as a valuable model species for monitoring the health effects of Cr(VI) in the environment and possibly as an indicator species to assess environmental human exposures and effects. Copyright © 2018 Elsevier B.V. All rights reserved.

  15. An Important Method in the Investigation of Vascular Pathologies: Endothelial Cell Culture

    Directory of Open Access Journals (Sweden)

    Yusufhan Yazır

    2012-12-01

    Full Text Available Endothelial cells line the interior surface of blood vessels and form an interface between circulating blood in the lumen and the rest of the vessel wall. Endothelial cells are involved in many aspects of vascular biology, including barrier function, vasoconstriction, coagulation and inflamation. The endothelial cells in different organs have different functions and surface phenotype. These cells express prostoglandin-I2, platelet activating factor, collagen, endothelin-1, laminin, fibronectin and growth factors including platelet derived growth factor, fibroblast growth factor. İn the cell culture, cells can be isolated, maintened and proliferate in the laboratory conditions. The techniques of the cell culture have allowed scientists to use the cells in vitro for experimental studies, such as the production of vaccine, antibody and enzime, drug research, cell-cell interactions. Human umbilical vein endothelial cell is a good source for endothelial cell, because it is cheaper, easy to find and has the basic features of the normal endothelial cells.

  16. Composite vascular grafts with high cell infiltration by co-electrospinning

    Energy Technology Data Exchange (ETDEWEB)

    Tan, Zhikai, E-mail: tanzk@hnu.edu.cn; Wang, Hongjie; Gao, Xiangkai; Liu, Tong; Tan, Yongjun

    2016-10-01

    There is an increasing demand for functional small-diameter vascular grafts (diameter < 6 mm) to be used in clinical arterial replacement. An ideal vascular graft should have appropriate biomechanical properties and be biocompatible. Electrospinning has become a popular polymer processing technique for vascular tissue engineering, but the grafts fabricated by electrospinning often have relatively small pores and low porosity, which limit cell infiltration into scaffolds and hinder the regeneration and remodeling of grafts. In the present study, we aimed to develop an efficient method to prepare electrospun composite vascular grafts comprising natural and synthetic materials. We fabricated grafts made of polycaprolactone, gelatin, and polyvinyl alcohol (PVA) by co-electrospinning, and the scaffolds were further functionalized by immobilizing heparin on them. The PVA fibers degraded rapidly in vivo and generated electrospun scaffolds with high porosity, which significantly enhanced cell proliferation and infiltration. The mechanical properties of the grafts are suitable for use in artery replacement. Heparin functionalization of the grafts yielded a good antithrombogenic effect, which was demonstrated in platelet adhesion tests. Moreover, in vitro and in vivo results demonstrated that the heparin release from the grafts enhanced the growth of endothelial cells, which is important for the endothelium of implanted grafts. The results of this study indicate that our method is effective and controllable for the fabrication of vascular grafts that meet the clinical requirements for blood vessel transplantation. - Highlights: • This study indicate an effective method for the fabrication of vascular grafts that meet the clinical requirements. • Co-electrospinning were used to fabricate grafts made of polycaprolactone (PCL), gelatin (GT), and polyvinyl alcohol (PVA). • PVA was used to create large pores within the hybrid scaffolds, thereby enhancing cell infiltration

  17. Composite vascular grafts with high cell infiltration by co-electrospinning

    International Nuclear Information System (INIS)

    Tan, Zhikai; Wang, Hongjie; Gao, Xiangkai; Liu, Tong; Tan, Yongjun

    2016-01-01

    There is an increasing demand for functional small-diameter vascular grafts (diameter < 6 mm) to be used in clinical arterial replacement. An ideal vascular graft should have appropriate biomechanical properties and be biocompatible. Electrospinning has become a popular polymer processing technique for vascular tissue engineering, but the grafts fabricated by electrospinning often have relatively small pores and low porosity, which limit cell infiltration into scaffolds and hinder the regeneration and remodeling of grafts. In the present study, we aimed to develop an efficient method to prepare electrospun composite vascular grafts comprising natural and synthetic materials. We fabricated grafts made of polycaprolactone, gelatin, and polyvinyl alcohol (PVA) by co-electrospinning, and the scaffolds were further functionalized by immobilizing heparin on them. The PVA fibers degraded rapidly in vivo and generated electrospun scaffolds with high porosity, which significantly enhanced cell proliferation and infiltration. The mechanical properties of the grafts are suitable for use in artery replacement. Heparin functionalization of the grafts yielded a good antithrombogenic effect, which was demonstrated in platelet adhesion tests. Moreover, in vitro and in vivo results demonstrated that the heparin release from the grafts enhanced the growth of endothelial cells, which is important for the endothelium of implanted grafts. The results of this study indicate that our method is effective and controllable for the fabrication of vascular grafts that meet the clinical requirements for blood vessel transplantation. - Highlights: • This study indicate an effective method for the fabrication of vascular grafts that meet the clinical requirements. • Co-electrospinning were used to fabricate grafts made of polycaprolactone (PCL), gelatin (GT), and polyvinyl alcohol (PVA). • PVA was used to create large pores within the hybrid scaffolds, thereby enhancing cell infiltration

  18. Hemorrhagic epithelioid and spindle cell hemangioma: a newly recognized, unique vascular tumor of bone.

    Science.gov (United States)

    Keel, S B; Rosenberg, A E

    1999-05-01

    Epithelioid vascular tumors of bone are uncommon and include epithelioid hemangioma, epithelioid hemangioendothelioma, and epithelioid angiosarcoma. It is important to distinguish among them because they have significantly different biologic potential and require different forms of therapy. In the current study the authors describe six cases of a distinct benign epithelioid and spindle cell vascular tumor of bone that, because of their unusual morphology, were confused with aggressive vascular neoplasms. Cases were retrieved from the surgical pathology files of the Department of Pathology or from the consultation files of one of the authors. Hematoxylin and eosin stained slides were examined. Immunohistochemistry was performed on two cases and electron microscopy was performed on one case. The tumors arose in the small bones of the hands and feet and the tibia. Three patients had multifocal bone disease at the time of presentation. Histologically, all lesions were comprised of lobules of spindle cells that grew focally in a fascicular pattern and were associated with abundant hemorrhage. Plump epithelioid cells were intermixed and were present focally in the interlobular areas as well, in which they lined larger, more well developed vascular spaces, often protruding into the vascular lumen in a "tombstone" fashion. Immunohistochemically and ultrastructurally the neoplastic cells had features of endothelium. One case was treated by amputation, one by resection, three by curettage, and one by curettage plus radiation therapy. None of the lesions was locally aggressive nor did any metastasize. The authors believe that hemorrhagic epithelioid and spindle cell hemangioma of bone is a histologically benign bone tumor. It should be distinguished from malignant epithelioid vascular tumors of bone, which have metastatic potential and need to be treated more aggressively.

  19. Endothelial vascular cell adhesion molecule 1 expression is suppressed by melanoma and carcinoma

    OpenAIRE

    1995-01-01

    Vascular cell adhesion molecule 1 (VCAM-1) mediates extravasation of circulating leukocytes into inflamed tissues, and presumably, plays a role in the immigration of cytotoxic effector lymphocytes into tumor metastases. Since metastases are rarely cleared by blood-borne cells from the immune system, we asked whether the tumor may escape host defense by interfering with the mechanism of effector cell extravasation. Here we show that in mice and humans, VCAM-1 expression is repressed on tumor-i...

  20. Vascular endothelium as a novel source of stem cells for bioengineering

    Science.gov (United States)

    Susienka, Michael J.; Medici, Damian

    2013-01-01

    Endothelial plasticity, the ability of endothelial cells to alter their lineage commitment to generate other cell types, is involved in many developmental and pathological processes. It was recently shown that vascular endothelial cells are converted to a mesenchymal stem cell phenotype through a process known as endothelial-mesenchymal transition (EndMT). EndMT is characterized as a morphological and phenotypical transformation of endothelial cells that has been implicated in cardiac development, cancer, fibrosis and heterotopic ossification. Here we describe the molecular and cellular basis for EndMT-dependent generation of endothelial-derived stem cells and their potential for tissue engineering and regenerative medicine. PMID:23603799

  1. Effect of melatonin or maternal nutrient restriction on vascularity and cell proliferation in the ovine placenta.

    Science.gov (United States)

    Eifert, Adam W; Wilson, Matthew E; Vonnahme, Kimberly A; Camacho, Leticia E; Borowicz, Pawel P; Redmer, Dale A; Romero, Sinibaldo; Dorsam, Sheri; Haring, Jodie; Lemley, Caleb O

    2015-02-01

    Previously we reported increased umbilical artery blood flow in ewes supplemented with melatonin from mid- to late-pregnancy, while maternal nutrient restriction decreased uterine artery blood flow. To further unravel these responses, this study was designed to assess placental cell proliferation and vascularity following supplementation with melatonin or maternal nutrient restriction. For the first experiment, 31 primiparous ewes were supplemented with 5mg of melatonin per day (MEL) or no melatonin (CON) and allocated to receive 100% (adequate fed; ADQ) or 60% (restricted; RES) of their nutrient requirements from day 50 to 130 of gestation. To examine melatonin receptor dependent effects, a second experiment was designed utilizing 14 primiparous ewes infused with vehicle, melatonin, or luzindole (melatonin receptor 1 and 2 antagonist) from day 62 to 90 of gestation. For experiment 1, caruncle concentrations of RNA were increased in MEL-RES compared to CON-RES. Caruncle capillary area density and average capillary cross-sectional area were decreased in MEL-RES compared to CON-RES. Cotyledon vascularity was not different across dietary treatments. For experiment 2, placental cellular proliferation and vascularity were not affected by infusion treatment. In summary, melatonin interacted with nutrient restriction to alter caruncle vascularity and RNA concentrations during late pregnancy. Although melatonin receptor antagonism alters feto-placental blood flow, these receptor dependent responses were not observed in placental vascularity. Moreover, placental vascularity measures do not fully explain the alterations in uteroplacental blood flow. Copyright © 2014 Elsevier B.V. All rights reserved.

  2. Vascular Graft Impregnation with Antibiotics: The Influence of High Concentrations of Rifampin, Vancomycin, Daptomycin, and Bacteriophage Endolysin HY-133 on Viability of Vascular Cells.

    Science.gov (United States)

    Herten, Monika; Idelevich, Evgeny A; Sielker, Sonja; Becker, Karsten; Scherzinger, Anna S; Osada, Nani; Torsello, Giovanni B; Bisdas, Theodosios

    2017-06-27

    BACKGROUND Rifampin-soaked synthetic prosthetic grafts have been widely used for prevention or treatment of vascular graft infections (VGIs). This in vitro study investigated the effect of the antibiotics daptomycin and vancomycin and the new recombinant bacteriophage endolysin HY-133 on vascular cells, as potential alternatives compared to rifampin. MATERIAL AND METHODS Primary human ECs, vascular smooth muscle cells (vSMC), and fibroblasts were cultivated in 96-well plates and incubated with rifampin, daptomycin, vancomycin, and endolysin HY-133 for 24 h. Subsequently, after washing, cell viability was determined by measuring mitochondrial ATP concentration. Antibiotics were used in their corresponding minimum and maximum serum concentrations, in decimal multiples and in maximum soaking concentration. The experiments were performed in triplicate. RESULTS The 10-fold max serum concentrations of rifampin, daptomycin, and vancomycin did not influence viability of EC and vSMC (100 µg/ml, p>0.170). Higher concentrations of rifampin (>1 mg/ml) significantly (pEndolysin did not display any cytotoxicity towards vascular cells. CONCLUSIONS Results of this in vitro study show the high cytotoxicity of rifampin against vascular cells, and may re-initiate the discussion about the benefit of prophylactic pre-soaking in high concentrations of rifampin. Further studies are necessary to determine the influence of rifampin on the restoration of vessel functionality versus its prophylactic effect against VGIs. Future use of recombinant phage endolysins for alternative prophylactic strategies needs further investigations.

  3. Vascular Cell Adhesion Molecule 1, Intercellular Adhesion Molecule 1, and Cluster of Differentiation 146 Levels in Patients with Type 2 Diabetes with Complications.

    Science.gov (United States)

    Hocaoglu-Emre, F Sinem; Saribal, Devrim; Yenmis, Guven; Guvenen, Guvenc

    2017-03-01

    Type 2 diabetes mellitus (T2DM) is a multisystemic, chronic disease accompanied by microvascular complications involving various complicated mechanisms. Intercellular adhesion molecule 1 (ICAM-1), vascular cell adhesion molecule 1 (VCAM-1), and cluster of differentiation-146 (CD146) are mainly expressed by endothelial cells, and facilitate the adhesion and transmigration of immune cells, leading to inflammation. In the present study, we evaluated the levels of soluble adhesion molecules in patients with microvascular complications of T2DM. Serum and whole blood samples were collected from 58 T2DM patients with microvascular complications and 20 age-matched healthy subjects. Levels of soluble ICAM-1 (sICAM-1) and soluble VCAM-1 (sVCAM-1) were assessed using enzyme-linked immunosorbent assay, while flow cytometry was used to determine CD146 levels. Serum sICAM-1 levels were lower in T2DM patients with microvascular complications than in healthy controls (Pmolecule levels were not correlated with the complication type. In the study group, most of the patients were on insulin therapy (76%), and 95% of them were receiving angiotensin-converting enzyme (ACE)-inhibitor agents. Insulin and ACE-inhibitors have been shown to decrease soluble adhesion molecule levels via various mechanisms, so we suggest that the decreased or unchanged levels of soluble forms of cellular adhesion molecules in our study group may have resulted from insulin and ACE-inhibitor therapy, as well as tissue-localized inflammation in patients with T2DM. Copyright © 2017 Korean Endocrine Society

  4. The healthy donor profile of immunoregulatory soluble mediators is altered by stem cell mobilization and apheresis.

    Science.gov (United States)

    Melve, Guro Kristin; Ersvaer, Elisabeth; Paulsen Rye, Kristin; Bushra Ahmed, Aymen; Kristoffersen, Einar K; Hervig, Tor; Reikvam, Håkon; Hatfield, Kimberley Joanne; Bruserud, Øystein

    2018-03-22

    Peripheral blood stem cells from healthy donors mobilized by granulocyte colony-stimulating factor (G-CSF) and thereafter harvested by leukapheresis are commonly used for allogeneic stem cell transplantation. Plasma levels of 38 soluble mediators (cytokines, soluble adhesion molecules, proteases, protease inhibitors) were analyzed in samples derived from healthy stem cell donors before G-CSF treatment and after 4 days, both immediately before and after leukapheresis. Donors could be classified into two main subsets based on their plasma mediator profile before G-CSF treatment. Seventeen of 36 detectable mediators were significantly altered by G-CSF; generally an increase in mediator levels was seen, including pro-inflammatory cytokines, soluble adhesion molecules and proteases. Several leukocyte- and platelet-released mediators were increased during apheresis. Both plasma and graft mediator profiles were thus altered and showed correlations to graft concentrations of leukocytes and platelets; these concentrations were influenced by the apheresis device used. Finally, the mediator profile of the allotransplant recipients was altered by graft infusion, and based on their day +1 post-transplantation plasma profile our recipients could be divided into two major subsets that differed in overall survival. G-CSF alters the short-term plasma mediator profile of healthy stem cell donors. These effects together with the leukocyte and platelet levels in the graft determine the mediator profile of the stem cell grafts. Graft infusion also alters the systemic mediator profile of the recipients, but further studies are required to clarify whether such graft-induced alterations have a prognostic impact. Copyright © 2018. Published by Elsevier Inc.

  5. Influence of Androgen Receptor in Vascular Cells on Reperfusion following Hindlimb Ischaemia.

    Directory of Open Access Journals (Sweden)

    Junxi Wu

    Full Text Available Studies in global androgen receptor knockout (G-ARKO and orchidectomised mice suggest that androgen accelerates reperfusion of the ischaemic hindlimb by stimulating angiogenesis. This investigation used novel, vascular cell-specific ARKO mice to address the hypothesis that the impaired hindlimb reperfusion in G-ARKO mice was due to loss of AR from cells in the vascular wall.Mice with selective deletion of AR (ARKO from vascular smooth muscle cells (SM-ARKO, endothelial cells (VE-ARKO, or both (SM/VE-ARKO were compared with wild type (WT controls. Hindlimb ischaemia was induced in these mice by ligation and removal of the femoral artery. Post-operative reperfusion was reduced in SM-ARKO and SM/VE-ARKO mice. Immunohistochemistry indicated that this was accompanied by a reduced density of smooth muscle actin-positive vessels but no change in the density of isolectin B4-positive vessels in the gastrocnemius muscle. Deletion of AR from the endothelium (VE-ARKO did not alter post-operative reperfusion or vessel density. In an ex vivo (aortic ring culture model of angiogenesis, AR was not detected in vascular outgrowths and angiogenesis was not altered by vascular ARKO or by exposure to dihydrotestosterone (DHT 10-10-10-7M; 6 days.These results suggest that loss of AR from vascular smooth muscle, but not from the endothelium, contributes to impaired reperfusion in the ischaemic hindlimb of G-ARKO. Impaired reperfusion was associated with reduced collateral formation rather than reduced angiogenesis.

  6. A role of TDIF peptide signaling in vascular cell differentiation is conserved among euphyllophytes

    Directory of Open Access Journals (Sweden)

    Yuki eHirakawa

    2015-11-01

    Full Text Available Peptide signals mediate a variety of cell-to-cell communication crucial for plant growth and development. During Arabidopsis thaliana vascular development, a CLE (CLAVATA3/EMBRYO SURROUNDING REGION-related family peptide hormone, TDIF (tracheary element differentiation inhibitory factor, regulates procambial cell fate by its inhibitory activity on xylem differentiation. To address if this activity is conserved among vascular plants, we performed comparative analyses of TDIF signaling in non-flowering vascular plants (gymnosperms, monilophytes and lycophytes. We identified orthologs of TDIF/CLE as well as its receptor TDR/PXY (TDIF RECEPTOR/PHLOEM INTERCALATED WITH XYLEM in Ginkgo biloba, Adiantum aethiopicum and Selaginella kraussiana by RACE-PCR. The predicted TDIF peptide sequences in seed plants and monilophytes were identical to that of A. thaliana TDIF. We examined the effects of exogenous CLE peptide-motif sequences of TDIF in these species. We found that liquid culturing of dissected leaves or shoots was useful for examining TDIF activity during vascular development. TDIF treatment suppressed xylem/tracheary element differentiation of procambial cells in G. bioloba and A. aethiopicum leaves. In contrast, neither TDIF nor putative endogenous TDIF inhibited xylem differentiation in developing shoots and rhizophores of S. kraussiana. These data suggest that activity of TDIF in vascular development is conserved among extant euphyllophytes. In addition to the conserved function, via liquid culturing of its bulbils, we found a novel inhibitory activity on root growth in the monilophyte Asplenium x lucrosum suggesting lineage-specific co-option of peptide signaling occurred during the evolution of vascular plant organs.

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

    African Journals Online (AJOL)

    ajl yemi

    2011-12-26

    Dec 26, 2011 ... migration period, transwell chambers were fixed by methyl alcohol and stained by hematoxylin, and then the non-migrating cells were removed with a cotton swab from the upper surface. The number of cells that had migrated to the lower surface of the membrane was determined per ×200 high power field.

  8. Mig-6 Gene Knockout Induces Neointimal Hyperplasia in the Vascular Smooth Muscle Cell

    Directory of Open Access Journals (Sweden)

    Ju Hee Lee

    2014-01-01

    Full Text Available Although advances in vascular interventions can reduce the mortality associated with cardiovascular disease, neointimal hyperplasia remains a clinically significant obstacle limiting the success of current interventions. Identification of signaling pathways involved in migration and proliferation of vascular smooth muscle cells (SMCs is an important approach for the development of modalities to combat this disease. Herein we investigate the role of an immediate early response gene, mitogen-inducible gene-6 (Mig-6, in the development of neointimal hyperplasia using vascular smooth muscle specific Mig-6 knockout mice. We induced endoluminal injury to one side of femoral artery by balloon dilatation in both Mig-6 knockout and control mice. Four weeks following injury, the artery of Mig-6 knockout mice demonstrated a 5.3-fold increase in the neointima/media ratio compared with control mice (P=0.04. In addition, Mig-6 knockout vascular SMCs displayed an increase in both cell migration and proliferation compared with wild-type SMCs. Taken together, our data suggest that Mig-6 plays a critical role in the development of atherosclerosis. This finding provides new insight into the development of more effective ways to treat and prevent neointimal hyperplasia, particularly in-stent restenosis after percutaneous vascular intervention.

  9. Implication of molecular vascular smooth muscle cell heterogeneity among arterial beds in arterial calcification.

    Directory of Open Access Journals (Sweden)

    Olivier Espitia

    Full Text Available Vascular calcification is a strong and independent predictive factor for cardiovascular complications and mortality. Our previous work identified important discrepancies in plaque composition and calcification types between carotid and femoral arteries. The objective of this study is to further characterize and understand the heterogeneity in vascular calcification among vascular beds, and to identify molecular mechanisms underlying this process. We established ECLAGEN biocollection that encompasses human atherosclerotic lesions and healthy arteries from different locations (abdominal, thoracic aorta, carotid, femoral, and infrapopliteal arteries for histological, cell isolation, and transcriptomic analysis. Our results show that lesion composition differs between these locations. Femoral arteries are the most calcified arteries overall. They develop denser calcifications (sheet-like, nodule, and are highly susceptible to osteoid metaplasia. These discrepancies may derive from intrinsic differences between SMCs originating from these locations, as microarray analysis showed specific transcriptomic profiles between primary SMCs isolated from each arterial bed. These molecular differences translated into functional disparities. SMC from femoral arteries showed the highest propensity to mineralize due to an increase in basal TGFβ signaling. Our results suggest that biological heterogeneity of resident vascular cells between arterial beds, reflected by our transcriptomic analysis, is critical in understanding plaque biology and calcification, and may have strong implications in vascular therapeutic approaches.

  10. ROBO4-Mediated Vascular Integrity Regulates the Directionality of Hematopoietic Stem Cell Trafficking

    Directory of Open Access Journals (Sweden)

    Stephanie Smith-Berdan

    2015-02-01

    Full Text Available Despite the use of hematopoietic stem cells (HSCs in clinical therapy for over half a century, the mechanisms that regulate HSC trafficking, engraftment, and life-long persistence after transplantation are unclear. Here, we show that the vascular endothelium regulates HSC trafficking into and out of bone marrow (BM niches. Surprisingly, we found that instead of acting as barriers to cellular entry, vascular endothelial cells, via the guidance molecule ROBO4, actively promote HSC translocation across vessel walls into the BM space. In contrast, we found that the vasculature inhibits the reverse process, as induced vascular permeability led to a rapid increase in HSCs in the blood stream. Thus, the vascular endothelium reinforces HSC localization to BM niches both by promoting HSC extravasation from blood-to-BM and by forming vascular barriers that prevent BM-to-blood escape. Our results uncouple the mechanisms that regulate the directionality of HSC trafficking and show that the vasculature can be targeted to improve hematopoietic transplantation therapies.

  11. Angiogenic T cell expansion correlates with severity of peripheral vascular damage in systemic sclerosis.

    Directory of Open Access Journals (Sweden)

    Mirko Manetti

    Full Text Available The mechanisms underlying endothelial cell injury and defective vascular repair in systemic sclerosis (SSc remain unclear. Since the recently discovered angiogenic T cells (Tang may have an important role in the repair of damaged endothelium, this study aimed to analyze the Tang population in relation to disease-related peripheral vascular features in SSc patients. Tang (CD3+CD31+CXCR4+ were quantified by flow cytometry in peripheral blood samples from 39 SSc patients and 18 healthy controls (HC. Circulating levels of the CXCR4 ligand stromal cell-derived factor (SDF-1α and proangiogenic factors were assessed in paired serum samples by immunoassay. Serial skin sections from SSc patients and HC were subjected to CD3/CD31 and CD3/CXCR4 double immunofluorescence. Circulating Tang were significantly increased in SSc patients with digital ulcers (DU compared either with SSc patients without DU or with HC. Tang levels were significantly higher in SSc patients with late nailfold videocapillaroscopy (NVC pattern than in those with early/active NVC patterns and in HC. No difference in circulating Tang was found when comparing either SSc patients without DU or patients with early/active NVC patterns and HC. In SSc peripheral blood, Tang percentage was inversely correlated to levels of SDF-1α and CD34+CD133+VEGFR-2+ endothelial progenitor cells (EPC, and positively correlated to levels of vascular endothelial growth factor and matrix metalloproteinase-9. Tang were frequently detected in SSc dermal perivascular inflammatory infiltrates. In summary, our findings demonstrate for the first time that Tang cells are selectively expanded in the circulation of SSc patients displaying severe peripheral vascular complications like DU. In SSc, Tang may represent a potentially useful biomarker reflecting peripheral vascular damage severity. Tang expansion may be an ineffective attempt to compensate the need for increased angiogenesis and EPC function. Further

  12. 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.

  13. Improvement of cell infiltration in electrospun polycaprolactone scaffolds for the construction of vascular grafts.

    Science.gov (United States)

    Wang, Kai; Zhu, Meifeng; Li, Ting; Zheng, Wenting; Li, Li; Xu, Mian; Zhao, Qiang; Kong, Deling; Wang, Lianyong

    2014-08-01

    The less-than-ideal cell infiltration resulting from inherently small pore size limits the application of electrospinning scaffold in tissue engineering and regeneration medicine. The present study aims to develop a porogenic method which can significantly increase pore size in electrospinning scaffold and enhance cell migration. With this method, composite scaffolds consisting of poly(epsilon-caprolactone) (PCL) fibers and poly(ethylene oxide) (PEO) microparticles were prepared by simultaneously electrospinning and electrospraying. Removal of the PEO microparticles from the composites generated large pores. In vitro culture of NIH3T3 cells and in vivo subcutaneous implantation both demonstrated that the porogenic scaffolds markedly facilitated cell infiltration. With the same technique, vascular grafts with alternative dense and loose layers were prepared by turning on or off electrospraying PEO. SEM showed that there was no a clear delamination between the loose and dense layers. The mechanical strength and burst pressure of these vascular grafts could meet the requirements of vascular implantation. In conclusion, electrospinning PCL fibers with electrospraying PEO microparticles may be an effective and controllable method to increase pore size in electrospinning scaffold and provides a useful tool for the fabrication of vascular grafts that meets the need of blood vessel replacement.

  14. Vascular smooth muscle cells in cultures on low density polyethylene modified with plasma discharge and biofunctionalization

    Czech Academy of Sciences Publication Activity Database

    Pařízek, Martin; Kasálková, N.; Bačáková, Lucie; Kolářová, K.; Lisá, Věra; Švorčík, V.

    2009-01-01

    Roč. 12, 89-91 (2009), s. 25-28 ISSN 1429-7248 R&D Projects: GA AV ČR(CZ) KAN400480701; GA AV ČR(CZ) 1QS500110564 Institutional research plan: CEZ:AV0Z50110509 Keywords : Ar plasma discharge * low density polyethylene * vascular smooth muscle cells Subject RIV: EI - Biotechnology ; Bionics

  15. Mast Cell Inhibition Improves Pulmonary Vascular Remodeling in Pulmonary Hypertension

    NARCIS (Netherlands)

    Bartelds, Beatrijs; van Loon, Rosa Laura E.; Mohaupt, Saffloer; Wijnberg, Hans; Dickinson, Michael G.; Takens, Janny; van Albada, Mirjam; Berger, Rolf M. F.; Boersma, B.

    Background: Pulmonary arterial hypertension (PAH) is a progressive angioproliferative disease with high morbidity and mortality. Although the histopathology is well described, its pathogenesis is largely unknown. We previously identified the increased presence of mast cells and their markers in a

  16. Peptide-modified PELCL electrospun membranes for regulation of vascular endothelial cells

    International Nuclear Information System (INIS)

    Zhou, Fang; Jia, Xiaoling; Yang, Yang; Yang, Qingmao; Gao, Chao; Zhao, Yunhui; Fan, Yubo; Yuan, Xiaoyan

    2016-01-01

    The efficiency of biomaterials used in small vascular repair depends greatly on their ability to interact with vascular endothelial cells (VECs). Rapid endothelialization of the vascular grafts is a promising way to prevent thrombosis and intimal hyperplasia. In this work, modification of electrospun membranes of poly(ethylene glycol)-b-poly(L-lactide-co-ε-caprolactone) (PELCL) by three different peptides for regulation of VECs were studied in order to obtain ideal bioactive biomaterials as small diameter vascular grafts. QK (a mimetic peptide to vascular endothelial growth factor), Arg-Glu-Asp-Val (REDV, a specific adhesive peptide to VECs) and Val-Ala-Pro-Gly (VAPG, a specific adhesive peptide to vascular smooth muscle cells) were investigated. Surface properties of the modified membranes and the response of VECs were verified. It was found that protein adsorption and platelet adhesion were effectively suppressed with the introduction of QK, REDV or VAPG peptides on the PELCL electrospun membranes. Both QK- and REDV-modified electrospun membranes could accelerate the proliferation of VECs in the first 9 days, and the QK-modified electrospun membrane promoted cell proliferation more significantly than the REDV-modified one. The REDV-modified PELCL membrane was the most favorable for VECs adhesion than QK- and VAPG-modified membranes. It was suggested that QK- or REDV-modified PELCL electrospun membranes may have great potential applications in cardiovascular biomaterials for rapid endothelialization in situ. - Highlights: • A series of peptide-modified PELCL electrospun membranes were prepared. • Hemocompatibility of the membranes was greatly improved by the modification. • QK-modified PELCL membrane promoted VECs proliferation more significantly. • REDV-modified PELCL membrane was the most favorable for VEC adhesion.

  17. Anti-TNF-α activity of Portulaca oleracea in vascular endothelial cells.

    Science.gov (United States)

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

    2012-01-01

    Vascular inflammation plays a key role in the pathogenesis and progression of atherosclerosis, a main complication of diabetes. The present study investigated whether an aqueous extract of Portulaca oleracea (AP) prevents the TNF-α-induced vascular inflammatory process in the human umbilical vein endothelial cell (HUVEC). The stimulation of TNF-α induced overexpression of adhesion molecules affects vascular cell adhesion molecule (VCAM)-1, intercellular adhesion molecule (ICAM)-1 and E-selectin for example. However, AP significantly suppressed TNF-α-induced over-expression of these adhesion molecules in a dose-dependent manner. In addition, pretreatment with AP dose-dependently reduced an increase of the adhesion of HL-60 cells to TNF-α-induced HUVEC. Furthermore, we observed that stimulation of TNF-α significantly increased intracellular reactive oxygen species (ROS) production. However, pretreatment with AP markedly blocked TNF-α-induced ROS production in a dose-dependent manner. The western blot and immunofluorescence analysis showed that AP inhibited the translocation of p65 NF-κB to the nucleus. In addition, AP suppressed the TNF-α-induced degradation of IκB-α and attenuated the TNF-α-induced NF-κB binding. AP also effectively reduced TNF-α-induced mRNA expressions of monocyte chemoattractant protein (MCP)-1 and interleukin (IL)-8 in a dose-dependent manner. Taken together, AP prevents the vascular inflammatory process through the inhibition of intracellular ROS production and NF-κB activation as well as the reduction of adhesion molecule expression in TNF-α-induced HUVEC. These results suggested that AP might have a potential therapeutic effect by inhibiting the vascular inflammation process in vascular diseases such as atherosclerosis.

  18. Novel mitochondria-targeted heat-soluble proteins identified in the anhydrobiotic Tardigrade improve osmotic tolerance of human cells.

    Directory of Open Access Journals (Sweden)

    Sae Tanaka

    Full Text Available Tardigrades are able to tolerate almost complete dehydration through transition to a metabolically inactive state, called "anhydrobiosis". Late Embryogenesis Abundant (LEA proteins are heat-soluble proteins involved in the desiccation tolerance of many anhydrobiotic organisms. Tardigrades, Ramazzottius varieornatus, however, express predominantly tardigrade-unique heat-soluble proteins: CAHS (Cytoplasmic Abundant Heat Soluble and SAHS (Secretory Abundant Heat Soluble proteins, which are secreted or localized in most intracellular compartments, except the mitochondria. Although mitochondrial integrity is crucial to ensure cellular survival, protective molecules for mitochondria have remained elusive. Here, we identified two novel mitochondrial heat-soluble proteins, RvLEAM and MAHS (Mitochondrial Abundant Heat Soluble, as potent mitochondrial protectants from Ramazzottius varieornatus. RvLEAM is a group3 LEA protein and immunohistochemistry confirmed its mitochondrial localization in tardigrade cells. MAHS-green fluorescent protein fusion protein localized in human mitochondria and was heat-soluble in vitro, though no sequence similarity with other known proteins was found, and one region was conserved among tardigrades. Furthermore, we demonstrated that RvLEAM protein as well as MAHS protein improved the hyperosmotic tolerance of human cells. The findings of the present study revealed that tardigrade mitochondria contain at least two types of heat-soluble proteins that might have protective roles in water-deficient environments.

  19. Adeno-associated virus vector transduction of vascular smooth muscle cells in vivo.

    Science.gov (United States)

    Richter, M; Iwata, A; Nyhuis, J; Nitta, Y; Miller, A D; Halbert, C L; Allen, M D

    2000-04-27

    Adeno-associated virus (AAV) vectors might offer solutions for restenosis and angiogenesis by transducing nondividing cells and providing long-term gene expression. We investigated the feasibility of vascular cell transduction by AAV vectors in an in vivo rabbit carotid artery model. Time course of gene expression, inflammatory reaction to the vector, and effects of varying viral titer, exposure time, and intraluminal pressures on gene expression were examined. Recombinant AAV vectors with an Rous sarcoma virus promoter and alkaline phosphatase reporter gene were injected intraluminally into transiently isolated carotid segments. Following transduction, gene expression increased significantly over 14 days and then remained stable to 28 days, the last time point examined. Medial vascular smooth muscle cells were the main cell type transduced even with an intact endothelial layer. Increasing the viral titer and intraluminal pressure both enhanced transduction efficiency to achieve a mean of 34 +/- 7% of the subintimal layer of smooth muscle cells expressing gene product. A mild inflammatory reaction, composed of T cells with only rare macrophages, with minimal intimal thickening was demonstrated in 40% of transduced vessels; inflammatory cells were not detected in sham-operated control arteries. These findings demonstrate that AAV is a promising vector for intravascular applications in coronary and peripheral vascular diseases.

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

    DEFF Research Database (Denmark)

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

    2006-01-01

    /hyaluronan complexes, an ECM network that has been suggested to be important during tissue repair. In this study we have analysed the presence of fibulin-2 in two different models of murine vascular lesions. We have also examined how the fibulin-2/versican network influences SMC migration. Methods: Presence of fibulin......Objective: The vascular extracellular matrix (ECM) can affect smooth muscle cell (SMC) adhesion, migration and proliferation-events that are important during the atherosclerotic process. Fibulin-2 is a member of the ECM protein family of fibulins and has been found to cross-link versican...... that regulates SMC migration during vessel wall repair....

  1. Obesity-induced vascular dysfunction and arterial stiffening requires endothelial cell arginase 1.

    Science.gov (United States)

    Bhatta, Anil; Yao, Lin; Xu, Zhimin; Toque, Haroldo A; Chen, Jijun; Atawia, Reem T; Fouda, Abdelrahman Y; Bagi, Zsolt; Lucas, Rudolf; Caldwell, Ruth B; Caldwell, Robert W

    2017-11-01

    Elevation of arginase activity has been linked to vascular dysfunction in diabetes and hypertension by a mechanism involving decreased nitric oxide (NO) bioavailability due to L-arginine depletion. Excessive arginase activity also can drive L-arginine metabolism towards the production of ornithine, polyamines, and proline, promoting proliferation of vascular smooth muscle cells and collagen formation, leading to perivascular fibrosis. We hypothesized that there is a specific involvement of arginase 1 expression within the vascular endothelial cells in this pathology. To test this proposition, we used models of type 2 diabetes and metabolic syndrome. Studies were performed using wild type (WT), endothelial-specific arginase 1 knockout (EC-A1-/-) and littermate controls(A1con) mice fed high fat-high sucrose (HFHS) or normal diet (ND) for 6 months and isolated vessels exposed to palmitate-high glucose (PA/HG) media. Some WT mice or isolated vessels were treated with an arginase inhibitor, ABH [2-(S)-amino-6-boronohexanoic acid. In WT mice, the HFHS diet promoted increases in body weight, fasting blood glucose, and post-prandial insulin levels along with arterial stiffening and fibrosis, elevated blood pressure, decreased plasma levels of L-arginine, and elevated L-ornithine. The HFHS diet or PA/HG treatment also induced increases in vascular arginase activity along with oxidative stress, reduced vascular NO levels, and impaired endothelial-dependent vasorelaxation. All of these effects except obesity and hypercholesterolemia were prevented or significantly reduced by endothelial-specific deletion of arginase 1 or ABH treatment. Vascular dysfunctions in diet-induced obesity are prevented by deletion of arginase 1 in vascular endothelial cells or arginase inhibition. These findings indicate that upregulation of arginase 1 expression/activity in vascular endothelial cells has an integral role in diet-induced cardiovascular dysfunction and metabolic syndrome. Published

  2. Invasion of Porphyromonas gingivalis strains into vascular cells and tissue

    Directory of Open Access Journals (Sweden)

    Ingar Olsen

    2015-08-01

    Full Text Available Porphyromonas gingivalis is considered a major pathogen in adult periodontitis and is also associated with multiple systemic diseases, for example, cardiovascular diseases. One of its most important virulence factors is invasion of host cells. The invasion process includes attachment, entry/internalization, trafficking, persistence, and exit. The present review discusses these processes related to P. gingivalis in cardiovascular cells and tissue. Although most P. gingivalis strains invade, the invasion capacity of strains and the mechanisms of invasion including intracellular trafficking among them differ. This is consistent with the fact that there are significant differences in the pathogenicity of P. gingivalis strains. P. gingivalis invasion mechanisms are also dependent on types of host cells. Although much is known about the invasion process of P. gingivalis, we still have little knowledge of its exit mechanisms. Nevertheless, it is intriguing that P. gingivalis can remain viable in human cardiovascular cells and atherosclerotic plaque and later exit and re-enter previously uninfected host cells.

  3. Vascular endothelial growth factor A and vascular endothelial growth factor receptor 2 expression in non-small cell lung cancer patients: relation to prognosis

    DEFF Research Database (Denmark)

    Bonnesen, Barbara; Pappot, Helle; Holmstav, Julie

    2009-01-01

    elements in neoplastic cells and their microenvironment have recently been and are continuously developed including drugs inhibiting the angiogenic system. Angiogenic factor vascular endothelial growth factor (VEGF) and its receptor vascular endothelial growth factor receptor 2 (VEGFR2) seem to play key......BACKGROUND: The majority of patients with non-small cell lung cancer (NSCLC) are diagnosed with advanced inoperable disease. While treatment with conventional chemotherapy has improved during the last decade the 5 years survival is still modest. Novel drugs, which selectively target aberrant...

  4. Do cell based tissue engineering products for meniscus regeneration influence vascularization?

    Science.gov (United States)

    Koch, Matthias; Ehrenreich, Tobias; Koehl, Gudrun; Pattappa, Girish; Pfeifer, Christian; Loibl, Markus; Müller, Michael; Nerlich, Michael; Angele, Peter; Zellner, Johannes

    2017-01-01

    Meniscus regeneration is observed within the peripheral, vascularized zone but decreases in the inner two thirds alongside the vascularization. Within this avascular area, cell-based tissue-engineering-approaches appear to be a promising strategy for the treatment of meniscal defects. Evaluation of the angiogenic potential of cell-based tissue-engineering-products for meniscus healing. Evaluation of angiogenesis induced by rabbit meniscus-pellets, meniscus-cells (MC) or mesenchymal stem-cells (MSC) in cell-based tissue-engineering-products within a rabbit meniscus-ring was performed using a transparent dorsal skin fold chamber in nude mice. Observations were undertaken during a 14 days period. Cell preconditioning differed between experimental groups. Immunohistochemical analysis of the regenerated tissue in the meniscus-ring induced by cell loaded composite scaffolds for differentiation and anti-angiogenic factors were performed. Meniscus-pellets and MSC-/MC-based tissue-engineering-products induced angiogenesis. An accelerated vascularization was detected in the group of meniscus-pellets derived from the vascularized zone compared to avascular meniscus-pellets. In terms of cell-based tissue-engineering-products, chondrogenic preconditioning resulted in significantly increased vessel growth. MSC-constructs showed an accelerated angiogenesis. Immunohistochemical evaluation showed a progressive differentiation and lower content for anti-angiogenic endostatin in the precultured group. Preconditioning of MC-/MSC-based tissue-engineering-products is a promising tool to influence the angiogenic potential of tissue-engineering-products and to adapt these properties according to the aimed tissue qualities.

  5. Atherogenic ω-6 Lipids Modulate PPAR- EGR-1 Crosstalk in Vascular Cells

    Directory of Open Access Journals (Sweden)

    Jia Fei

    2011-01-01

    Full Text Available Atherogenic ω-6 lipids are physiological ligands of peroxisome proliferator-activated receptors (PPARs and elicit pro- and antiatherogenic responses in vascular cells. The objective of this study was to investigate if ω-6 lipids modulated the early growth response-1 (Egr-1/PPAR crosstalk thereby altering vascular function. Rat aortic smooth muscle cells (RASMCs were exposed to ω-6 lipids, linoleic acid (LA, or its oxidized form, 13-HPODE (OxLA in the presence or absence of a PPARα antagonist (MK886 or PPARγ antagonist (GW9662 or PPAR-specific siRNA. Our results demonstrate that ω-6 lipids, induced Egr-1 and monocyte chemotactic protein-1 (MCP-1 mRNA and protein levels at the acute phase (1–4 hrs when PPARα was downregulated and at subacute phase (4–12 hrs by modulating PPARγ, thus resulting in altered monocyte adhesion to RASMCs. We provide novel insights into the mechanism of action of ω-6 lipids on Egr-1/PPAR interactions in vascular cells and their potential in altering vascular function.

  6. Composite vascular grafts with high cell infiltration by co-electrospinning.

    Science.gov (United States)

    Tan, Zhikai; Wang, Hongjie; Gao, Xiangkai; Liu, Tong; Tan, Yongjun

    2016-10-01

    There is an increasing demand for functional small-diameter vascular grafts (diameterElectrospinning has become a popular polymer processing technique for vascular tissue engineering, but the grafts fabricated by electrospinning often have relatively small pores and low porosity, which limit cell infiltration into scaffolds and hinder the regeneration and remodeling of grafts. In the present study, we aimed to develop an efficient method to prepare electrospun composite vascular grafts comprising natural and synthetic materials. We fabricated grafts made of polycaprolactone, gelatin, and polyvinyl alcohol (PVA) by co-electrospinning, and the scaffolds were further functionalized by immobilizing heparin on them. The PVA fibers degraded rapidly in vivo and generated electrospun scaffolds with high porosity, which significantly enhanced cell proliferation and infiltration. The mechanical properties of the grafts are suitable for use in artery replacement. Heparin functionalization of the grafts yielded a good antithrombogenic effect, which was demonstrated in platelet adhesion tests. Moreover, in vitro and in vivo results demonstrated that the heparin release from the grafts enhanced the growth of endothelial cells, which is important for the endothelium of implanted grafts. The results of this study indicate that our method is effective and controllable for the fabrication of vascular grafts that meet the clinical requirements for blood vessel transplantation. Copyright © 2016 Elsevier B.V. All rights reserved.

  7. Adipose-derived stem cell differentiation as a basic tool for vascularized adipose tissue engineering.

    Science.gov (United States)

    Volz, Ann-Cathrin; Huber, Birgit; Kluger, Petra J

    2016-01-01

    The development of in vitro adipose tissue constructs is highly desired to cope with the increased demand for substitutes to replace damaged soft tissue after high graded burns, deformities or tumor removal. To achieve clinically relevant dimensions, vascularization of soft tissue constructs becomes inevitable but still poses a challenge. Adipose-derived stem cells (ASCs) represent a promising cell source for the setup of vascularized fatty tissue constructs as they can be differentiated into adipocytes and endothelial cells in vitro and are thereby available in sufficiently high cell numbers. This review summarizes the currently known characteristics of ASCs and achievements in adipogenic and endothelial differentiation in vitro. Further, the interdependency of adipogenesis and angiogenesis based on the crosstalk of endothelial cells, stem cells and adipocytes is addressed at the molecular level. Finally, achievements and limitations of current co-culture conditions for the construction of vascularized adipose tissue are evaluated. Copyright © 2016 International Society of Differentiation. Published by Elsevier B.V. All rights reserved.

  8. Human umbilical cord blood-derived mesenchymal stem cells promote vascular growth in vivo.

    Directory of Open Access Journals (Sweden)

    Santiago Roura

    Full Text Available Stem cell therapies are promising strategies to regenerate human injured tissues, including ischemic myocardium. Here, we examined the acquisition of properties associated with vascular growth by human umbilical cord blood-derived mesenchymal stem cells (UCBMSCs, and whether they promoted vascular growth in vivo. UCBMSCs were induced in endothelial cell-specific growth medium (EGM-2 acquiring new cell markers, increased Ac-LDL uptake, and migratory capacity as assessed by qRT-PCR, Western blotting, indirect immunofluorescence, and invasion assays. Angiogenic and vasculogenic potentials could be anticipated by in vitro experiments showing self organization into Matrigel-mediated cell networks, and activation of circulating angiogenic-supportive myeloid cells. In mice, following subcutaneous co-injection with Matrigel, UCBMSCs modified to co-express bioluminescent (luciferases and fluorescent proteins were demonstrated to participate in the formation of new microvasculature connected with the host circulatory system. Response of UCBMSCs to ischemia was explored in a mouse model of acute myocardial infarction (MI. UCBMSCs transplanted using a fibrin patch survived 4 weeks post-implantation and organized into CD31(+network structures above the infarcted myocardium. MI-treated animals showed a reduced infarct scar and a larger vessel-occupied area in comparison with MI-control animals. Taken together, the presented results show that UCBMSCs can be induced in vitro to acquire angiogenic and vasculogenic properties and contribute to vascular growth in vivo.

  9. Characterization of ionizing radiation-induced unfolded protein response in human vascular endothelial cells

    International Nuclear Information System (INIS)

    Kim, Eun Ju; Lee, Yoon Jin; Kang, Seong Man

    2013-01-01

    Misfolded or unfolded proteins within the endoplasmic reticulum (ER stress), viral infection, or amino acid deprivation induce eukaryotic translation initiation factor 2α phosphorylation (eIF2α) in eukaryotic cells, repressing global protein synthesis coincident with preferential translation of activating transcription factor 4 (ATF4). ATF4 is a transcriptional activator of genes involved in amino acid metabolism, cellular redox homeostasis, and regulation of apoptosis. When the eIF2α/ATF4 pathway is initiated by ER stress, the pathway is referred toas the unfolded protein response (UPR). In addition to DNA, proteins may be initial and important targets of ionizing radiation (IR), and the damaged protein can trigger ER stress pathway. Recent investigations suggested that IR induces ER stress followed by UPR in various cell types including intestinal epithelial cells. We conducted this study to determine whether IR can activate UPR in human vascular endothelial cells. Our data have shown that IR increased PERK-dependent eIF2α phosphorylation accompanied by induction in ATF4 protein levels in human vascular endothelial cells without alterations in expressions of XBP-1s and GRP78. Based on these data, we suggest that IR selectively activates PERK branch of unfolded protein response in human vascular endothelial cells

  10. Characterization of ionizing radiation-induced unfolded protein response in human vascular endothelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Eun Ju; Lee, Yoon Jin; Kang, Seong Man [Laboratory of Radiation Effect, Korea Institute of Radiological and Medical Sciences, Seoul (Korea, Republic of)

    2013-04-15

    Misfolded or unfolded proteins within the endoplasmic reticulum (ER stress), viral infection, or amino acid deprivation induce eukaryotic translation initiation factor 2α phosphorylation (eIF2α) in eukaryotic cells, repressing global protein synthesis coincident with preferential translation of activating transcription factor 4 (ATF4). ATF4 is a transcriptional activator of genes involved in amino acid metabolism, cellular redox homeostasis, and regulation of apoptosis. When the eIF2α/ATF4 pathway is initiated by ER stress, the pathway is referred toas the unfolded protein response (UPR). In addition to DNA, proteins may be initial and important targets of ionizing radiation (IR), and the damaged protein can trigger ER stress pathway. Recent investigations suggested that IR induces ER stress followed by UPR in various cell types including intestinal epithelial cells. We conducted this study to determine whether IR can activate UPR in human vascular endothelial cells. Our data have shown that IR increased PERK-dependent eIF2α phosphorylation accompanied by induction in ATF4 protein levels in human vascular endothelial cells without alterations in expressions of XBP-1s and GRP78. Based on these data, we suggest that IR selectively activates PERK branch of unfolded protein response in human vascular endothelial cells.

  11. Endothelial cell chimerism after renal transplantation and vascular rejection.

    NARCIS (Netherlands)

    Lagaaij, E.L.; Cramer-Knijnenburg, G.F.; Kemenade, F.J. van; Es, L.A. van; Bruijn, J.A.; Krieken, J.H.J.M. van

    2001-01-01

    BACKGROUND: The blood vessels of a transplanted organ are the interface between donor and recipient. The endothelium in the blood vessels is thought to be the major target for graft rejection. Endothelial cells of a transplanted organ are believed to remain of donor origin after transplantation. We

  12. Vascular Cell Senescence Contributes to Blood-Brain Barrier Breakdown

    NARCIS (Netherlands)

    Yamazaki, Y.; Baker, D.J.; Tachibana, M.; Liu, C.C.; Deursen, J.M.A. van; Brott, T.G.; Bu, G.; Kanekiyo, T.

    2016-01-01

    BACKGROUND AND PURPOSE: Age-related changes in the cerebrovasculature, including blood-brain barrier (BBB) disruption, are emerging as potential risks for diverse neurological conditions. Because the accumulation of senescent cells in tissues is increasingly recognized as a critical step leading to

  13. Succinobucol induces apoptosis in vascular smooth muscle cells

    Czech Academy of Sciences Publication Activity Database

    Midwinter, R.G.; Maghzal, G.; Dennis, J.M.; Wu, B.J.; Cai, H.; Kapralov, A.A.; Belikova, N.A.; Tyurina, Y.Y.; Dong, L. F.; Khachigian, L.; Neužil, Jiří; Kagan, V.E.; Stocker, R.

    2012-01-01

    Roč. 52, č. 5 (2012), s. 871-879 ISSN 0891-5849 R&D Projects: GA ČR(CZ) GAP301/10/1937 Institutional research plan: CEZ:AV0Z50520701 Keywords : reactive oxygen species * free radicals * apoptosis Subject RIV: EA - Cell Biology Impact factor: 5.271, year: 2012

  14. Chaperone-assisted thermostability engineering of a soluble T cell receptor using phage display

    DEFF Research Database (Denmark)

    Gunnarsen, Kristin S; Kristinsson, Solveig G; Justesen, Sune

    2013-01-01

    We here report a novel phage display selection strategy enabling fast and easy selection of thermostabilized proteins. The approach is illustrated with stabilization of an aggregation-prone soluble single chain T cell receptor (scTCR) characteristic of the murine MOPC315 myeloma model. Random...... mutation scTCR phage libraries were prepared in E. coli over-expressing the periplasmic chaperone FkpA, and such over-expression during library preparation proved crucial for successful downstream selection. The thermostabilized scTCR(mut) variants selected were produced in high yields and isolated...

  15. Water Soluble Aluminum Paste Using Polyvinyl Alcohol for Silicon Solar Cells

    Directory of Open Access Journals (Sweden)

    Abdullah Uzum

    2015-01-01

    Full Text Available Screen-printing aluminum is still dominantly used in the solar cell fabrication process. Ethyl cellulose is one of the main contents of screen-printing pastes that require dichloromethane for its cleaning process, a substance renowned for being extremely toxic and threatening to the human body. Developing environmental friendly aluminum pastes is essential in order to provide an alternative to the commercial pastes. In this work, new, nontoxic polyvinyl alcohol-based aluminum pastes are introduced. Polyvinyl alcohol was used as a soluble polymer that can be synthesized without saponification and that is also soluble in water. Three different pastes were developed using different recipes including many aluminum particle sizes varying from 3.0 to 45 μm, aluminum oxide with particle sizes between 35 and 50 μm, and acetic acid. Evaluation of the pastes was carried out by Scanning Electron Microscope (SEM image analysis, sheet resistance measurements, and fabricating silicon solar cells using each paste. Solar cells with 15.6% efficiency were fabricated by nonvacuum processing on CZ-Si p-type wafers using developed aluminum pastes on the back side.

  16. Water soluble glucose derivative of thiocarbohydrazone acts as ionophore with cytotoxic effects on tumor cells.

    Science.gov (United States)

    Bonaccorso, Carmela; Grasso, Giulia; Musso, Nicolò; Barresi, Vincenza; Condorelli, Daniele F; La Mendola, Diego; Rizzarelli, Enrico

    2018-05-01

    A novel water-soluble ionophore based on the thiocarbohydrazone moiety conjugated with glucose (GluTch) was synthesized through a simple two-step procedure. Structural elucidation was carried out in water solution by means of various spectroscopic techniques (NMR, UV-Vis, and CD), electrospray ionization mass spectrometry and density functional theory calculations. The flexible nature of the thiocarbohydrazone moiety of the new glycoderivative compound induced both different coordination motifs and stoichiometry towards copper and zinc. Cytotoxicity assays of the ligands on the human normal keratinocyte NCTC-2544, MDA-MB-231 breast cancer and PC-3 human prostate adenocarcinoma cell lines demonstrated that i) higher activity on cancer cells growth inhibition compared to a normal cell line; ii) the introduction of the glucose unit does not alter the cytotoxic activity of the underivatized ionophore ligand and iii) the presence of copper ion improves the activity of the thiocarbohydrazones. Copyright © 2018 Elsevier Inc. All rights reserved.

  17. ER Alpha Rapid Signaling Is Required for Estrogen Induced Proliferation and Migration of Vascular Endothelial Cells.

    Directory of Open Access Journals (Sweden)

    Qing Lu

    Full Text Available Estrogen promotes the proliferation and migration of vascular endothelial cells (ECs, which likely underlies its ability to accelerate re-endothelialization and reduce adverse remodeling after vascular injury. In previous studies, we have shown that the protective effects of E2 (the active endogenous form of estrogen in vascular injury require the estrogen receptor alpha (ERα. ERα transduces the effects of estrogen via a classical DNA binding, "genomic" signaling pathway and via a more recently-described "rapid" signaling pathway that is mediated by a subset of ERα localized to the cell membrane. However, which of these pathways mediates the effects of estrogen on endothelial cells is poorly understood. Here we identify a triple point mutant version of ERα (KRR ERα that is specifically defective in rapid signaling, but is competent to regulate transcription through the "genomic" pathway. We find that in ECs expressing wild type ERα, E2 regulates many genes involved in cell migration and proliferation, promotes EC migration and proliferation, and also blocks the adhesion of monocytes to ECs. ECs expressing KRR mutant ERα, however, lack all of these responses. These observations establish KRR ERα as a novel tool that could greatly facilitate future studies into the vascular and non-vascular functions of ERα rapid signaling. Further, they support that rapid signaling through ERα is essential for many of the transcriptional and physiological responses of ECs to E2, and that ERα rapid signaling in ECs, in vivo, may be critical for the vasculoprotective and anti-inflammatory effects of estrogen.

  18. ER Alpha Rapid Signaling Is Required for Estrogen Induced Proliferation and Migration of Vascular Endothelial Cells

    Science.gov (United States)

    Lu, Qing; Schnitzler, Gavin R.; Ueda, Kazutaka; Iyer, Lakshmanan K.; Diomede, Olga I.; Andrade, Tiffany; Karas, Richard H.

    2016-01-01

    Estrogen promotes the proliferation and migration of vascular endothelial cells (ECs), which likely underlies its ability to accelerate re-endothelialization and reduce adverse remodeling after vascular injury. In previous studies, we have shown that the protective effects of E2 (the active endogenous form of estrogen) in vascular injury require the estrogen receptor alpha (ERα). ERα transduces the effects of estrogen via a classical DNA binding, “genomic” signaling pathway and via a more recently-described “rapid” signaling pathway that is mediated by a subset of ERα localized to the cell membrane. However, which of these pathways mediates the effects of estrogen on endothelial cells is poorly understood. Here we identify a triple point mutant version of ERα (KRR ERα) that is specifically defective in rapid signaling, but is competent to regulate transcription through the “genomic” pathway. We find that in ECs expressing wild type ERα, E2 regulates many genes involved in cell migration and proliferation, promotes EC migration and proliferation, and also blocks the adhesion of monocytes to ECs. ECs expressing KRR mutant ERα, however, lack all of these responses. These observations establish KRR ERα as a novel tool that could greatly facilitate future studies into the vascular and non-vascular functions of ERα rapid signaling. Further, they support that rapid signaling through ERα is essential for many of the transcriptional and physiological responses of ECs to E2, and that ERα rapid signaling in ECs, in vivo, may be critical for the vasculoprotective and anti-inflammatory effects of estrogen. PMID:27035664

  19. Paraquat induces oxidative stress and neuronal cell death; neuroprotection by water-soluble Coenzyme Q10

    International Nuclear Information System (INIS)

    McCarthy, S.; Somayajulu, M.; Sikorska, M.; Borowy-Borowski, H.; Pandey, S.

    2004-01-01

    Neuronal cell death induced by oxidative stress is correlated with numerous neurodegenerative diseases, including Alzheimer's disease (AD), Parkinson's disease (PD), and stroke. The causes of sporadic forms of age-related neurodegenerative diseases are still unknown. Recently, a correlation between paraquat exposure and neurodegenerative diseases has been observed. Paraquat, a nonselective herbicide, was once widely used in North America and is still routinely used in Taiwan. We have used differentiated Human Neuroblastoma (SHSY-5Y) cells as an in vitro model to study the mechanism of cell death induced by paraquat. We observed that paraquat-induced oxidative stress in differentiated SHSY-5Y cells as indicated by an increase in the production of cellular reactive oxygen species (ROS). Furthermore, apoptosis was evident as indicated by cellular and nuclear morphology and DNA fragmentation. Interestingly, pretreatment of SHSY-5Y cells with water-soluble Coenzyme Q 10 (CoQ 10 ) before paraquat exposure inhibited ROS generation. Pretreatment with CoQ 10 also significantly reduced the number of apoptotic cells and DNA fragmentation. We also analyzed the effect of paraquat and CoQ 10 on isolated mitochondria. Our results indicated that treatment with paraquat induced the generation of ROS from isolated mitochondria and depolarization of the inner mitochondrial membrane. Pretreatment with CoQ 10 was able to inhibit ROS generation from isolated mitochondria as well as the collapse of mitochondrial membrane potential. Our results indicate that water-soluble CoQ 10 can prevent oxidative stress and neuronal damage induced by paraquat and therefore, can be used for the prevention and therapy of neurodegenerative diseases caused by environmental toxins

  20. Insulin resistance in vascular endothelial cells promotes intestinal tumour formation

    DEFF Research Database (Denmark)

    Wang, X; Häring, M-F; Rathjen, Thomas

    2017-01-01

    The risk of several cancers, including colorectal cancer, is increased in patients with obesity and type 2 diabetes, conditions characterised by hyperinsulinaemia and insulin resistance. Because hyperinsulinaemia itself is an independent risk factor for cancer development, we examined tissue...... did not change intestinal tumour number or size distribution on either a low or high-fat diet. We therefore asked whether cells in the tumour stroma might explain the association between tumour formation and insulin resistance. To this end, we generated Apc(Min/+) mice with loss of insulin receptors...... and increased the frequency of neutrophils in tumours. We conclude that although insulin is mitogenic for intestinal tumour cells in vitro, impaired insulin action in the tumour microenvironment may be more important in conditions where hyperinsulinaemia is secondary to insulin resistance. Insulin resistance...

  1. Slow and sustained nitric oxide releasing compounds inhibit multipotent vascular stem cell proliferation and differentiation without causing cell death

    Energy Technology Data Exchange (ETDEWEB)

    Curtis, Brandon M.; Leix, Kyle Alexander [Department of Chemistry, Central Michigan University, Mount Pleasant, MI 48859 (United States); Ji, Yajing [Department of Biomedical Science and Medicine, Michigan State University, East Lansing, MI 48824 (United States); Glaves, Richard Samuel Elliot [Department of Biology, Central Michigan University, Mount Pleasant, MI 48859 (United States); Ash, David E. [Department of Chemistry, Central Michigan University, Mount Pleasant, MI 48859 (United States); Mohanty, Dillip K., E-mail: Mohan1dk@cmich.edu [Department of Chemistry, Central Michigan University, Mount Pleasant, MI 48859 (United States)

    2014-07-18

    Highlights: • Multipotent vascular stem cells (MVSCs) proliferate and differentiate. • Nitric oxide inhibits proliferation of MVSCs. • Nitric oxide inhibits MVSC differentiation to mesenchymal-like stem cells (MSCs). • Smooth muscle cells (SMCs) neither de-differentiate nor proliferate. - Abstract: Atherosclerosis is the leading cause of cerebral and myocardial infarction. It is believed that neointimal growth common in the later stages of atherosclerosis is a result of vascular smooth muscle cell (SMC) de-differentiation in response to endothelial injury. However, the claims of the SMC de-differentiation theory have not been substantiated by monitoring the fate of mature SMCs in response to such injuries. A recent study suggests that atherosclerosis is a consequence of multipotent vascular stem cell (MVSC) differentiation. Nitric oxide (NO) is a well-known mediator against atherosclerosis, in part because of its inhibitory effect on SMC proliferation. Using three different NO-donors, we have investigated the effects of NO on MVSC proliferation. Results indicate that NO inhibits MVSC proliferation in a concentration dependent manner. A slow and sustained delivery of NO proved to inhibit proliferation without causing cell death. On the other hand, larger, single-burst NO concentrations, inhibits proliferation, with concurrent significant cell death. Furthermore, our results indicate that endogenously produced NO inhibits MVSC differentiation to mesenchymal-like stem cells (MSCs) and subsequently to SMC as well.

  2. Slow and sustained nitric oxide releasing compounds inhibit multipotent vascular stem cell proliferation and differentiation without causing cell death

    International Nuclear Information System (INIS)

    Curtis, Brandon M.; Leix, Kyle Alexander; Ji, Yajing; Glaves, Richard Samuel Elliot; Ash, David E.; Mohanty, Dillip K.

    2014-01-01

    Highlights: • Multipotent vascular stem cells (MVSCs) proliferate and differentiate. • Nitric oxide inhibits proliferation of MVSCs. • Nitric oxide inhibits MVSC differentiation to mesenchymal-like stem cells (MSCs). • Smooth muscle cells (SMCs) neither de-differentiate nor proliferate. - Abstract: Atherosclerosis is the leading cause of cerebral and myocardial infarction. It is believed that neointimal growth common in the later stages of atherosclerosis is a result of vascular smooth muscle cell (SMC) de-differentiation in response to endothelial injury. However, the claims of the SMC de-differentiation theory have not been substantiated by monitoring the fate of mature SMCs in response to such injuries. A recent study suggests that atherosclerosis is a consequence of multipotent vascular stem cell (MVSC) differentiation. Nitric oxide (NO) is a well-known mediator against atherosclerosis, in part because of its inhibitory effect on SMC proliferation. Using three different NO-donors, we have investigated the effects of NO on MVSC proliferation. Results indicate that NO inhibits MVSC proliferation in a concentration dependent manner. A slow and sustained delivery of NO proved to inhibit proliferation without causing cell death. On the other hand, larger, single-burst NO concentrations, inhibits proliferation, with concurrent significant cell death. Furthermore, our results indicate that endogenously produced NO inhibits MVSC differentiation to mesenchymal-like stem cells (MSCs) and subsequently to SMC as well

  3. Taurine prevents beta-glycerophosphate-induced calcification in cultured rat vascular smooth muscle cells.

    Science.gov (United States)

    Li, Juxiang; Zhang, Baohong; Huang, Zhiyu; Wang, Shuhen; Tang, Chaoshu; Du, Junbao

    2004-05-01

    Vascular calcification is an ectopic calcification that commonly occurs in atherosclerosis. Because taurine was previously shown to protect against cardiovascular diseases, the effect of taurine on vascular calcification was evaluated in calcified vascular smooth muscle cells (VSMCs) of rat in vitro in the present study. Osteoblastic differentiation, calcification, and proliferation in VSMCs were detected in the presence and absence of taurine. Alkaline phosphatase (ALP), cellular calcium content, and (45)Ca accumulation were measured as the indicators of osteoblastic differentiation and calcification. Incubation of VSMCs with Beta-glycerophosphate for 10 days induced an osteoblast-like morphological change. The activity of ALP was enhanced. Calcium content and (45)Ca uptake were increased in these cells. Calcification of these VSMCs was demonstrated with Beta-glycerophosphate treatment. In association with these alterations, cell proliferation, detected by cell counting, [(3)H]thymidine ([(3)H]TdR), and [(3)H]leucine ([(3)H]Leu) incorporation, was also increased in these calcified VSMCs. Taurine at 20 mmol/l decreased calcium content, (45)Ca(2+) uptake, and ALP activity both after early and late treatment, in which a reduction of the cell count, [(3)H"]TdR, and [(3)H]Leu incorporation of calcified VSMCs was also noted. Compared with the calcified group, morphological changes in the VSMCs of the early-treated group were deferred. These results demonstrated that calcification of VSMCs could be alleviated by taurine. Taurine treatment appeared to be more beneficial when the treatment was started earlier.

  4. Soluble ephrin a1 is necessary for the growth of HeLa and SK-BR3 cells

    Directory of Open Access Journals (Sweden)

    Bazowski Jessa

    2010-10-01

    Full Text Available Abstract Background Ephrin A1 (EFNA1 is a member of the A-type ephrin family of cell surface proteins that function as ligands for the A-type Eph receptor tyrosine kinase family. In malignancy, the precise role of EFNA1 and its preferred receptor, EPHA2, is controversial. Several studies have found that EFNA1 may suppress EPHA2-mediated oncogenesis, or enhance it, depending on cell type and context. However, little is known about the conditions that influence whether EFNA1 promotes or suppresses tumorigenicity. EFNA1 exists in a soluble form as well as a glycophosphatidylinositol (GPI membrane attached form. We investigated whether the contradictory roles of EFNA1 in malignancy might in part be related to the existence of both soluble and membrane attached forms of EFNA1 and potential differences in the manner in which they interact with EPHA2. Results Using a RNAi strategy to reduce the expression of endogenous EFNA1 and EPHA2, we found that both EFNA1 and EPHA2 are required for growth of HeLa and SK-BR3 cells. The growth defects could be rescued by conditioned media from cells overexpressing soluble EFNA1. Interestingly, we found that overexpression of the membrane attached form of EFNA1 suppresses growth of HeLa cells in 3D but not 2D. Knockdown of endogenous EFNA1, or overexpression of full-length EFNA1, resulted in relocalization of EPHA2 from the cell surface to sites of cell-cell contact. Overexpression of soluble EFNA1 however resulted in more EPHA2 distributed on the cell surface, away from cell-cell contacts, and promoted the growth of HeLa cells. Conclusions We conclude that soluble EFNA1 is necessary for the transformation of HeLa and SK-BR3 cells and participates in the relocalization of EPHA2 away from sites of cell-cell contact during transformation.

  5. Regulation of vascular endothelial cell polarization and migration by Hsp70/Hsp90-organizing protein.

    Science.gov (United States)

    Li, Jingyu; Sun, Xiaodong; Wang, Zaizhu; Chen, Li; Li, Dengwen; Zhou, Jun; Liu, Min

    2012-01-01

    Hsp70/Hsp90-organizing protein (HOP) is a member of the co-chaperone family, which directly binds to chaperones to regulate their activities. The participation of HOP in cell motility and endothelial cell functions remains largely unknown. In this study, we demonstrate that HOP is critically involved in endothelial cell migration and angiogenesis. Tube formation and capillary sprouting experiments reveal that depletion of HOP expression significantly inhibits vessel formation from endothelial cells. Wound healing and transwell migration assays show that HOP is important for endothelial cell migration. By examination of centrosome reorientation and membrane ruffle dynamics, we find that HOP plays a crucial role in the establishment of cell polarity in response to migratory stimulus. Furthermore, our data show that HOP interacts with tubulin and colocalizes with microtubules in endothelial cells. These findings indicate HOP as a novel regulator of angiogenesis that functions through promoting vascular endothelial cell polarization and migration.

  6. TRAIL-coated lipid-nanoparticles overcome resistance to soluble recombinant TRAIL in non-small cell lung cancer cells

    Science.gov (United States)

    De Miguel, Diego; Gallego-Lleyda, Ana; María Ayuso, José; Erviti-Ardanaz, Sandra; Pazo-Cid, Roberto; del Agua, Celia; José Fernández, Luis; Ochoa, Ignacio; Anel, Alberto; Martinez-Lostao, Luis

    2016-05-01

    Purpose. Non-small cell lung cancer (NSCLC) is one the types of cancer with higher prevalence and mortality. Apo2-Ligand/TRAIL is a TNF family member able to induce apoptosis in tumor cells but not in normal cells. It has been tested in clinical trials against different types of human cancer including NSCLC. However, results of clinical trials have shown a limited efficacy of TRAIL-based therapies. Recently we have demonstrated that artificial lipid nanoparticles coated with bioactive Apo2L/TRAIL (LUV-TRAIL) greatly improved TRAIL cytotoxic ability being capable of killing chemoresistant hematological cancer cells. In the present work we have extended the study to NSCLC. Methods/patients. LUV-TRAIL-induced cytotoxicity was assessed on different NSCLC cell lines with different sensitivity to soluble TRAIL and on primary human tumor cells from three patients suffering from NSCLC cancer. We also tested LUV-TRAIL-cytotoxic ability in combination with several anti-tumor agents. Results. LUV-TRAIL exhibited a greater cytotoxic effect compared to soluble TRAIL both in A549 cells and primary human NSCLC cells. LUV-TRAIL-induced cell death was dependent on caspase-8 and caspase-3 activation. Moreover, combination of LUV-TRAIL with other anti-tumor agents such as flavopiridol, and SNS-032 clearly enhanced LUV-TRAIL-induced cytotoxicity against NSCLC cancer cells. Conclusion. The novel formulation of TRAIL based on displaying it on the surface of lipid nanoparticles greatly increases its anti-tumor activity and has clinical potential in cancer treatment.

  7. TRAIL-coated lipid-nanoparticles overcome resistance to soluble recombinant TRAIL in non-small cell lung cancer cells

    International Nuclear Information System (INIS)

    De Miguel, Diego; Gallego-Lleyda, Ana; Erviti-Ardanaz, Sandra; Anel, Alberto; Martinez-Lostao, Luis; Ayuso, José María; Fernández, Luis José; Ochoa, Ignacio; Pazo-Cid, Roberto; Del Agua, Celia

    2016-01-01

    Purpose. Non-small cell lung cancer (NSCLC) is one the types of cancer with higher prevalence and mortality. Apo2-Ligand/TRAIL is a TNF family member able to induce apoptosis in tumor cells but not in normal cells. It has been tested in clinical trials against different types of human cancer including NSCLC. However, results of clinical trials have shown a limited efficacy of TRAIL-based therapies. Recently we have demonstrated that artificial lipid nanoparticles coated with bioactive Apo2L/TRAIL (LUV-TRAIL) greatly improved TRAIL cytotoxic ability being capable of killing chemoresistant hematological cancer cells. In the present work we have extended the study to NSCLC. Methods/patients. LUV-TRAIL-induced cytotoxicity was assessed on different NSCLC cell lines with different sensitivity to soluble TRAIL and on primary human tumor cells from three patients suffering from NSCLC cancer. We also tested LUV-TRAIL-cytotoxic ability in combination with several anti-tumor agents. Results. LUV-TRAIL exhibited a greater cytotoxic effect compared to soluble TRAIL both in A549 cells and primary human NSCLC cells. LUV-TRAIL-induced cell death was dependent on caspase-8 and caspase-3 activation. Moreover, combination of LUV-TRAIL with other anti-tumor agents such as flavopiridol, and SNS-032 clearly enhanced LUV-TRAIL-induced cytotoxicity against NSCLC cancer cells. Conclusion. The novel formulation of TRAIL based on displaying it on the surface of lipid nanoparticles greatly increases its anti-tumor activity and has clinical potential in cancer treatment. (paper)

  8. Posttranslational nitrotyrosination of alpha-tubulin induces cell cycle arrest and inhibits proliferation of vascular smooth muscle cells

    Czech Academy of Sciences Publication Activity Database

    Phung, A.D.; Souček, Karel; Kubala, Lukáš; Harper, R.W.; Bulinski, J.Ch.; Eiserich, J.P.

    2006-01-01

    Roč. 85, č. 12 (2006), s. 1241-1252 ISSN 0171-9335 Institutional research plan: CEZ:AV0Z50040507 Keywords : vascular smooth muscle cells * microtubules * tubulin tyrosine ligase Subject RIV: BO - Biophysics Impact factor: 3.039, year: 2006

  9. A spleen tyrosine kinase inhibitor attenuates the proliferation and migration of vascular smooth muscle cells

    Directory of Open Access Journals (Sweden)

    Hyang‑Hee Seo

    Full Text Available Abstract Background Pathologic vascular smooth muscle cell (VSMC proliferation and migration after vascular injury promotes the development of occlusive vascular disease. Therefore, an effective chemical agent to suppress aberrant proliferation and migration of VSMCs can be a potential therapeutic modality for occlusive vascular disease such as atherosclerosis and restenosis. To find an anti-proliferative chemical agent for VSMCs, we screened an in-house small molecule library, and the selected small molecule was further validated for its anti-proliferative effect on VSMCs using multiple approaches, such as cell proliferation assays, wound healing assays, transwell migration assays, and ex vivo aortic ring assay. Results Among 43 initially screened small molecule inhibitors of kinases that have no known anti-proliferative effect on VSMCs, a spleen tyrosine kinase (Syk inhibitor (BAY61-3606 showed significant anti-proliferative effect on VSMCs. Further experiments indicated that BAY61 attenuated the VSMC proliferation in both concentration- and time-dependent manner, and it also significantly suppressed the migration of VSMCs as assessed by both wound healing assays and transwell assays. Additionally, BAY61 suppressed the sprouting of VSMCs from endothelium-removed aortic rings. Conclusion The present study identified a Syk kinase inhibitor as a potent VSMC proliferation and migration inhibitor and warrants further studies to elucidate its underlying molecular mechanisms, such as its primary target, and to validate its in vivo efficacy as a therapeutic agent for restenosis and atherosclerosis.

  10. Vascular endothelial growth factor 165-transfected adipose-derived mesenchymal stem cells promote vascularization-assisted fat transplantation.

    Science.gov (United States)

    Jun-Jiang, Chen; Huan-Jiu, Xi

    2016-06-01

    To investigate the effect of vascular endothelial growth factor 165 (VEGF165) and adipose-derived mesenchymal stem cells (ASCs) in promoting the survival of fat grafts, and to provide new methods and theoretical evidence for increasing the survival rate of autologous fat particle grafts. The VEGF165 gene was recombined with the target fragment, and the recombinant gene was introduced into adenovirus pAdEasy-1 system; the virus was then packaged and the titer was detected. The control group received the same processing. ASCs were cultured and subcultured, and then identified with immunohistochemistry and adipogenic differentiation assay. The subsequent experiments were performed in three groups: the VEGF165 gene-virus group, blank virus group, and control group. After the viral solution was transfected into the ASCs, the viral transfection efficiency was detected using a tracing factor, EGFP. The expression of VEGF165 mRNA and protein in the transfected cells were determined. The proliferation of ASCs in each group was detected with the MTT assay. (1) Recombinant adenoviral vector was constructed successfully in the two groups and the packaging was identified. The viral titer was 2.0 × 10(8) pfu/ml and 1.9 × 10(8) pfu/ml, which was in line with the requirements of the subsequent transfection experiments. (2) Immunohistochemistry and adipogenic differentiation results showed that the culture of ASCs was successful, and the cultured cells could serve as seed cells in this experiment. (3) The RT-PCR analysis showed that the relative optical density of VEGF165 mRNA expression was 0.76 ± 0.05 in the experimental group, and there were statistically significant differences compared with the values obtained for the other two groups (P < 0.05). (4) The western blot analysis showed that the relative optical density of VEGF165 protein expression in the experimental group was significantly higher than that in the other two groups (P < 0.05). (5) The proliferation of ASCs was

  11. The role of perivascular adipose tissue in vascular smooth muscle cell growth.

    Science.gov (United States)

    Miao, Chao-Yu; Li, Zhi-Yong

    2012-02-01

    Adipose tissue is the largest endocrine organ, producing various adipokines and many other substances. Almost all blood vessels are surrounded by perivascular adipose tissue (PVAT), which has not received research attention until recently. This review will discuss the paracrine actions of PVAT on the growth of underlying vascular smooth muscle cells (VSMCs). PVAT can release growth factors and inhibitors. Visfatin is the first identified growth factor derived from PVAT. Decreased adiponectin and increased tumour necrosis factor-α in PVAT play a pathological role for neointimal hyperplasia after endovascular injury. PVAT-derived angiotensin II, angiotensin 1-7, reactive oxygen species, complement component 3, NO and H(2) S have a paracrine action on VSMC contraction, endothelial or fibroblast function; however, their paracrine actions on VSMC growth remain to be directly verified. Factors such as monocyte chemoattractant protein-1, interleukin-6, interleukin-8, leptin, resistin, plasminogen activator inhibitor type-1, adrenomedullin, free fatty acids, glucocorticoids and sex hormones can be released from adipose tissue and can regulate VSMC growth. Most of them have been verified for their secretion by PVAT; however, their paracrine functions are unknown. Obesity, vascular injury, aging and infection may affect PVAT, causing adipocyte abnormality and inflammatory cell infiltration, inducing imbalance of PVAT-derived growth factors and inhibitors, leading to VSMC growth and finally resulting in development of proliferative vascular disease, including atherosclerosis, restenosis and hypertension. In the future, using cell-specific gene interventions and local treatments may provide definitive evidence for identification of key factor(s) involved in PVAT dysfunction-induced vascular disease and thus may help to develop new therapies. This article is part of a themed section on Fat and Vascular Responsiveness. To view the other articles in this section visit http

  12. 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.

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

    International Nuclear Information System (INIS)

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

    2013-01-01

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

  14. Bim Regulates Alloimmune-Mediated Vascular Injury Through Effects on T Cell Activation and Death

    Science.gov (United States)

    von Rossum, Anna; Enns, Winnie; Shi, Yu P.; MacEwan, Grace E.; Malekesmaeli, Mehrnoush; Brinkman, Ryan; Choy, Jonathan C.

    2014-01-01

    Objective Bim is a pro-apoptotic Bcl-2 protein known to down-regulate immune responses and to also be required for antigen-induced T cell activation. However, it is not known how the effect of Bim on these offsetting processes determines the outcome of allogeneic immune responses. We have defined the role of Bim in regulating alloantigen-driven T cell responses in a model of vascular rejection. Approach and Results Bim was required for proliferation of CD4 and CD8 T cells, and for IL-2 production, in T cells stimulated with alloantigen in vitro. Moreover, a partial reduction in Bim expression was sufficient to attenuate T cell activation whereas a complete elimination of Bim was required to prevent CD4 T cell death in response to cytokine withdrawl. When alloimmune-mediated vascular rejection was examined using an aortic interposition model, there was significantly less intimal thickening in Bim+/−, but not Bim−/−, graft recipients. T cell proliferation in response to allograft arteries was significantly reduced in both Bim+/− and Bim−/− mice, but cell death was attenuated only in Bim−/− animals. Conclusions Bim controls both T cell activation and death in response to alloantigen stimulation. These processes act cooperatively to determine the outcome of immune responses in allograft arteries. PMID:24700126

  15. Bim regulates alloimmune-mediated vascular injury through effects on T-cell activation and death.

    Science.gov (United States)

    von Rossum, Anna; Enns, Winnie; Shi, Yu P; MacEwan, Grace E; Malekesmaeli, Mehrnoush; Brinkman, Ryan; Choy, Jonathan C

    2014-06-01

    Bim is a proapoptotic Bcl-2 protein known to downregulate immune responses and to also be required for antigen-induced T-cell activation. However, it is not known how the effect of Bim on these offsetting processes determines the outcome of allogeneic immune responses. We have defined the role of Bim in regulating alloantigen-driven T-cell responses in a model of vascular rejection. Bim was required for proliferation of CD4 and CD8 T cells, and for interleukin-2 production, in T cells stimulated with alloantigen in vitro. Moreover, a partial reduction in Bim expression was sufficient to attenuate T-cell activation, whereas a complete elimination of Bim was required to prevent CD4 T-cell death in response to cytokine withdrawl. When alloimmune-mediated vascular rejection was examined using an aortic interposition model, there was significantly less intimal thickening in Bim(+/-), but not Bim(-/-), graft recipients. T-cell proliferation in response to allograft arteries was significantly reduced in both Bim(+/-) and Bim(-/-) mice, but cell death was attenuated only in Bim(-/-) animals. Bim controls both T-cell activation and death in response to alloantigen stimulation. These processes act cooperatively to determine the outcome of immune responses in allograft arteries. © 2014 American Heart Association, Inc.

  16. Unique responses of stem cell-derived vascular endothelial and mesenchymal cells to high levels of glucose.

    Directory of Open Access Journals (Sweden)

    Emily Keats

    Full Text Available Diabetes leads to complications in selected organ systems, and vascular endothelial cell (EC dysfunction and loss is the key initiating and perpetuating step in the development of these complications. Experimental and clinical studies have shown that hyperglycemia leads to EC dysfunction in diabetes. Vascular stem cells that give rise to endothelial progenitor cells (EPCs and mesenchymal progenitor cells (MPCs represent an attractive target for cell therapy for diabetic patients. Whether these vascular stem/progenitor cells succumb to the adverse effects of high glucose remains unknown. We sought to determine whether adult vascular stem/progenitor cells display cellular activation and dysfunction upon exposure to high levels of glucose as seen in diabetic complications. Mononuclear cell fraction was prepared from adult blood and bone marrow. EPCs and MPCs were derived, characterized, and exposed to either normal glucose (5 mmol/L or high glucose levels (25 mmol/L. We then assayed for cell activity and molecular changes following both acute and chronic exposure to high glucose. Our results show that high levels of glucose do not alter the derivation of either EPCs or MPCs. The adult blood-derived EPCs were also resistant to the effects of glucose in terms of growth. Acute exposure to high glucose levels increased caspase-3 activity in EPCs (1.4x increase and mature ECs (2.3x increase. Interestingly, MPCs showed a transient reduction in growth upon glucose challenge. Our results also show that glucose skews the differentiation of MPCs towards the adipocyte lineage while suppressing other mesenchymal lineages. In summary, our studies show that EPCs are resistant to the effects of high levels of glucose, even following chronic exposure. The findings further show that hyperglycemia may have detrimental effects on the MPCs, causing reduced growth and altering the differentiation potential.

  17. Human adipose-derived stem cells promote vascularization of collagen-based scaffolds transplanted into nude mice.

    Science.gov (United States)

    Cherubino, Mario; Valdatta, Luigi; Balzaretti, Riccardo; Pellegatta, Igor; Rossi, Federica; Protasoni, Marina; Tedeschi, Alessandra; Accolla, Roberto S; Bernardini, Giovanni; Gornati, Rosalba

    2016-04-01

    After in vivo implantation of cell-loaded devices, only the cells close to the capillaries can obtain nutrients to maintain their functions. It is known that factors secreted by stem cells, rather than stem cells themselves, are fundamental to guarantee new vascularization in the area of implant. To investigate this possibility, we have grafted mice with Bilayer and Flowable Integra(®) scaffolds, loaded or not with human adipose-derived stem cells. Our results support the therapeutic potential of human adipose-derived stem cells to induce new vascular networks of engineered organs and tissues. This finding suggests that our approach can help to form new vascular networks that allow sufficient vascularization of engineered organs and tissues in cases of difficult wound healing due to ischemic conditions.

  18. Photosensitizing activity of water- and lipid-soluble phthalocyanines on prokaryotic and eukaryotic microbial cells.

    Science.gov (United States)

    Bertoloni, G; Rossi, F; Valduga, G; Jori, G; Ali, H; van Lier, J E

    1992-01-01

    The photosensitizing activity of lipophilic zinc-phthalocyanine (Zn-Pc) and its water-soluble sulphonated derivative (Zn-PcS) towards Streptococcus faecium and Candida albicans was studied and correlated with the amount of cell-bound photosensitizer. With both micro-organisms Zn-PcS was more tightly bound in larger amounts than Zn-Pc in the protoplasts of the cytoplasmic membrane. As a consequence, the photoinduced damage in S. faecium initially involved membrane proteins, while DNA was modified only upon prolonged irradiation. For C. albicans only Zn-PcS showed a preferential affinity for the spheroplasts and the decrease in cell survival was not accompanied by detectable modifications of the electrophoretic pattern of membrane proteins. The photoinduced ultrastructural alteration of both micro-organisms suggests damage at membrane level. This would indicate the involvement of different targets in bacteria and yeast for phthalocyanine photosensitization.

  19. In Vitro Endothelialization of Biodegradable Vascular Grafts Via Endothelial Progenitor Cell Seeding and Maturation in a Tubular Perfusion System Bioreactor

    OpenAIRE

    Melchiorri, Anthony J.; Bracaglia, Laura G.; Kimerer, Lucas K.; Hibino, Narutoshi; Fisher, John P.

    2016-01-01

    A critical challenge to the success of biodegradable vascular grafts is the establishment of a healthy endothelium. To establish this monolayer of endothelial cells (ECs), a variety of techniques have been developed, including cell seeding. Vascular grafts may be seeded with relevant cell types and allowed to mature before implantation. Due to the low proliferative ability of adult ECs and issues with donor site morbidity, there has been increasing interest in using endothelial progenitor cel...

  20. Identification of genes associated with the effect of inflammation on the neurotransmission of vascular smooth muscle cell

    OpenAIRE

    Gan, Shujie; Qiu, Shenlong; Feng, Yiwen; Zhang, Yanping; Qian, Qin; Wan, Zhong; Tang, Jingdong

    2017-01-01

    Vascular smooth muscle cell (VSMC) accumulation and hypertrophy are common in vascular disorders, and inflammation has a crucial role in the development of these diseases. To investigate the effect of inflammation on the neurotransmission of VSMC, bioinformatic analysis was performed, following next generation sequencing. Genes of lipopolysaccharide (LPS)-treated A7r5 cells and phosphate-buffered saline (PBS)-treated A7r5 cells were sequenced via next generation sequencing, and each assay was...

  1. Improvement of endothelial progenitor outgrowth cell (EPOC)-mediated vascularization in gelatin-based hydrogels through pore size manipulation.

    Science.gov (United States)

    Fu, Jiayin; Wiraja, Christian; Muhammad, Hamizan B; Xu, Chenjie; Wang, Dong-An

    2017-08-01

    In addition to chemical compositions, physical properties of scaffolds, such as pore size, can also influence vascularization within the scaffolds. A larger pore has been shown to improve host vascular tissue invasion into scaffolds. However, the influence of pore sizes on vascularization by endothelial cells directly encapsulated in hydrogels remains unknown. In this study, micro-cavitary hydrogels with different pore sizes were created in gelatin-methacrylate hydrogels with dissolvable gelatin microspheres (MS) varying in sizes. The effect of pore sizes on vascular network formation by endothelial progenitor outgrowth cells (EPOCs) encapsulated in hydrogels was then investigated both in vitro and in vivo. When cultured in vitro, vascular networks were formed around pore structures in micro-cavitary hydrogels. The middle pore size supported best differentiation of EPOCs and thus best hydrogel vascularization in vitro. When implantation in vivo, functional connections between encapsulated EPOCs and host vasculature micro-cavitary hydrogels were established. Vascularization in vivo was promoted best in hydrogels with the large pore size due to the increased vascular tissue invasion. These results highlight the difference between in vitro and in vivo culture conditions and indicate that pore sizes shall be designed for in vitro and in vivo hydrogel vascularization respectively. Pore sizes for hydrogel vascularization in vitro shall be middle ones and pore sizes for hydrogel vascularization in vivo shall be large ones. This study reveals that the optimal pore size for hydrogel vascularization in vitro and in vivo is different. The middle pore size supported best differentiation of EPOCs and thus best hydrogel vascularization in vitro, while vascularization in vivo was promoted best in hydrogels with the large pore size due to the increased vascular tissue invasion. These results highlight the difference between in vitro and in vivo culture conditions and indicate that

  2. Low-dose effects of Bisphenol A on human primary vascular endothelial cells and colon cancer cells

    OpenAIRE

    Varandas, Edna Soraia Gregório Ribeiro Varandas

    2014-01-01

    Doutoramento em Biologia - Instituto Superior de Agronomia Bisphenol A (BPA) is an extensively utilized endocrine disruptor for which human exposure is considered generalized through ingestion. Information regarding BPA effects on vascular and digestive tract tissues is scarce. Therefore, in this work primary Human Umbilical Vein Endothelial Cells (HUVEC) and human colon adenocarcinona cell line HT29 were used to evaluate BPA effects at two distinct low-dose concentrations r...

  3. Single-cell profiling reveals heterogeneity and functional patterning of GPCR expression in the vascular system.

    Science.gov (United States)

    Kaur, H; Carvalho, J; Looso, M; Singh, P; Chennupati, R; Preussner, J; Günther, S; Albarrán-Juárez, J; Tischner, D; Classen, S; Offermanns, S; Wettschureck, N

    2017-06-16

    G-protein-coupled receptor (GPCR) expression is extensively studied in bulk cDNA, but heterogeneity and functional patterning of GPCR expression in individual vascular cells is poorly understood. Here, we perform a microfluidic-based single-cell GPCR expression analysis in primary smooth muscle cells (SMC) and endothelial cells (EC). GPCR expression is highly heterogeneous in all cell types, which is confirmed in reporter mice, on the protein level and in human cells. Inflammatory activation in murine models of sepsis or atherosclerosis results in characteristic changes in the GPCR repertoire, and we identify functionally relevant subgroups of cells that are characterized by specific GPCR patterns. We further show that dedifferentiating SMC upregulate GPCRs such as Gpr39, Gprc5b, Gprc5c or Gpr124, and that selective targeting of Gprc5b modulates their differentiation state. Taken together, single-cell profiling identifies receptors expressed on pathologically relevant subpopulations and provides a basis for the development of new therapeutic strategies in vascular diseases.

  4. Recent advances in understanding the roles of vascular endothelial cells in allergic inflammation.

    Science.gov (United States)

    Shoda, Tetsuo; Futamura, Kyoko; Orihara, Kanami; Emi-Sugie, Maiko; Saito, Hirohisa; Matsumoto, Kenji; Matsuda, Akio

    2016-01-01

    Allergic disorders commonly involve both chronic tissue inflammation and remodeling caused by immunological reactions to various antigens on tissue surfaces. Due to their anatomical location, vascular endothelial cells are the final responders to interact with various exogenous factors that come into contact with the epithelial surface, such as pathogen-associated molecular patterns (PAMPs) and antigens. Recent studies have shed light on the important roles of endothelial cells in the development and exacerbation of allergic disorders. For instance, endothelial cells have the greatest potential to produce several key molecules that are deeply involved in allergic inflammation, such as periostin and thymus and activation-regulated chemokine (TARC/CCL17). Additionally, endothelial cells were recently shown to be important functional targets for IL-33--an essential regulator of allergic inflammation. Notably, almost all endothelial cell responses and functions involved in allergic inflammation are not suppressed by corticosteroids. These corticosteroid-refractory endothelial cell responses and functions include TNF-α-associated angiogenesis, leukocyte adhesion, IL-33-mediated responses and periostin and TARC production. Therefore, these unique responses and functions of endothelial cells may be critically involved in the pathogenesis of various allergic disorders, especially their refractory processes. Here, we review recent studies, including ours, which have elucidated previously unknown pathophysiological roles of vascular endothelial cells in allergic inflammation and discuss the possibility of endothelium-targeted therapy for allergic disorders. Copyright © 2015 The Authors. Production and hosting by Elsevier B.V. All rights reserved.

  5. Changes in adipose tissue stromal-vascular cells in primary culture due to porcine sera

    International Nuclear Information System (INIS)

    Jewell, D.E.; Hausman, G.J.

    1986-01-01

    This study was conducted to determine the response of rat stromal-vascular cells to pig sea. Sera were collected from unselected contemporary (lean) and high backfat thickness selected (obese) pigs. Sera from obese pigs were collected either by exsanguination or cannulation. sera from lean pigs during the growing phase (45 kg) and the fattening phase (100-110 kg) were collected. Stromal-vascular cells derived rom rat inguinal tissue were cultured on either 25 cm 2 flasks, collagen-coated coverslips or petri dishes. Cell proliferation was measured by [ 3 H]-thymidine incorporation during the fourth day of culture. Coverslip cultures were used for histochemical analysis. Petri dish cultures were used for analysis of Sn-glycerol-3-phosphate dehydrogenase (GPDH) activity. All cells were plated for 24 hours in media containing 10 fetal bovine sera. Test media contained 2.5, 5.0, 10.0% sera. Sera from obese pigs increased GPDH activity and fat cell production when compared to the lean controls. The increased concentration of sera increased esterase activity and lipid as measured with oil red O. The sera from obese pigs collected at slaughter stimulated more fat cell production than obese sera collected by cannulation. These studies show there are adipogenic factors in obese pigs sera which promote fat cell development in primary cell culture

  6. Alkali-soluble pectin is the primary target of Aluminum immobilization in root border cells of pea (Pisum sativum

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    Mei Qu

    2016-09-01

    Full Text Available We investigated the hypothesis that a discrepancy of Al binding in cell wall constituents determines Al mobility in root border cells (RBCs of pea (Pisum sativum, which provides protection for RBCs and root apices under Al toxicity. Plants of pea (Pisum sativum L. ‘Zhongwan no. 6’ were subjected to Al treatments under mist culture. The concentration of Al in RBCs was much higher than that in the root apex. The Al content in RBCs surrounding one root apex (104 RBCs was approximately 24.5 % of the total Al in the root apex (0–2.5 mm, indicating a shielding role of RBCs for the root apex under Al toxicity. Cell wall analysis showed that Al accumulated predominantly in alkali-soluble pectin (pectin 2 of RBCs. This could be attributed to a significant increase of uronic acids under Al toxicity, higher capacity of Al adsorption in alkali-soluble pectin (5.3-fold higher than that of chelate-soluble pectin, and lower ratio of Al desorption from alkali-soluble pectin (8.5% compared with chelate-soluble pectin (68.5%. These results indicate that alkali-soluble pectin is the primary target of Al immobilization in RBCs of pea, which impairs Al access to the intracellular space of RBCs and mobility to root apices, and therefore protects root apices and RBCs from Al toxicity.

  7. The irradiation effects on the cytoskeletons of C3H/He mouse mammary tumor cells and vascular basement membrane in relation to vascular invasion

    International Nuclear Information System (INIS)

    Kobayashi, Michiya

    1988-01-01

    We investigated the short-term effects of a single high-dose radiation upon transplanted MM46 tumor cells in mice by means of immunohistochemistry and electron microscopy. The irradiation induced: 1) giant cell formation from the 3rd day, 2) arrest of tumor cell mitosis in prophase and metaphase due to the disorganization of the mitotic spindles, 3) changes in immunoreactivity of laminin and cytoskeletons, and 4) multilayering of the vascular basal lamina and perivascular fibrosis. The above findings suggest a decrease in tumor cell compliance, growth and invasiveness and the potentiation of defensive host responses against vascular invasion after irradiation. The analysis of the temporal sequences of the events indicates that the time lapse between the optimal host response, tumor growth and invasion constitutes a critical period. (author)

  8. Nuclear IL-33 regulates soluble ST2 receptor and IL-6 expression in primary human arterial endothelial cells and is decreased in idiopathic pulmonary arterial hypertension

    Energy Technology Data Exchange (ETDEWEB)

    Shao, Dongmin [Section of Vascular Biology, National Heart and Lung Institute, Imperial College London, London (United Kingdom); Perros, Frédéric [Faculté de Médecine, Université Paris-Sud, Paris, Clamart (France); Caramori, Gaetano [Dipartimento di Scienze Mediche, Sezione di Medicina Interna e Cardiorespiratoria, Centro Interdipartimentale per lo Studio delle Malattie Infiammatorie delle Vie Aeree e Patologie Fumo-Correlate, University of Ferrara, Ferrara (Italy); Meng, Chao [Section of Vascular Biology, National Heart and Lung Institute, Imperial College London, London (United Kingdom); Department of Geriatrics, Renji Hospital, School of Medicine, Shanghai Jiaotong University, Shanghai (China); Dormuller, Peter [Faculté de Médecine, Université Paris-Sud, Paris, Clamart (France); Chou, Pai-Chien [Airways Disease, National Heart and Lung Institute (United Kingdom); Church, Colin [Scottish Pulmonary Vascular Unit, University of Glasgow (United Kingdom); Papi, Alberto; Casolari, Paolo [Dipartimento di Scienze Mediche, Sezione di Medicina Interna e Cardiorespiratoria, Centro Interdipartimentale per lo Studio delle Malattie Infiammatorie delle Vie Aeree e Patologie Fumo-Correlate, University of Ferrara, Ferrara (Italy); Welsh, David; Peacock, Andrew [Scottish Pulmonary Vascular Unit, University of Glasgow (United Kingdom); Humbert, Marc [Faculté de Médecine, Université Paris-Sud, Paris, Clamart (France); Adcock, Ian M. [Airways Disease, National Heart and Lung Institute (United Kingdom); Wort, Stephen J., E-mail: s.wort@imperial.ac.uk [Section of Vascular Biology, National Heart and Lung Institute, Imperial College London, London (United Kingdom)

    2014-08-15

    Highlights: • Nuclear IL-33 expression is reduced in vascular endothelial cells from PAH patients. • Knockdown of IL-33 leads to increased IL-6 and sST2 mRNA expression. • IL-33 binds homeobox motifs in target gene promoters and recruits repressor proteins. - Abstract: Idiopathic pulmonary arterial hypertension (IPAH) is an incurable condition leading to right ventricular failure and death and inflammation is postulated to be associated with vascular remodelling. Interleukin (IL)-33, a member of the “alarmin” family can either act on the membrane ST2 receptor or as a nuclear repressor, to regulate inflammation. We show, using immunohistochemistry, that IL-33 expression is nuclear in the vessels of healthy subjects whereas nuclear IL-33 is markedly diminished in the vessels of IPAH patients. This correlates with reduced IL-33 mRNA expression in their lung. In contrast, serum levels of IL-33 are unchanged in IPAH. However, the expression of the soluble form of ST2, sST2, is enhanced in the serum of IPAH patients. Knock-down of IL-33 in human endothelial cells (ECs) using siRNA is associated with selective modulation of inflammatory genes involved in vascular remodelling including IL-6. Additionally, IL-33 knock-down significantly increased sST2 release from ECs. Chromatin immunoprecipitation demonstrated that IL-33 bound multiple putative homeodomain protein binding motifs in the proximal and distal promoters of ST2 genes. IL-33 formed a complex with the histone methyltransferase SUV39H1, a transcriptional repressor. In conclusion, IL-33 regulates the expression of IL-6 and sST2, an endogenous IL-33 inhibitor, in primary human ECs and may play an important role in the pathogenesis of PAH through recruitment of transcriptional repressor proteins.

  9. Combined strategy of endothelial cells coating, Sertoli cells coculture and infusion improves vascularization and rejection protection of islet graft.

    Directory of Open Access Journals (Sweden)

    Yang Li

    Full Text Available Improving islet graft revascularization and inhibiting rejection become crucial tasks for prolonging islet graft survival. Endothelial cells (ECs are the basis of islet vascularization and Sertoli cells (SCs have the talent to provide nutritional support and exert immunosuppressive effects. We construct a combined strategy of ECs coating in the presence of nutritious and immune factors supplied by SCs in a co-culture system to investigate the effect of vascularization and rejection inhibition for islet graft. In vivo, the combined strategy improved the survival and vascularization as well as inhibited lymphocytes and inflammatory cytokines. In vitro, we found the combinatorial strategy improved the function of islets and the effect of ECs-coating on islets. Combined strategy treated islets revealed higher levels of anti-apoptotic signal molecules (Bcl-2 and HSP-32, survival and function related molecules (PDX-1, Ki-67, ERK1/2 and Akt and demonstrated increased vascular endothelial growth factor receptor 2 (KDR and angiogenesis signal molecules (FAk and PLC-γ. SCs effectively inhibited the activation of lymphocyte stimulated by islets and ECs. Predominantly immunosuppressive cytokines could be detected in culture supernatants of the SCs coculture group. These results suggest that ECs-coating and Sertoli cells co-culture or infusion synergistically enhance islet survival and function after transplantation.

  10. Vascular Calcification: Is it rather a Stem/Progenitor Cells Driven Phenomenon?

    Directory of Open Access Journals (Sweden)

    Aleksandra Leszczynska

    2018-02-01

    Full Text Available Vascular calcification (VC has witnessed a surge of interest. Vasculature is virtually an omnipresent organ and has a notably high capacity for repair throughout embryonic and adult life. Of the vascular diseases, atherosclerosis is a leading cause of morbidity and mortality on account of ectopic cartilage and bone formation. Despite the identification of a number of risk factors, all the current theories explaining pathogenesis of VC in atherosclerosis are far from complete. The most widely accepted response to injury theory and smooth muscle transdifferentiation to explain the VC observed in atherosclerosis is being challenged. Recent focus on circulating and resident progenitor cells in the vasculature and their role in atherogenesis and VC has been the driving force behind this review. This review discusses intrinsic cellular players contributing to fate determination of cells and tissues to form ectopic cartilage and bone formation.

  11. Vascular regulation of glioma stem-like cells: a balancing act.

    Science.gov (United States)

    Brooks, Lucy J; Parrinello, Simona

    2017-12-01

    Glioblastoma (GBM) are aggressive and therapy-resistant brain tumours driven by glioma stem-like cells (GSCs). GSC behaviour is controlled by the microenvironment, or niche, in which the cells reside. It is well-established that the vasculature is a key component of the GSC niche, which drives maintenance in the tumour bulk and invasion at the margin. Emerging evidence now indicates that the specific properties of the vasculature within these two regions impose different functional states on resident GSCs, generating distinct subpopulations. Here, we review these recent findings, focusing on the mechanisms that underlie GSC/vascular communication. We further discuss how plasticity enables GSCs to respond to vascular changes by interconverting bidirectionally between states, and address the therapeutic implications of this dynamic response. Copyright © 2017 The Authors. Published by Elsevier Ltd.. All rights reserved.

  12. Detection and Quantification of Vascular Endothelial Growth Factor Receptor Tyrosine Kinases in Primary Human Endothelial Cells.

    Science.gov (United States)

    Fearnley, Gareth W; Wheatcroft, Stephen B; Ponnambalam, Sreenivasan

    2015-01-01

    Proteins differ widely in their pattern of expression depending on organism, tissue, and regulation in response to changing conditions. In the mammalian vasculature, the endothelium responds to vascular endothelial growth factors (VEGFs) via membrane-bound receptor tyrosine kinases (VEGFRs) to modulate many aspects of vascular physiology including vasculogenesis, angiogenesis, and blood pressure. Studies on VEGFR biology are thus dependent on detecting expression levels in different cell types and evaluating how changes in protein levels correlate with changing conditions including circulating VEGF levels. Here, we present a robust immunoblot-based protocol for detecting and quantifying VEGFRs in human endothelial cells. Using internal and external standards, we can rapidly evaluate receptor copy number and assess how this is altered in response to the cellular environment.

  13. MIXED HYALINE VASCULAR AND PLASMA CELL TYPE CASTLEMAN’S DISEASE: REPORT OF A CASE

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    F. Asgarani

    2006-05-01

    Full Text Available Castleman’s disease (angiofollicular lymphoid hyperplasia includes a heterogeneous group of lymphoproliferative disorders. The cause of this disease remains uncertain. There are two types of localized Castleman’s disease: the more common hyaline vascular and the plasma cell types. Mixed variant is an uncommon localized lesion in general population. The lesions can occur in any part of the body that contains lymphoid tissue, although seventy percent are found in the anterior mediastinum. We report a thirty years old boy with Castleman’s disease who presented with fever, anorexia, weight loss,sweating, anemia and abdominal mass. The histologic examination of the biopsy specimens revealed a mixed hyaline vascular and plasma cell type of Castleman’s disease.

  14. Sulforaphane inhibits restenosis by suppressing inflammation and the proliferation of vascular smooth muscle cells.

    Science.gov (United States)

    Kwon, Jin-Sook; Joung, Hosouk; Kim, Yong Sook; Shim, Young-Sun; Ahn, Youngkeun; Jeong, Myung Ho; Kee, Hae Jin

    2012-11-01

    Sulforaphane, a naturally occurring organosulfur compound in broccoli, has chemopreventive properties in cancer. However, the effects of sulforaphane in vascular diseases have not been examined. We therefore aimed to investigate the effects of sulforaphane on vascular smooth muscle cell (VSMC) proliferation and neointimal formation and the related mechanisms. The expression of vascular cell adhesion molecule 1 (VCAM-1) and intercellular adhesion molecule 1 (ICAM-1) was examined in VSMCs. The nuclear translocation of nuclear factor-κB (NF-κB) and GATA6 expression was examined in VSMCs and in a carotid artery injury model by Western blot and immunohistochemistry. We also investigated whether local delivery of sulforaphane affected neointimal formation. Sulforaphane inhibited the mRNA and protein expression of VCAM-1 induced by tumor necrosis factor (TNF)-α in VSMCs. Treatment of VSMCs with sulforaphane blocked TNF-α-induced IκBα degradation and NF-κB p65 and GATA6 expression. Furthermore, NF-κB p65 and GATA6 expression were reduced in sulforaphane-treated carotid injury sections. Notably, binding of GATA6 to the VCAM-1 promoter was dramatically reduced by sulforaphane. The MTT, BrdU incorporation, and in vitro scratch assays revealed that the proliferation and migration of VSMCs were reduced by sulforaphane. Furthermore, local administration of sulforaphane significantly reduced neointima formation 14 days after vascular injury in rats. Our results indicate that sulforaphane inhibits neointima formation via targeting of adhesion molecules through the suppression of NF-κB/GATA6. Furthermore, sulforaphane regulates migration and proliferation in VSMCs. Sulforaphane may be a potential therapeutic agent for preventing restenosis after vascular injury. Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.

  15. Differential uptake and cross-presentation of soluble and necrotic cell antigen by human DC subsets.

    Science.gov (United States)

    Chiang, Meng-Chieh; Tullett, Kirsteen M; Lee, Yoke Seng; Idris, Adi; Ding, Yitian; McDonald, Kylie J; Kassianos, Andrew; Leal Rojas, Ingrid M; Jeet, Varinder; Lahoud, Mireille H; Radford, Kristen J

    2016-02-01

    Cross-presentation is the mechanism by which exogenous Ag is processed for recognition by CD8(+) T cells. Murine CD8α(+) DCs are specialized at cross-presenting soluble and cellular Ag, but in humans this process is poorly characterized. In this study, we examined uptake and cross-presentation of soluble and cellular Ag by human blood CD141(+) DCs, the human equivalent of mouse CD8α(+) DCs, and compared them with human monocyte-derived DCs (MoDCs) and blood CD1c(+) DC subsets. MoDCs were superior in their capacity to internalize and cross-present soluble protein whereas CD141(+) DCs were more efficient at ingesting and cross-presenting cellular Ag. Whilst cross-presentation by CD1c(+) DCs and CD141(+) DCs was dependent on the proteasome, and hence cytosolic translocation, cross-presentation by MoDCs was not. Inhibition of endosomal acidification enhanced cross-presentation by CD1c(+) DCs and MoDCs but not by CD141(+) DCs. These data demonstrate that CD1c(+) DCs, CD141(+) DCs, and MoDCs are capable of cross-presentation; however, they do so via different mechanisms. Moreover, they demonstrate that human CD141(+) DCs, like their murine CD8α(+) DC counterparts, are specialized at cross-presenting cellular Ag, most likely mediated by an enhanced capacity to ingest cellular Ag combined with subtle changes in lysosomal pH during Ag processing and use of the cytosolic pathway. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  16. Soluble histone H2AX is induced by DNA replication stress and sensitizes cells to undergo apoptosis

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    Duensing Stefan

    2008-07-01

    Full Text Available Abstract Background Chromatin-associated histone H2AX is a key regulator of the cellular responses to DNA damage. However, non-nucleosomal functions of histone H2AX are poorly characterized. We have recently shown that soluble H2AX can trigger apoptosis but the mechanisms leading to non-chromatin-associated H2AX are unclear. Here, we tested whether stalling of DNA replication, a common event in cancer cells and the underlying mechanism of various chemotherapeutic agents, can trigger increased soluble H2AX. Results Transient overexpression of H2AX was found to lead to a detectable fraction of soluble H2AX and was associated with increased apoptosis. This effect was enhanced by the induction of DNA replication stress using the DNA polymerase α inhibitor aphidicolin. Cells manipulated to stably express H2AX did not contain soluble H2AX, however, short-term treatment with aphidicolin (1 h resulted in detectable amounts of H2AX in the soluble nuclear fraction and enhanced apoptosis. Similarly, soluble endogenous H2AX was detected under these conditions. We found that excessive soluble H2AX causes chromatin aggregation and inhibition of ongoing gene transcription as evidenced by the redistribution and/or loss of active RNA polymerase II as well as the transcriptional co-activators CBP and p300. Conclusion Taken together, these results show that DNA replication stress rapidly leads to increased soluble H2AX and that non-chromatin-associated H2AX can sensitize cells to undergo apoptosis. Our findings encourage further studies to explore H2AX and the cellular pathways that control its expression as anti-cancer drug targets.

  17. Cloning of Soluble Human Stem Cell Factor in pET-26b(+ Vector

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    Salman Asghari

    2014-03-01

    Full Text Available Purpose: Stem cell factor (SCF plays an important role in the survival, proliferation and differentiation of hematopoietic stem cells and progenitor cells. Potential therapeutic applications of SCF include hematopoietic stem cell mobilization, exvivo stem/progenitor cell expansion, gene therapy, and immunotherapy. Considering the cost and problem in accessibility of this product in Iran, clears the importance of indigenizing production of rhSCF. In the present work, we describe the construction of the soluble rhSCF expression vector in pET-26b (+ with periplasmic localization potential. Methods: Following PCR amplification of human SCF ORF, it is cloned in pET-26b (+ vector in NcoI and XhoI sites. The recombinant construct was transformed into BL21 (DE3 Ecoli strains. Results: The construction of recombinant vector was verified by colony PCR and sequence analysis of pET26b-hSCF vector. Sequence analyses proved that human SCF ORF has been inserted into NcoI and XhoI site with correct orientation downstream of strong T7 promotor and showed no nucleotide errors. Conclusion: The SCF ORF was successfully cloned in pET-26b (+ expression vector and is ready for future production of SCF protein.

  18. Cloning of Soluble Human Stem Cell Factor in pET-26b(+) Vector.

    Science.gov (United States)

    Asghari, Salman; Shekari Khaniani, Mahmoud; Darabi, Masood; Mansoori Derakhshan, Sima

    2014-01-01

    Stem cell factor (SCF) plays an important role in the survival, proliferation and differentiation of hematopoietic stem cells and progenitor cells. Potential therapeutic applications of SCF include hematopoietic stem cell mobilization, exvivo stem/progenitor cell expansion, gene therapy, and immunotherapy. Considering the cost and problem in accessibility of this product in Iran, clears the importance of indigenizing production of rhSCF. In the present work, we describe the construction of the soluble rhSCF expression vector in pET-26b (+) with periplasmic localization potential. Following PCR amplification of human SCF ORF, it is cloned in pET-26b (+) vector in NcoI and XhoI sites. The recombinant construct was transformed into BL21 (DE3) Ecoli strains. The construction of recombinant vector was verified by colony PCR and sequence analysis of pET26b-hSCF vector. Sequence analyses proved that human SCF ORF has been inserted into NcoI and XhoI site with correct orientation downstream of strong T7 promotor and showed no nucleotide errors. The SCF ORF was successfully cloned in pET-26b (+) expression vector and is ready for future production of SCF protein.

  19. 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.

  20. Prominent Vascularization Capacity of Mesenchymal Stem Cells in Collagen-Gold Nanocomposites.

    Science.gov (United States)

    Hsieh, Shu-Chen; Chen, Hui-Jye; Hsu, Shan-Hui; Yang, Yi-Chin; Tang, Cheng-Ming; Chu, Mei-Yun; Lin, Pei-Ying; Fu, Ru-Huei; Kung, Mei-Lang; Chen, Yun-Wen; Yeh, Bi-Wen; Hung, Huey-Shan

    2016-10-26

    The ideal characteristics of surface modification on the vascular graft for clinical application would be with excellent hemocompatibility, endothelialization capacity, and antirestenosis ability. Here, Fourier transform infrared spectroscopy (FTIR), surface enhanced Raman spectroscopy (SERS), atomic force microscopy (AFM), contact angle (θ) measurement, and thermogravimetric analysis (TGA) were used to evaluate the chemical and mechanical properties of collagen-gold nanocomposites (collagen+Au) with 17.4, 43.5, and 174 ppm of Au and suggested that the collagen+Au with 43.5 ppm of Au had better biomechanical properties and thermal stability than pure collagen. Besides, stromal-derived factor-1α (SDF-1α) at 50 ng/mL promoted the migration of mesenchymal stem cells (MSCs) on collagen+Au material through the α5β3 integrin/endothelial oxide synthase (eNOS)/metalloproteinase (MMP) signaling pathway which can be abolished by the knockdown of vascular endothelial growth factor (VEGF). The potentiality of collagen+Au with MSCs for vascular regeneration was evaluated by our in vivo rat model system. Artery tissues isolated from an implanted collagen+Au-coated catheter with MSCs expressed substantial CD-31 and α-SMA, displayed higher antifibrotic ability, antithrombotic activity, as well as anti-inflammatory response than all other materials. Our results indicated that the implantation of collagen+Au-coated catheters with MSCs could be a promising strategy for vascular regeneration.

  1. Vascular endothelial growth factor A-stimulated signaling from endosomes in primary endothelial cells.

    Science.gov (United States)

    Fearnley, Gareth W; Smith, Gina A; Odell, Adam F; Latham, Antony M; Wheatcroft, Stephen B; Harrison, Michael A; Tomlinson, Darren C; Ponnambalam, Sreenivasan

    2014-01-01

    The vascular endothelial growth factor A (VEGF-A) is a multifunctional cytokine that stimulates blood vessel sprouting, vascular repair, and regeneration. VEGF-A binds to VEGF receptor tyrosine kinases (VEGFRs) and stimulates intracellular signaling leading to changes in vascular physiology. An important aspect of this phenomenon is the spatiotemporal coordination of VEGFR trafficking and intracellular signaling to ensure that VEGFR residence in different organelles is linked to downstream cellular outputs. Here, we describe a series of assays to evaluate the effects of VEGF-A-stimulated intracellular signaling from intracellular compartments such as the endosome-lysosome system. These assays include the initial isolation and characterization of primary human endothelial cells, performing reverse genetics for analyzing protein function; methods used to study receptor trafficking, signaling, and proteolysis; and assays used to measure changes in cell migration, proliferation, and tubulogenesis. Each of these assays has been exemplified with studies performed in our laboratories. In conclusion, we describe necessary techniques for studying the role of VEGF-A in endothelial cell function. © 2014 Elsevier Inc. All rights reserved.

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

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    Sherin Samuel

    2017-04-01

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

  3. Vascular Endothelial Growth Factor Receptor 3 Controls Neural Stem Cell Activation in Mice and Humans

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    Jinah Han

    2015-02-01

    Full Text Available Neural stem cells (NSCs continuously produce new neurons within the adult mammalian hippocampus. NSCs are typically quiescent but activated to self-renew or differentiate into neural progenitor cells. The molecular mechanisms of NSC activation remain poorly understood. Here, we show that adult hippocampal NSCs express vascular endothelial growth factor receptor (VEGFR 3 and its ligand VEGF-C, which activates quiescent NSCs to enter the cell cycle and generate progenitor cells. Hippocampal NSC activation and neurogenesis are impaired by conditional deletion of Vegfr3 in NSCs. Functionally, this is associated with compromised NSC activation in response to VEGF-C and physical activity. In NSCs derived from human embryonic stem cells (hESCs, VEGF-C/VEGFR3 mediates intracellular activation of AKT and ERK pathways that control cell fate and proliferation. These findings identify VEGF-C/VEGFR3 signaling as a specific regulator of NSC activation and neurogenesis in mammals.

  4. Glial cell line-derived neurotrophic factor induced the differentiation of amniotic fluid-derived stem cells into vascular endothelial-like cells in vitro.

    Science.gov (United States)

    Zhang, Ruyu; Lu, Ying; Li, Ju; Wang, Jia; Liu, Caixia; Gao, Fang; Sun, Dong

    2016-02-01

    Amniotic fluid-derived stem cells (AFSCs) are a novel source of stem cells that are isolated and cultured from second trimester amniocentesis. Glial cell line-derived neurotrophic factor (GDNF) acts as a tissue morphogen and regulates stem cell proliferation and differentiation. This study investigated the effect of an adenovirus-mediated GDNF gene, which was engineered into AFSCs, on the cells' biological properties and whether GDNF in combination with AFSCs can be directionally differentiated into vascular endothelial-like cells in vitro. AFSCs were isolated and cultured using the plastic adherence method in vitro and identified by the transcription factor Oct-4, which is the primary marker of pluripotent stem cells. AFSCs were efficiently transfected by a GFP-labeled plasmid system of an adenovirus vector carrying the GDNF gene (Ad-GDNF-GFP). Transfected AFSCs stably expressed GDNF. Transfected AFSCs were cultured in endothelial growth medium-2 containing vascular endothelial growth factor. After 1 week, AFSCs were positive for von Willebrand factor (vWF) and CD31, which are markers of endothelial cells, and the recombinant GDNF group was significantly higher than undifferentiated controls and the GFP only group. These results demonstrated that AFSCs differentiated into vascular endothelial-like cells in vitro, and recombinant GDNF promoted differentiation. The differentiation-induced AFSCs may be used as seed cells to provide a new manner of cell and gene therapies for transplantation into the vascular injury site to promote angiogenesis.

  5. Vacuolar Sorting Receptor-Mediated Trafficking of Soluble Vacuolar Proteins in Plant Cells

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    Hyangju Kang

    2014-08-01

    Full Text Available Vacuoles are one of the most prominent organelles in plant cells, and they play various important roles, such as degradation of waste materials, storage of ions and metabolites, and maintaining turgor. During the past two decades, numerous advances have been made in understanding how proteins are specifically delivered to the vacuole. One of the most crucial steps in this process is specific sorting of soluble vacuolar proteins. Vacuolar sorting receptors (VSRs, which are type I membrane proteins, are involved in the sorting and packaging of soluble vacuolar proteins into transport vesicles with the help of various accessory proteins. To date, large amounts of data have led to the development of two different models describing VSR-mediated vacuolar trafficking that are radically different in multiple ways, particularly regarding the location of cargo binding to, and release from, the VSR and the types of carriers utilized. In this review, we summarize current literature aimed at elucidating VSR-mediated vacuolar trafficking and compare the two models with respect to the sorting signals of vacuolar proteins, as well as the molecular machinery involved in VSR-mediated vacuolar trafficking and its action mechanisms.

  6. Bacterial CpG-DNA activates dendritic cells in vivo: T helper cell-independent cytotoxic T cell responses to soluble proteins.

    Science.gov (United States)

    Sparwasser, T; Vabulas, R M; Villmow, B; Lipford, G B; Wagner, H

    2000-12-01

    Receptors for conserved molecular patterns associated with microbial pathogens induce synthesis of co-stimulatory molecules and cytokines in immature dendritic cells (DC), as do antigen-reactive CD4 T helper cells via CD40 signaling. Once activated, antigen-presenting DC may activate CD8 T cell responses in a T helper cell-independent fashion. Using immunostimulatory CpG-oligonucleotides (ODN) mimicking bacterial CpG-DNA, we tested whether CpG-DNA bypasses the need for T helper cells in CTL responses towards proteins by directly activating antigen-presenting DC to transit into professional APC. We describe that immature DC in situ constitutively process soluble proteins and generate CD8 T cell determinants yet CD8 T cell responses remain abortive. Induction of primary antigen-specific CD8 cytotoxic T lymphocyte (CTL)-mediated responses becomes initiated in wild-type as well as T helper cell-deficient mice, provided soluble protein and CpG-ODN are draining into the same lymph node. Specifically we show that CpG-ODN trigger antigen-presenting immature DC within the draining lymph node to acutely up-regulate co-stimulatory molecules and produce IL-12. These results provide new insights for generating in vivo efficient CTL responses to soluble proteins which may influence vaccination strategies.

  7. An analysis on the roles of angiogenesis-related factors including serum vitamin D, soluble endoglin (sEng), soluble fms-like tyrosine kinase 1 (sFlt1), and vascular endothelial growth factor (VEGF) in the diagnosis and severity of late-onset preeclampsia.

    Science.gov (United States)

    Cim, Numan; Kurdoglu, Mertihan; Ege, Serhat; Yoruk, Ibrahim; Yaman, Gorkem; Yildizhan, Recep

    2017-07-01

    The aim of this study was to evaluate the roles of proangiogenic factors including serum vitamin D and vascular endothelial growth factor (VEGF) and anti-angiogenic factors including soluble endoglin (sEng) and soluble fms-like tyrosine kinase 1 (sFlt1) in the diagnosis and severity of late-onset preeclampsia. The study was conducted at Yuzuncu Yil University Research and Education Hospital Department of Gynecology and Obstetrics. The study included a patient group of 40 women with late-onset preeclampsia who were pregnant at ≥32 weeks of gestation according to the last menstrual period (LMP) or ultrasonographic fetal biometric measurement and a control group of 40 healthy pregnant women who presented to our clinic for routine pregnancy examination and were at the same age and gestational period with those in the patient group. The two groups were compared in terms of maternal age, gravida, parity, week of gestation, systolic/diastolic blood pressure, total protein in spot urine sample, 24-h urine protein, white blood cell (WBC), hemoglobin (Hgb), platelet count, urea, creatinine, liver function tests (AST, ALT, LDH), vitamin D 3 , 25(OH) vitamin D 3 , 1,25(OH) vitamin D 3 , sEng, sFlt1, and VEGF levels, mode of delivery, the infant APGAR score at 1 and 5 min after delivery, and infant weight at delivery. The groups were similar in terms of age, gravida, parity, week of gestation, serum vitamin D 3 , 25(OH) vitamin D 3 , 1,25(OH) 2 vitamin D 3 and VEGF levels, and infant weight at delivery (p > 0.05). Systolic/diastolic blood pressure, total protein in spot urine sample, 24-h urine protein, WBC, Hgb, serum urea, creatine, AST, ALT, and LDH were significantly higher in the preeclamptic group compared to the healthy group (p preeclampsia compared to the women with mild preeclampsia (p preeclampsia (p > 0.05). Both sEng and sFlt1 levels are remarkably high in patients with late-onset preeclampsia; however, only sEng may be a useful tool in the

  8. 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.

  9. Carbohydrate-related genes and cell wall biosynthesis in vascular tissues of loblolly pine (Pinus taeda).

    Science.gov (United States)

    Nairn, Campbell J; Lennon, Denise M; Wood-Jones, Alicia; Nairn, Alison V; Dean, Jeffrey F D

    2008-07-01

    Loblolly pine (Pinus taeda L.), the most widely planted tree species in the United States, is an important source of wood and wood fibers for a multitude of consumer products. Wood fibers are primarily composed of secondary cell walls, and cellulose, hemicelluloses and lignin are major components of wood. Fiber morphology and cell wall composition are important determinants of wood properties. We used comparative genomics to identify putative genes for cellulose and hemicellulose synthesis in loblolly pine that are homologous to genes implicated in cell wall synthesis in angiosperms. Sequences encoding putative secondary cell wall cellulose synthase genes, cellulose synthase-like genes, a membrane-bound endoglucanase gene, a sucrose synthase gene, a UDP-glucose pyrophosphorylase gene and GDP-mannose pyrophosphorylase genes were identified in expressed sequence tag (EST) collections from loblolly pine. Full-length coding sequences were obtained from cDNA clones isolated from a library constructed from developing xylem. Phylogenetic relationships between the genes from loblolly pine and angiosperm taxa were examined and transcriptional profiling in vascular tissues was conducted by real-time quantitative, reverse transcriptase-polymerase chain reaction. The putative cell wall synthesis genes were expressed at high levels in vascular tissues and a subset was differentially regulated in xylem and phloem tissues. Inferred phylogenetic relationships and expression patterns for the genes from loblolly pine were consistent with roles in synthesis of complex carbohydrates of the cell wall. These studies suggest functional conservation of homologous wood formation genes in gymnosperm and angiosperm taxa.

  10. Mobilization of endothelial precursor cells: systemic vascular response to musculoskeletal trauma.

    LENUS (Irish Health Repository)

    Laing, A J

    2012-02-03

    Postnatal vasculogenesis, the process by which vascular committed bone marrow stem cells or endothelial precursor cells (EPC) migrate, differentiate, and incorporate into the nacent endothelium contributing to physiological and pathological neovascularization, has stimulated much interest. Its contribution to tumor nonvascularization, wound healing, and revascularization associated with skeletal and cardiac muscles ischaemia is established. We evaluated the mobilization of EPCs in response to musculoskeletal trauma. Blood from patients (n = 15) following AO type 42a1 closed diaphyseal tibial fractures was analyzed for CD34 and AC133 cell surface marker expression. Immunomagnetically enriched CD34+ mononuclear cell (MNC(CD34+)) populations were cultured and examined for phenotypic and functional vascular endothelial differentiation. Circulating MNC(CD34+) levels increased sevenfold by day 3 postinjury. Circulating MNC(AC133+) increased 2.5-fold. Enriched MNC(CD34+) populations from day 3 samples in culture exhibited cell cluster formation with sprouting spindles. These cells bound UEA-1 and incorporated fluorescent DiI-Ac-LDL intracellularily. Our findings suggest a systemic provascular response is initiated in response to musculoskeletal trauma. Its therapeutic manipulation may have implications for the potential enhancement of fracture healing.

  11. Recellularization of biological heart valves with human vascular cells: in vitro hemocompatibility assessment.

    Science.gov (United States)

    Schopka, Simon; Schmid, Franz-Xaver; Hirt, Stephan; Birnbaum, Dietrich E; Schmid, Christof; Lehle, Karla

    2009-01-01

    Coverage of cardiovascular bioprostheses with autologous endothelium is used for the purpose of improving blood compatibility. The aim of our study was to analyze endothelialization potential of glutaraldehyde-fixed heart valves, cellular functions of seeded endothelial cells (EC), and the impact of a two-stage seeding protocol using human vascular fibroblasts (FB) and EC from saphenous veins (HSVEC) on cellular functional properties in vitro. Adherence and morphology of adhered cells were assessed by scanning electronic microscopy and immunohistochemistry. Reproducible, complete surface coverage with EC was established on decellularized and glutaraldehyde-fixed bovine pericardium. Analyzing functional properties of cells directly adhered to biomaterial revealed nonproliferative cells, which were capable of inflammatory stimulation in terms of TNF-induced increase in interleukin-6 secretion and adhesion of inflammatory cells. Furthermore, EC showed sustained antithrombotic properties quantified by platelet adhesion onto EC and prostacyclin secretion by EC. Preseeding with vascular fibroblasts using a two-stage seeding protocol induced EC proliferation and improved inflammatory and anti-thrombotic functions. Cardiovascular biomaterials differ significantly in their potential to allow for adhesion of human EC. Successfully endothelialized biomaterial, however, revealed cellular properties which are likely to be favorable to improving performance of biomaterials. Two-stage seeding adds regenerative potential and improves cell functions of adherent EC. (c) 2008 Wiley Periodicals, Inc.

  12. Sulforaphane reduces vascular inflammation in mice and prevents TNF-α-induced monocyte adhesion to primary endothelial cells through interfering with the NF-κB pathway.

    Science.gov (United States)

    Nallasamy, Palanisamy; Si, Hongwei; Babu, Pon Velayutham Anandh; Pan, Dengke; Fu, Yu; Brooke, Elizabeth A S; Shah, Halley; Zhen, Wei; Zhu, Hong; Liu, Dongmin; Li, Yunbo; Jia, Zhenquan

    2014-08-01

    Sulforaphane, a naturally occurring isothiocyanate present in cruciferous vegetables, has received wide attention for its potential to improve vascular function in vitro. However, its effect in vivo and the molecular mechanism of sulforaphane at physiological concentrations remain unclear. Here, we report that a sulforaphane concentration as low as 0.5 μM significantly inhibited tumor necrosis factor-α (TNF-α)-induced adhesion of monocytes to human umbilical vein endothelial cells, a key event in the pathogenesis of atherosclerosis both in static and under flow conditions. Such physiological concentrations of sulforaphane also significantly suppressed TNF-α-induced production of monocyte chemotactic protein-1 and adhesion molecules including soluble vascular adhesion molecule-1 and soluble E-selectin, key mediators in the regulation of enhanced endothelial cell-monocyte interaction. Furthermore, sulforaphane inhibited TNF-α-induced nuclear factor (NF)-κB transcriptional activity, Inhibitor of NF-κB alpha (IκBα) degradation and subsequent NF-κB p65 nuclear translocation in endothelial cells, suggesting that sulforaphane can inhibit inflammation by suppressing NF-κB signaling. In an animal study, sulforaphane (300 ppm) in a mouse diet significantly abolished TNF-α-increased ex vivo monocyte adhesion and circulating adhesion molecules and chemokines in C57BL/6 mice. Histology showed that sulforaphane treatment significantly prevented the eruption of endothelial lining in the intima layer of the aorta and preserved elastin fibers' delicate organization, as shown by Verhoeff-van Gieson staining. Immunohistochemistry studies showed that sulforaphane treatment also reduced vascular adhesion molecule-1 and monocyte-derived F4/80-positive macrophages in the aorta of TNF-α-treated mice. In conclusion, sulforaphane at physiological concentrations protects against TNF-α-induced vascular endothelial inflammation, in both in vitro and in vivo models. This anti

  13. Bacteria-induced release of white cell--and platelet-derived vascular endothelial growth factor in vitro

    DEFF Research Database (Denmark)

    Nielsen, Hans Jørgen; Werther, K; Mynster, T

    2001-01-01

    BACKGROUND AND OBJECTIVES: Poor prognosis after resection of primary colorectal cancer may be related to the combination of perioperative blood transfusion and subsequent development of infectious complications. White blood cell--and platelet-derived cancer growth substances, including vascular...

  14. Construction and analysis of SSH cDNA library of human vascular endothelial cells related to gastrocarcinoma

    OpenAIRE

    Liu, Yong-Bo; Wei, Zhao-Xia; Li, Li; Li, Hang-Sheng; Chen, Hui; Li, Xiao-Wen

    2003-01-01

    AIM: To construct subtracted cDNA libraries of human vascular endothelial cells (VECs) related to gastrocarcinoma using suppression substractive hybridization (SSH) and to analyze cDNA libraries of gastrocarcinoma and VECs in Cancer Gene Anatomy Project (CGAP) database.

  15. Designing a highly active soluble PQQ-glucose dehydrogenase for efficient glucose biosensors and biofuel cells

    Energy Technology Data Exchange (ETDEWEB)

    Durand, Fabien [Universite de Bordeaux, Centre de Recherche Paul Pascal (CRPP), UPR 8641, Avenue Albert Schweitzer, 33600 Pessac (France); Stines-Chaumeil, Claire [Universite de Bordeaux, CNRS, Institut de Biochimie et de Genetique Cellulaires, 1 rue Camille Saint Saens, 33077 Bordeaux Cedex (France); Flexer, Victoria [Universite de Bordeaux, Centre de Recherche Paul Pascal (CRPP), UPR 8641, Avenue Albert Schweitzer, 33600 Pessac (France); Andre, Isabelle [Universite de Toulouse, INSA, UPS, INP, LISBP, 135 Avenue de Rangueil, F-31077 Toulouse (France); CNRS, UMR5504, F-31400 Toulouse (France); INRA, UMR 792 Ingenierie des Systemes Biologiques et des Procedes, F-31400 Toulouse (France); Mano, Nicolas, E-mail: mano@crpp-bordeaux.cnrs.fr [Universite de Bordeaux, Centre de Recherche Paul Pascal (CRPP), UPR 8641, Avenue Albert Schweitzer, 33600 Pessac (France)

    2010-11-26

    Research highlights: {yields} A new mutant of PQQ-GDH designed for glucose biosensors application. {yields} First mutant of PQQ-GDH with higher activity for D-glucose than the Wild type. {yields} Position N428 is a key point to increase the enzyme activity. {yields} Molecular modeling shows that the N428 C mutant displays a better interaction for PQQ than the WT. -- Abstract: We report for the first time a soluble PQQ-glucose dehydrogenase that is twice more active than the wild type for glucose oxidation and was obtained by combining site directed mutagenesis, modelling and steady-state kinetics. The observed enhancement is attributed to a better interaction between the cofactor and the enzyme leading to a better electron transfer. Electrochemical experiments also demonstrate the superiority of the new mutant for glucose oxidation and make it a promising enzyme for the development of high-performance glucose biosensors and biofuel cells.

  16. In vivo vascularization of cell sheets provided better long-term tissue survival than injection of cell suspension.

    Science.gov (United States)

    Takeuchi, Ryohei; Kuruma, Yosuke; Sekine, Hidekazu; Dobashi, Izumi; Yamato, Masayuki; Umezu, Mitsuo; Shimizu, Tatsuya; Okano, Teruo

    2016-08-01

    Cell sheets have shown a remarkable ability for repairing damaged myocardium in clinical and preclinical studies. Although they demonstrate a high degree of viability as engrafted cells in vivo, the reason behind their survivability is unclear. In this study, the survival and vascularization of rat cardiac cell sheets transplanted in the subcutaneous tissue of athymic rats were investigated temporally. The cell sheets showed significantly higher survival than cell suspensions for up to 12 months, using an in vivo bioluminescence imaging system to detect luciferase-positive transplanted cells. Terminal deoxynucleotidyl transferase dUTP nick-end labelling (TUNEL) assay also showed a smaller number of apoptotic cells in the cell sheets than in the cell suspensions at 1 day. Rapid vascular formation and maturation were observed inside the cell sheets using an in vivo imaging system. Leaky vessels appeared at 6 h, red blood cells flowing through functional vessels appeared at 12 h, and morphologically matured vessels appeared at 7 days. In addition, immunostaining of cell sheets with nerve/glial antigen-2 (NG2) showed that vessel maturity increased over time. Interestingly, these results correlated with the dynamics of cell sheet mRNA expression. Genes related to endothelial cells (ECs) proliferation, migration and vessel sprouting were highly expressed within 1 day, and genes related to pericyte recruitment and vessel maturation were highly expressed at 3 days or later. This suggested that the cell sheets could secrete appropriate angiogenic factors in a timely way after transplantation, and this ability might be a key reason for their high survival. Copyright © 2014 John Wiley & Sons, Ltd. Copyright © 2014 John Wiley & Sons, Ltd.

  17. Soluble Mediators in Platelet Concentrates Modulate Dendritic Cell Inflammatory Responses in an Experimental Model of Transfusion.

    Science.gov (United States)

    Perros, Alexis J; Christensen, Anne-Marie; Flower, Robert L; Dean, Melinda M

    2015-10-01

    The transfusion of platelet concentrates (PCs) is widely used to treat thrombocytopenia and severe trauma. Ex vivo storage of PCs is associated with a storage lesion characterized by partial platelet activation and the release of soluble mediators, such as soluble CD40 ligand (sCD40L), RANTES, and interleukin (IL)-8. An in vitro whole blood culture transfusion model was employed to assess whether mediators present in PC supernatants (PC-SNs) modulated dendritic cell (DC)-specific inflammatory responses (intracellular staining) and the overall inflammatory response (cytometric bead array). Lipopolysaccharide (LPS) was included in parallel cultures to model the impact of PC-SNs on cell responses following toll-like receptor-mediated pathogen recognition. The impact of both the PC dose (10%, 25%) and ex vivo storage period was investigated [day 2 (D2), day 5 (D5), day 7 (D7)]. PC-SNs alone had minimal impact on DC-specific inflammatory responses and the overall inflammatory response. However, in the presence of LPS, exposure to PC-SNs resulted in a significant dose-associated suppression of the production of DC IL-12, IL-6, IL-1α, tumor necrosis factor-α (TNF-α), and macrophage inflammatory protein (MIP)-1β and storage-associated suppression of the production of DC IL-10, TNF-α, and IL-8. For the overall inflammatory response, IL-6, TNF-α, MIP-1α, MIP-1β, and inflammatory protein (IP)-10 were significantly suppressed and IL-8, IL-10, and IL-1β significantly increased following exposure to PC-SNs in the presence of LPS. These data suggest that soluble mediators present in PCs significantly suppress DC function and modulate the overall inflammatory response, particularly in the presence of an infectious stimulus. Given the central role of DCs in the initiation and regulation of the immune response, these results suggest that modulation of the DC inflammatory profile is a probable mechanism contributing to transfusion-related complications.

  18. Real-time measurements of endogenous CO production from vascular cells using an ultrasensitive laser sensor

    Science.gov (United States)

    Morimoto, Y.; Durante, W.; Lancaster, D. G.; Klattenhoff, J.; Tittel, F. K.

    2001-01-01

    Carbon monoxide (CO) has been implicated as a biological messenger molecule analogous to nitric oxide. A compact gas sensor based on a midinfrared laser absorption spectroscopy was developed for direct and real-time measurement of trace levels (in approximate pmol) of CO release by vascular cells. The midinfrared light is generated by difference frequency mixing of two nearinfrared lasers in a nonlinear optical crystal. A strong infrared absorption line of CO (4.61 microm) is chosen for convenient CO detection without interference from other gas species. The generation of CO from cultured vascular smooth muscle cells was detected every 20 s without any chemical modification to the CO. The sensitivity of the sensor reached 6.9 pmol CO. CO synthesis was measured from untreated control cells (0.25 nmol per 10(7) cells/h), sodium nitroprusside-treated cells (0.29 nmol per 10(7) cells/h), and hemin-treated cells (0.49 nmol per 10(7) cells/h). The sensor also detected decreases in CO production after the addition of the heme oxygenase (HO) inhibitor tin protoporphyrin-IX (from 0.49 to 0.02 nmol per 10(7) cells/h) and increases after the administration of the HO substrate hemin (from 0.27 to 0.64 nmol per 10(7) cells/h). These results demonstrate that midinfrared laser absorption spectroscopy is a useful technique for the noninvasive and real-time detection of trace levels of CO from biological tissues.

  19. Dimethylfumarate attenuates restenosis after acute vascular injury by cell-specific and Nrf2-dependent mechanisms

    Directory of Open Access Journals (Sweden)

    Chang Joo Oh

    2014-01-01

    Full Text Available Excessive proliferation of vascular smooth muscle cells (VSMCs and incomplete re-endothelialization is a major clinical problem limiting the long-term efficacy of percutaneous coronary angioplasty. We tested if dimethylfumarate (DMF, an anti-psoriasis drug, could inhibit abnormal vascular remodeling via NF−E2-related factor 2 (Nrf2-NAD(PH quinone oxidoreductase 1 (NQO1 activity. DMF significantly attenuated neointimal hyperplasia induced by balloon injury in rat carotid arteries via suppression of the G1 to S phase transition resulting from induction of p21 protein in VSMCs. Initially, DMF increased p21 protein stability through an enhancement in Nrf2 activity without an increase in p21 mRNA. Later on, DMF stimulated p21 mRNA expression through a process dependent on p53 activity. However, heme oxygenase-1 (HO-1 or NQO1 activity, well-known target genes induced by Nrf2, were dispensable for the DMF induction of p21 protein and the effect on the VSMC proliferation. Likewise, DMF protected endothelial cells from TNF-α-induced apoptosis and the dysfunction characterized by decreased eNOS expression. With knock-down of Nrf2 or NQO1, DMF failed to prevent TNF-α-induced cell apoptosis and decreased eNOS expression. Also, CD31 expression, an endothelial specific marker, was restored in vivo by DMF. In conclusion, DMF prevented abnormal proliferation in VSMCs by G1 cell cycle arrest via p21 upregulation driven by Nrf2 and p53 activity, and had a beneficial effect on TNF-α-induced apoptosis and dysfunction in endothelial cells through Nrf2–NQO1 activity suggesting that DMF might be a therapeutic drug for patients with vascular disease.

  20. Hypothyroidism Affects Vascularization and Promotes Immune Cells Infiltration into Pancreatic Islets of Female Rabbits

    Science.gov (United States)

    Rodríguez-Castelán, Julia; Martínez-Gómez, Margarita; Castelán, Francisco; Cuevas, Estela

    2015-01-01

    Thyroidectomy induces pancreatic edema and immune cells infiltration similarly to that observed in pancreatitis. In spite of the controverted effects of hypothyroidism on serum glucose and insulin concentrations, the number and proliferation of Langerhans islet cells as well as the presence of extracellular matrix are affected depending on the islet size. In this study, we evaluated the effect of methimazole-induced hypothyroidism on the vascularization and immune cells infiltration into islets. A general observation of pancreas was also done. Twelve Chinchilla-breed female adult rabbits were divided into control (n = 6) and hypothyroid groups (n = 6, methimazole, 0.02% in drinking water for 30 days). After the treatment, rabbits were sacrificed and their pancreas was excised, histologically processed, and stained with Periodic Acid-Schiff (PAS) or Masson's Trichrome techniques. Islets were arbitrarily classified into large, medium, and small ones. The external and internal portions of each islet were also identified. Student-t-test and Mann-Whitney-U test or two-way ANOVAs were used to compare variables between groups. In comparison with control rabbits, hypothyroidism induced a strong infiltration of immune cells and a major presence of collagen and proteoglycans in the interlobular septa. Large islets showed a high vascularization and immune cells infiltration. The present results show that hypothyroidism induces pancreatitis and insulitis. PMID:26175757

  1. Metabolomic profiling of cellular responses to carvedilol enantiomers in vascular smooth muscle cells.

    Directory of Open Access Journals (Sweden)

    Mingxuan Wang

    Full Text Available Carvedilol is a non-selective β-blocker indicated in the treatment of hypertension and heart failure. Although the differential pharmacological effects of individual Carvedilol enantiomer is supported by preceding studies, the cellular response to each enantiomer is not well understood. Here we report the use of GC-MS metabolomic profiling to study the effects of Carvedilol enantiomers on vascular smooth muscle cells (A7r5 and to shed new light on molecular events underlying Carvedilol treatment. The metabolic analysis revealed alternations in the levels of 8 intracellular metabolites and 5 secreted metabolites in A7r5 cells incubated separately with S- and R-Carvedilol. Principal component analysis of the metabolite data demonstrated the characteristic metabolic signatures in S- and R-Carvedilol-treated cells. A panel of metabolites, including L-serine, L-threonine, 5-oxoproline, myristic acid, palmitic acid and inositol are closely correlated to the vascular smooth muscle contraction. Our findings reveal the differentiating metabolites for A7r5 cells incubated with individual enantiomer of Carvedilol, which opens new perspectives to employ metabolic profiling platform to study chiral drug-cell interactions and aid their incorporation into future improvement of β-blocker therapy.

  2. Role of Galectin-3 in Bone Cell Differentiation, Bone Pathophysiology and Vascular Osteogenesis

    Directory of Open Access Journals (Sweden)

    Carla Iacobini

    2017-11-01

    Full Text Available Galectin-3 is expressed in various tissues, including the bone, where it is considered a marker of chondrogenic and osteogenic cell lineages. Galectin-3 protein was found to be increased in the differentiated chondrocytes of the metaphyseal plate cartilage, where it favors chondrocyte survival and cartilage matrix mineralization. It was also shown to be highly expressed in differentiating osteoblasts and osteoclasts, in concomitance with expression of osteogenic markers and Runt-related transcription factor 2 and with the appearance of a mature phenotype. Galectin-3 is expressed also by osteocytes, though its function in these cells has not been fully elucidated. The effects of galectin-3 on bone cells were also investigated in galectin-3 null mice, further supporting its role in all stages of bone biology, from development to remodeling. Galectin-3 was also shown to act as a receptor for advanced glycation endproducts, which have been implicated in age-dependent and diabetes-associated bone fragility. Moreover, its regulatory role in inflammatory bone and joint disorders entitles galectin-3 as a possible therapeutic target. Finally, galectin-3 capacity to commit mesenchymal stem cells to the osteoblastic lineage and to favor transdifferentiation of vascular smooth muscle cells into an osteoblast-like phenotype open a new area of interest in bone and vascular pathologies.

  3. "ApoptomiRs" in vascular cells: their role in physiological and pathological angiogenesis.

    Science.gov (United States)

    Quintavalle, Cristina; Garofalo, Michela; Croce, Carlo M; Condorelli, Gerolama

    2011-10-01

    MicroRNAs (miRNAs) have emerged as crucial players regulating the magnitude of gene expression in a variety of organisms. This class of short (22 nucleotides) noncoding RNA molecules have been shown to participate in almost every cellular process investigated so far, and their deregulation is observed in different human pathologies including cancer, heart disease, and neurodegeneration. These new molecular regulators have been identified also in endothelial cells (ECs), and their role in the regulation of different aspects of the angiogenic process has been recently investigated in a variety of laboratories. The current review focuses on the research progress regarding the roles of miRNAs in vascular pathology and their potential therapeutic applications for vascular diseases associated with abnormal angiogenesis, such as cancer. Copyright © 2011 Elsevier Inc. All rights reserved.

  4. Early outgrowth cells release soluble endocrine antifibrotic factors that reduce progressive organ fibrosis.

    Science.gov (United States)

    Yuen, Darren A; Connelly, Kim A; Zhang, Yanling; Advani, Suzanne L; Thai, Kerri; Kabir, Golam; Kepecs, David; Spring, Christopher; Smith, Christopher; Batruch, Ihor; Kosanam, Hari; Advani, Andrew; Diamandis, Eleftherios; Marsden, Philip A; Gilbert, Richard E

    2013-11-01

    Adult bone marrow-derived cells can improve organ function in chronic disease models, ostensibly by the release of paracrine factors. It has, however, been difficult to reconcile this prevailing paradigm with the lack of cell retention within injured organs and their rapid migration to the reticuloendothelial system. Here, we provide evidence that the salutary antifibrotic effects of bone marrow-derived early outgrowth cells (EOCs) are more consistent with an endocrine mode of action, demonstrating not only the presence of antifibrotic factors in the plasma of EOC-treated rats but also that EOC conditioned medium (EOC-CM) potently attenuates both TGF-β- and angiotensin II-induced fibroblast collagen production in vitro. To examine the therapeutic relevance of these findings in vivo, 5/6 subtotally nephrectomized rats, a model of chronic kidney and heart failure characterized by progressive fibrosis of both organs, were randomized to receive i.v. injections of EOC-CM, unconditioned medium, or 10(6) EOCs. Rats that received unconditioned medium developed severe kidney injury with cardiac diastolic dysfunction. In comparison, EOC-CM-treated rats demonstrated substantially improved renal and cardiac function and structure, mimicking the changes found in EOC-treated animals. Mass spectrometric analysis of EOC-CM identified proteins that regulate cellular functions implicated in fibrosis. These results indicate that EOCs secrete soluble factor(s) with highly potent antifibrotic activity, that when injected intravenously replicate the salutary effects of the cells themselves. Together, these findings suggest that an endocrine mode of action may underlie the effectiveness of cell therapy in certain settings and portend the possibility for systemic delivery of cell-free therapy. © AlphaMed Press.

  5. The clinical impact of MTHFR polymorphism on the vascular complications of sickle cell disease

    Directory of Open Access Journals (Sweden)

    F. Moreira Neto

    2006-10-01

    Full Text Available Sickle cell disease (SCD is one of the most common inherited diseases in the world and the patients present notorious clinical heterogeneity. It is known that patients with SCD present activation of the blood coagulation and fibrinolytic systems, especially during vaso-occlusive crises, but also during the steady state of the disease. We determined if the presence of the factor V gene G1691A mutation (factor V Leiden, the prothrombin gene G20210A variant, and methylenetetrahydrofolate reductase (MTHFR C677T polymorphism may be risk factors for vascular complications in individuals with SCD. We studied 53 patients with SCD (60% being women, 29 with SS (sickle cell anemia; 28 years, range: 13-52 years and 24 with SC (sickle-hemoglobin C disease; 38.5 years, range: 17-72 years hemoglobinopathy. Factor V Leiden, MTHFR C677T polymorphism, and prothrombin G20210A variant were identified by PCR followed by further digestion of the PCR product with specific endonucleases. The following vascular complications were recorded: stroke, retinopathy, acute thoracic syndrome, and X-ray-documented avascular necrosis. Only one patient was heterozygous for factor V Leiden (1.8% and there was no prothrombin G20210A variant. MTHFR 677TT polymorphism was detected in 1 patient (1.8% and the heterozygous form 677TC was observed in 18 patients (34%, 9 with SS and 9 with SC disease, a prevalence similar to that reported by others. No association was detected between the presence of the MTHFR 677T allele and other genetic modulation factors, such as alpha-thalassemia, ß-globin gene haplotype and fetal hemoglobin. The presence of the MTHFR 677T allele was associated with the occurrence of vascular complications in SCD, although this association was not significant when each complication was considered separately. In conclusion, MTHFR C677T polymorphism might be a risk factor for vascular complications in SCD.

  6. Metal-free Phtalocyanine and 5-Aminolevulenic Acid in Photodynamic Treatment of Human Vascular Cells

    Science.gov (United States)

    Udartseva, Olga O.; Andreeva, Elena R.; Buravkova, Ludmila B.; Tararak, Eduard M.

    2010-05-01

    Originally developed as a tumor therapy, now photodynamic therapy (PDT) may become a useful tool for treatment of cardiovascular diseases. Different cell types are involved in this vascular pathology, and these cells possess different susceptibility to PDT. In this study we screened the effects of two new photosensitizers (PtS and ALA) on human vascular cells. Human macrophages (Mph), aorta endothelial (HAEC) and smooth muscle (SMC) cells were obtained and cultured as described elsewhere. 2-10 ug/ml PtS was added to culture medium 24 h before PDT. ALA was added in 2-10 mM concentration in serum-free culture medium. Then cells were washed carefully and illuminated with 692-nm (PtS) or 633-nm (ALA) light. Cellular viability was measured with MTT-test. Except the case of use 5-10 mM ALA, either photosensitizer accumulation alone or laser illumination alone did not affect cells. Illumination of PtS or ALA-loaded cells (1-20 J/cm2) impaired cellular viability in dose-dependent manner. LD90 for different vascular cells with PtS were as follows: HAEC -1 J/cm2, SMC -2 J/cm2, Mph -5 J/cm2. HAEC and some Mph were unsusceptible to ALA-PDT. SMC LD90 with ALA was 20 J/cm2. Effects of ALA-PDT depended on protoporphyrin IX (PpIX) formation in cells. HAEC didn't accumulate PpIX and were non-sensitive to ALA-PDT. PpIX formation in Mph changed individually according to donor. Illumination of ALA-loaded Mph with low PpIX formation did not affect cells. However LD90 for Mph with high PpIX formation comprised 20 J/cm2. All cell types were more susceptible to PtS-PDT compared to ALA-PDT. Among tested photosensitizers PtS was the most effective one. HAEC were the most susceptible to PtS-PDT.

  7. Renal Cell Toxicity of Water-Soluble Coal Extracts from the Gulf Coast

    Science.gov (United States)

    Ojeda, A. S.; Ford, S.; Ihnat, M.; Gallucci, R. M.; Philp, P. R.

    2017-12-01

    In the Gulf Coast, many rural residents rely on private well water for drinking, cooking, and other domestic needs. A large portion of this region contains lignite coal deposits within shallow aquifers that potentially leach organic matter into the water supply. It is proposed that the organic matter leached from low-rank coal deposits contributes to the development of kidney disease, however, little work has been done to investigate the toxicity of coal extracts. In this study, human kidney cells (HK-2) were exposed to water-soluble extracts of Gulf Coast Coals to assess toxicity. Cell viability was measured by direct counts of total and necrotic cells. A dose-response curve was used to generate IC50 values, and the extracts showed significant toxicity that ranged from 0.5% w/v to 3% w/v IC50. The most toxic extract was from Louisiana where coal-derived organic material has been previously linked to high incidents of renal pelvic cancer (RPC). Although the toxic threshold measured in this study is significantly higher than the concentration of organic matter in the groundwater, typically <5 mg/L (0.005% w/v), residents in the affected areas may consume contaminated water over a lifetime. It is possible that the cumulative toxic effects of coal-derived material contribute to the development of disease.

  8. Insulin sensitizers prevent fine particulate matter-induced vascular insulin resistance and changes in endothelial progenitor cell homeostasis.

    Science.gov (United States)

    Haberzettl, Petra; McCracken, James P; Bhatnagar, Aruni; Conklin, Daniel J

    2016-06-01

    Exposure to fine particular matter (PM2.5) increases the risk of developing cardiovascular disease and Type 2 diabetes. Because blood vessels are sensitive targets of air pollutant exposure, we examined the effects of concentrated ambient PM2.5 (CAP) on vascular insulin sensitivity and circulating levels of endothelial progenitor cells (EPCs), which reflect cardiovascular health. We found that CAP exposure for 9 days decreased insulin-stimulated Akt phosphorylation in the aorta of mice maintained on control diet. This change was accompanied by the induction of IL-1β and increases in the abundance of cleaved IL-18 and p10 subunit of Casp-1, consistent with the activation of the inflammasome pathway. CAP exposure also suppressed circulating levels of EPCs (Flk-1(+)/Sca-1(+) cells), while enhancing the bone marrow abundance of these cells. Although similar changes in vascular insulin signaling and EPC levels were observed in mice fed high-fat diet, CAP exposure did not exacerbate diet-induced changes in vascular insulin resistance or EPC homeostasis. Treatment with an insulin sensitizer, metformin or rosiglitazone, prevented CAP-induced vascular insulin resistance and NF-κB and inflammasome activation and restored peripheral blood and bone marrow EPC levels. These findings suggest that PM2.5 exposure induces diet-independent vascular insulin resistance and inflammation and prevents EPC mobilization, and that this EPC mobilization defect could be mediated by vascular insulin resistance. Impaired vascular insulin sensitivity may be an important mechanism underlying PM2.5-induced vascular injury, and pharmacological sensitization to insulin action could potentially prevent deficits in vascular repair and mitigate vascular inflammation due to exposure to elevated levels of ambient air pollution. Copyright © 2016 the American Physiological Society.

  9. Beta-type transforming growth factor specifies organizational behavior in vascular smooth muscle cell cultures.

    Science.gov (United States)

    Majack, R A

    1987-07-01

    In culture, vascular smooth muscle cells (SMC) grow in a "hill-and-valley" (multilayered) pattern of organization. We have studied the growth, behavioral organization, and biosynthetic phenotype of rat aortic SMC exposed to purified platelet-derived growth regulatory molecules. We show that multilayered growth is not a constitutive feature of cultured SMC, and that beta-type transforming growth factor (TGF-beta) is the primary determinant of multilayered growth and the hill-and-valley pattern of organization diagnostic for SMC in culture. TGF-beta inhibited, in a dose-dependent manner, the serum- or platelet-derived growth factor-mediated proliferation of these cells in two-dimensional culture, but only when cells were plated at subconfluent densities. The ability of TGF-beta to inhibit SMC growth was inversely correlated to plating cell density. When SMC were plated at monolayer density (5 X 10(4) cells/cm2) to allow maximal cell-to-cell contact, TGF-beta potentiated cell growth. This differential response of SMC to TGF-beta may contribute to the hill-and-valley pattern of organization. Unlike its effect on other cell types, TGF-beta did not enhance the synthesis of fibronectin or its incorporation into the extracellular matrix. However, the synthesis of a number of other secreted proteins was altered by TGF-beta treatment. SMC treated with TGF-beta for 4 or 8 h secreted markedly enhanced amounts of an Mr 38,000-D protein doublet whose synthesis is known to be increased by heparin (another inhibitor of SMC growth), suggesting metabolic similarities between heparin- and TGF-beta-mediated SMC growth inhibition. The data suggest that TGF-beta may play an important and complex regulatory role in SMC proliferation and organization during development and after vascular injury.

  10. Direct measurement of actual levels of nitric oxide (NO in cell culture conditions using soluble NO donors

    Directory of Open Access Journals (Sweden)

    Weilue He

    2016-10-01

    Full Text Available Applying soluble nitric oxide (NO donors is the most widely used method to expose cells of interest to exogenous NO. Because of the complex equilibria that exist between components in culture media, the donor compound and NO itself, it is very challenging to predict the dose and duration of NO cells actually experience. To determine the actual level of NO experienced by cells exposed to soluble NO donors, we developed the CellNO Trap, a device that allows continuous, real-time monitoring of the level of NO adherent cells produce and/or experience in culture without the need to alter cell culturing procedures. Herein, we directly measured the level of NO that cells grown in the CellNO Trap experienced when soluble NO donors were added to solutions in culture wells and we characterized environmental conditions that effected the level of NO in in vitro culture conditions. Specifically, the dose and duration of NO generated by the soluble donors S-nitroso-N-acetylpenicillamine (SNAP, S-nitrosoglutathione (GSNO, S-nitrosocysteine (CysNO and the diazeniumdiolate diethyltriamine (DETA/NO were investigated in both phosphate buffered saline (PBS and cell culture media. Other factors that were studied that potentially affect the ultimate NO level achieved with these donors included pH, presence of transition metals (ion species, redox level, presence of free thiol and relative volume of media. Then murine smooth muscle cell (MOVAS with different NO donors but with the same effective concentration of available NO were examined and it was demonstrated that the cell proliferation ratio observed does not correlate with the half-lives of NO donors characterized in PBS, but does correlate well with the real-time NO profiles measured under the actual culture conditions. This data demonstrates the dynamic characteristic of the NO and NO donor in different biological systems and clearly illustrates the importance of tracking individual NO profiles under the actual

  11. Vascular effects of multiwalled carbon nanotubes in dyslipidemic ApoE-/- mice and cultured endothelial cells

    DEFF Research Database (Denmark)

    Cao, Yi; Jacobsen, Nicklas Raun; Danielsen, Pernille Høgh

    2014-01-01

    , oxidative stress, DNA repair, and vascular activation response genes. The level of oxidatively damaged DNA in lung tissue was unaltered, probably due to increased DNA repair capacities. Despite upregulation of inflammatory genes in the liver, effects on systemic cytokines and lipid peroxidation were minimal....... The exposure to MWCNTs in cultured human endothelial cells increased the expression of cell adhesion molecules (ICAM1 and VCAM1). In cocultures, there was increased adhesion of monocytes to endothelial cells after exposure to MWCNT. The exposure to both types of MWCNT was also associated with increased lipid...... accumulation in monocytic-derived foam cells, which was dependent on concomitant oxidative stress because the antioxidant N-acetylcysteine inhibited the lipid accumulation. Collectively, our results indicate that exposure to MWCNT is associated with accelerated progression of atherosclerosis, which could...

  12. Arachidonic metabolism and radiation toxicity in cultures of vascular endothelial cells

    International Nuclear Information System (INIS)

    Eldor, A.; Vlodavsky, I.; Fuks, Z.; Matzner, Y.; Rubin, D.B.

    1989-01-01

    The authors conclude that the observed changes in eicosanoid production by vascular endothelial cells exposed to ionizing irradiation may be relevant to the pathogenesis of post-radiation injury in small and large blood vessels. Anomalies of PGI 2 production may lead to thrombosis and accelerated arteriosclerosis which are observed in irradiated vessels. The generation of potent cells may greatly facilitate inflammation in irradiated vessels. The model of irradiated cultured endothelial cells may also be useful for the study of various methods and agents aimed at reducing the radiation induced damage to blood vessels. Evaluation of the capacity of cultured endothelial cells to produce eicosanoids may serve as an appropriate index for the metabolic damage induced by radiation. (author)

  13. 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.

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

    International Nuclear Information System (INIS)

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

    2013-01-01

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

  15. Arsenite enhances tumor necrosis factor-α-induced expression of vascular cell adhesion molecule-1

    International Nuclear Information System (INIS)

    Tsou, T.-C.; Yeh, Szu Ching; Tsai, E.-M.; Tsai, F.-Y.; Chao, H.-R.; Chang, Louis W.

    2005-01-01

    Epidemiological studies demonstrated a high association of vascular diseases with arsenite exposure. We hypothesize that arsenite potentiates the effect of proinflammatory cytokines on vascular endothelial cells, and hence contributes to atherosclerosis. In this study, we investigated the effect of arsenite and its induction of glutathione (GSH) on vascular cell adhesion molecule-1 (VCAM-1) protein expression in human umbilical vein endothelial cells (HUVECs) in response to tumor necrosis factor-α (TNF-α), a typical proinflammatory cytokine. Our study demonstrated that arsenite pretreatment potentiated the TNF-α-induced VCAM-1 expression with up-regulations of both activator protein-1 (AP-1) and nuclear factor-κB (NF-κB). To elucidate the role of GSH in regulation of AP-1, NF-κB, and VCAM-1 expression, we employed L-buthionine (S,R)-sulfoximine (BSO), a specific γ-glutamylcysteine synthetase (γ-GCS) inhibitor, to block intracellular GSH synthesis. Our investigation revealed that, by depleting GSH, arsenite attenuated the TNF-α-induced VCAM-1 expression as well as a potentiation of AP-1 and an attenuation of NF-κB activations by TNF-α. Moreover, we found that depletion of GSH would also attenuate the TNF-α-induced VCAM-1 expression with a down-regulation of the TNF-α-induced NF-κB activation and without significant effect on AP-1. On the other hand, the TNF-α-induced VCAM-1 expression could be completely abolished by inhibition of AP-1 or NF-κB activity, suggesting that activation of both AP-1 and NF-κB was necessary for VCAM-1 expression. In summary, we demonstrate that arsenite enhances the TNF-α-induced VCAM-1 expression in HUVECs via regulation of AP-1 and NF-κB activities in a GSH-sensitive manner. Our present study suggested a potential mechanism for arsenite in the induction of vascular inflammation and vascular diseases via modulating the actions of proinflammatory cytokines

  16. Suppression of Retinal Neovascularization in vivo by Inhibition of Vascular Endothelial Growth Factor (VEGF) Using Soluble VEGF-Receptor Chimeric Proteins

    Science.gov (United States)

    Aiello, Lloyd Paul; Pierce, Eric A.; Foley, Eliot D.; Takagi, Hitoshi; Chen, Helen; Riddle, Lavon; Ferrara, Napoleone; King, George L.; Smith, Lois E. H.

    1995-11-01

    The majority of severe visual loss in the United States results from complications associated with retinal neovascularization in patients with ischemic ocular diseases such as diabetic retinopathy, retinal vein occlusion, and retinopathy of prematurity. Intraocular expression of the angiogenic protein vascular endothelial growth factor (VEGF) is closely correlated with neovascularization in these human disorders and with ischemia-induced retinal neovascularization in mice. In this study, we evaluated whether in vivo inhibition of VEGF action could suppress retinal neovascularization in a murine model of ischemic retinopathy. VEGF-neutralizing chimeric proteins were constructed by joining the extracellular domain of either human (Flt) or mouse (Flk) high-affinity VEGF receptors with IgG. Control chimeric proteins that did not bind VEGF were also used. VEGF-receptor chimeric proteins eliminated in vitro retinal endothelial cell growth stimulation by either VEGF (P hypoxic conditioned medium (P < 0.005) without affecting growth under nonstimulated conditions. Control proteins had no effect. To assess in vivo response, animals with bilateral retinal ischemia received intravitreal injections of VEGF antagonist in one eye and control protein in the contralateral eye. Retinal neovascularization was quantitated histologically by a masked protocol. Retinal neovascularization in the eye injected with human Flt or murine Flk chimeric protein was reduced in 100% (25/25; P < 0.0001) and 95% (21/22; P < 0.0001) of animals, respectively, compared to the control treated eye. This response was evident after only a single intravitreal injection and was dose dependent with suppression of neovascularization noted after total delivery of 200 ng of protein (P < 0.002). Reduction of histologically evident neovascular nuclei per 6-um section averaged 47% ± 4% (P < 0.001) and 37% ± 2% (P < 0.001) for Flt and Flk chimeric proteins with maximal inhibitory effects of 77% and 66

  17. Anti-Inflammatory effect of Buddleja officinalis on vascular inflammation in human umbilical vein endothelial cells.

    Science.gov (United States)

    Lee, Yun Jung; Moon, Mi Kyoung; Hwang, Sun Mi; Yoon, Jung Joo; Lee, So Min; Seo, Kwan Soo; Kim, Jin Sook; Kang, Dae Gill; Lee, Ho Sub

    2010-01-01

    Vascular inflammation process has been suggested to be an important risk factor in the initiation and development of atherosclerosis. In this study, we investigated whether and by what mechanisms an aqueous extract of Buddleja officinalis (ABO) inhibited the expressions of cellular adhesion molecules, which are relevant to inflammation and atherosclerosis. Pretreatment of human umbilical vein endothelial cells (HUVEC) with ABO (1-10 microg/ml) for 18 hours dose-dependently inhibited TNF-alpha-induced adhesion U937 monocytic cells, as well as mRNA and protein expressions of vascular cell adhesion molecule-1 (VCAM-1), and intercellular cell adhesion molecule-1 (ICAM-1). Pretreatment with ABO also blocked TNF-alpha-induced ROS formation. Nuclear factor-kappa B (NF-kappaB) is required in the transcription of these adhesion molecule genes. Western blot analysis revealed that ABO inhibits the translocation of the p65 subunit of NF-kappaB to the nucleus. ABO inhibited the TNF-alpha-induced degradation of IkappaB-alpha, an inhibitor of NF-kappaB, by inhibiting the phosphorylation of IkappaB-alpha in HUVEC. Taken together, ABO could reduce cytokine-induced endothelial adhesiveness throughout down-regulating intracellular ROS production, NF-kappaB, and adhesion molecule expression in HUVEC, suggesting that the natural herb Buddleja officinalis may have potential implications in atherosclerosis.

  18. A water-soluble derivative of propolis augments the cytotoxic activity of natural killer cells.

    Science.gov (United States)

    Takeda, Kazuyoshi; Nagamatsu, Katashi; Okumura, Ko

    2018-05-23

    Propolis, a resinous material collected from numerous plants by honeybees, has historically been used as a health-promoting food. Recently, due to its potential anti-tumor effects, use of propolis has been proposed as an adjuvant therapy to chemotherapy; however, the effects of propolis on immune responses remain unclear. In this study, we examined the effects of the oral ingestion of propolis on natural killer (NK) cell activity, which is important in immune surveillance against cancer and viral infections. In addition, we assessed the effects of the major components of the water-soluble powder derivative of propolis (WPP). C57BL/6 (B6) wild-type (WT) and RAG 2-deficient (RAG -/- ) mice and BALB/c WT, interferon (IFN)-γ-deficient (IFN-γ -/- ), IFN-γ receptor-deficient (IFN-γR -/- ) and RAG -/- mice were orally administered WPP or its major components. NK cell populations and cytotoxic activity were then examined by flow cytometry and 51 Cr release assay, respectively. While the cytotoxic activity of NK cells was increased following administration of 100 mg/kg/day of WPP for 7 days or 200 or 500 mg/kg/day of WPP for 4 days in WT mice, the proportions of NK cell populations were unaltered. Similar activation of NK cell cytotoxicity was observed when RAG -/- , but not IFN-γ -/- or IFN-γR -/- , mice were orally administered 200 mg/kg/day of WPP for 4 days. Oral ingestion of artepillin C or p-coumaric acid, but not drupanin, augmented NK cell cytotoxicity in a manner similar to WPP and to the mixture of these three components. These results suggest that oral ingestion of WPP enhances NK cell cytotoxic activity, but not proliferation, in a manner dependent on IFN-γ and without the contribution of acquired immune responses. Further, artepillin C or p-coumaric acid, but not drupanin, may be the components responsible for this augmentation of NK cell cytotoxicity. These findings suggest the possible utility of WPP as a therapeutic for prevention of cancer

  19. High Aldehyde Dehydrogenase Activity Identifies a Subset of Human Mesenchymal Stromal Cells with Vascular Regenerative Potential.

    Science.gov (United States)

    Sherman, Stephen E; Kuljanin, Miljan; Cooper, Tyler T; Putman, David M; Lajoie, Gilles A; Hess, David A

    2017-06-01

    During culture expansion, multipotent mesenchymal stromal cells (MSCs) differentially express aldehyde dehydrogenase (ALDH), an intracellular detoxification enzyme that protects long-lived cells against oxidative stress. Thus, MSC selection based on ALDH-activity may be used to reduce heterogeneity and distinguish MSC subsets with improved regenerative potency. After expansion of human bone marrow-derived MSCs, cell progeny was purified based on low versus high ALDH-activity (ALDH hi ) by fluorescence-activated cell sorting, and each subset was compared for multipotent stromal and provascular regenerative functions. Both ALDH l ° and ALDH hi MSC subsets demonstrated similar expression of stromal cell (>95% CD73 + , CD90 + , CD105 + ) and pericyte (>95% CD146 + ) surface markers and showed multipotent differentiation into bone, cartilage, and adipose cells in vitro. Conditioned media (CDM) generated by ALDH hi MSCs demonstrated a potent proliferative and prosurvival effect on human microvascular endothelial cells (HMVECs) under serum-free conditions and augmented HMVEC tube-forming capacity in growth factor-reduced matrices. After subcutaneous transplantation within directed in vivo angiogenesis assay implants into immunodeficient mice, ALDH hi MSC or CDM produced by ALDH hi MSC significantly augmented murine vascular cell recruitment and perfused vessel infiltration compared with ALDH l ° MSC. Although both subsets demonstrated strikingly similar mRNA expression patterns, quantitative proteomic analyses performed on subset-specific CDM revealed the ALDH hi MSC subset uniquely secreted multiple proangiogenic cytokines (vascular endothelial growth factor beta, platelet derived growth factor alpha, and angiogenin) and actively produced multiple factors with chemoattractant (transforming growth factor-β, C-X-C motif chemokine ligand 1, 2, and 3 (GRO), C-C motif chemokine ligand 5 (RANTES), monocyte chemotactic protein 1 (MCP-1), interleukin [IL]-6, IL-8) and matrix

  20. Txnip ablation reduces vascular smooth muscle cell inflammation and ameliorates atherosclerosis in apolipoprotein E knockout mice.

    Science.gov (United States)

    Byon, Chang Hyun; Han, Tieyan; Wu, Judy; Hui, Simon T

    2015-08-01

    Inflammation of vascular smooth muscle cells (VSMC) is intimately linked to atherosclerosis and other vascular inflammatory disease. Thioredoxin interacting protein (Txnip) is a key regulator of cellular sulfhydryl redox and a mediator of inflammasome activation. The goals of the present study were to examine the impact of Txnip ablation on inflammatory response to oxidative stress in VSMC and to determine the effect of Txnip ablation on atherosclerosis in vivo. Using cultured VSMC, we showed that ablation of Txnip reduced cellular oxidative stress and increased protection from oxidative stress when challenged with oxidized phospholipids and hydrogen peroxide. Correspondingly, expression of inflammatory markers and adhesion molecules were diminished in both VSMC and macrophages from Txnip knockout mice. The blunted inflammatory response was associated with a decrease in NF-ĸB nuclear translocation. Loss of Txnip in VSMC also led to a dramatic reduction in macrophage adhesion to VSMC. In vivo data from Txnip-ApoE double knockout mice showed that Txnip ablation led to 49% reduction in atherosclerotic lesion in the aortic root and 71% reduction in the abdominal aorta, compared to control ApoE knockout mice. Our data show that Txnip plays an important role in oxidative inflammatory response and atherosclerotic lesion development in mice. The atheroprotective effect of Txnip ablation implicates that modulation of Txnip expression may serve as a potential target for intervention of atherosclerosis and inflammatory vascular disease. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

  1. Mononuclear Phagocyte-Derived Microparticulate Caspase-1 Induces Pulmonary Vascular Endothelial Cell Injury

    Science.gov (United States)

    Mitra, Srabani

    2015-01-01

    Lung endothelial cell apoptosis and injury occurs throughout all stages of acute lung injury (ALI/ARDS) and impacts disease progression. Lung endothelial injury has traditionally been focused on the role of neutrophil trafficking to lung vascular integrin receptors induced by proinflammatory cytokine expression. Although much is known about the pathogenesis of cell injury and death in ALI/ARDS, gaps remain in our knowledge; as a result of which there is currently no effective pharmacologic therapy. Enzymes known as caspases are essential for completion of the apoptotic program and secretion of pro-inflammatory cytokines. We hypothesized that caspase-1 may serve as a key regulator of human pulmonary microvascular endothelial cell (HPMVEC) apoptosis in ALI/ARDS. Our recent experiments confirm that microparticles released from stimulated monocytic cells (THP1) induce lung endothelial cell apoptosis. Microparticles pretreated with the caspase-1 inhibitor, YVAD, or pan-caspase inhibitor, ZVAD, were unable to induce cell death of HPMVEC, suggesting the role of caspase-1 or its substrate in the induction of HPMVEC cell death. Neither un-induced microparticles (control) nor direct treatment with LPS induced apoptosis of HPMVEC. Further experiments showed that caspase-1 uptake into HPMVEC and the induction of HPMVEC apoptosis was facilitated by caspase-1 interactions with microparticulate vesicles. Altering vesicle integrity completely abrogated apoptosis of HPMVEC suggesting an encapsulation requirement for target cell uptake of active caspase-1. Taken together, we confirm that microparticle centered caspase-1 can play a regulator role in endothelial cell injury. PMID:26710067

  2. Mononuclear Phagocyte-Derived Microparticulate Caspase-1 Induces Pulmonary Vascular Endothelial Cell Injury.

    Science.gov (United States)

    Mitra, Srabani; Wewers, Mark D; Sarkar, Anasuya

    2015-01-01

    Lung endothelial cell apoptosis and injury occurs throughout all stages of acute lung injury (ALI/ARDS) and impacts disease progression. Lung endothelial injury has traditionally been focused on the role of neutrophil trafficking to lung vascular integrin receptors induced by proinflammatory cytokine expression. Although much is known about the pathogenesis of cell injury and death in ALI/ARDS, gaps remain in our knowledge; as a result of which there is currently no effective pharmacologic therapy. Enzymes known as caspases are essential for completion of the apoptotic program and secretion of pro-inflammatory cytokines. We hypothesized that caspase-1 may serve as a key regulator of human pulmonary microvascular endothelial cell (HPMVEC) apoptosis in ALI/ARDS. Our recent experiments confirm that microparticles released from stimulated monocytic cells (THP1) induce lung endothelial cell apoptosis. Microparticles pretreated with the caspase-1 inhibitor, YVAD, or pan-caspase inhibitor, ZVAD, were unable to induce cell death of HPMVEC, suggesting the role of caspase-1 or its substrate in the induction of HPMVEC cell death. Neither un-induced microparticles (control) nor direct treatment with LPS induced apoptosis of HPMVEC. Further experiments showed that caspase-1 uptake into HPMVEC and the induction of HPMVEC apoptosis was facilitated by caspase-1 interactions with microparticulate vesicles. Altering vesicle integrity completely abrogated apoptosis of HPMVEC suggesting an encapsulation requirement for target cell uptake of active caspase-1. Taken together, we confirm that microparticle centered caspase-1 can play a regulator role in endothelial cell injury.

  3. Mononuclear Phagocyte-Derived Microparticulate Caspase-1 Induces Pulmonary Vascular Endothelial Cell Injury.

    Directory of Open Access Journals (Sweden)

    Srabani Mitra

    Full Text Available Lung endothelial cell apoptosis and injury occurs throughout all stages of acute lung injury (ALI/ARDS and impacts disease progression. Lung endothelial injury has traditionally been focused on the role of neutrophil trafficking to lung vascular integrin receptors induced by proinflammatory cytokine expression. Although much is known about the pathogenesis of cell injury and death in ALI/ARDS, gaps remain in our knowledge; as a result of which there is currently no effective pharmacologic therapy. Enzymes known as caspases are essential for completion of the apoptotic program and secretion of pro-inflammatory cytokines. We hypothesized that caspase-1 may serve as a key regulator of human pulmonary microvascular endothelial cell (HPMVEC apoptosis in ALI/ARDS. Our recent experiments confirm that microparticles released from stimulated monocytic cells (THP1 induce lung endothelial cell apoptosis. Microparticles pretreated with the caspase-1 inhibitor, YVAD, or pan-caspase inhibitor, ZVAD, were unable to induce cell death of HPMVEC, suggesting the role of caspase-1 or its substrate in the induction of HPMVEC cell death. Neither un-induced microparticles (control nor direct treatment with LPS induced apoptosis of HPMVEC. Further experiments showed that caspase-1 uptake into HPMVEC and the induction of HPMVEC apoptosis was facilitated by caspase-1 interactions with microparticulate vesicles. Altering vesicle integrity completely abrogated apoptosis of HPMVEC suggesting an encapsulation requirement for target cell uptake of active caspase-1. Taken together, we confirm that microparticle centered caspase-1 can play a regulator role in endothelial cell injury.

  4. Comparative cytotoxicity and genotoxicity of soluble and particulate hexavalent chromium in human and hawksbill sea turtle (Eretmochelys imbricata) skin cells.

    Science.gov (United States)

    Young, Jamie L; Wise, Sandra S; Xie, Hong; Zhu, Cairong; Fukuda, Tomokazu; Wise, John Pierce

    2015-12-01

    Chromium is both a global marine pollutant and a known human health hazard. In this study, we compare the cytotoxicity and genotoxicity of both soluble and particulate chromate in human and hawksbill sea turtle (Eretmochelys imbricata) skin fibroblasts. Our data show that both soluble and particulate Cr(VI) induce concentration-dependent increases in cytotoxicity, genotoxicity, and intracellular Cr ion concentrations in both human and hawksbill sea turtle fibroblasts. Based on administered concentration, particulate and soluble Cr(VI) were more cytotoxic and clastogenic to human cells than sea turtle cells. When the analysis was based on the intracellular concentration of Cr, the data showed that the response of both species was similar. The one exception was the cytotoxicity of intracellular Cr ions from soluble Cr(VI), which caused more cytotoxicity in sea turtle cells (LC50=271μM) than that of human cells (LC50=471μM), but its clastogenicity was similar between the two species. Thus, adjusting for differences in uptake indicated that the explanation for the difference in potency was mostly due to uptake rather than differently affected mechanisms. Overall these data indicate that sea turtles may be a useful sentinel for human health responses to marine pollution. Copyright © 2015 Elsevier Inc. All rights reserved.

  5. Comparative cytotoxicity and genotoxicity of soluble and particulate hexavalent chromium in human and hawksbill sea turtle (Eretmochelys imbricate) skin cells

    Science.gov (United States)

    Young, Jamie L.; Wise, Sandra S.; Xie, Hong; Zhu, Cairong; Fukuda, Tomokazu; Wise, John Pierce

    2015-01-01

    Chromium is both a global marine pollutant and a known human health hazard. In this study, we compare the cytotoxicity and genotoxicity of both soluble and particulate chromate in human and hawksbill sea turtle (Eretmochelys imbricata) skin fibroblasts. Our data show that both soluble and particulate Cr(VI) induce concentration-dependent increases in cytotoxicity, genotoxicity, and intracellular Cr ion concentrations in both human and hawksbill sea turtle fibroblasts. Based on administered concentration, particulate and soluble Cr(VI) were more cytotoxic and clastogenic to human cells than sea turtle cells. When the analysis was based on the intracellular concentration of Cr, the data showed the response of both species was similar. The one exception was the cytotoxicity of intracellular Cr ions from soluble Cr(VI), which caused more cytotoxicity in sea turtle cells (LC50=271 uM) that human cells (LC50=471 uM), but its clastogenicity was similar between the two species. Thus, adjusting for differences in uptake indicated the explanation for the difference in potency was mostly due to uptake rather than differently affected mechanisms. Overall these data indicate sea turtles may be a useful sentinel for human health responses to marine pollution. PMID:26440299

  6. Soluble heat shock protein 70 members in patients undergoing allogeneic hematopoietic cell transplantation.

    Science.gov (United States)

    Mirza, Nora; Prokop, Lea; Kowalewski, Daniel; Gouttefangeas, Cécile; Faul, Christoph; Bethge, Wolfgang A; Vogel, Wichard; Kanz, Lothar; Rammensee, Hans-Georg; Haen, Sebastian P

    2016-05-01

    Heat shock proteins (HSP) are highly conserved immunogenic proteins serving as potent danger signals. They are upregulated under stress conditions like fever and hypoxia. Extracellular HSP are involved in antigen presentation, cytokine release and maturation of antigen presenting cells. The release of the inducible members of the HSP70 family, Hsp72 and Hsp70B', into the serum of 20 patients undergoing allogeneic hematopoietic cell transplantation and 20 healthy donors was evaluated using enzyme linked immunosorbent assay (ELISA) kits. Eight patients (40%) did not receive anti-thymocyte globulin (ATG) for prophylaxis of graft versus host disease (GvHD). These patients had no detectable or low serum levels of Hsp72 (n=3, 0.03 to 1.92ng/ml) which were in line with levels detected in 20 healthy individuals (p=0.07). Measurable HSP was not associated with any medication or transplantation-related procedures. In twelve patients (60%) receiving ATG, detected high levels of HSP reflected cross-reactivity of the rabbit-derived ATG with the anti-rabbit antibody used in the ELISA. Assumed HSP70 expression detected such ELISA has to be regarded carefully after ATG application. Neither radiochemotherapy, nor inflammation or sepsis during aplasia induced HSP70 release into the serum. Thus, soluble HSP70 may not be involved in the pathogenesis of acute GvHD. Copyright © 2016 Elsevier B.V. All rights reserved.

  7. Differential pattern of cell-surface and soluble TREM-1 between sepsis and SIRS.

    Science.gov (United States)

    Oku, Reiko; Oda, Shigeto; Nakada, Taka-aki; Sadahiro, Tomohito; Nakamura, Masataka; Hirayama, Yoh; Abe, Ryuzo; Tateishi, Yoshihisa; Ito, Michihiro; Iseki, Toru; Hirasawa, Hiroyuki

    2013-01-01

    Triggering receptor expressed on myeloid cells-1 (TREM-1) was reported to play a key roll in amplification of production of inflammatory cytokines. TREM-1 is suggested to be a specific biomarker for sepsis for this reason, but the clinical significance of TREM-1 has not been elucidated. We investigated TREM-1 expression on the cell-surface, and plasma levels of soluble TREM-1 (sTREM-1) in patients with non-infectious systemic inflammatory response syndrome (SIRS) and sepsis admitted to the ICU. Thirty-five patients with SIRS and 21 patients with sepsis admitted to ICU were subjected to the study. TREM-1 expressions on the surfaces of monocytes and neutrophils were measured by flow cytometry. Plasma sTREM-1 level and serum interleukin (IL)-6 level were measured. Septic patients had decreased TREM-1 expression, clearly on neutrophils or to a lesser extent on monocyte compared to SIRS patients on ICU admission (neutrophils pSIRS patients (pSIRS/sepsis patients (pSIRS patients. sTREM-1 may be useful to evaluate disease severity and outcome of patients with SIRS or sepsis. Copyright © 2012 Elsevier Ltd. All rights reserved.

  8. Soluble Triggering Receptor Expressed on Myeloid Cells-1 as a Novel Marker for Abdominal Sepsis.

    Science.gov (United States)

    Song, Xiaofei; Song, Yucheng; Zhang, Xuedong; Xue, Huanzhou

    2017-07-01

    The aim of the study was to investigate the concentration and diagnostic significance of soluble triggering receptor expressed on myeloid cells-1 (sTREM-1) in acute abdominal conditions. Plasma specimens were obtained from 68 patients with abdominal sepsis, 60 patients with systemic inflammatory response syndrome (SIRS), and 60 healthy individuals. The sepsis group was divided into the survival and death groups according to the 28-d outcome. Plasma sTREM-1, procalcitonin (PCT), C-reactive protein (CRP), and white blood cell (WBC) count were measured. A receiver operating characteristic curve (ROC) was used to compare the diagnostic values of sTREM-1, PCT, CRP, and WBC count. In addition, the correlation between plasma sTREM-1 and the Acute Physiology and Chronic Health Evaluation (APACHE) II score in the sepsis group was assessed by Spearman correlation analysis. The plasma concentration of sTREM-1 in the sepsis group was significantly higher than that in the SIRS and healthy groups (both p sepsis vs. SIRS showed that the area under the curve of sTREM-1 (0.82) was greater than that of PCT (0.77), CRP (0.72), and WBC count (0.70). Additionally, in the sepsis group, the plasma sTREM-1 concentration correlated positively with the APACHE II score (r = 0.41; p sepsis.

  9. Toxic and transcriptional responses of PC12 cells to soluble tungsten alloy surrogates

    Directory of Open Access Journals (Sweden)

    V.H. Adams

    2015-01-01

    Full Text Available There is increasing evidence that metals have a role in the etiology of diverse neurological diseases. This study used PC12 cells as an in vitro model to examine the toxicity of tungsten alloys that have important military applications. Initially, the relative concentrations of tungsten (W, nickel (Ni, and cobalt (Co mobilized from pellets of a weapons-grade tungsten alloy incubated in physiologically relevant solutions were determined. Dosing solutions of soluble metal salts that were equivalent in ratio to those mobilized from these alloy pellets were used to treat nerve growth factor (NGF differentiated PC12 cells. Treatments consisted of single (W, Ni or Co, paired (W/Ni, W/Co or Ni/Co or complete (W/Ni/Co metal exposures for 24 h followed by measurement of cytotoxicity, viability, and microarray analysis to examine their impact on survival and viability, global gene expression, and biological processes. Gene expression changed dramatically with addition of NGF. Addition of Ni or Co either singly or in combination further impacted gene expression. An observed additive effect of Ni and Co on gene expression was unaffected by the addition of W. The work showed that tungsten, as found in this tungsten alloy, had minimal relative toxicity as compared to the other alloy components when used either alone or in combination.

  10. Fibroblast growth factor 23 inhibits osteoblastic gene expression and induces osteoprotegerin in vascular smooth muscle cells.

    Science.gov (United States)

    Nakahara, Takehiro; Kawai-Kowase, Keiko; Matsui, Hiroki; Sunaga, Hiroaki; Utsugi, Toshihiro; Iso, Tatsuya; Arai, Masashi; Tomono, Shouichi; Kurabayashi, Masahiko

    2016-10-01

    Elevated fibroblast growth factor 23 (FGF23) levels are associated with cardiovascular mortality in patients with chronic kidney disease. However, both clinical and basic research have demonstrated conflicting evidence regarding the pathophysiological role of FGF23 in vascular calcification. The aim of this study was to determine the role of FGF23 in the osteoblastic gene expression in vascular smooth muscle cells (SMCs). We transduce human aortic SMCs (HASMCs) expressing klotho and FGF receptors with the adenovirus expressing human FGF23 (Ad-FGF23). We observed significant decreases in the expression of osteoblast-marker genes including BMP2, BMP4, MSX2, RUNX2 and ALP, as well as reduced calcification. Notably, Ad-FGF23 increased mRNA and protein levels of osteoprotegerin (OPG), and human OPG promoter was activated by FGF23. Moreover, in HASMCs overexpressing klotho, FGF23 upregulated OPG expression, whereas depletion of klotho by siRNA attenuated FGF23-induced OPG expression. Furthermore, in 73 consecutive patients with type 2 diabetes mellitus undergoing cardiac computed tomography to determine coronary calcium scores (CCSs), serum FGF23 levels were positively correlated with OPG independent of phosphate and estimated glomerular filtration rate (eGFR, r = 0.65, p < 0.01). Serum FGF23 levels were significantly elevated in patients with high CCSs (≧100) compared to those with low CCSs (<100). Our in vitro results indicate that FGF23 suppresses osteoblastic gene expression and induces OPG expression in HASMCs. Together with our cross-sectional clinical assessment, the present study lends support to our hypothesis that FGF23 counteracts osteogenic conversion of vascular SMCs as a part of a compensatory mechanism to mitigate vascular calcification. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

  11. Basigin mediates pulmonary hypertension by promoting inflammation and vascular smooth muscle cell proliferation.

    Science.gov (United States)

    Satoh, Kimio; Satoh, Taijyu; Kikuchi, Nobuhiro; Omura, Junichi; Kurosawa, Ryo; Suzuki, Kota; Sugimura, Koichiro; Aoki, Tatsuo; Nochioka, Kotaro; Tatebe, Shunsuke; Miyamichi-Yamamoto, Saori; Miura, Masanobu; Shimizu, Toru; Ikeda, Shohei; Yaoita, Nobuhiro; Fukumoto, Yoshihiro; Minami, Tatsuro; Miyata, Satoshi; Nakamura, Kazufumi; Ito, Hiroshi; Kadomatsu, Kenji; Shimokawa, Hiroaki

    2014-09-26

    Cyclophilin A (CyPA) is secreted from vascular smooth muscle cells (VSMCs) by oxidative stress and promotes VSMC proliferation. However, the role of extracellular CyPA and its receptor Basigin (Bsg, encoded by Bsg) in the pathogenesis of pulmonary hypertension (PH) remains to be elucidated. To determine the role of CyPA/Bsg signaling in the development of PH. In the pulmonary arteries of patients with PH, immunostaining revealed strong expression of CyPA and Bsg. The pulmonary arteries of CyPA(±) and Bsg(±) mice exposed to normoxia did not differ in morphology compared with their littermate controls. In contrast, CyPA(±) and Bsg(±) mice exposed to hypoxia for 4 weeks revealed significantly reduced right ventricular systolic pressure, pulmonary artery remodeling, and right ventricular hypertrophy compared with their littermate controls. These features were unaltered by bone marrow reconstitution. To further evaluate the role of vascular Bsg, we harvested pulmonary VSMCs from Bsg(+/+) and Bsg(±) mice. Proliferation was significantly reduced in Bsg(±) compared with Bsg(+/+) VSMCs. Mechanistic studies demonstrated that Bsg(±) VSMCs revealed reduced extracellular signal-regulated kinase 1/2 activation and less secretion of cytokines/chemokines and growth factors (eg, platelet-derived growth factor-BB). Finally, in the clinical study, plasma CyPA levels in patients with PH were increased in accordance with the severity of pulmonary vascular resistance. Furthermore, event-free curve revealed that high plasma CyPA levels predicted poor outcome in patients with PH. These results indicate the crucial role of extracellular CyPA and vascular Bsg in the pathogenesis of PH. © 2014 American Heart Association, Inc.

  12. 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

  13. [Effect of paeonol on LPS-induced rat vascular endothelial cell adhesion reaction].

    Science.gov (United States)

    Chen, Jun-Jun; Dai, Min; Chen, Peng

    2013-03-01

    To observe the effect of Paeonol (Pae) on lipopolysaccharide (LPS) induced rat mononuclear cells (MCs) adhesion to vascular endothelial cells (VECs) and provide basis foundation for inflammatary mechanisms of Pae against atherosclerosis. Rat vascular endothelial cells were isolated with tissue predigested adherent method. LPS was used as stimulator to induce VEC injury. Serum containing Pae obtained from healthy rats which were given Pae in intragastric. RP-HPLC method was used for detecting the concentration of Pae in serum. MTT assay was used to determine the protective effect of Pae on injured VECs. Rose Bengal Staining was used to detect the effect of Pae on LPS-induced MCs adhesion to VECs. LPS induced rat MCs adhesion to VECs. The effect was the strongest when the concentration was 10 ng/mL and incubated with VECs for 5 h. Pae in concentration of 2. 5,5 and 10 microg/mL and incubated for 24 h could effectively inhibit the adhesion and improve the survival rate of LPS injured VECs significantly. LPS can damage VECs. Pae could protect VECs from LPS injury via inhibiting MCs adhesion to VECs and improving the VEC survival rate.

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

    International Nuclear Information System (INIS)

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

    2007-01-01

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

  15. Brain Tumor Tropism of Transplanted Human Neural Stem Cells Is Induced by Vascular Endothelial Growth Factor

    Directory of Open Access Journals (Sweden)

    Nils Ole Schmidt

    2005-06-01

    Full Text Available The transplantation of neural stem cells (NSCs offers a new potential therapeutic approach as a cell-based delivery system for gene therapy in brain tumors. This is based on the unique capacity of NSCs to migrate throughout the brain and to target invading tumor cells. However, the signals controlling the targeted migration of transplanted NSCs are poorly defined. We analyzed the in vitro and in vivo effects of angiogenic growth factors and protein extracts from surgical specimens of brain tumor patients on NSC migration. Here, we demonstrate that vascular endothelial growth factor (VEGF is able to induce a long-range attraction of transplanted human NSCs from distant sites in the adult brain. Our results indicate that tumorupregulated VEGF and angiogenic-activated microvasculature are relevant guidance signals for NSC tropism toward brain tumors.

  16. Quantitation of soluble aggregates in recombinant monoclonal antibody cell culture by pH-gradient protein A chromatography.

    Science.gov (United States)

    Pan, Hai; Chen, Ken; Pulisic, Matt; Apostol, Izydor; Huang, Gang

    2009-05-15

    Monoclonal antibodies (mAbs) produced from mammalian cell culture may contain significant amounts of dimers and higher order aggregates. Quantitation of soluble aggregates in the cell culture is time-consuming and labor-intensive, usually involving a purification step to remove the impurities that interfere with the subsequent size exclusion chromatography (SEC) analysis. We have developed a novel pH-gradient protein A chromatography for rapid, non-size based separation of the aggregates in mAb cell culture samples. Our results demonstrate that this method has excellent correlation with SEC and can be applied to both human immunoglobulin gamma 1 (IgG1) and IgG2 antibodies. This approach can be useful in the quantitation of soluble aggregates in crude cell culture samples.

  17. Unique cell type-specific junctional complexes in vascular endothelium of human and rat liver sinusoids.

    Directory of Open Access Journals (Sweden)

    Cyrill Géraud

    Full Text Available Liver sinusoidal endothelium is strategically positioned to control access of fluids, macromolecules and cells to the liver parenchyma and to serve clearance functions upstream of the hepatocytes. While clearance of macromolecular debris from the peripheral blood is performed by liver sinusoidal endothelial cells (LSECs using a delicate endocytic receptor system featuring stabilin-1 and -2, the mannose receptor and CD32b, vascular permeability and cell trafficking are controlled by transcellular pores, i.e. the fenestrae, and by intercellular junctional complexes. In contrast to blood vascular and lymphatic endothelial cells in other organs, the junctional complexes of LSECs have not yet been consistently characterized in molecular terms. In a comprehensive analysis, we here show that LSECs express the typical proteins found in endothelial adherens junctions (AJ, i.e. VE-cadherin as well as α-, β-, p120-catenin and plakoglobin. Tight junction (TJ transmembrane proteins typical of endothelial cells, i.e. claudin-5 and occludin, were not expressed by rat LSECs while heterogenous immunreactivity for claudin-5 was detected in human LSECs. In contrast, junctional molecules preferentially associating with TJ such as JAM-A, B and C and zonula occludens proteins ZO-1 and ZO-2 were readily detected in LSECs. Remarkably, among the JAMs JAM-C was considerably over-expressed in LSECs as compared to lung microvascular endothelial cells. In conclusion, we show here that LSECs form a special kind of mixed-type intercellular junctions characterized by co-occurrence of endothelial AJ proteins, and of ZO-1 and -2, and JAMs. The distinct molecular architecture of the intercellular junctional complexes of LSECs corroborates previous ultrastructural findings and provides the molecular basis for further analyses of the endothelial barrier function of liver sinusoids under pathologic conditions ranging from hepatic inflammation to formation of liver metastasis.

  18. Magnesium reduces calcification in bovine vascular smooth muscle cells in a dose-dependent manner

    Science.gov (United States)

    Peter, Mirjam E.; Sevinc Ok, Ebru; Celenk, Fatma Gul; Yilmaz, Mumtaz; Steppan, Sonja; Asci, Gulay; Ok, Ercan; Passlick-Deetjen, Jutta

    2012-01-01

    Background. Vascular calcification (VC), mainly due to elevated phosphate levels, is one major problem in patients suffering from chronic kidney disease. In clinical studies, an inverse relationship between serum magnesium and VC has been reported. However, there is only few information about the influence of magnesium on calcification on a cellular level available. Therefore, we investigated the effect of magnesium on calcification induced by β-glycerophosphate (BGP) in bovine vascular smooth muscle cells (BVSMCs). Methods. BVSMCs were incubated with calcification media for 14 days while simultaneously increasing the magnesium concentration. Calcium deposition, transdifferentiation of cells and apoptosis were measured applying quantification of calcium, von Kossa and Alizarin red staining, real-time reverse transcription–polymerase chain reaction and annexin V staining, respectively. Results. Calcium deposition in the cells dramatically increased with addition of BGP and could be mostly prevented by co-incubation with magnesium. Higher magnesium levels led to inhibition of BGP-induced alkaline phosphatase activity as well as to a decreased expression of genes associated with the process of transdifferentiation of BVSMCs into osteoblast-like cells. Furthermore, estimated calcium entry into the cells decreased with increasing magnesium concentrations in the media. In addition, higher magnesium concentrations prevented cell damage (apoptosis) induced by BGP as well as progression of already established calcification. Conclusions. Higher magnesium levels prevented BVSMC calcification, inhibited expression of osteogenic proteins, apoptosis and further progression of already established calcification. Thus, magnesium is influencing molecular processes associated with VC and may have the potential to play a role for VC also in clinical situations. PMID:21750166

  19. A soluble form of Siglec-9 provides an antitumor benefit against mammary tumor cells expressing MUC1 in transgenic mice

    International Nuclear Information System (INIS)

    Tomioka, Yukiko; Morimatsu, Masami; Nishijima, Ken-ichi; Usui, Tatsufumi; Yamamoto, Sayo; Suyama, Haruka; Ozaki, Kinuyo; Ito, Toshihiro

    2014-01-01

    Highlights: • Tumor-associated antigen MUC1 binds to Siglec-9. • Soluble Siglec-9 reduced proliferation of MUC1-positive tumor in transgenic mice. • Soluble Siglec-9 and MUC1 on tumor cells were colocalized in transgenic mice. • MUC1 expression on tumor cells were reduced in soluble Siglec-9 transgenic mice. - Abstract: Tumor-associated MUC1 binds to Siglec-9, which is expected to mediate tumor cell growth and negative immunomodulation. We hypothesized that a soluble form of Siglec-9 (sSiglec-9) competitively inhibits a binding of MUC1 to its receptor molecules like human Siglec-9, leading to provide antitumor benefit against MUC1-expressing tumor, and generated transgenic mouse lines expressing sSiglec-9 (sSiglec-9 Tg). When mammary tumor cells expressing MUC1 were intraperitoneally transplanted into sSiglec-9 Tg, tumor proliferation was slower with the lower histological malignancy as compared with non-transgenic mice. The sSiglec-9 was detected in the ascites caused by the tumor in the sSiglec-9 Tg, and sSiglec-9 and MUC1 were often colocalized on surfaces of the tumor cells. PCNA immunohistochemistry also revealed the reduced proliferation of the tumor cells in sSiglec-9 Tg. In sSiglec-9 Tg with remarkable suppression of tumor proliferation, MUC1 expressions were tend to be reduced. In the ascites of sSiglec-9 Tg bearing the tumor, T cells were uniformly infiltrated, whereas aggregations of degenerative T cells were often observed in the non-transgenic mice. These results suggest that sSiglec-9 has an antitumor benefit against MUC1-expressing tumor in the transgenic mice, which may avoid the negative immunomodulation and/or suppress tumor-associated MUC1 downstream signal transduction, and subsequent tumor proliferation

  20. Soluble CD163, a marker of Kupffer cell activation, is related to portal hypertension in patients with liver cirrhosis

    DEFF Research Database (Denmark)

    Grønbaek, H; Sandahl, T D; Mortensen, C

    2012-01-01

    BACKGROUND: Activation of Kupffer cells may be involved in the pathogenesis of portal hypertension by release of vasoconstrictive substances and fibrosis due to co-activation of hepatic stellate cells. AIM: To study soluble plasma (s) CD163, a specific marker of activated macrophages...... for HVPG. These findings support a primary role of macrophage activation in portal hypertension, and may indicate a target for biological intervention....

  1. A soluble form of Siglec-9 provides an antitumor benefit against mammary tumor cells expressing MUC1 in transgenic mice

    Energy Technology Data Exchange (ETDEWEB)

    Tomioka, Yukiko, E-mail: ytomi@muses.tottori-u.ac.jp [Division of Disease Model Innovation, Institute for Genetic Medicine, Hokkaido University, Sapporo 060-0815 (Japan); Avian Zoonosis Research Center, Faculty of Agriculture, Tottori University, Tottori 680-8553 (Japan); Morimatsu, Masami, E-mail: mmorimat@vetmed.hokudai.ac.jp [Division of Disease Model Innovation, Institute for Genetic Medicine, Hokkaido University, Sapporo 060-0815 (Japan); Laboratory of Laboratory Animal Science and Medicine, Department of Disease Control, Graduate School of Veterinary Medicine, Hokkaido University, Sapporo 060-0818 (Japan); Nishijima, Ken-ichi, E-mail: nishijma@nubio.nagoya-u.ac.jp [Department of Biotechnology, Graduate School of Engineering, Nagoya University, Nagoya 464-8603 (Japan); Usui, Tatsufumi, E-mail: usutatsu@muses.tottori-u.ac.jp [Avian Zoonosis Research Center, Faculty of Agriculture, Tottori University, Tottori 680-8553 (Japan); Yamamoto, Sayo, E-mail: ysayo@anim.med.kyushu-u.ac.jp [Center of Biomedical Research, Research Center for Human Disease Modeling, Graduate School of Medical Sciences, Kyushu University, Fukuoka 812-8582 (Japan); Suyama, Haruka, E-mail: sharuka@anim.med.kyushu-u.ac.jp [Center of Biomedical Research, Research Center for Human Disease Modeling, Graduate School of Medical Sciences, Kyushu University, Fukuoka 812-8582 (Japan); Ozaki, Kinuyo, E-mail: k-ozaki@anim.med.kyushu-u.ac.jp [Center of Biomedical Research, Research Center for Human Disease Modeling, Graduate School of Medical Sciences, Kyushu University, Fukuoka 812-8582 (Japan); Ito, Toshihiro, E-mail: toshiito@muses.tottori-u.ac.jp [Avian Zoonosis Research Center, Faculty of Agriculture, Tottori University, Tottori 680-8553 (Japan); and others

    2014-07-18

    Highlights: • Tumor-associated antigen MUC1 binds to Siglec-9. • Soluble Siglec-9 reduced proliferation of MUC1-positive tumor in transgenic mice. • Soluble Siglec-9 and MUC1 on tumor cells were colocalized in transgenic mice. • MUC1 expression on tumor cells were reduced in soluble Siglec-9 transgenic mice. - Abstract: Tumor-associated MUC1 binds to Siglec-9, which is expected to mediate tumor cell growth and negative immunomodulation. We hypothesized that a soluble form of Siglec-9 (sSiglec-9) competitively inhibits a binding of MUC1 to its receptor molecules like human Siglec-9, leading to provide antitumor benefit against MUC1-expressing tumor, and generated transgenic mouse lines expressing sSiglec-9 (sSiglec-9 Tg). When mammary tumor cells expressing MUC1 were intraperitoneally transplanted into sSiglec-9 Tg, tumor proliferation was slower with the lower histological malignancy as compared with non-transgenic mice. The sSiglec-9 was detected in the ascites caused by the tumor in the sSiglec-9 Tg, and sSiglec-9 and MUC1 were often colocalized on surfaces of the tumor cells. PCNA immunohistochemistry also revealed the reduced proliferation of the tumor cells in sSiglec-9 Tg. In sSiglec-9 Tg with remarkable suppression of tumor proliferation, MUC1 expressions were tend to be reduced. In the ascites of sSiglec-9 Tg bearing the tumor, T cells were uniformly infiltrated, whereas aggregations of degenerative T cells were often observed in the non-transgenic mice. These results suggest that sSiglec-9 has an antitumor benefit against MUC1-expressing tumor in the transgenic mice, which may avoid the negative immunomodulation and/or suppress tumor-associated MUC1 downstream signal transduction, and subsequent tumor proliferation.

  2. Fluorine-19 Labeling of Stromal Vascular Fraction Cells for Clinical Imaging Applications.

    Science.gov (United States)

    Rose, Laura C; Kadayakkara, Deepak K; Wang, Guan; Bar-Shir, Amnon; Helfer, Brooke M; O'Hanlon, Charles F; Kraitchman, Dara L; Rodriguez, Ricardo L; Bulte, Jeff W M

    2015-12-01

    Stromal vascular fraction (SVF) cells are used clinically for various therapeutic targets. The location and persistence of engrafted SVF cells are important parameters for determining treatment failure versus success. We used the GID SVF-1 platform and a clinical protocol to harvest and label SVF cells with the fluorinated ((19)F) agent CS-1000 as part of a first-in-human phase I trial (clinicaltrials.gov identifier NCT02035085) to track SVF cells with magnetic resonance imaging during treatment of radiation-induced fibrosis in breast cancer patients. Flow cytometry revealed that SVF cells consisted of 25.0% ± 15.8% CD45+, 24.6% ± 12.5% CD34+, and 7.5% ± 3.3% CD31+ cells, with 2.1 ± 0.7 × 10⁵ cells per cubic centimeter of adipose tissue obtained. Fluorescent CS-1000 (CS-ATM DM Green) labeled 87.0% ± 13.5% of CD34+ progenitor cells compared with 47.8% ± 18.5% of hematopoietic CD45+ cells, with an average of 2.8 ± 2.0 × 10¹² ¹⁹F atoms per cell, determined using nuclear magnetic resonance spectroscopy. The vast majority (92.7% ± 5.0%) of CD31+ cells were also labeled, although most coexpressed CD34. Only 16% ± 22.3% of CD45-/CD31-/CD34- (triple-negative) cells were labeled with CS-ATM DM Green. After induction of cell death by either apoptosis or necrosis, >95% of ¹⁹F was released from the cells, indicating that fluorine retention can be used as a surrogate marker for cell survival. Labeled-SVF cells engrafted in a silicone breast phantom could be visualized with a clinical 3-Tesla magnetic resonance imaging scanner at a sensitivity of approximately 2 × 10⁶ cells at a depth of 5 mm. The current protocol can be used to image transplanted SVF cells at clinically relevant cell concentrations in patients. Stromal vascular fraction (SVF) cells harvested from adipose tissue offer great promise in regenerative medicine, but methods to track such cell therapies are needed to ensure correct administration and monitor survival. A clinical protocol was

  3. In Vivo FRET Imaging of Tumor Endothelial Cells Highlights a Role of Low PKA Activity in Vascular Hyperpermeability.

    Science.gov (United States)

    Yamauchi, Fumio; Kamioka, Yuji; Yano, Tetsuya; Matsuda, Michiyuki

    2016-09-15

    Vascular hyperpermeability is a pathological hallmark of cancer. Previous in vitro studies have elucidated roles of various signaling molecules in vascular hyperpermeability; however, the activities of such signaling molecules have not been examined in live tumor tissues for technical reasons. Here, by in vivo two-photon excitation microscopy with transgenic mice expressing biosensors based on Förster resonance energy transfer, we examined the activity of protein kinase A (PKA), which maintains endothelial barrier function. The level of PKA activity was significantly lower in the intratumoral endothelial cells than the subcutaneous endothelial cells. PKA activation with a cAMP analogue alleviated the tumor vascular hyperpermeability, suggesting that the low PKA activity in the endothelial cells may be responsible for the tumor-tissue hyperpermeability. Because the vascular endothelial growth factor (VEGF) receptor is a canonical inducer of vascular hyperpermeability and a molecular target of anticancer drugs, we examined the causality between VEGF receptor activity and the PKA activity. Motesanib, a kinase inhibitor for VEGF receptor, activated tumor endothelial PKA and reduced the vascular permeability in the tumor. Conversely, subcutaneous injection of VEGF decreased endothelial PKA activity and induced hyperpermeability of subcutaneous blood vessels. Notably, in cultured human umbilical vascular endothelial cells, VEGF activated PKA rather than decreasing its activity, highlighting the remarkable difference between its actions in vitro and in vivo These data suggested that the VEGF receptor signaling pathway increases vascular permeability, at least in part, by reducing endothelial PKA activity in the live tumor tissue. Cancer Res; 76(18); 5266-76. ©2016 AACR. ©2016 American Association for Cancer Research.

  4. Erythropoietin Attenuates Pulmonary Vascular Remodeling in Experimental Pulmonary Arterial Hypertension through Interplay between Endothelial Progenitor Cells and Heme Oxygenase

    OpenAIRE

    van Loon, Rosa Laura E; Bartelds, Beatrijs; Wagener, Frank A D T G; Affara, Nada; Mohaupt, Saffloer; Wijnberg, Hans; Pennings, Sebastiaan W C; Takens, Janny; Berger, Rolf M F

    2015-01-01

    BACKGROUND: Pulmonary arterial hypertension (PAH) is a pulmonary vascular disease with a high mortality, characterized by typical angio-proliferative lesions. Erythropoietin (EPO) attenuates pulmonary vascular remodeling in PAH. We postulated that EPO acts through mobilization of endothelial progenitor cells (EPCs) and activation of the cytoprotective enzyme heme oxygenase-1 (HO-1). METHODS: Rats with flow-associated PAH, resembling pediatric PAH, were treated with HO-1 inducer EPO in the pre...

  5. Erythropoietin Attenuates Pulmonary Vascular Remodeling in Experimental Pulmonary Arterial Hypertension through Interplay between Endothelial Progenitor Cells and Heme Oxygenase

    OpenAIRE

    van Loon, Rosa Laura E.; Bartelds, Beatrijs; Wagener, Frank A. D. T. G.; Affara, Nada; Mohaupt, Saffloer; Wijnberg, Hans; Pennings, Sebastiaan W. C.; Takens, Janny; Berger, Rolf M. F.

    2015-01-01

    Background Pulmonary arterial hypertension (PAH) is a pulmonary vascular disease with a high mortality, characterized by typical angio-proliferative lesions. Erythropoietin (EPO) attenuates pulmonary vascular remodeling in PAH. We postulated that EPO acts through mobilization of endothelial progenitor cells (EPCs) and activation of the cytoprotective enzyme heme oxygenase-1 (HO-1). Methods Rats with flow-associated PAH, resembling pediatric PAH, were treated with HO-1 inducer EPO i...

  6. Estradiol-mediated ERK phosphorylation and apoptosis in vascular smooth muscle cells requires GPR 30.

    Science.gov (United States)

    Ding, Qingming; Gros, Robert; Limbird, Lee E; Chorazyczewski, Jozef; Feldman, Ross D

    2009-11-01

    Recent studies suggest that the rapid and nongenomic effects of estradiol may be mediated through the G protein-coupled receptor dubbed GPR30 receptor. The present study examines the role of GPR30 versus a classical estrogen receptor (ERalpha) in mediating the growth regulatory effects of estradiol. GPR30 is readily detectable in freshly isolated vascular tissue but barely detectable in cultured vascular smooth muscle cells (VSMC). In freshly isolated aortic tissue, estradiol stimulated extracellular signal-regulated kinases (ERK) phosphorylation. In contrast, in cultured VSMC, where GPR30 expression is significantly reduced, estradiol inhibits ERK phosphorylation. Transfer of the genes encoding GPR30 led to estradiol stimulation of ERK phosphorylation, which is opposite the effects of estradiol in the primary culture of VSMCs. Transduction of the mineralocorticoid receptor (MR) had no effect on estradiol effects on ERK. Estradiol-mediated stimulation of ERK subsequent to heterologous GPR30 expression was pertussis toxin sensitive and phosphoinositide 3-kinase (PI3 kinase) dependent; under these conditions, estradiol also inhibited protein kinase A (PKA). In contrast, in the absence of GPR30 expression in cultured VSMC, estradiol stimulated PKA activity and inhibited ERK phosphorylation. To determine the functional effect of GPR30 (vs. estrogen receptor expression), we assessed estradiol-mediated apoptosis. In the absence of GPR30 expression, estradiol inhibited apoptosis. This effect was enhanced with ERalpha expression. In contrast, with GPR30 expression, estradiol stimulated apoptosis in an ERK-dependent manner. Thus the effect of estradiol on vascular smooth muscle cell apoptosis is likely dependent on the balance between ER-mediated PKA activation and GPR30-mediated PKA inhibition and PI3 kinase activation. Taken together, we postulate that modulation of GPR30 expression or activity may be an important determinant of the effects of estradiol in the vasculature.

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

    International Nuclear Information System (INIS)

    Moon, Chang Yoon; Ku, Cheol Ryong; Cho, Yoon Hee; Lee, Eun Jig

    2012-01-01

    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.

  8. 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.

  9. Modulating the vascular behavior of metastatic breast cancer cells by curcumin treatment

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    Anna Lisa ePalange

    2012-11-01

    Full Text Available The spreading of tumor cells to secondary sites (tumor metastasis is a complex process that involves multiple, sequential steps. Vascular adhesion and extravasation of circulating tumor cells (CTCs is one, critical step. Curcumin, a natural compound extracted from Curcuma longa, is known to have anti-tumoral, anti-proliferative, anti-inflammatory properties and affect the expression of cell adhesion molecules, mostly by targeting the NF-κB transcription factor. Here, upon treatment with Curcumin, the vascular behavior of three different estrogen receptor negative (ER– breast adenocarcinoma cell lines (SK-BR-3, MDA-MB-231, MDA-MB-468 is analyzed using a microfluidic system. First, the dose response to curcumin is characterized at 24, 48 and 72h using a XTT assay. For all three cell lines, an IC50 larger than 20 µM is observed at 72 h; whereas no significant reduction in cell viability is detected for curcumin concentrations up to 10 µM. Upon 24 h treatment at 10 µM of curcumin, SK-BR3 and MDA-MB-231 cells show a decrease in adhesion propensity of 40% (p = 0.02 and 47% (p = 0.001, respectively. No significant change is documented for the less metastatic MDA-MB-468 cells. All three treated cell lines show a 20% increase in rolling velocity from 48.3 to 58.7 µm/s in SK-BR-3, from 64.1 to 73.77 µm/s in MDA-MB-231 and from 57.5 to 74.4 µm/s in MDA-MB-468. Collectively, these results suggest that mild curcumin treatments could limit the metastatic potential of these adenocarcinoma cell lines, possibly by altering the expression of adhesion molecules, and the organization and stiffness of the cell cytoskeleton. Future studies will elucidate the biophysical mechanisms regulating this curcumin-induced behavior and further explore the clinical relevance of these findings.

  10. Modulating the vascular behavior of metastatic breast cancer cells by curcumin treatment

    International Nuclear Information System (INIS)

    Palange, Anna L.; Mascolo, Daniele Di; Singh, Jaykrishna; Franceschi, Maria S. De; Carallo, Claudio; Gnasso, Agostino; Decuzzi, Paolo

    2012-01-01

    The spreading of tumor cells to secondary sites (tumor metastasis) is a complex process that involves multiple, sequential steps. Vascular adhesion and extravasation of circulating tumor cells (CTCs) is one, critical step. Curcumin, a natural compound extracted from Curcuma longa, is known to have anti-tumoral, anti-proliferative, anti-inflammatory properties and affect the expression of cell adhesion molecules, mostly by targeting the NF-κB transcription factor. Here, upon treatment with curcumin, the vascular behavior of three different estrogen receptor negative (ER – ) breast adenocarcinoma cell lines (SK-BR-3, MDA-MB-231, MDA-MB-468) is analyzed using a microfluidic system. First, the dose response to curcumin is characterized at 24, 48, and 72 h using a XTT assay. For all three cell lines, an IC 50 larger than 20 µM is observed at 72 h; whereas no significant reduction in cell viability is detected for curcumin concentrations up to 10 µM. Upon 24 h treatment at 10 µM of curcumin, SK-BR3 and MDA-MB-231 cells show a decrease in adhesion propensity of 40% (p = 0.02) and 47% (p = 0.001), respectively. No significant change is documented for the less metastatic MDA-MB-468 cells. All three treated cell lines show a 20% increase in rolling velocity from 48.3 to 58.7 µm/s in SK-BR-3, from 64.1 to 73.77 µm/s in MDA-MB-231, and from 57.5 to 74.4 µm/s in MDA-MB-468. Collectively, these results suggest that mild curcumin treatments could limit the metastatic potential of these adenocarcinoma cell lines, possibly by altering the expression of adhesion molecules, and the organization and stiffness of the cell cytoskeleton. Future studies will elucidate the biophysical mechanisms regulating this curcumin-induced behavior and further explore the clinical relevance of these findings.

  11. [Effects on proliferation ability of vascular smooth muscle cells by static and/or dynamic cell culture: utility of pre-seeding technique for dynamic cell culture].

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    Yokomuro, Hiroki; Ozawa, Tsukasa; Fujii, Takeshiro; Shiono, Noritsugu; Watanabe, Yoshinori; Yoshihara, Katsunori; Koyama, Nobuya; Okada, Mitsumasa

    2007-11-01

    Conventional biomaterials are not viable, do not grow, and do not provide contractile effects in cardiac tissue. Foreign synthetic material may become thrombogenic or infected. The most recent cardiac constructs consist of biodegradable material which has the potential to solve these problems. However, dynamic three-dimensional cell culture is necessary because conventional culture is limited to construct tough biografts. Vascular smooth muscle cells derived from rat aorta were seeded to poly-L-lactide-epsilon-capro-lactone copolymer in three groups; static culture group (static cell seeding + static cell culture), dynamic culture group (dynamic cell seeding + dynamic cell culture), and pre-seeding group [static cell seeding and culture for 1 week (pre-seeding) + dynamic cell culture]. The dynamic cell culture system used an original spinner flask. The pre-seeding technique used static cell seeding and culture before dynamic culture. The three groups were evaluated by cell proliferation and histologic studies. Vascular smooth muscle cells could be proliferated in/on the biodegradable materials. The pre-seeding group cells grew much more efficiently than the other groups. Very few cells were found in the biodegradable materials with the dynamic groups. However, there were many cells in the materials with the static culture group and pre-seeding group, especially the pre-seeding group. Dynamic culture is useful for constructing tough biografts by the pre-seeding technique.

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

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    Lin Sheng

    2012-06-01

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

  13. Disturbance of copper homeostasis is a mechanism for homocysteine-induced vascular endothelial cell injury.

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    Daoyin Dong

    Full Text Available Elevation of serum homocysteine (Hcy levels is a risk factor for cardiovascular diseases. Previous studies suggested that Hcy interferes with copper (Cu metabolism in vascular endothelial cells. The present study was undertaken to test the hypothesis that Hcy-induced disturbance of Cu homeostasis leads to endothelial cell injury. Exposure of human umbilical vein endothelial cells (HUVECs to concentrations of Hcy at 0.01, 0.1 or 1 mM resulted in a concentration-dependent decrease in cell viability and an increase in necrotic cell death. Pretreatment of the cells with a final concentration of 5 µM Cu in cultures prevented the effects of Hcy. Hcy decreased intracellular Cu concentrations. HPLC-ICP-MS analysis revealed that Hcy caused alterations in the distribution of intracellular Cu; more Cu was redistributed to low molecular weight fractions. ESI-Q-TOF detected the formation of Cu-Hcy complexes. Hcy also decreased the protein levels of Cu chaperone COX17, which was accompanied by a decrease in the activity of cytochrome c oxidase (CCO and a collapse of mitochondrial membrane potential. These effects of Hcy were all preventable by Cu pretreatment. The study thus demonstrated that Hcy disturbs Cu homeostasis and limits the availability of Cu to critical molecules such as COX17 and CCO, leading to mitochondrial dysfunction and endothelial cell injury.

  14. COX-2 mediates PM2.5-induced apoptosis and inflammation in vascular endothelial cells.

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    Yin, Jie; Xia, Weiwei; Li, Yuanyuan; Guo, Chuchu; Zhang, Yue; Huang, Songming; Jia, Zhanjun; Zhang, Aihua

    2017-01-01

    Emerging evidence demonstrated that particulate matter 2.5 (PM2.5) exposure served as an important risk factor of cardiovascular diseases. Some studies also reported that COX-2/mPGES-1/PGE2 cascade played a pathogenic role in vascular injury. However, the relationship between the PM2.5 exposure and the activation of COX-2/mPGES-1/PGE2 cascade in endothelial cells is still unknown. In the present study, mouse aorta endothelial cells were exposed to PM2.5. Strikingly, following the PM2.5 treatment, we observed dose- and time-dependent upregulation of COX-2 at both protein and mRNA levels as determined by Western blotting and qRT-PCR, respectively. However, COX-1 mRNA expression was not affected by PM2.5 treatment. Next, we examined mPGES-1 expression. As expected, mPGES-1 protein was markedly increased by PM2.5 exposure in line with a significant increment of PGE2 release in medium. At the same time, we observed a dose-dependent upregulation of another two PGE2 synthases of mPGES-2 and cPGES determined by qRT-PCR. Inhibition of COX-2 by using a specific COX-2 inhibitor NS-398 markedly blocked cell apoptosis, inflammation, and PGE2 secretion. Taken together, these results suggested that PM2.5 could activate inflammatory axis of COX-2/PGES/PGE2 in vascular endothelial cells to promote cell apoptosis and inflammatory response.

  15. Manipulating the Plasticity of Smooth Muscle Cells to Regulate Vascular Calcification

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    Kelsey M. McArthur

    2017-11-01

    Full Text Available Cardiovascular complications are one of the leading causes of death in patients with kidney disease or diabetes. Vascular calcification (VC was once considered a passive process resulting from elevated calcium-phosphate interactions, but is now considered an active cell-mediated process. VC can affect quality of life because healthy arteries harden analogously to bone development leading to hypertension and compromised structural integrity. Based on previous literature, the in vitro model was developed by culturing human primary aortic smooth muscle cells with 3-mmol inorganic phosphate (Pi and sodium to induce calcification. The in vitro model was then used to prompt VC and promote the genetic switching from healthy smooth muscle cells to osteoblast-like cells through manipulation of the cells’ plasticity. The in vitro model examined the Wnt signaling pathway in VC and Sclerostin’s ability to block activation of the pathway. Atomic absorption spectroscopy, Western blot, and Polymerase chain reaction (PCR analysis revealed that the model was capable of inducing VC, up-regulating the osteogenic differentiation markers runt-related transcription factor 2 (Runx2 and bone morphogenetic protein 2 (BMP2, and down-regulating α-smooth muscle actin activity. Under the same methods, it was revealed that Sclerostin was capable of recovering α-smooth muscle actin activity in calcification media and able to down-regulate the osteogenic differentiation marker Runx2. This study proved the effectiveness of the in vitro model to induce calcification of healthy vascular smooth muscle cells and Sclerostin’s ability to be used as a potential therapeutic target for VC.

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

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    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.

  17. METACASPASE9 modulates autophagy to confine cell death to the target cells during Arabidopsis vascular xylem differentiation

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    Sacha Escamez

    2016-02-01

    Full Text Available We uncovered that the level of autophagy in plant cells undergoing programmed cell death determines the fate of the surrounding cells. Our approach consisted of using Arabidopsis thaliana cell cultures capable of differentiating into two different cell types: vascular tracheary elements (TEs that undergo programmed cell death (PCD and protoplast autolysis, and parenchymatic non-TEs that remain alive. The TE cell type displayed higher levels of autophagy when expression of the TE-specific METACASPASE9 (MC9 was reduced using RNAi (MC9-RNAi. Misregulation of autophagy in the MC9-RNAi TEs coincided with ectopic death of the non-TEs, implying the existence of an autophagy-dependent intercellular signalling from within the TEs towards the non-TEs. Viability of the non-TEs was restored when AUTOPHAGY2 (ATG2 was downregulated specifically in MC9-RNAi TEs, demonstrating the importance of autophagy in the spatial confinement of cell death. Our results suggest that other eukaryotic cells undergoing PCD might also need to tightly regulate their level of autophagy to avoid detrimental consequences for the surrounding cells.

  18. Mitochondrial fission induced by platelet-derived growth factor regulates vascular smooth muscle cell bioenergetics and cell proliferation

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    Joshua K. Salabei

    2013-01-01

    Full Text Available Vascular smooth muscle cells (VSMCs develop a highly proliferative and synthetic phenotype in arterial diseases. Because such phenotypic changes are likely integrated with the energetic state of the cell, we hypothesized that changes in cellular metabolism regulate VSMC plasticity. VSMCs were exposed to platelet-derived growth factor-BB (PDGF and changes in mitochondrial morphology, proliferation, contractile protein expression, and mitochondrial metabolism were examined. Exposure of VSMCs to PDGF resulted in mitochondrial fragmentation and a 50% decrease in the abundance of mitofusin 2. Synthetic VSMCs demonstrated a 20% decrease in glucose oxidation, which was accompanied by an increase in fatty acid oxidation. Results of mitochondrial function assays in permeabilized cells showed few changes due to PDGF treatment in mitochondrial respiratory chain capacity and coupling. Treatment of VSMCs with Mdivi-1—an inhibitor of mitochondrial fission—inhibited PDGF-induced mitochondrial fragmentation by 50% and abolished increases in cell proliferation; however, it failed to prevent PDGF-mediated activation of autophagy and removal of contractile proteins. In addition, treatment with Mdivi-1 reversed changes in fatty acid and glucose oxidation associated with the synthetic phenotype. These results suggest that changes in mitochondrial morphology and bioenergetics underlie the hyperproliferative features of the synthetic VSMC phenotype, but do not affect the degradation of contractile proteins. Mitochondrial fragmentation occurring during the transition to the synthetic phenotype could be a therapeutic target for hyperproliferative vascular disorders.

  19. Diagnostic value of soluble triggering receptor expressed on myeloid cells in paediatric sepsis: a systematic review.

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    Pontrelli, Giuseppe; De Crescenzo, Franco; Buzzetti, Roberto; Calò Carducci, Francesca; Jenkner, Alessandro; Amodio, Donato; De Luca, Maia; Chiurchiù, Sara; Davies, Elin Haf; Simonetti, Alessandra; Ferretti, Elena; Della Corte, Martina; Gramatica, Luca; Livadiotti, Susanna; Rossi, Paolo

    2016-04-27

    Differential diagnosis between sepsis and non-infectious inflammatory disorders demands improved biomarkers. Soluble Triggering Receptor Expression on Myeloid cells (sTREM-1) is an activating receptor whose role has been studied throughout the last decade. We performed a systematic review to evaluate the accuracy of plasma sTREM-1 levels in the diagnosis of sepsis in children with Systemic Inflammatory Response Syndrome (SIRS). A literature search of PubMed, Cochrane Central Register of Controlled Trials, Cumulative Index to Nursing and Allied Health Literature (CINAHL) and ISI Web of Knowledge databases was performed using specific search terms. Studies were included if they assessed the diagnostic accuracy of plasma sTREM-1 for sepsis in paediatric patients with SIRS. Data on sensitivity, specificity, positive predictive value, negative predictive value, area under receiver operating characteristic curve were extracted. The methodological quality of each study was assessed using a checklist based on the Quality Assessment Tool for Diagnostic Accuracy Studies. Nine studies comprising 961 patients were included, four of which were in newborns, three in children and two in children with febrile neutropenia. Some data from single studies support a role of sTREM-1 as a diagnostic tool in pediatric sepsis, but cannot be considered conclusive, because a quantitative synthesis was not possible, due to heterogeneity in studies design. This systematic review suggests that available data are insufficient to support a role for sTREM in the diagnosis and follow-up of paediatric sepsis.

  20. Telmisartan activates endothelial nitric oxide synthase via Ser1177 phosphorylation in vascular endothelial cells.

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    Masahiro Myojo

    Full Text Available Because endothelial nitric oxide synthase (eNOS has anti-inflammatory and anti-arteriosclerotic functions, it has been recognized as one of the key molecules essential for the homeostatic control of blood vessels other than relaxation of vascular tone. Here, we examined whether telmisartan modulates eNOS function through its pleiotropic effect. Administration of telmisartan to mice significantly increased the phosphorylation level of eNOS (Ser1177 in the aortic endothelium, but administration of valsartan had no effect. Similarly, telmisartan treatment of human umbilical vein endothelial cells significantly increased the phosphorylation levels of AMP-activated protein kinase (Thr172 and eNOS and the concentration of intracellular guanosine 3',5'-cyclic monophosphate (cGMP. Furthermore, pretreatment with a p38 mitogen-activated protein kinase (p38 MAPK inhibitor suppressed the increased phosphorylation level of eNOS and intracellular cGMP concentration. These data show that telmisartan increases eNOS activity through Ser1177 phosphorylation in vascular endothelial cells mainly via p38 MAPK signaling.

  1. Inhibitory effect of curcumin in human endometriosis endometrial cells via downregulation of vascular endothelial growth factor.

    Science.gov (United States)

    Cao, Hong; Wei, Yu-Xi; Zhou, Qi; Zhang, Ying; Guo, Xiao-Peng; Zhang, Jun

    2017-10-01

    Endometriosis, which affects up to 10% of women of reproductive age, is defined as endometrial-like gland and stroma tissue growths outside the uterine cavity. Despite increasing research efforts, there are no current effective treatment methods for this disease, therefore investigations for therapeutic strategies are of primary concern. In preliminary work, the authors demonstrated that curcumin inhibits endometriosis in vivo. The present in vitro study aimed to investigate the association between endometriotic stromal cells and curcumin and to clarify the underlying mechanism of action. A total of 14 patients with endometriosis were enrolled in the present study. The purity of endometrial stromal cell cultures was proven by standard immunofluorescent staining of vimentin. The cell proliferation and curcumin effects on endometrial stromal cells were assessed by the MTT assay and Hematoxylin and Eosin staining. For cell cycle analysis, phase distribution was detected by flow cytometry. Vascular endothelial growth factor (VEGF) protein expression was examined using immunohistochemistry staining. Apoptosis was assessed using Annexin V‑fluorescein isothiocyanate staining. The results indicated that the treatment of curcumin decreased human ectopic and eutopic stromal cell growth. Following treatment with curcumin, human endometriotic stromal cells demonstrated an increased percentage of G1‑phase cells and decreased percentages of S‑phase cells, particularly in the group treated with 50 µmol/l curcumin. Treatment with curcumin additionally decreased expression of VEGF. The data provide evidence that curcumin reduces cell survival in human endometriotic stromal cells, and this may be mediated via downregulation of the VEGF signaling pathway.

  2. The adipose tissue-derived stromal vascular fraction cells from lipedema patients: Are they different?

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    Priglinger, Eleni; Wurzer, Christoph; Steffenhagen, Carolin; Maier, Julia; Hofer, Victoria; Peterbauer, Anja; Nuernberger, Sylvia; Redl, Heinz; Wolbank, Susanne; Sandhofer, Matthias

    2017-07-01

    Lipedema is a hormone-related disease of women characterized by enlargement of the extremities caused by subcutaneous deposition of adipose tissue. In healthy patients application of autologous adipose tissue-derived cells has shown great potential in several clinical studies for engrafting of soft tissue reconstruction in recent decades. The majority of these studies have used the stromal vascular fraction (SVF), a heterogeneous cell population containing adipose-derived stromal/stem cells (ASC), among others. Because cell identity and regenerative properties might be affected by the health condition of patients, we characterized the SVF cells of 30 lipedema patients in comparison to 22 healthy patients. SVF cells were analyzed regarding cell yield, viability, adenosine triphosphate content, colony forming units and proliferative capacity, as well as surface marker profile and differentiation potential in vitro. Our results demonstrated a significantly enhanced SVF cell yield isolated from lipedema compared with healthy patients. In contrast, the adipogenic differentiation potential of SVF cells isolated from lipedema patients was significantly reduced compared with healthy patients. Interestingly, expression of the mesenchymal marker CD90 and the endothelial/pericytic marker CD146 was significantly enhanced when isolated from lipedema patients. The enhanced number of CD90 + and CD146 + cells could explain the increased cell yield because the other tested surface marker were not reduced in lipedema patients. Because the cellular mechanism and composition in lipedema is largely unknown, our findings might contribute to a better understanding of its etiology. Copyright © 2017 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

  3. ANGIOTENSIN II REGULATES ADRENAL VASCULAR TONE THROUGH ZONA GLOMERULOSA CELL-DERIVED EETS AND DHETS

    Science.gov (United States)

    Kopf, Phillip G.; Gauthier, Kathryn M.; Zhang, David X.; Falck, John R.; Campbell, William B.

    2011-01-01

    Elevated concentrations of aldosterone are associated with several cardiovascular diseases. Angiotensin II increases aldosterone secretion and adrenal blood flow. This concurrent increase in steroidogenesis and adrenal blood flow is not understood. We investigated the role of zona glomerulosa cells in the regulation of vascular tone of bovine adrenal cortical arteries by angiotensin II. Zona glomerulosa cells enhance endothelium-dependent relaxations to angiotensin II. The zona glomerulosa cell-dependent relaxations to angiotensin II are unchanged by removing the endothelium-dependent response to angiotensin II. These zona glomerulosa cell-mediated relaxations are ablated by cytochrome P450 inhibition, epoxyeicostrienoic acid antagonism, and potassium channel blockade. Analysis of zona glomerulosa cell epoxyeicosatrienoic acid production by liquid chromatography/mass spectrometry demonstrates an increase in epoxyeicosatrienoic and dihydroxyeicosatrienoic acids with angiotensin II stimulation. These epoxyeicosatrienoic and dihydroxyeicosatrienoic acids produced similar concentration-dependent relaxations of adrenal arteries, which were attenuated by epoxyeicosatrienoic acid antagonism. Whole cell potassium current of adrenal artery smooth muscle cells were increased by angiotensin II stimulation in the presence of zona glomerulosa cells but decreased in the absence of zona glomerulosa cells. This increase in potassium current was abolished by iberiotoxin. Similarly, 14,15-epoxyeicosatrienoic acid induced concentration-dependent increases in potassium current, which was abolished by iberiotoxin. Zona glomerulosa cell aldosterone release is not directly altered by epoxyeicosatrienoic acids. These data suggest that angiotensin II stimulates zona glomerulosa cells to release epoxyeicosatrienoic and dihydroxyeicosatrienoic acids, resulting in potassium channel activation and relaxation of adrenal arteries. This provides a mechanism by which Ang II concurrently increases

  4. Cell-mediated immune responses to Plasmodium falciparum purified soluble antigens in sickle-cell trait subjects.

    Science.gov (United States)

    Bayoumi, R A; Abu-Zeid, Y A; Abdulhadi, N H; Saeed, B O; Theander, T G; Hviid, L; Ghalib, H W; Nugud, A H; Jepsen, S; Jensen, J B

    1990-08-01

    To determine the possible differences in the immune response to Plasmodium falciparum between sickle-cell trait (Hb AS) and normal haemoglobin (Hb AA) individuals, we examined 35 Hb AS and 24 Hb AA subjects matched for age and microenvironment. Their age was 2-55 years and all lived in a malaria endemic area 300 km south of Khartoum. Antibodies to ring-infected erythrocyte surface antigen (Pf155/RESA) and to circumsporozoite (CS) protein (anti-NANP40) indicated equal exposure to falciparum malaria. Peripheral blood mononuclear cells (BMNCs) from 20/35 (57%) Hb AS subjects compared with 10/24 (42%) Hb AA subjects, responded to affinity-purified P. falciparum soluble antigens (SPAg). Of those responding to SPAg, 9 (26%) Hb AS subjects and only two (8%) Hb AA subjects had high responses. The mean proliferative response to SPAg of BMNCs from Hb AS individuals was significantly higher than in Hb AA individuals (P less than 0.025). Responses of BMNCs to PPD and PHA were also higher among Hb AS individuals and correlated positively with responses to SPAg. These findings support the hypotheses that the sickle-cell trait protects individuals from P. falciparum infections, at least in part, by modulating the immune response.

  5. Proteomic profiling of extracellular vesicles released from vascular smooth muscle cells during initiation of phosphate-induced mineralization.

    Science.gov (United States)

    Chaudhary, Sandeep C; Khalid, Sana; Smethurst, Victoria; Monier, Daisy; Mobley, James; Huet, Alexis; Conway, James F; Napierala, Dobrawa

    2018-02-22

    Elevated serum phosphate is one of the major factors contributing to vascular calcification. Studies suggested that extracellular vesicles released from vascular smooth muscle cells significantly contribute to the initiation and progression of this pathology. Recently, we have demonstrated that elevated phosphate stimulates release of extracellular vesicles from osteogenic cells at the initiation of the mineralization process. Here, we used MOVAS cell line as an in vitro model of vascular calcification to examine whether vascular smooth muscle cells respond to high phosphate levels in a similar way and increase formation of extracellular vesicles. Vesicles residing in extracellular matrix as well as vesicles released to culture medium were evaluated by nanoparticle tracking analyses. In addition, using mass spectrometry and protein profiling, protein composition of extracellular vesicles released by MOVAS cells under standard growth conditions and upon exposure to high phosphate was compared. Significant increase of the number of extracellular vesicles was detected after 72 hours of exposure of cells to high phosphate. Elevated phosphate levels also affected protein composition of extracellular vesicles released from MOVAS cells. Finally, the comparative analyses of proteins in extracellular vesicles isolated from extracellular matrix and from conditioned medium identified significant differences in protein composition in these two groups of extracellular vesicles. In conclusion, results of this study demonstrate that exposure of MOVAS cells to high phosphate levels stimulates the release of extracellular vesicles and changes their protein composition.

  6. Implication of Soluble Forms of Cell Adhesion Molecules in Infectious Disease and Tumor: Insights from Transgenic Animal Models

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    Etsuro Ono

    2018-01-01

    Full Text Available Cell adhesion molecules (CAMs are surface ligands, usually glycoproteins, which mediate cell-to-cell adhesion. They play a critical role in maintaining tissue integrity and mediating migration of cells, and some of them also act as viral receptors. It has been known that soluble forms of the viral receptors bind to the surface glycoproteins of the viruses and neutralize them, resulting in inhibition of the viral entry into cells. Nectin-1 is one of important CAMs belonging to immunoglobulin superfamily and herpesvirus entry mediator (HVEM is a member of the tumor necrosis factor (TNF receptor family. Both CAMs also act as alphaherpesvirus receptor. Transgenic mice expressing the soluble form of nectin-1 or HVEM showed almost complete resistance against the alphaherpesviruses. As another CAM, sialic acid-binding immunoglobulin-like lectins (Siglecs that recognize sialic acids are also known as an immunoglobulin superfamily member. Siglecs play an important role in the regulation of immune cell functions in infectious diseases, inflammation, neurodegeneration, autoimmune diseases and cancer. Siglec-9 is one of Siglecs and capsular polysaccharide (CPS of group B Streptococcus (GBS binds to Siglec-9 on neutrophils, leading to suppress host immune response and provide a survival advantage to the pathogen. In addition, Siglec-9 also binds to tumor-produced mucins such as MUC1 to lead negative immunomodulation. Transgenic mice expressing the soluble form of Siglec-9 showed significant resistance against GBS infection and remarkable suppression of MUC1 expressing tumor proliferation. This review describes recent developments in the understanding of the potency of soluble forms of CAMs in the transgenic mice and discusses potential therapeutic interventions that may alter the outcomes of certain diseases.

  7. Binding, internalization, and degradation of atrial natriuretic peptide in cultured vascular smooth muscle cells of rat

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    Hirata, Y.; Takata, S.; Tomita, M.; Takaichi, S.

    1985-11-15

    Binding, internalization, and degradation of /sup 125/I-labeled-rat atrial natriuretic peptide (rANP) were studied in cultured rat aortic vascular smooth muscle cells (VSMC). At 37 degrees C, /sup 125/I-labeled-rANP rapidly bound to VSMCs, but the cell-bound radioactivity rapidly decreased upon subsequent incubation, while the binding was slow at 4 degrees C, reaching to an apparent equilibrium after 6 hrs. The cell-bound /sup 125/I-labeled-rANP at 37 degrees C is rapidly dissociated from VSMC (t 1/2: approximately 40 min) with the appearance of degradaded product(s) of radioligand in the medium, whereas the degradation was minimal at 4 degrees C. This degradative process was blocked by inhibitors of metabolic energy production (azide, dinitrophenol), inhibitors of lysosomal cathepsins (leupeptin, pepstatin), and lysosomotropic agents (NH/sub 4/Cl, chloroquine, lidocaine, methylamine, dansylcadaverine), but not by inhibitors of serine or thiol proteases. /sup 125/I-labeled-rANP initially bound to the cell-surface was rapidly internalized, and delivered to lysosomal structures, which was confirmed by autoradiographic studies. These data indicate that rANP, after binding to the cell-surface receptors, is rapidly internalized into the cells through receptor-mediated endocytosis, and subsequently degradaded by lysosomal hydrolases.

  8. Bisphenol A Disrupts Transcription and Decreases Viability in Aging Vascular Endothelial Cells

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    Edna Ribeiro-Varandas

    2014-09-01

    Full Text Available Bisphenol A (BPA is a widely utilized endocrine disruptor capable of mimicking endogenous hormones, employed in the manufacture of numerous consumer products, thereby interfering with physiological cellular functions. Recent research has shown that BPA alters epigenetic cellular mechanisms in mammals and may be correlated to enhanced cellular senescence. Here, the effects of BPA at 10 ng/mL and 1 µg/mL, concentrations found in human samples, were analyzed on HT29 human colon adenocarcinona cell line and Human Umbilical Vein Endothelial Cells (HUVEC. Quantitative Real-Time Polymerase Chain Reaction (qRT-PCR transcriptional analysis of the Long Interspersed Element-1 (LINE-1 retroelement showed that BPA induces global transcription deregulation in both cell lines, although with more pronounced effects in HUVEC cells. Whereas there was an increase in global transcription in HT29 exclusively after 24 h of exposure, this chemical had prolonged effects on HUVEC. Immunoblotting revealed that this was not accompanied by alterations in the overall content of H3K9me2 and H3K4me3 epigenetic marks. Importantly, cell viability assays and transcriptional analysis indicated that prolonged BPA exposure affects aging processes in senescent HUVEC. To our knowledge this is the first report that BPA interferes with senescence in primary vascular endothelial cells, therefore, suggesting its association to the etiology of age-related human pathologies, such as atherosclerosis.

  9. Bisphenol A Disrupts Transcription and Decreases Viability in Aging Vascular Endothelial Cells

    Science.gov (United States)

    Ribeiro-Varandas, Edna; Pereira, H. Sofia; Monteiro, Sara; Neves, Elsa; Brito, Luísa; Boavida Ferreira, Ricardo; Viegas, Wanda; Delgado, Margarida

    2014-01-01

    Bisphenol A (BPA) is a widely utilized endocrine disruptor capable of mimicking endogenous hormones, employed in the manufacture of numerous consumer products, thereby interfering with physiological cellular functions. Recent research has shown that BPA alters epigenetic cellular mechanisms in mammals and may be correlated to enhanced cellular senescence. Here, the effects of BPA at 10 ng/mL and 1 µg/mL, concentrations found in human samples, were analyzed on HT29 human colon adenocarcinona cell line and Human Umbilical Vein Endothelial Cells (HUVEC). Quantitative Real-Time Polymerase Chain Reaction (qRT-PCR) transcriptional analysis of the Long Interspersed Element-1 (LINE-1) retroelement showed that BPA induces global transcription deregulation in both cell lines, although with more pronounced effects in HUVEC cells. Whereas there was an increase in global transcription in HT29 exclusively after 24 h of exposure, this chemical had prolonged effects on HUVEC. Immunoblotting revealed that this was not accompanied by alterations in the overall content of H3K9me2 and H3K4me3 epigenetic marks. Importantly, cell viability assays and transcriptional analysis indicated that prolonged BPA exposure affects aging processes in senescent HUVEC. To our knowledge this is the first report that BPA interferes with senescence in primary vascular endothelial cells, therefore, suggesting its association to the etiology of age-related human pathologies, such as atherosclerosis. PMID:25207595

  10. [Comparison of electrophysiological properties of vascular smooth muscle cells in different arterioles in guinea pig].

    Science.gov (United States)

    Ma, Ke-Tao; Li, Xin-Zhi; Li, Li; Zhang, Zhi-Ping; Zhao, Lei; Zhu, He; Si, Jun-Qiang

    2010-10-25

    Arterioles are major contributors to the control of systemic blood pressure and local blood flow. In this study, we compared electrophysiological properties of vascular smooth muscle cells (VSMCs) in anterior inferior cerebellar artery (AICA), mesenteric artery (MA) and spiral modiolar artery (SMA) by intracellular microelectrode recording and whole-cell patch clamp recording techniques. Results were shown as below: (1) Intracellular microelectrode recordings were made from VSMCs in AICA, MA and SMA with resting potentials of (-68±1.8) (n=65), (-71±2.4) (n=80) and (-66±2.9) mV (n=58), respectively. There was no significant difference in resting potentials among arterioles. (2) The membrane capacitance and membrane conductance in situ cells were much larger than those in dispersed smooth muscle cells by whole-cell recording techniques, and there was significant difference among arterioles, which were in the order: MA>AICA>SMA. After application of gap junction blocker 2-APB (100 μmol/L), the membrane capacitance and membrane conductance in situ cells were very close with those in single smooth muscle cells. (3) The I/V relation of whole-cell current of dissociated smooth muscle cells (AICA, MA and SMA) showed a prominent outward rectification, and the currents were substantially inhibited by 1 mmol/L 4-AP or 10 mmol/L TEA. When the command voltage was +40 mV, the current densities of VSMCs in AICA, MA and SMA were (26±2.0), (24±1.7) and (18±1.3) pA/pF respectively. SMA showed significant difference in the current density from AICA and MA respectively. These results suggest that the electrophysiological properties of coupling strength of gap junction and current density of smooth muscle cells are different among arterioles in the guinea pig.

  11. PML-RARa modulates the vascular signature of extracellular vesicles released by acute promyelocytic leukemia cells.

    Science.gov (United States)

    Fang, Yi; Garnier, Delphine; Lee, Tae Hoon; D'Asti, Esterina; Montermini, Laura; Meehan, Brian; Rak, Janusz

    2016-01-01

    Oncogenic transformation is believed to impact the vascular phenotype and microenvironment in cancer, at least in part, through mechanisms involving extracellular vesicles (EVs). We explored these questions in the context of acute promyelocytic leukemia cells (NB4) expressing oncogenic fusion protein, PML-RARa and exquisitely sensitive to its clinically used antagonist, the all-trans retinoic acid (ATRA). We report that NB4 cells produce considerable numbers of EVs, which are readily taken up by cultured endothelial cells triggering their increased survival. NB4 EVs contain PML-RARa transcript, but no detectable protein, which is also absent in endothelial cells upon the vesicle uptake, thereby precluding an active intercellular trafficking of this oncogene in this setting. ATRA treatment changes the emission profile of NB4-related EVs resulting in preponderance of smaller vesicles, an effect that occurs in parallel with the onset of cellular differentiation. ATRA also increases IL-8 mRNA and protein content in NB4 cells and their EVs, while decreasing the levels of VEGF and tissue factor (TF). Endothelial cell uptake of NB4-derived EVs renders these cells more TF-positive and procoagulant, and this effect is diminished by pre-treatment of EV donor cells with ATRA. Profiling angiogenesis-related transcripts in intact and ATRA-treated APL cells and their EVs reveals multiple differences attributable to cellular responses and EV molecular packaging. These observations point to the potential significance of changes in the angiogenic signature and activity associated with EVs released from tumor cells subjected to targeted therapy.

  12. Artemisinin attenuates platelet-derived growth factor BB-induced migration of vascular smooth muscle cells.

    Science.gov (United States)

    Lee, Kang Pa; Park, Eun-Seok; Kim, Dae-Eun; Park, In-Sik; Kim, Jin Tack; Hong, Heeok

    2014-10-01

    Artemisinin (AT), an active compound in Arternisia annua, is well known as an anti-malaria drug. It is also known to have several effects including anti-oxidant, anti-inflammation, and anti-cancer activities. To date, the effect of AT on vascular disorders has not been studied. In this study, we investigated the effects of AT on the migration and proliferation of vascular smooth muscle cells (VSMC) stimulated by platelet-derived growth factor BB (PDGF-BB). Aortic smooth muscle cells were isolated from Sprague-Dawley rats. PDGF-BB stimulated VSMC migration was measured by the scratch wound healing assay and the Boyden chamber assay. Cell viability was determined by using an EZ-Cytox Cell Viability Assay Kit. The production of reactive oxygen species (ROS) in PDGF-BB stimulated VSMC was measured through H2DCF-DA staining. We also determined the expression levels of signal proteins relevant to ROS, including measures of extracellular signal-regulated kinase (ERK) 1/2 measured by western blot analysis and matrix metalloproteinase (MMP) 9 measured by reverse transcription-polymerase chain reaction (RT-PCR). AT (10 µM and 30 µM) significantly reduced the proliferation and migration of PDGF-BB stimulated VSMC in a dose-dependent manner. The production of ROS, normally induced by PDGF-BB, is reduced by treatment with AT at both concentrations. PDGF-BB stimulated VSMC treated with AT (10 µM and 30 µM) have reduced phosphorylation of ERK1/2 and inhibited MMP9 expression compared to untreated PDGF-BB stimulated VSMC. We suggest, based on these results, that AT may exert an anti-atherosclerotic effect on PDGF-BB stimulated VSMCs by inhibiting their proliferation and migration through down-regulation of ERK1/2 and MMP9 phosphorylation.

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

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    Castellot John J

    2003-11-01

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

  14. Zoledronate upregulates MMP-9 and -13 in rat vascular smooth muscle cells by inducing oxidative stress

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    Arun MZ

    2016-04-01

    Full Text Available Mehmet Zuhuri Arun,1 Buket Reel,1 Graciela B Sala-Newby,2 Mark Bond,2 Aikaterini Tsaousi,2 Perry Maskell,2 Andrew C Newby21Department of Pharmacology, Faculty of Pharmacy, Ege University, Izmir, Turkey; 2Bristol Heart Institute, University of Bristol, Bristol Royal Infirmary, Bristol, UK Background: Bisphosphonates, including zoledronate, target osteoclasts and are widely used in the treatment of osteoporosis and other bone resorption diseases, despite side effects that include damaging the stomach epithelium. Beneficial and adverse effects on other organ systems, including the cardiovascular system, have also been described and could impact on the use of bisphosphonates as therapeutic agents. Vascular smooth muscle cells (VSMCs are major constituents of the normal vascular wall and have a key role in intimal thickening and atherosclerosis, in part by secreting MMPs that remodel the extracellular matrix and cleave cell surface proteins or secreted mediators. In this study, we investigated the effects of zoledronate on MMP expression.Methods: Rat VSMCs were stimulated by PDGF (50 ng/mL plus TNF-α (10 ng/mL or left unstimulated for a further 24 hours in serum-free medium. In other series of experiments, cells were pre-treated either with SC-514 (50 µM or with apocynin (20 nM for 2 hours, then zoledronate (100 µM was added into 2% fetal calf serum containing medium for 24 hours.Results and discussion: Using isolated rat VSMCs in culture, zoledronate (100 µM increased MMP-9 and -13 mRNA expressions but inhibited MMP-2 expression. MMP-9 and MMP-13 up-regulation was shown to depend on the NF-κB pathway; and this was activated by zoledronate. Furthermore, zoledronate elevated the levels of reactive oxygen species detected by either dichlorofluorescein in isolated VSMCs or lucigenin enhanced chemiluminescence in rat aortic rings in vitro. Apocynin, an inhibitor of NADPH oxidase, reversed NF-κB activation and MMP-9 and MMP-13 up-regulation by

  15. The New Role of CD163 in the Differentiation of Bone Marrow Stromal Cells into Vascular Endothelial-Like Cells

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    Wei Lu

    2016-01-01

    Full Text Available Bone marrow stromal cells (BMSCs can differentiate into vascular endothelial cells (VECs. It is regarded as an important solution to cure many diseases, such as ischemic diseases and diabetes. However, the mechanisms underlying BMSC differentiation into VECs are not well understood. Recent reports showed that CD163 expression was associated with angiogenesis. In this study, overexpression of CD163 in BMSCs elevated the protein level of the endothelial-associated markers CD31, Flk-1, eNOS, and VE-cadherin, significantly increased the proportion of Alexa Fluor 488-acetylated-LDL-positive VECs, and promoted angiogenesis on Matrigel. Furthermore, we demonstrated that CD163 acted downstream homeobox containing 1 (Hmbox1 and upstream fibroblast growth factor 2 (FGF-2. These data suggested that CD163 was involved in Hmbox1/CD163/FGF-2 signal pathway in BMSC differentiation into vascular endothelial-like cells. We found a new signal pathway and a novel target for further investigating the gene control of BMSC differentiation into a VEC lineage.

  16. Degree of anemia, indirect markers of hemolysis, and vascular complications of sickle cell disease in Africa.

    Science.gov (United States)

    Dubert, Marie; Elion, Jacques; Tolo, Aissata; Diallo, Dapa Aly; Diop, Saliou; Diagne, Ibrahima; Sanogo, Ibrahima; Belinga, Suzanne; Guifo, Odette; Wamba, Guillaume; Ngo Sack, Françoise; Boidy, Kouakou; Kamara, Ismael; Traore, Youssouf; Diakite, Cheick Oumar; Gbonon, Valérie; Faye, Blaise Felix; Seck, Moussa; Deme Ly, Indou; Chelo, David; N'Guetta, Roland; Diop, Ibrahima Bara; Gaye, Bamba; Jouven, Xavier; Ranque, Brigitte

    2017-11-16

    The hyperhemolysis paradigm that describes overlapping "hyperhemolytic-endothelial dysfunction" and "high hemoglobin-hyperviscous" subphenotypes of sickle cell disease (SCD) patients is based on North American studies. We performed a transversal study nested in the CADRE cohort to analyze the association between steady-state hemolysis and vascular complications of SCD among sub-Saharan African patients. In Mali, Cameroon, and Ivory Coast, 2407 SCD patients (1751 SS or sickle β-zero-thalassemia [Sβ 0 ], 495 SC, and 161 sickle β + -thalassemia [Sβ + ]), aged 3 years old and over, were included at steady state. Relative hemolytic intensity was estimated from a composite index derived from principal component analysis, which included bilirubin levels or clinical icterus, and lactate dehydrogenase levels. We assessed vascular complications (elevated tricuspid regurgitant jet velocity [TRV], microalbuminuria, leg ulcers, priapism, stroke, and osteonecrosis) by clinical examination, laboratory tests, and echocardiography. After adjustment for age, sex, country, and SCD phenotype, a low hemoglobin level was significantly associated with TRV and microalbuminuria in the whole population and with leg ulcers in SS-Sβ 0 adults. A high hemolysis index was associated with microalbuminuria in the whole population and with elevated TRV, microalbuminuria, and leg ulcers in SS-Sβ 0 adults, but these associations were no longer significant after adjustment for hemoglobin level. In conclusion, severe anemia at steady state in SCD patients living in West and Central Africa is associated with elevated TRV, microalbuminuria, and leg ulcers, but these vascular complications are not independently associated with indirect markers of increased hemolysis. Other mechanisms leading to anemia, including malnutrition and infectious diseases, may also play a role in the development of SCD vasculopathy. © 2017 by The American Society of Hematology.

  17. Effect of sulfated polysaccharides from Laminaria japonica on vascular endothelial cells in psychological stress rats.

    Science.gov (United States)

    Li, Jing; Wang, Shengyong; Yang, Xiaomei; Pang, Guangbao; Zheng, Hua; Shen, Bin; Li, Guanhong; Shi, Dianchun; Wang, Jienian; Feng, Liaoyun; Li, Mulan; Wei, Wuying; Qin, Wu; Xie, Lu

    2014-01-01

    Laminaria japonica is a popular seafood and medicinal plant in China. Laminaria japonica is used in traditional Chinese medicine to treat and prevent hypertension and edema. The vascular protective activity and mechanism of sulfated polysaccharides were studied in adrenalin-induced vascular endothelial damage in rats after psychological stress (PS). Vehicle (sham and PS groups), sulfated polysaccharide from Laminaria japonica (LP; 1mg/kg and 5mg/kg) and enoxaparin sodium (1IU/kg, reference drug) were all administered for 10 days. Behavioral changes were recorded. Plasma levels of adrenalin, cortisol, monoamine oxidase (MAO), semicarbazide-sensitive amine oxidase (SSAO), formaldehyde, H2O2, nitric oxide (NO), endothelin-1 (ET-1), 6-keto-prostaglandin F1a (6-keto-PGF1a), and thromboxane B2 (TXB2) were measured. Endothelium-dependent relaxation of the thoracic aorta was measured and transmission electron microscopy of aortic vessels was performed. Adrenalin metabolites in plasma were significantly lower (P<0.01) in rats after LP administration compared with those in the PS groups. The normalized ratios of plasma NO/ET-1 and 6-keto-PGF1a/TXB2 were maintained and endothelium-dependent relaxation of the aorta was greatly enhanced after LP treatment (P<0.05). Morphological alterations were observed in vascular endothelial cells (VECs) in PS rats, with a higher number of lysosomes and vague mitochondrial cristae compared with those in the sham group. However, these histopathological changes were markedly alleviated after LP treatment. This study shows a protective effect of LP on VECs in PS rats. LP can regulate plasma levels of NO, ET-1, and 6-keto-PGF1a, enhance endothelium-dependent relaxation, and alleviate histopathological changes of lysosomes and mitochondria in VECs. The potential mechanism of LP on VECs in PS rats is related to its function of reducing metabolites of adrenalin. © 2013 Published by Elsevier Ireland Ltd.

  18. Modification of MCF-10A Cells with Pioglitazone and Serum-Rich Growth Medium Increases Soluble Factors in the Conditioned Medium, Likely Reducing BT-474 Cell Growth

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    Kalpanah Nadarajan

    2012-05-01

    Full Text Available In the present study, we aimed to preincubate MCF-10A cells with pioglitazone and/or serum-rich growth media and to determine adhesive and non-adhesive interactions of the preincubated MCF-10A cells with BT-474 cells. For this purpose, the MCF-10A cells were preincubated with pioglitazone and/or serum-rich growth media, at appropriate concentrations, for 1 week. The MCF-10A cells preincubated with pioglitazone and/or serum-rich growth media were then co-cultured adhesively and non-adhesively with BT-474 cells for another week. Co-culture of BT-474 cells with the preincubated MCF-10A cells, both adhesively and non-adhesively, reduced the growth of the cancer cells. The inhibitory effect of the preincubated MCF-10A cells against the growth of BT-474 cells was likely produced by increasing levels of soluble factors secreted by the preincubated MCF-10A cells into the conditioned medium, as immunoassayed by ELISA. However, only an elevated level of a soluble factor distinguished the conditioned medium collected from the MCF-10A cells preincubated with pioglitazone and serum-rich growth medium than that with pioglitazone alone. This finding was further confirmed by the induction of the soluble factor transcript expression in the preincubated MCF-10A cells, as determined using real-time PCR, for the above phenomenon. Furthermore, modification of the MCF-10A cells through preincubation did not change the morphology of the cells, indicating that the preincubated cells may potentially be injected into mammary fat pads to reduce cancer growth in patients or to be used for others cell-mediated therapy.

  19. Synergistic effect of histamine and TNF-α on monocyte adhesion to vascular endothelial cells.

    Science.gov (United States)

    Chen, Chong; Khismatullin, Damir B

    2013-04-01

    The histamine level is high during allergic attacks, and patients with allergy may have chronic inflammatory conditions at which tumor necrosis factor (TNF)-α is extensively released by macrophages. Here, in vitro static and microfluidic flow assays were conducted to investigate the combined influence of histamine and TNF-α on adhesion of monocytic THP-1 cells to human umbilical vein endothelial cells (HUVEC). In a static assay, histamine stimulation of TNF-α-activated HUVEC elevated the number of attached THP-1 cells. In a flow assay, the number of crawling and firmly adherent THP-1 cells was higher on TNF-α + histamine activated HUVEC than on HUVEC activated by TNF-α alone. This synergistic effect of histamine and TNF-α is caused by the increased endothelial surface expression of intercellular adhesion molecule-1, vascular cell adhesion molecule-1, and E-selectin. Since the exposure of TNF-α-activated endothelium to histamine favors monocyte recruitment, it may be a serious risk factor for atherosclerosis and other inflammatory disorders.

  20. Heparan sulfate chains potentiate cadmium cytotoxicity in cultured vascular endothelial cells.

    Science.gov (United States)

    Fujiwara, Yasuyuki; Yamamoto, Chika; Yoshida, Eiko; Kumagai, Yoshito; Kaji, Toshiyuki

    2016-02-01

    The monolayer of vascular endothelial cells, which is rich in heparan sulfate chains, is an important target of cadmium cytotoxicity. To investigate the effects of heparan sulfate chains on cadmium cytotoxicity, bovine aortic endothelial cells were cultured in the presence of cadmium, with or without exogenous heparan sulfate. The following results were obtained: (1) Heparan sulfate chains potentiated cadmium cytotoxicity. (2) Such a potentiation did not occur in bovine aortic smooth muscle cells. (3) Heparin chains as well as heparan sulfate chains potentiated cadmium cytotoxicity, while other glycosaminoglycan chains failed to exhibit such an activity. (4) The disaccharide units of heparan sulfate chains did not potentiate cadmium cytotoxicity in the endothelial cells. (5) Heparan sulfate chains did not potentiate mercury and arsenite cytotoxicity. (6) Fibroblast growth factor-2 (FGF-2) also potentiated cadmium cytotoxicity in the endothelial cells. (7) Heparan sulfate chains significantly increased intracellular cadmium accumulation by inducing the expression of metallothionein. Taken together, these results suggest that heparan sulfate chains activate FGF-2, which in turn elevates the expression and/or activity of metal transporter(s) that facilitate cadmium influx from the extracellular space into the cytoplasm.

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

    International Nuclear Information System (INIS)

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

    1986-01-01

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

  2. 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.

  3. Resveratrol induces vascular smooth muscle cell differentiation through stimulation of SirT1 and AMPK.

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    Anne Marie Thompson

    Full Text Available 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 protein expression, contractile morphology and contraction in collagen gels. Resveratrol induced VSMC differentiation through stimulation of SirT1 and AMPK. We made the novel finding that low or high dose resveratrol had an initially different mechanism on induction of differentiation. We found that low dose resveratrol stimulated differentiation through SirT1-mediated activation of AKT, whereas high dose resveratrol stimulated differentiation through AMPK-mediated inhibition of the mTORC1 pathway, allowing activation of AKT. The health effects of resveratrol in cardiovascular diseases, cancer and longevity are an area of active research. We have demonstrated a supplemental avenue where-by resveratrol may promote health by maintaining and enhancing plasticity of the vasculature.

  4. VEGFR signaling during lymphatic vascular development: From progenitor cells to functional vessels.

    Science.gov (United States)

    Secker, Genevieve A; Harvey, Natasha L

    2015-03-01

    Lymphatic vessels are an integral component of the cardiovascular system, serving important roles in fluid homeostasis, lipid absorption, and immune cell trafficking. Defining the mechanisms by which the lymphatic vasculature is constructed and remodeled into a functional vascular network not only provides answers to fascinating biological questions, but is fundamental to understanding how lymphatic vessel growth and development goes awry in human pathologies. While long recognized as dysfunctional in lymphedema and exploited as a route of tumor metastasis, recent work has highlighted important roles for lymphatic vessels in modulating immune responses, regulating salt-sensitive hypertension and important for lung inflation at birth. Substantial progress in our understanding of the signaling pathways important for development and morphogenesis of the lymphatic vasculature has been made in recent years. Here, we review advances in our knowledge of the best characterized of these signaling pathways, that involving the vascular endothelial growth factor (VEGF) family members VEGF-C and VEGF-D, together with their receptors VEGFR2 and VEGFR3. Recent work has defined multiple levels at which signal transduction by means of this key axis is regulated; these include control of ligand processing and bioavailability, modulation of receptor activation by interacting proteins, and regulation of receptor endocytosis and trafficking. © 2014 Wiley Periodicals, Inc.

  5. Proliferation of smooth muscle cells at sites distant from vascular injury

    International Nuclear Information System (INIS)

    Reidy, M.A.

    1990-01-01

    This study investigated the phenomenon that injury at one specific site in blood vessels induces cell replication at distant vascular sites. A polyethylene tube was inserted via the common carotid into rat aortic arch, which caused focal endothelial cell loss and formation of platelet thrombi. In a similar fashion, a polyethylene tube was placed into the lower abdominal aorta via a femoral artery. All animals received 3H-thymidine continuously for 2 weeks, after which time segments of the aorta distant from the polyethylene tubing were processed for autoradiography. These sites showed no loss of endothelium or adherent platelets, and yet the smooth muscle and endothelial cell replications were significantly elevated as compared to control aortas. There was no significant change in blood pressure during these experiments and no increase in smooth muscle cell ploidy. When the polyethylene tubing was left in situ for 2 months, no increased replication of the smooth muscle cells was observed during the last 2 weeks of the experiment, and at this time the aorta adjacent to the tubing was completely re-endothelialized. Finally, the mitogenic activity of plasma from these animals was tested in vitro. At the time of a significant increase of in vivo cell replication (2 weeks), the mitogenic activity of the plasma from animals with the indwelling tubing was similar to that of the control animals. In summary, these data show that injury at one discrete arterial site leads to general cell proliferation in the same vessel, and the data would support the possibility that cell communication initiates this response

  6. Ultrathin Metal Films with Defined Topographical Structures as In Vitro Cell Culture Platforms for Unveiling Vascular Cell Behaviors.

    Science.gov (United States)

    Jun, Indong; Chung, Yong-Woo; Park, Jimin; Han, Hyung-Seop; Park, Jaeho; Kim, Saeromi; Lee, Hyunjung; Kim, Sang Hoon; Han, Jun-Hyun; Kim, Hyunjung; Seok, Hyun-Kwang; Kim, Yu-Chan; Jeon, Hojeong

    2016-09-01

    Implanted material surfaces make direct contact with body tissues to work on its own purpose. Therefore, studies of the surface properties of implantable materials that determine cell fate are very important for successful implantation. Although numerous studies have addressed the relationship between cells and material surfaces, nonmetallic surfaces and metallic surfaces likely produce different cellular responses because of their intrinsic differences in surface energy, roughness, and chemical composition. Moreover, given the nontransparent property of metal materials, which hampers the real-time imaging of cellular behavior, a detailed cellular-level analysis at the metal-tissue interface has not been performed. In this study, metal-based cell culture platforms (MCPs) with defined microscale topographical patterns are developed using a combination of photolithography and direct current magnetron sputtering techniques. The MCPs allow to observe vascular cells on metals in real time and identify the selective regulation of human aortic smooth muscle cells and Human umbilical vein endothelial cells (HUVECs) by metallic surface topography. Additionally, atomic force microscopy, contact angles, and energy-dispersive X-ray spectroscopy analyses show that the MCPs exhibit nearly identical chemical properties with their bulk counterparts, demonstrating that MCPs can be utilized as an in vitro cell culture platform system for understanding the cellular behavior on metal substrates. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  7. Human iPS cell-derived insulin producing cells form vascularized organoids under the kidney capsules of diabetic mice.

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    Sudhanshu P Raikwar

    Full Text Available Type 1 diabetes (T1D is caused by autoimmune disease that leads to the destruction of pancreatic β-cells. Transplantation of cadaveric pancreatic organs or pancreatic islets can restore normal physiology. However, there is a chronic shortage of cadaveric organs, limiting the treatment of the majority of patients on the pancreas transplantation waiting list. Here, we hypothesized that human iPS cells can be directly differentiated into insulin producing cells (IPCs capable of secreting insulin. Using a series of pancreatic growth factors, we successfully generated iPS cells derived IPCs. Furthermore, to investigate the capability of these cells to secrete insulin in vivo, the differentiated cells were transplanted under the kidney capsules of diabetic immunodeficient mice. Serum glucose levels gradually declined to either normal or near normal levels over 150 days, suggesting that the IPCs were secreting insulin. In addition, using MRI, a 3D organoid appeared as a white patch on the transplanted kidneys but not on the control kidneys. These organoids showed neo-vascularization and stained positive for insulin and glucagon. All together, these data show that a pancreatic organ can be created in vivo providing evidence that iPS cells might be a novel option for the treatment of T1D.

  8. Brazilin Ameliorates High Glucose-Induced Vascular Inflammation via Inhibiting ROS and CAMs Production in Human Umbilical Vein Endothelial Cells

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    Thanasekaran Jayakumar

    2014-01-01

    Full Text Available Vascular inflammatory process has been suggested to play a key role in the initiation and progression of atherosclerosis, a major complication of diabetes mellitus. Recent studies have shown that brazilin exhibits antihepatotoxic, antiplatelet, cancer preventive, or anti-inflammatory properties. Thus, we investigated whether brazilin suppresses vascular inflammatory process induced by high glucose (HG in cultured human umbilical vein endothelial cells (HUVEC. HG induced nitrite production, lipid peroxidation, and intracellular reactive oxygen species formation in HUVEC cells, which was reversed by brazilin. Western blot analysis revealed that brazilin markedly inhibited HG-induced phosphorylation of endothelial nitric oxide synthase. Besides, we investigated the effects of brazilin on the MAPK signal transduction pathway because MAPK families are associated with vascular inflammation under stress. Brazilin blocked HG-induced phosphorylation of extracellular signal-regulated kinase and transcription factor NF-κB. Furthermore, brazilin concentration-dependently attenuated cell adhesion molecules (ICAM-1 and VCAM-1 expression induced by various concentrations of HG in HUVEC. Taken together, the present data suggested that brazilin could suppress high glucose-induced vascular inflammatory process, which may be closely related with the inhibition of oxidative stress, CAMs expression, and NF-κB activation in HUVEC. Our findings may highlight a new therapeutic intervention for the prevention of vascular diseases.

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

    International Nuclear Information System (INIS)

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

    1989-01-01

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

  10. Shear stress with appropriate time-step and amplification enhances endothelial cell retention on vascular grafts.

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    Liu, Haifeng; Gong, Xianghui; Jing, Xiaohui; Ding, Xili; Yao, Yuan; Huang, Yan; Fan, Yubo

    2017-11-01

    Endothelial cells (ECs) are sensitive to changes in shear stress. The application of shear stress to ECs has been well documented to improve cell retention when placed into a haemodynamically active environment. However, the relationship between the time-step and amplification of shear stress on EC functions remains elusive. In the present study, human umbilical cord veins endothelial cells (HUVECs) were seeded on silk fibroin nanofibrous scaffolds and were preconditioned by shear stress at different time-steps and amplifications. It is shown that gradually increasing shear stress with appropriate time-steps and amplification could improve EC retention, yielding a complete endothelial-like monolayer both in vitro and in vivo. The mechanism of this improvement is mediated, at least in part, by an upregulation of integrin β1 and focal adhesion kinase (FAK) expression, which contributed to fibronectin (FN) assembly enhancement in ECs in response to the shear stress. A modest gradual increase in shear stress was essential to allow additional time for ECs to gradually acclimatize to the changing environment, with the goal of withstanding the physiological levels of shear stress. This study recognized that the time-steps and amplifications of shear stress could regulate EC tolerance to shear stress and the anti-thrombogenicity function of engineered vascular grafts via an extracellular cell matrix-specific, mechanosensitive signalling pathway and might prevent thrombus formation in vivo. Copyright © 2016 John Wiley & Sons, Ltd. Copyright © 2016 John Wiley & Sons, Ltd.

  11. Dysregulation of Vascular Endothelial Progenitor Cells Lung-Homing in Subjects with COPD

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    Brittany M. Salter

    2016-01-01

    Full Text Available Chronic obstructive pulmonary disease (COPD is characterized by fixed airflow limitation and progressive decline of lung function and punctuated by occasional exacerbations. The disease pathogenesis may involve activation of the bone marrow stimulating mobilization and lung-homing of progenitor cells. We investigated the hypothesis that lower circulating numbers of vascular endothelial progenitor cells (VEPCs are a consequence of increased lung-sequestration in COPD. Nonatopic, current or ex-smokers with diagnosed COPD and nonatopic, nonsmoking normal controls were enrolled. Blood and induced sputum extracted primitive hemopoietic progenitors (HPCs and VEPC were enumerated by flow cytometry. Migration and adhesive responses to fibronectin were assessed. In sputum, VEPC numbers were significantly greater in COPD compared to normal controls. In blood, VEPCs were significantly lower in COPD versus normal controls. There were no differences in HPC levels between the two groups in either compartment. Functionally, there was a greater migrational responsiveness of progenitors from COPD subjects to stromal cell-derived factor-1alpha (SDF-1α compared to normal controls. This was associated with greater numbers of CXCR4+ progenitors in sputum from COPD. Increased migrational responsiveness of progenitor cells may promote lung-homing of VEPC in COPD which may disrupt maintenance and repair of the airways and contribute to COPD disease pathogenesis.

  12. Vascular Platform to Define Hematopoietic Stem Cell Factors and Enhance Regenerative Hematopoiesis

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    Michael G. Poulos

    2015-11-01

    Full Text Available Hematopoietic stem cells (HSCs inhabit distinct microenvironments within the adult bone marrow (BM, which govern the delicate balance between HSC quiescence, self-renewal, and differentiation. Previous reports have proposed that HSCs localize to the vascular niche, comprised of endothelium and tightly associated perivascular cells. Herein, we examine the capacity of BM endothelial cells (BMECs to support ex vivo and in vivo hematopoiesis. We demonstrate that AKT1-activated BMECs (BMEC-Akt1 have a unique transcription factor/cytokine profile that supports functional HSCs in lieu of complex serum and cytokine supplementation. Additionally, transplantation of BMEC-Akt1 cells enhanced regenerative hematopoiesis following myeloablative irradiation. These data demonstrate that BMEC-Akt1 cultures can be used as a platform for the discovery of pro-HSC factors and justify the utility of BMECs as a cellular therapy. This technical advance may lead to the development of therapies designed to decrease pancytopenias associated with myeloablative regimens used to treat a wide array of disease states.

  13. Pathological aging of the vascular endothelium: are endothelial progenitor cells the sentinels of the cardiovascular system?

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    Thorin-Trescases, Nathalie; Voghel, Guillaume; Gendron, Marie-Eve; Krummen, Stephane; Farhat, Nada; Drouin, Annick; Perrault, Louis P; Thorin, Eric

    2005-10-01

    Aging is associated with vascular endothelial dysfunction, which ultimately leads to atherosclerosis. On the other hand, it is clear that in young patients with risk factors for cardiovascular diseases (CVD), endothelial dysfunction is an early marker of the ongoing atherogenic process. It is therefore tempting to speculate that risk factors for CVD accelerate the aging process. The aging of an endothelial cell (EC) is not chronological but rather dependent on its replication rate. ECs have a finite number of divisions and enter replicative senescence after exhaustion of this potential. Telomere attrition is believed to be responsible for this phenomenon. Upon reaching a critical minimal telomere length, ECs enter a nondividing state of replicative senescence. Recently, endothelial progenitor cells originating from the bone marrow have been isolated from the circulation. They integrate into the endothelial layer of the vessel and contribute to healing, ischemic repair and angiogenesis. A completely new field of investigation is now open. Are endothelial progenitor cells sensitive to the aging process? Do they prevent endothelial dysfunction? Are they the ultimate shield against the damages induced by risk factors for CVD? There are no definite answers to these questions, but the potential of these cells is tremendous and understanding their physiology is essential.

  14. Periplasmic expression of soluble single chain T cell receptors is rescued by the chaperone FkpA

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    Bogen Bjarne

    2010-02-01

    Full Text Available Abstract Background Efficient expression systems exist for antibody (Ab molecules, which allow for characterization of large numbers of individual Ab variants. In contrast, such expression systems have been lacking for soluble T cell receptors (TCRs. Attempts to generate bacterial systems have generally resulted in low yields and material which is prone to aggregation and proteolysis. Here we present an optimized periplasmic bacterial expression system for soluble single chain (sc TCRs. Results The effect of 1 over-expression of the periplasmic chaperon FkpA, 2 culture conditions and 3 molecular design was investigated. Elevated levels of FkpA allowed periplasmic soluble scTCR expression, presumably by preventing premature aggregation and inclusion body formation. Periplasmic expression enables disulphide bond formation, which is a prerequisite for the scTCR to reach its correct fold. It also enables quick and easy recovery of correctly folded protein without the need for time-consuming downstream processing. Expression without IPTG induction further improved the periplasmic expression yield, while addition of sucrose to the growth medium showed little effect. Shaker flask yield of mg levels of active purified material was obtained. The Vαβ domain orientation was far superior to the Vβα domain orientation regarding monomeric yield of functionally folded molecules. Conclusion The general expression regime presented here allows for rapid production of soluble scTCRs and is applicable for 1 high yield recovery sufficient for biophysical characterization and 2 high throughput screening of such molecules following molecular engineering.

  15. Bee venom phospholipase A2 as a membrane-binding vector for cell surface display or internalization of soluble proteins.

    Science.gov (United States)

    Babon, Aurélie; Wurceldorf, Thibault; Almunia, Christine; Pichard, Sylvain; Chenal, Alexandre; Buhot, Cécile; Beaumelle, Bruno; Gillet, Daniel

    2016-06-15

    We showed that bee venom phospholipase A2 can be used as a membrane-binding vector to anchor to the surface of cells a soluble protein fused to its C-terminus. ZZ, a two-domain derivative of staphylococcal protein A capable of binding constant regions of antibodies was fused to the C-terminus of the phospholipase or to a mutant devoid of enzymatic activity. The fusion proteins bound to the surface of cells and could themselves bind IgGs. Their fate depended on the cell type to which they bound. On the A431 carcinoma cell line the proteins remained exposed on the cell surface. In contrast, on human dendritic cells the proteins were internalized into early endosomes. Copyright © 2015 Elsevier Ltd. All rights reserved.

  16. Copper reverses cardiomyocyte hypertrophy through vascular endothelial growth factor-mediated reduction in the cell size.

    Science.gov (United States)

    Zhou, Yang; Jiang, Youchun; Kang, Y James

    2008-07-01

    Previous studies have shown that dietary copper supplementation reversed heart hypertrophy induced by pressure overload in a mouse model. The present study was undertaken to understand the cellular basis of copper-induced regression of cardiac hypertrophy. Primary cultures of neonatal rat cardiomyocytes were treated with phenylephrine (PE) at a final concentration of 100 microM in cultures for 48 h to induce cellular hypertrophy. The hypertrophied cardiomyocytes were exposed to copper sulfate at a final concentration of 5 microM in cultures for additional 24 h. This copper treatment reduced the size of the hypertrophied cardiomyocytes, as measured by flow cytometry, protein content in cells, cell volume and cardiomyocyte hypertrophy markers including beta-myosin heavy chain protein, skeletal alpha-actin, and atrial natriuretic peptide. Cell cycle analysis and cell sorting of p-histone-3 labeled cardiomyocytes indicated that cell division was not involved in the copper-induced regression of cardiomyocyte hypertrophy. Copper also inhibited PE-induced apoptosis, determined by a TUNEL assay. Because copper stimulates vascular endothelial growth factor (VEGF) production through activation of hypoxia-inducible transcription factor, an anti-VEGF antibody at a final concentration of 2 ng/ml in cultures was used and shown to blunt copper-induced regression of cell hypertrophy. Conversely, VEGF alone at a final concentration of 0.2 microg/ml reversed cell hypertrophy as the same as copper did. This study demonstrates that both copper and VEGF reduce the size of hypertrophied cardiomyocytes, and copper regression of cardiac hypertrophy is VEGF-dependent.

  17. Schedule-Dependent Antiangiogenic and Cytotoxic Effects of Chemotherapy on Vascular Endothelial and Retinoblastoma Cells.

    Science.gov (United States)

    Winter, Ursula; Mena, Hebe A; Negrotto, Soledad; Arana, Eloisa; Pascual-Pasto, Guillem; Laurent, Viviana; Suñol, Mariona; Chantada, Guillermo L; Carcaboso, Angel M; Schaiquevich, Paula

    2016-01-01

    Current treatment of retinoblastoma involves using the maximum dose of chemotherapy that induces tumor control and is tolerated by patients. The impact of dose and schedule on the cytotoxicity of chemotherapy has not been studied. Our aim was to gain insight into the cytotoxic and antiangiogenic effect of the treatment scheme of chemotherapy used in retinoblastoma by means of different in vitro models and to evaluate potential effects on multi-drug resistance proteins. Two commercial and two patient-derived retinoblastoma cell types and two human vascular endothelial cell types were exposed to increasing concentrations of melphalan or topotecan in a conventional (single exposure) or metronomic (7-day continuous exposure) treatment scheme. The concentration of chemotherapy causing a 50% decrease in cell proliferation (IC50) was determined by MTT and induction of apoptosis was evaluated by flow cytometry. Expression of ABCB1, ABCG2 and ABCC1 after conventional or metronomic treatments was assessed by RT-qPCR. We also evaluated the in vivo response to conventional (0.6 mg/kg once a week for 2 weeks) and metronomic (5 days a week for 2 weeks) topotecan in a retinoblastoma xenograft model. Melphalan and topotecan were cytotoxic to both retinoblastoma and endothelial cells after conventional and metronomic treatments. A significant decrease in the IC50 (median, 13-fold; range: 3-23) was observed following metronomic chemotherapy treatment in retinoblastoma and endothelial cell types compared to conventional treatment (p0.05). In mice, continuous topotecan lead to significantly lower tumor volumes compared to conventional treatment after 14 days of treatment (pretinoblastoma and endothelial cells to both chemotherapy agents with lower IC50 values compared to short-term treatment. These findings were validated in an in vivo model. None of the dosing modalities induced multidrug resistance mechanisms while apoptosis was the mechanism of cell death after both treatment

  18. Analysis of Active Components in Salvia Miltiorrhiza Injection Based on Vascular Endothelial Cell Protection

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    Shen Jie

    2014-09-01

    Full Text Available Correlation analysis based on chromatograms and pharmacological activities is essential for understanding the effective components in complex herbal medicines. In this report, HPLC and measurement of antioxidant properties were used to describe the active ingredients of Salvia miltiorrhiza injection (SMI. HPLC results showed that tanshinol, protocatechuic aldehyde, rosmarinic acid, salvianolic acid B, protocatechuic acid and their metabolites in rat serum may contribute to the efficacy of SMI. Assessment of antioxidant properties indicated that differences in the composition of serum powder of SMI caused differences in vascular endothelial cell protection. When bivariate correlation was carried out it was found that salvianolic acid B, tanshinol and protocatechuic aldehyde were active components of SMI because they were correlated to antioxidant properties.

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

    Science.gov (United States)

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

    2017-07-28

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

  20. Regulation of ERK5 by insulin and angiotensin-II in vascular smooth muscle cells

    International Nuclear Information System (INIS)

    Sharma, Girish; Goalstone, Marc Lee

    2007-01-01

    ERK5 is involved in proliferation of vascular smooth muscle cells (VSMC). The proliferative actions of insulin and angiotensin-II (A-II) in VSMC are mediated in part by ERK1/2. We hypothesized that insulin and A-II also regulate ERK5 activity in VSMC. Acute treatment ( 50 for insulin treatment effecting ERK1/2 and ERK5 phosphorylation was 1.5 and 0.1 nM, whereas the EC 50 for A-II was 2 nM, each. Insulin plus A-II induced an additive effect only on ERK5 phosphorylation. Inhibition of insulin- and A-II-stimulated phosphorylation of ERK5 and ERK1/2 by PD98059 and Wortmannin exhibited differential and time-dependent effects. Taken together, these data indicate that insulin and A-II regulate the activity of ERK5, but different from that seen for ERK1/2

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

    International Nuclear Information System (INIS)

    Luan Zhaoxia; Lan Xiaoli

    2003-01-01

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

  2. Bartonella henselae: subversion of vascular endothelial cell functions by translocated bacterial effector proteins.

    Science.gov (United States)

    Pulliainen, Arto Tapio; Dehio, Christoph

    2009-03-01

    Bartonella henselae (Bh) is a worldwide distributed zoonotic pathogen. Depending on the immune status of the infected individual this bacterium can cause a wide spectrum of clinical manifestations, ranging from cat scratch disease (CSD) to bacillary angiomatosis (BA) and bacillary peliosis (BP). BA and BP are characterized by tumor-like lesions at the skin or in the inner organs, respectively. These structures display pathological sprouting of capillaries with enlarged and hyperproliferated vascular endothelial cells (ECs) that are frequently found in close association with bacteria. Here we review the cellular changes observed upon Bh infection of ECs in vitro and outline the role of the VirB type IV secretion system (T4SS) and its translocated effector proteins in the modulation of EC signalling cascades. The current model how this virulence system could contribute to the vasoproliferative activity of Bh is described.

  3. Vascular smooth cell proliferation in perfusion culture of porcine carotid arteries

    International Nuclear Information System (INIS)

    Liao, Dan; Lin, Peter H.; Yao Qizhi; Chen Changyi

    2008-01-01

    Objective of this study was to develop a novel in vitro artery culture system to study vascular smooth muscle cell (SMC) proliferation of porcine carotid arteries in response to injury, basic fibroblast growth factor (FGF2), and FGF2 conjugated with cytotoxin saporin (SAP). Perfusion-cultured porcine carotid arteries remained contractile in response to norepinephrine and relaxant to acetylcholine for up to 96 h. SMC proliferation of cultured arteries was detected by bromodeoxyuridine incorporation in both non-injured and balloon-injured arteries. In the inner layer of the vessel wall near the lumen, SMC proliferation were less than 10% in uninjured vessels, 66% in injured vessels, 80% in injured vessels with FGF2 treatment, and 5% in injured vessels with treatment of FGF2-SAP. Thus, the cultured porcine carotid arteries were viable; and the injury stimulated SMC proliferation, which was significantly enhanced by FGF2 and inhibited by FGF2-SAP

  4. Collagenase-resistant collagen promotes mouse aging and vascular cell senescence.

    Science.gov (United States)

    Vafaie, Faran; Yin, Hao; O'Neil, Caroline; Nong, Zengxuan; Watson, Alanna; Arpino, John-Michael; Chu, Michael W A; Wayne Holdsworth, David; Gros, Robert; Pickering, J Geoffrey

    2014-02-01

    Collagen fibrils become resistant to cleavage over time. We hypothesized that resistance to type I collagen proteolysis not only marks biological aging but also drives it. To test this, we followed mice with a targeted mutation (Col1a1(r/r) ) that yields collagenase-resistant type I collagen. Compared with wild-type littermates, Col1a1(r/r) mice had a shortened lifespan and developed features of premature aging including kyphosis, weight loss, decreased bone mineral density, and hypertension. We also found that vascular smooth muscle cells (SMCs) in the aortic wall of Col1a1(r/r) mice were susceptible to stress-induced senescence, displaying senescence-associated ß-galactosidase (SA-ßGal) activity and upregulated p16(INK4A) in response to angiotensin II infusion. To elucidate the basis of this pro-aging effect, vascular SMCs from twelve patients undergoing coronary artery bypass surgery were cultured on collagen derived from Col1a1(r/r) or wild-type mice. This revealed that mutant collagen directly reduced replicative lifespan and increased stress-induced SA-ßGal activity, p16(INK4A) expression, and p21(CIP1) expression. The pro-senescence effect of mutant collagen was blocked by vitronectin, a ligand for αvß3 integrin that is presented by denatured but not native collagen. Moreover, inhibition of αvß3 with echistatin or with αvß3-blocking antibody increased senescence of SMCs on wild-type collagen. These findings reveal a novel aging cascade whereby resistance to collagen cleavage accelerates cellular aging. This interplay between extracellular and cellular compartments could hasten mammalian aging and the progression of aging-related diseases. © 2013 The Authors. Aging Cell published by the Anatomical Society and John Wiley & Sons Ltd.

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

    International Nuclear Information System (INIS)

    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

    Highlights: ► Anti-atherogenic effect of PL was examined using partial carotid ligation model in ApoE KO mice. ► PL prevented atherosclerotic plaque development, VSMCs proliferation, and NF-κB activation. ► Piperlongumine reduced vascular smooth muscle cell activation through PDGF-Rβ and NF-κB-signaling. ► 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-κB) 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 Cγ1, extracellular signal-regulated kinases 1 and 2 and Akt. Lastly, PL significantly attenuated activation of NF-κB—a downstream transcriptional regulator in PDGF receptor signaling, in response to PDGF-BB stimulation. In conclusion, our findings demonstrate a novel, therapeutic mechanism by which PL suppresses atherosclerosis plaque formation in vivo.

  6. Mitochondrial factor and cell cytokines associate with TCM syndrome scale in vascular dementia patients.

    Science.gov (United States)

    Sun, Linjuan

    2018-01-01

    Mitochondrial factor and cell cytokines play important roles in the incidence of vascular dementia (VD), but their correlations with inflammatory and mitochondrial factors and the role of both in the kidney essence deficiency pattern and phlegm turbidity blocking orifice pattern are not clear. This study was aimed at studying the correlations between the serum mitochondrial factor and cell cytokines with TCM Syndrome Scale in vascular dementia. According to the inclusion criteria we collected 108 vascular dementia patients which were divided into the kidney essence deficiency pattern and phlegm turbidity blocking orifice pattern based on the TCM Syndrome Scale. We measured serum tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), interleukin-18 (IL-18) concentration using an enzyme-linked immunosorbent assay (ELISA) and serum malondialdelyde (MDA) and superoxide dismutase (SOD) was quantified according to instructions of kits from the 108 patients (45 in kidney essence deficiency pattern, 63 in phlegm turbidity blocking orifice pattern). The scale scores were assessed using TCM Syndrome Scale, MMSE, Hachinski, Barthel, BBS, CDR. There was a significant difference on the scores of CDR and the factors of the memory, judgment, social affairs, personal care, family and hobbies among the two groups, The means of kidney essence deficiency group was higher than that of phlegm turbidity blocking orifice pattern group. IL-18 and SOD in the phlegm turbidity blocking orifice group was higher than those in the kidney essence deficiency pattern; IL-6 in phlegm turbidity blocking orifice pattern group was lower than that in the kidney essence deficiency pattern. By logistic regression analysis, we demonstrated that high concentration of IL-6, TNF-α, and MDA were associated with increased TCM syndrome scores in two groups, while IL-6, IL-18, TNF-α, SOD were associated with decreased MMSE, Barthel. Our study support the notion that IL-6 plays a more important role in the

  7. Hypoxia and the Presence of Human Vascular Endothelial Cells Affect Prostate Cancer Cell Invasion and Metabolism

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    Ellen Ackerstaff

    2007-12-01

    Full Text Available Tumor progression and metastasis are influenced by hypoxia, as well as by interactions between cancer cells and components of the stroma, such as endothelial cells. Here, we have used a magnetic resonance (MRcompatible invasion assay to further understand the effects of hypoxia on human prostate cancer cell invasion and metabolism in the presence and absence of human umbilical vein endothelial cells (HUVECs. Additionally, we compared endogenous activities of selected proteases related to invasion in PC-3 cells and HUVECs, profiled gene expression of PC-3 cells by microarray, evaluated cell proliferation of PC-3 cells and HUVECs by flow cytometry, under hypoxic and oxygenated conditions. The invasion of less-invasive DU-145 cells was not affected by either hypoxia or the presence of HUVECs. However, hypoxia significantly decreased the invasion of PC-3 cells. This hypoxia-induced decrease was attenuated by the presence of HUVECs, whereas under oxygenated conditions, HUVECs did not alter the invasion of PC-3 cells. Cell metabolism changed distinctly with hypoxia and invasion. The endogenous activity of selected extracellular proteases, although altered by hypoxia, did not fully explain the hypoxia-induced changes in invasion. Gene expression profiling indicated that hypoxia affects multiple cellular functions and pathways.

  8. Cancer cell-soluble factors reprogram mesenchymal stromal cells to slow cycling, chemoresistant cells with a more stem-like state

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    Ahmed El-Badawy

    2017-11-01

    Full Text Available Abstract Background Mesenchymal stem cells (MSCs play different roles in modulating tumor progression, growth, and metastasis. MSCs are recruited to the tumor site in large numbers and subsequently have an important microenvironmental role in modulating tumor progression and drug sensitivity. However, the effect of the tumor microenvironment on MSC plasticity remains poorly understood. Herein, we report a paracrine effect of cancer cells, in which they secrete soluble factors that promote a more stem-like state in bone marrow mesenchymal stem cells (BM-MSCs. Methods The effect of soluble factors secreted from MCF7, Hela, and HepG2 cancer cell lines on BM-MSCs was assessed using a Transwell indirect coculture system. After 5 days of coculture, BM-MSCs were characterized by flow cytometry for surface marker expression, by qPCR for gene expression profile, and by confocal immunofluorescence for marker expression. We then measured the sensitivity of cocultured BM-MSCs to chemotherapeutic agents, their cell cycle profile, and their response to DNA damage. The sphere formation, invasive properties, and in-vivo performance of BM-MSCs after coculture with cancer cells were also measured. Results Indirect coculture of cancer cells and BM-MSCs, without direct cell contact, generated slow cycling, chemoresistant spheroid stem cells that highly expressed markers of pluripotency, cancer cells, and cancer stem cells (CSCs. They also displayed properties of a side population and enhanced sphere formation in culture. Accordingly, these cells were termed cancer-induced stem cells (CiSCs. CiSCs showed a more mesenchymal phenotype that was further augmented upon TGF-β stimulation and demonstrated a high expression of the β-catenin pathway and ALDH1A1. Conclusions These findings demonstrate that MSCs, recruited to the tumor microenvironment in large numbers, may display cellular plasticity, acquire a more stem-like state, and acquire some properties of CSCs upon

  9. Brief secondhand smoke exposure depresses endothelial progenitor cells activity and endothelial function: sustained vascular injury and blunted nitric oxide production.

    Science.gov (United States)

    Heiss, Christian; Amabile, Nicolas; Lee, Andrew C; Real, Wendy May; Schick, Suzaynn F; Lao, David; Wong, Maelene L; Jahn, Sarah; Angeli, Franca S; Minasi, Petros; Springer, Matthew L; Hammond, S Katharine; Glantz, Stanton A; Grossman, William; Balmes, John R; Yeghiazarians, Yerem

    2008-05-06

    This study sought to analyze the effects of acute secondhand smoke (SHS) exposure on the number and function of endothelial progenitor cells (EPCs) over 24 h. Secondhand smoke increases the risk of vascular disease and is a major public health concern, but the mechanism(s) of action are not fully understood. Healthy nonsmokers (age SEM 30.3 +/- 1.3 years, n = 10) were exposed to 30 min of SHS yielding cotinine levels commonly observed in passive smokers and to smokefree air on 2 separate days. Measurements were taken before exposure (baseline), immediately after (0 h), and at 1 h, 2.5 h, and 24 h after. The EPCs (CD133(+)/KDR(+), CD34(+)/KDR(+)) and endothelial microparticles (EMPs: CD31(+)/CD41(-), CD144(+), CD62e(+)) were determined in blood using flow cytometry. The EPC chemotaxis toward vascular endothelial growth factor was measured. Endothelial function was assessed as flow-mediated dilation (FMD) using ultrasound. Secondhand smoke exposure increased EPCs and plasma vascular endothelial growth factor and completely abolished EPC chemotaxis during 24 h after exposure. Secondhand smoke increased EMPs and decreased FMD. Although FMD returned to baseline at 2.5 h, EMPs and vascular endothelial growth factor levels remained elevated at 24 h, suggesting endothelial activation and injury with functional impairment of the vascular endothelium. Exposure to smokefree air had no effect. Incubation of EPCs from nonexposed subjects with plasma isolated from SHS-exposed subjects in vitro decreased chemotaxis by blockade of vascular endothelial growth factor-stimulated nitric oxide production. Brief exposure to real-world levels of SHS leads to sustained vascular injury characterized by mobilization of dysfunctional EPCs with blocked nitric oxide production. Our results suggest that SHS not only affects the vascular endothelium, but also the function of EPCs.

  10. Seasonal changes in cell mediated immune responses to soluble Plasmodium falciparum antigens in children with haemoglobin AA and haemoglobin AS

    DEFF Research Database (Denmark)

    Abu-Zeid, Y A; Abdulhadi, N H; Theander, T G

    1992-01-01

    In this longitudinal study peripheral blood mononuclear cells (PBMC) were obtained before and during the malaria season from healthy HbAA and HbAS children. Cells were compared for proliferation in response to stimulation by soluble Plasmodium falciparum antigens (SPAg) or purified derivative...... of tuberculin (PPD). The lymphoproliferative responses to SPAg of the paired PBMC samples showed 2 distinct seasonal changes in relation to the haemoglobin phenotype. In HbAA children, the lymphoproliferative responses to SPAg were suppressed during the malaria season. In contrast, they were enhanced in Hb...

  11. Non-vascular smooth muscle cells in the human choroid: distribution, development and further characterization

    Science.gov (United States)

    May, Christian Albrecht

    2005-01-01

    To characterize further non-vascular smooth muscle cells (NVSMC) in the choroid of the human eye, extensive morphological studies were performed including a three-dimensional distribution of NVSMC in the adult human eye and their appearance during development. Whole mounts and sections through the choroid and sclera of eyes of 42 human donors (between the 13th week of gestation and 89 years of age) were stained with antibodies against smooth muscle actin and other markers for smooth muscle cells. On the basis of their morphological localization, three groups of NVSMC could be distinguished in the adult eyes: (a) a semicircular arrangement of NVSMC in the suprachoroid and inner sclera, around the entry of posterior ciliary arteries and nerves; (b) NVSMC parallel to the vessels in the posterior eye segment between the point of entry of the posterior ciliary arteries and the point of exit of the vortex veins; and (c) a dense plaque-like arrangement of NVSMC in the suprachoroid, overlying the foveal region. The last of these groups showed most pronounced interindividual differences. During development, the first NVSMC to be observed at the 20th week of gestation belonged to group b. A complete NVSMC network was first observed in a 6-year-old donor eye. All three groups stained positive for smoothelin, caldesmon and calponin in all localizations. The NVSMC show a distinct distribution that might reflect different aspects of their function in the choroid and suprachoroid. All cells could be histochemically characterized as truly contractile. PMID:16191166

  12. Brugia malayi microfilariae adhere to human vascular endothelial cells in a C3-dependent manner.

    Directory of Open Access Journals (Sweden)

    Jan-Hendrik Schroeder

    2017-05-01

    Full Text Available Brugia malayi causes the human tropical disease, lymphatic filariasis. Microfilariae (Mf of this nematode live in the bloodstream and are ingested by a feeding mosquito vector. Interestingly, in a remarkable co-evolutionary adaptation, Mf appearance in the peripheral blood follows a circadian periodicity and reaches a peak when the mosquito is most likely to feed. For the remaining hours, the majority of Mf sequester in the lung capillaries. This circadian phenomenon has been widely reported and is likely to maximise parasite fitness and optimise transmission potential. However, the mechanism of Mf sequestration in the lungs remains largely unresolved. In this study, we demonstrate that B. malayi Mf can, directly adhere to vascular endothelial cells under static conditions and under flow conditions, they can bind at high (but not low flow rates. High flow rates are more likely to be experienced diurnally. Furthermore, a non-periodic nematode adheres less efficiently to endothelial cells. Strikingly C3, the central component of complement, plays a crucial role in the adherence interaction. These novel results show that microfilariae have the ability to bind to endothelial cells, which may explain their sequestration in the lungs, and this binding is increased in the presence of inflammatory mediators.

  13. Ethanol interferes with thrombin mediated changes in the morphology and cytoskeleton of human vascular endothelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Pratt, K.J.; Rubin, R.; Hoek, J.; Williams, S.K. (Thomas Jefferson Univ., Philadelphia, PA (United States))

    1991-03-15

    The effect of physiological concentrations of ethanol (EtOH) on the response of human vascular endothelial cells (EC) to thrombin was examined Treatment of EC with EtOH concentrations of 20-85mM for 2-10 min. produced no significant changes in the morphology of 3- and 4-day monolayers established on fibronectin coated polystyrene. When examined immunofluorescently no significantly changes in the microfilament or microtubule structures were seen. Exposure of EC monolayers to 0.5 and 1 U/ml of thrombin for 1-60 minutes causes a concentration and time dependent monolayer retraction, evidenced by a general decrease in cell size, increase in visible gaps in the monolayer and redistribution of the microtubule and microfilament networks. Pretreatment of EC monolayers with EtOH for 3-5 minutes prior to addition of thrombin prevents the changes seen with thrombin alone. Immunofluorescent examination of the microfilament and microtubule structures suggests than EtOH may act in part via the microtubule network, which appears to be disorganized/disrupted when the EC are exposed to EtOH and then thrombin. Colchicine studies show that EC which have been pretreated with EtOH respond to colchicine differently then cells which have not previously seen EtOH. These data suggest that EtOH may alter EC monolayer responsiveness either by indirect changes which are reflected in cytoskeletal disorganization or possibly by direct influence on the cytoskeleton.

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

    Directory of Open Access Journals (Sweden)

    Lingfang Li

    2016-10-01

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

  15. Acetylbritannilactone Modulates Vascular Endothelial Growth Factor Signaling and Regulates Angiogenesis in Endothelial Cells.

    Directory of Open Access Journals (Sweden)

    Jingshan Zhao

    Full Text Available The present study was conducted to determine the effects of 1-O-acetylbritannilactone (ABL, a compound extracted from Inula britannica L., on vascular endothelial growth factor (VEGF signaling and angiogenesis in endothelial cells (ECs. We showed that ABL promotes VEGF-induced cell proliferation, growth, migration, and tube formation in cultured human ECs. Furthermore, the modulatory effect of ABL on VEGF-induced Akt, MAPK p42/44, and p38 phosphorylation, as well as on upstream VEGFR-2 phosphorylation, were associated with VEGF-dependent Matrigel angiogenesis in vivo. In addition, animals treated with ABL (26 mg/kg/day recovered blood flow significantly earlier than control animals, suggesting that ABL affects ischemia-mediated angiogenesis and arteriogenesis in vivo. Finally, we demonstrated that ABL strongly reduced the levels of VEGFR-2 on the cell surface, enhanced VEGFR-2 endocytosis, which consistent with inhibited VE-cadherin, a negative regulator of VEGF signaling associated with VEGFR-2 complex formation, but did not alter VE-cadherin or VEGFR-2 expression in ECs. Our results suggest that ABL may serve as a novel therapeutic intervention for various cardiovascular diseases, including chronic ischemia, by regulating VEGF signaling and modulating angiogenesis.

  16. Acetylbritannilactone Modulates Vascular Endothelial Growth Factor Signaling and Regulates Angiogenesis in Endothelial Cells.

    Science.gov (United States)

    Zhao, Jingshan; Niu, Honglin; Li, Aiying; Nie, Lei

    2016-01-01

    The present study was conducted to determine the effects of 1-O-acetylbritannilactone (ABL), a compound extracted from Inula britannica L., on vascular endothelial growth factor (VEGF) signaling and angiogenesis in endothelial cells (ECs). We showed that ABL promotes VEGF-induced cell proliferation, growth, migration, and tube formation in cultured human ECs. Furthermore, the modulatory effect of ABL on VEGF-induced Akt, MAPK p42/44, and p38 phosphorylation, as well as on upstream VEGFR-2 phosphorylation, were associated with VEGF-dependent Matrigel angiogenesis in vivo. In addition, animals treated with ABL (26 mg/kg/day) recovered blood flow significantly earlier than control animals, suggesting that ABL affects ischemia-mediated angiogenesis and arteriogenesis in vivo. Finally, we demonstrated that ABL strongly reduced the levels of VEGFR-2 on the cell surface, enhanced VEGFR-2 endocytosis, which consistent with inhibited VE-cadherin, a negative regulator of VEGF signaling associated with VEGFR-2 complex formation, but did not alter VE-cadherin or VEGFR-2 expression in ECs. Our results suggest that ABL may serve as a novel therapeutic intervention for various cardiovascular diseases, including chronic ischemia, by regulating VEGF signaling and modulating angiogenesis.

  17. 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.

  18. 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.

  19. Establishment of a vascular endothelial cell-reactive type II NKT cell clone from a rat model of autoimmune vasculitis.

    Science.gov (United States)

    Iinuma, Chihiro; Waki, Masashi; Kawakami, Ai; Yamaguchi, Madoka; Tomaru, Utano; Sasaki, Naomi; Masuda, Sakiko; Matsui, Yuki; Iwasaki, Sari; Baba, Tomohisa; Kasahara, Masanori; Yoshiki, Takashi; Paletta, Daniel; Herrmann, Thomas; Ishizu, Akihiro

    2015-02-01

    We previously generated a rat model that spontaneously developed small vessel vasculitis (SVV). In this study, a T cell clone reactive with rat vascular endothelial cells (REC) was established and named VASC-1. Intravenous injection of VASC-1 induced SVV in normal recipients. VASC-1 was a TCRαβ/CD3-positive CD4/CD8 double-negative T cell clone with expression of NKG2D. The cytokine mRNA profile under unstimulated condition was positive for IL-4 and IFN-γ but negative for IL-2 and IL-10. After interaction with REC, the mRNA expression of IL-2, IL-5 and IL-6 was induced in VASC-1, which was inhibited by blocking of CD1d on the REC surface. Although the protein levels of these cytokines seemed to be lower than the detection limit in the culture medium, IFN-γ was detectable. The production of IFN-γ from the VASC-1 stimulated with LPS-pre-treated REC was inhibited by the CD1d blockade on the REC. These findings indicated VASC-1 as an NKT cell clone. The NKT cell pool includes two major subsets, namely types I and II. Type I NKT cells are characterized by expression of semi-invariant TCRs and the potential to bind to marine sponge-derived α-galactosylceramide (α-GalCer) loaded on CD1d; whereas, type II NKT cells do not manifest these characteristics. VASC-1 exhibited a usage of TCR other than the type I invariant TCR α chain and did not bind to α-GalCer-loaded CD1d; therefore, it was determined as a type II NKT cell clone. The collective evidence suggested that REC-reactive type II NKT cells could be involved in the pathogenesis of SVV in rats. © The Japanese Society for Immunology. 2014. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

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

    Energy Technology Data Exchange (ETDEWEB)

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

    2012-08-01

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

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

    Czech Academy of Sciences Publication Activity Database

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

    2013-01-01

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

  2. Fluorescent water-Soluble Probes Based on Ammonium Cation Peg Substituted Perylenepisimides: Synthesis, Photophysical Properties, and Live Cell Images

    Science.gov (United States)

    Yang, Wei; Cai, Jiaxuan; Zhang, Shuchen; Yi, Xuegang; Gao, Baoxiang

    2018-01-01

    To synthesize perylenbisimides (PBI) fluorescent probes that will improve the water-soluble ability and the cytocompatibility, the synthesis and properties of fluorescent water-soluble probes based on dendritic ammonium cation polyethylene glycol (PEG) substituted perylenebisimides(GPDIs) are presented. As we expected, with increased ammonium cation PEG, the aggregation of the PBI in an aqueous solution is completely suppressed by the hydrophilic ammonium cation PEG groups. And the fluorescence quantum yield increases from 25% for GPDI-1 to 62% for GPDI-2. When incubated with Hela cells for 48 h, the viabilities are 71% (for GPDI-1) and 76% (for GPDI-2). Live cell imaging shows that these probes are efficiently internalized by HeLa cells. The study of the photophysical properties indicated increasing the ammonium cation PEG generation can increase the fluorescence quantum yield. Live cell imaging shows that with the ammonium cation PEG chains of perylenebisimides has high biocompatibility. The exceptionally low cytotoxicity is ascribed to the ammonium cation PEG chains, which protect the dyes from nonspecifically interacting with the extracellular proteins. Live cell imaging shows that ammonium cations PEG chains can promote the internalization of these probes.

  3. Oxidized low density lipoprotein increases RANKL level in human vascular cells. Involvement of oxidative stress

    Energy Technology Data Exchange (ETDEWEB)

    Mazière, Cécile, E-mail: maziere.cecile@chu-amiens.fr [Biochemistry Laboratory, South Hospital University, René Laennec Avenue, Amiens 80000 (France); Salle, Valéry [Internal Medicine, North Hospital University, Place Victor Pauchet, Amiens 80000 (France); INSERM U1088 (EA 4292), SFR CAP-Santé (FED 4231), University of Picardie – Jules Verne (France); Gomila, Cathy; Mazière, Jean-Claude [Biochemistry Laboratory, South Hospital University, René Laennec Avenue, Amiens 80000 (France)

    2013-10-18

    Highlights: •Oxidized LDL enhances RANKL level in human smooth muscle cells. •The effect of OxLDL is mediated by the transcription factor NFAT. •UVA, H{sub 2}O{sub 2} and buthionine sulfoximine also increase RANKL level. •All these effects are observed in human fibroblasts and endothelial cells. -- Abstract: Receptor Activator of NFκB Ligand (RANKL) and its decoy receptor osteoprotegerin (OPG) have been shown to play a role not only in bone remodeling but also in inflammation, arterial calcification and atherosclerotic plaque rupture. In human smooth muscle cells, Cu{sup 2+}-oxidized LDL (CuLDL) 10–50 μg/ml increased reactive oxygen species (ROS) and RANKL level in a dose-dependent manner, whereas OPG level was not affected. The lipid extract of CuLDL reproduced the effects of the whole particle. Vivit, an inhibitor of the transcription factor NFAT, reduced the CuLDL-induced increase in RANKL, whereas PKA and NFκB inhibitors were ineffective. LDL oxidized by myeloperoxidase (MPO-LDL), or other pro-oxidant conditions such as ultraviolet A (UVA) irradiation, incubation with H{sub 2}O{sub 2} or with buthionine sulfoximine (BSO), an inhibitor of glutathione synthesis{sub ,} also induced an oxidative stress and enhanced RANKL level. The increase in RANKL in pro-oxidant conditions was also observed in fibroblasts and endothelial cells. Since RANKL is involved in myocardial inflammation, vascular calcification and plaque rupture, this study highlights a new mechanism whereby OxLDL might, by generation of an oxidative stress, exert a deleterious effect on different cell types of the arterial wall.

  4. PRELIMINARY CLINICAL RESULTS WITH LIPOASPIRATE STROMAL VASCULAR CELL FRACTION IN TREATMENT OF PATIENTS WITH KNEE OSTEOARTHRITIS

    Directory of Open Access Journals (Sweden)

    E. Ya. Shevela

    2017-01-01

    Full Text Available The paper presents results of clinical application of autologous stromal-vascular fraction (SVF cells in patients with degenerative osteoarthritis (OA of the knee, grade II and III (Kellgren–Lawrence scale.We recruited six patients with knee OA (3 men and 3 women; median age 64 years with mean disease duration of 7 years. All the patients were administered a single intra-articular injection of autologous nucleated SVF cells at an average dose of 16.8±0.9 × 106 per joint (a total of 11 joints. The patients did not experience any serious side effects (allergic, toxic or inflammatory related to the knee injection. Patient surveys at 1 month after SVF administration revealed a decrease in the severity of pain, as measured by a visual analog scale (VAS and a specialized 100-point scale KOOS (subscale "pain" (p < 0.05 on both scales. Moreover, the patients reported improvement in the joint functions and quality of life related to affected joints on a KOOS scale (p < 0.05. These positive clinical changes persisted during 6 month follow up. Significant improvements were noted in ultrasound findings, with increased thickness of the cartilage layer at 3 months (in 73% of cases and at 6 months (in 82%. Our pilot study demonstrated the safety and tolerability of intra-articular injection of autologous SVF cells in patients with moderate to severe OA. The results obtained also indicate a significant antiinflammatory effect of autologous adipose tissue SVF cells, which is manifested at the early stages of cell therapy. Our further investigations will be focused on exploring the SVF stimulatory effects on regeneration of damaged joints.

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

    NARCIS (Netherlands)

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

    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

  6. Engraftment of donor mesenchymal stem cells in chimeric BXSB includes vascular endothelial cells and hepatocytes

    Directory of Open Access Journals (Sweden)

    Jones OY

    2011-12-01

    Full Text Available Olcay Y Jones1, Faysal Gok2, Elisabeth J Rushing3, Iren Horkayne-Szakaly4, Atif A Ahmed51Department of Pediatrics, Walter Reed National Military Medical Center, Bethesda, MD, USA; 2Department of Pediatrics, Gulhane Military Medical Academy, Ankara, Turkey; 3Institut für Neuropathologie, Universitäts Spital Zürich, Zürich, Switzerland; 4Department of Neuropathology, Armed Forces Institute of Pathology, Washington, DC, USA; 5Department of Pathology and Laboratory Medicine, Children's Mercy Hospitals and Clinics, Kansas City, MO, USAAbstract: Somatic tissue engraftment was studied in BXSB mice treated with mesenchymal stem cell transplantation. Hosts were conditioned with nonlethal radiation prior to introducing donor cells from major histocompatibility complex-matched green fluorescent protein transgenic mice. Transplant protocols differed for route of injection, ie, intravenous (i.v. versus intraperitoneal (i.p., and source of mesenchymal stem cells, ie, unfractionated bone marrow cells, ex vivo expanded mesenchymal stem cells, or bone chips. Tissue chimerism was determined after short (10–12 weeks or long (62 weeks posttransplant follow-up by immunohistochemistry for green fluorescent protein. Engraftment of endothelial cells was seen in several organs including liver sinusoidal cells in i.v. treated mice with ex vivo expanded mesenchymal stem cells or with unfractionated bone marrow cells. Periportal engraftment of liver hepatocytes, but not engraftment of endothelial cells, was found in mice injected i.p. with bone chips. Engraftment of adipocytes was a common denominator in both i.v. and i.p. routes and occurred during early phases post-transplant. Disease control was more robust in mice that received both i.v. bone marrow and i.p. bone chips compared to mice that received i.v. bone marrow alone. Thus, the data support potential use of mesenchymal stem cell transplant for treatment of severe lupus. Future studies are needed to optimize

  7. The inhibitory effect of simvastatin and aspirin on histamine responsiveness in human vascular endothelial cells.

    Science.gov (United States)

    Absi, Mais; Bruce, Jason I; Ward, Donald T

    2014-04-01

    Statins and aspirin deliver well-established cardiovascular benefits resulting in their increased use as combined polypills to decrease risk of stroke and heart disease. However, the direct endothelial effect of combined statin/aspirin cotreatment remains unclear. Histamine is an inflammatory mediator that increases vascular permeability, and so we examined the effect of treating human umbilical vein endothelial cells (HUVECs) for 24 h with 1 μM simvastatin and 100 μM aspirin on histamine responsiveness. Subsequent histamine (1 μM) challenge increased intracellular calcium (Ca(2+)i) concentration, an effect that was significantly inhibited by combined simvastatin/aspirin pretreatment but not when then the compounds were given separately, even at 10-fold higher concentrations. In contrast, the Ca(2+)i mobilization response to ATP challenge (10 μM) was not inhibited by combined simvastatin/aspirin pretreatment. The H1 receptor antagonist pyrilamine significantly inhibited both histamine-induced Ca(2+)i mobilization and extracellular signal-regulated kinase (ERK) activation, whereas ranitidine (H2 receptor antagonist) was without effect. However, combined simvastatin/aspirin pretreatment failed to decrease H1 receptor protein expression ruling out receptor downregulation as the mechanism of action. Histamine-induced ERK activation was also inhibited by atorvastatin pretreatment, while simvastatin further inhibited histamine-induced vascular endothelial cadherin phosphorylation as well as altered HUVEC morphology and inhibited actin polymerization. Therefore, in addition to the known therapeutic benefits of statins and aspirin, here we provide initial cellular evidence that combined statin/aspirin treatment inhibits histamine responsiveness in HUVECs.

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

    Science.gov (United States)

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

    2013-01-01

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

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

    Directory of Open Access Journals (Sweden)

    Ming Zhang

    2014-01-01

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

  10. Magnolol inhibits migration of vascular smooth muscle cells via cytoskeletal remodeling pathway to attenuate neointima formation

    International Nuclear Information System (INIS)

    Karki, Rajendra; Kim, Seong-Bin; Kim, Dong-Wook

    2013-01-01

    Background: Increased proliferation and migration of vascular smooth muscle cells (VSMCs) contribute importantly to the formation of both atherosclerotic and restenotic lesions. The objective of this study was to investigate the effect of magnolol on VSMC migration. Methods: The proteolytic activity of matrix metalloproteinases (MMPs) in tumor necrosis factor alpha (TNF-α) stimulated VSMCs was performed by gelatin zymography. VSMC migration was assessed by wound healing and Boyden chamber methods. Collagen induced VSMC adhesion was determined by spectrofluorimeter and stress fibers formation was evaluated by fluorescence microscope. The expression of signaling molecules involved in stress fibers formation was determined by western blot. The phosphorylation of myosin light chain (MLC20) was determined by urea-glycerol polyacrylamide gel electrophoresis. Immunohistochemistry was performed to determine the expression of β1-integrin and collagen type I in the injured carotid arteries of rats on day 35 after vascular injury. Results: VSMC migration was strongly inhibited by magnolol without affecting MMPs expression. Also, magnolol inhibited β1-integrin expression, FAK phosphorylation and RhoA and Cdc42 activation to inhibit the collagen induced stress fibers formation. Moreover, magnolol inhibited the phosphorylation of MLC20. Our in vivo results showed that magnolol inhibited β1-integrin expression, collagen type I deposition and FAK phosphorylation in injured carotid arteries without affecting MMP-2 activity. Conclusions: Magnolol inhibited VSMC migration via inhibition of cytoskeletal remodeling pathway to attenuate neointima formation. General significance: This study provides a rationale for further evaluation of magnolol for the management of atherosclerosis and restenosis. - Highlights: • Magnolol strongly inhibited migration of VSMCs. • Magnolol inhibited stress fibers formation. • MLC20 phosphorylation was also inhibited by magnolol. • Anti

  11. 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

  12. An increase in vascular endothelial growth factor (VEGF and VEGF soluble receptor-1 (sFlt-1 are associated with early recurrent spontaneous abortion.

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    Lihong Pang

    Full Text Available Recurrent spontaneous abortion (RSA is a health problem that affects approximately 1% to 5% reproductive age woman. Yet, in around half of these patients, the mechanism for RSA is unexplained. Recent studies have indicated that placental ischemia/hypoxia and endothelial dysfunction are important factors in miscarriage. Other studies have indicated that the level and expression of soluble FMS-like tyrosine kinase-1 (sFlt1 is increased under a hypoxic environment. However, decreased sFlt-1 in the maternal circulation during the first trimester has recently been proposed as a potential marker for identifying risk of pregnancy loss. In this prospective study clinical samples were obtained within a short time after the fetal death, protein expression and maternal serum levels of sFlt1 were assessed and compared to samples taken from those with normal pregnancies. Our results indicate that levels of VEGF and sFlt-1 are both increased in women during early pregnancy compared women that are not pregnant (p<0.05 indicating that VEGF and sFlt-1 are both associated with pregnancy. More importantly, we detected a significant (p<0.05 increase in sFlt1 and VEGF levels and expression in the RSA patients who suffered subsequent miscarriages compare to controls. These results demonstrate that there is likely a relationship between VEGF, sFlt-1 and RSA suggesting that the high levels and over expression of sFlt-1 and VEGF might be associated with the pathogenesis of RSA.

  13. Fli+ etsrp+ hemato-vascular progenitor cells proliferate at the lateral plate mesoderm during vasculogenesis in zebrafish.

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    Chang Zoon Chun

    2011-02-01

    Full Text Available Vasculogenesis, the de novo formation of blood vessels from precursor cells is critical for a developing embryo. However, the signals and events that dictate the formation of primary axial vessels remain poorly understood.In this study, we use ets-related protein-1 (etsrp, which is essential for vascular development, to analyze the early stages of vasculogenesis in zebrafish. We found etsrp(+ cells of the head, trunk and tail follow distinct developmental sequences. Using a combination of genetic, molecular and chemical approaches, we demonstrate that fli(+etsrp(+ hemato-vascular progenitors (FEVPs are proliferating at the lateral plate mesoderm (LPM. The Shh-VEGF-Notch-Hey2 signaling pathway controls the proliferation process, and experimental modulation of single components of this pathway alters etsrp(+ cell numbers at the LPM.This study for the first time defines factors controlling proliferation, and cell numbers of pre-migratory FEVPs in zebrafish.

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

    OpenAIRE

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

    2015-01-01

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

  15. A Novel Mammary Fat Pad Transplantation Technique to Visualize the Vessel Generation of Vascular Endothelial Stem Cells.

    Science.gov (United States)

    Yu, Qing Cissy; Song, Wenqian; Lai, Dengwen; Zeng, Yi Arial

    2017-08-03

    Endothelial cells (ECs) are the fundamental building blocks of the vascular architecture and mediate vascular growth and remodeling to ensure proper vessel development and homeostasis. However, studies on endothelial lineage hierarchy remain elusive due to the lack of tools to gain access as well as to directly evaluate their behavior in vivo. To address this shortcoming, a new tissue model to study angiogenesis using the mammary fat pad has been developed. The mammary gland develops mostly in the postnatal stages, including puberty and pregnancy, during which robust epithelium proliferation is accompanied by extensive vascular remodeling. Mammary fat pads provide space, matrix, and rich angiogenic stimuli from the growing mammary epithelium. Furthermore, mammary fat pads are located outside the peritoneal cavity, making them an easily accessible grafting site for assessing the angiogenic potential of exogenous cells. This work also describes an efficient tracing approach using fluorescent reporter mice to specifically label the targeted population of vascular endothelial stem cells (VESCs) in vivo. This lineage tracing method, coupled with subsequent tissue whole-mount microscopy, enable the direct visualization of targeted cells and their descendants, through which the proliferation capability can be quantified and the differentiation commitment can be fate-mapped. Using these methods, a population of bipotent protein C receptor (Procr) expressing VESCs has recently been identified in multiple vascular systems. Procr + VESCs, giving rise to both new ECs and pericytes, actively contribute to angiogenesis during development, homeostasis, and injury repair. Overall, this manuscript describes a new mammary fat pad transplantation and in vivo lineage tracing techniques that can be used to evaluate the stem cell properties of VESCs.

  16. Microparticle Shedding by Erythrocytes, Monocytes and Vascular Smooth Muscular Cells Is Reduced by Aspirin in Diabetic Patients.

    Science.gov (United States)

    Chiva-Blanch, Gemma; Suades, Rosa; Padró, Teresa; Vilahur, Gemma; Peña, Esther; Ybarra, Juan; Pou, Jose M; Badimon, Lina

    2016-07-01

    Diabetes mellitus is associated with an enhanced risk for cardiovascular disease and its prevalence is increasing. Diabetes induces metabolic stress on blood and vascular cells, promoting platelet activation and vascular dysfunction. The level of vascular cell activation can be measured by the number and phenotype of microparticles found in the circulation. The aim of this study was to investigate the effect of a platelet-inhibitory dose of aspirin on the number and type of microparticles shed to the circulation. Forty-three diabetic patients were enrolled in the study and received a daily dose of 100mg of aspirin for 10 days to cover the average platelet life-span in the circulation. Before and after the intervention period, circulating microparticles were characterized and quantified by flow cytometry. Type 1 diabetic patients had about twice the number of tissue factor-positive circulating microparticles (derived both from platelets and monocytes) and endothelial-derived E-selectin positive microparticles than type 2 diabetic patients. Aspirin therapy significantly inhibited platelets since cyclooxygenase 1 derived thromboxane generation levels were reduced by 99%. Microparticles derived from erythrocytes, activated monocytes, and smooth muscle cells were significantly reduced after 10 days of aspirin administration. These results indicate that: a) vascular and blood cells in type 1 diabetic patients are exposed to more sustained stress shown by their specific microparticle origin and levels; b) aspirin therapy inhibits vascular wall cell activation and microparticle shedding, and c) the effects of aspirin are similar in type 1 and 2 diabetes. Copyright © 2016 Sociedad Española de Cardiología. Published by Elsevier España, S.L.U. All rights reserved.

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

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    Weissmann Norbert

    2005-11-01

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

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

    International Nuclear Information System (INIS)

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

    2004-01-01

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

  19. Suppression of vascular endothelial growth factor expression by cannabinoids in a canine osteosarcoma cell line

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    Figueiredo AS

    2013-07-01

    Full Text Available Andreza S Figueiredo,1 Hiram J García-Crescioni,1 Sandra C Bulla,1 Matthew K Ross,2 Chelsea McIntosh,1 Kari Lunsford,3 Camilo Bulla11Department of Pathobiology and Population Medicine, 2Department of Basic Sciences, 3Department of Clinical Sciences and Animal Health Center, College of Veterinary Medicine, Mississippi State University, Mississippi State, MS, USAAbstract: Vascular endothelial growth factor (VEGF is a key regulator in both physiologic and pathologic angiogenesis, and cannabinoids decrease VEGF release in human and murine cancer cells. The aim of this study was to assess the in vitro effects of a synthetic cannabinoid, WIN-55,212-2, on the expression of the proangiogenic factor VEGF-A in the canine osteosarcoma cell line 8. After analysis of gene expression by quantitative real-time polymerase chain reaction, the compound decreased VEGF-A expression by 35% ± 10% (P < 0.0001 as compared with the control. This synthetic cannabinoid shows promise as a potential inhibitor of angiogenesis, and further studies are warranted to investigate its in vivo effects and to explore the potential of this and related compounds as adjuvant cancer therapy in the dog.Keywords: dog, cancer, angiogenesis, cannabinoids

  20. Nutritional milieu of isolated stromal vascular cells determines their proliferative, adipogenic, and lipogenic capacity in vitro.

    Science.gov (United States)

    Kadegowda, Anil K G; Wright, Asher; Duckett, Susan K

    2014-01-01

    The objective was to determine the effect of nutritional milieu of isolated stromal vascular (SV) cells on proliferative capacity of preadipocytes, and adipogenic and lipogenic capacity in adipocytes in vitro. Proliferation of the preadipocytes increased over time with 48 and 72 h being greater than 24 h; however, preadipocytes from steers supplemented with corn (LC) had lower proliferation rates compared with those without corn grain supplementation (L) at 72 h. Adipocyte cultures isolated from LC group had higher mean diameter on d 4 and 6, and higher mean volume on d 0, 4, 6, and 12 of culture. Adipocytes from steers supplemented with corn grain (LC) had lower expression of key adipogenic genes during extended days in culture. The results show that prior nutritional treatment of the donor animal used to isolate SV cultures alters their proliferative, adipogenic, and lipogenic capacity in culture. These differences may be related to lower induction/expression of AP2 gene in the adipose cultures from corn supplemented group. Corn grain supplementation to steers grazing legumes could have stimulated more active adipogenic progenitor cells to differentiate, which would leave fewer behind in the SV pool for subsequent isolation.

  1. Effect of a rosmarinic acid supplemented hemodialysis fluid on inflammation of human vascular endothelial cells

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    W-J. Wang

    2017-10-01

    Full Text Available Chronic systemic inflammation and repetitive damage of vascular endothelia by incompatible dialysis system are probable causes of cardiovascular disease in patients on dialysis. The present study aimed to assess in vitro biocompatibility and anti-inflammatory effect of hemodialysis fluid supplemented with rosmarinic acid (RA using human umbilical vein endothelial cells (HUVEC. HUVECs (5×106 cells/mL were pre-exposed to 1 μg/mL of lipopolysaccharides (LPS and incubated with RA-supplemented hemodialysis fluid (HDF. Cytotoxicity was assessed qualitatively by morphologic assessment and quantitatively by MTT assay. Expressions of proinflammatory mediators were assessed using quantitative real-time PCR and production of NO was quantified. Phosphorylation of AKT and nuclear localization of nuclear factor kappa B (NF-κB were examined using western blotting. Exposure of HUVECs to RA-supplemented HDF had no influence on morphology and viability. Inhibition of proinflammatory mediator production in HUVECs by RA supplementation to HDF was significant in a dose-dependent manner. Exposure to RA-supplemented HDF resulted in a decrease in nitric oxide synthase expression and reduction of NO production in LPS-stimulated HUVECs. RA supplementation of HDF suppressed Akt activation in LPS-stimulated HUVECs. In addition, the level of cellular IκB was increased in parallel to a reduced nuclear translocation of NF-κB in LPS-induced endothelial cells. Our results suggest that RA-supplemented HDF is biocompatible and significantly suppressed inflammation induced in endothelial cells. In this respect, the use of HDF supplemented with RA could alleviate inflammation and improve long-term treatment of patients with renal failure on dialysis. Further clinical studies are required to confirm the effects.

  2. Global Remodeling of the Vascular Stem Cell Niche in Bone Marrow of Diabetic Patients

    Science.gov (United States)

    Spinetti, Gaia; Cordella, Daniela; Fortunato, Orazio; Sangalli, Elena; Losa, Sergio; Gotti, Ambra; Carnelli, Franco; Rosa, Francesco; Riboldi, Stefano; Sessa, Fausto; Avolio, Elisa; Beltrami, Antonio Paolo; Emanueli, Constanza; Madeddu, Paolo

    2013-01-01

    Rationale The impact of diabetes mellitus on bone marrow (BM) structure is incompletely understood. Objective Investigate the effect of type-2 diabetes mellitus (T2DM) on BM microvascular and hematopoietic cell composition in patients without vascular complications. Methods and Results Bone samples were obtained from T2DM patients and nondiabetic controls (C) during hip replacement surgery and from T2DM patients undergoing amputation for critical limb ischemia. BM composition was assessed by histomorphometry, immunostaining, and flow cytometry. Expressional studies were performed on CD34pos immunosorted BM progenitor cells (PCs). Diabetes mellitus causes a reduction of hematopoietic tissue, fat deposition, and microvascular rarefaction, especially when associated with critical limb ischemia. Immunohistochemistry documented increased apoptosis and reduced abundance of CD34pos-PCs in diabetic groups. Likewise, flow cytometry showed scarcity of BM PCs in T2DM and T2DM+critical limb ischemia compared with C, but similar levels of mature hematopoietic cells. Activation of apoptosis in CD34pos-PCs was associated with upregulation and nuclear localization of the proapoptotic factor FOXO3a and induction of FOXO3a targets, p21 and p27kip1. Moreover, microRNA-155, which regulates cell survival through inhibition of FOXO3a, was downregulated in diabetic CD34pos-PCs and inversely correlated with FOXO3a levels. The effect of diabetes mellitus on anatomic and molecular end points was confirmed when considering background covariates. Furthermore, exposure of healthy CD34pos-PCs to high glucose reproduced the transcriptional changes induced by diabetes mellitus, with this effect being reversed by forced expression of microRNA-155. Conclusions We provide new anatomic and molecular evidence for the damaging effect of diabetes mellitus on human BM, comprising microvascular rarefaction and shortage of PCs attributable to activation of proapoptotic pathway. PMID:23250986

  3. Early diagnosis of diabetic vascular complications: impairment of red blood cell deformability

    Science.gov (United States)

    Shin, Sehyun; Ku, Yunhee; Park, Cheol-Woo; Suh, Jang-Soo

    2006-02-01

    Reduced deformability of red blood cells (RBCs) may play an important role on the pathogenesis of chronic vascular complications of diabetes mellitus. However, available techniques for measuring RBC deformability often require washing process after each measurement, which is not optimal for day-to-day clinical use at point of care. The objectives of the present study are to develop a device and to delineate the correlation of impaired RBC deformability with diabetic nephropathy. We developed a disposable ektacytometry to measure RBC deformability, which adopted a laser diffraction technique and slit rheometry. The essential features of this design are its simplicity (ease of operation and no moving parts) and a disposable element which is in contact with the blood sample. We studied adult diabetic patients divided into three groups according to diabetic complications. Group I comprised 57 diabetic patients with normal renal function. Group II comprised 26 diabetic patients with chronic renal failure (CRF). Group III consisted of 30 diabetic subjects with end-stage renal disease (ESRD) on hemodialysis. According to the renal function for the diabetic groups, matched non-diabetic groups were served as control. We found substantially impaired red blood cell deformability in those with normal renal function (group I) compared to non-diabetic control (P = 0.0005). As renal function decreases, an increased impairment in RBC deformability was found. Diabetic patients with chronic renal failure (group II) when compared to non-diabetic controls (CRF) had an apparently greater impairment in RBC deformability (P = 0.07). The non-diabetic cohort (CRF), on the other hand, manifested significant impairment in red blood cell deformability compared to healthy control (P = 0.0001). The newly developed slit ektacytometer can measure the RBC deformability with ease and accuracy. In addition, progressive impairment in cell deformability is associated with renal function loss in all

  4. Erythropoietin attenuates pulmonary vascular remodeling in experimental pulmonary arterial hypertension through interplay between endothelial progenitor cells and heme-oxygenase

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    Rosa L.E. Loon

    2015-08-01

    Full Text Available BackgroundPulmonary arterial hypertension (PAH is a pulmonary vascular disease with a high mortality, characterized by typical angio-proliferative lesions. Erythropoietin (EPO attenuates pulmonary vascular remodeling in PAH. We postulated that EPO acts through mobilization of endothelial progenitor cells (EPCs and activation of the cytoprotective enzyme heme oxygenase-1 (HO1.MethodsRats with flow-associated PAH, resembling pediatric PAH, were treated with HO-1 inducer EPO in the presence or absence of the selective HO-activity-inhibitor tin-mesoporphyrin (SnMP. HO-activity, circulating EPCs and pulmonary vascular lesions were assessed after 3 weeks.ResultsIn PAH-rats, circulating EPCs were decreased and HO-activity was increased compared to control. EPO-treatment restored circulating EPCs and improved pulmonary vascular remodeling, as shown by a reduced wall thickness and occlusion rate of the intra-acinar vessels. Inhibition of HO-activity with SnMP aggravated PAH. Moreover, SnMP treatment abrogated EPO-induced amelioration of pulmonary vascular remodeling, while surprisingly further increasing circulating EPCs as compared with EPO alone.ConclusionsIn experimental PAH, EPO treatment restored the number of circulating EPC’s to control level, improved pulmonary vascular remodeling, and showed important interplay with HO-activity. Inhibition of increased HO-activity in PAH-rats exacerbated progression of pulmonary vascular remodeling, despite the presence of restored numbers of circulating EPC’s. We suggest that both EPO-induced HO1 and EPCs are promising targets to ameliorate the pulmonary vasculature in PAH.

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

    Science.gov (United States)

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

    2013-04-01

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

  6. Inhibition of myeloperoxidase decreases vascular oxidative stress and increases vasodilatation in sickle cell disease mice.

    Science.gov (United States)

    Zhang, Hao; Xu, Hao; Weihrauch, Dorothee; Jones, Deron W; Jing, Xigang; Shi, Yang; Gourlay, David; Oldham, Keith T; Hillery, Cheryl A; Pritchard, Kirkwood A

    2013-11-01

    Activated leukocytes and polymorphonuclear neutrophils (PMN) release myeloperoxidase (MPO), which binds to endothelial cells (EC), is translocated, and generates oxidants that scavenge nitric oxide (NO) and impair EC function. To determine whether MPO impairs EC function in sickle cell disease (SCD), control (AA) and SCD mice were treated with N-acetyl-lysyltyrosylcysteine-amide (KYC). SCD humans and mice have high plasma MPO and soluble L-selectin (sL-selectin). KYC had no effect on MPO but decreased plasma sL-selectin and malondialdehyde in SCD mice. MPO and 3-chlorotyrosine (3-ClTyr) were increased in SCD aortas. KYC decreased MPO and 3-ClTyr in SCD aortas to the levels in AA aortas. Vasodilatation in SCD mice was impaired. KYC increased vasodilatation in SCD mice more than 2-fold, to ∼60% of levels in AA mice. KYC inhibited MPO-dependent 3-ClTyr formation in EC proteins. SCD mice had high plasma alanine transaminase (ALT), which tended to decrease in KYC-treated SCD mice (P = 0.07). KYC increased MPO and XO/XDH and decreased 3-ClTyr and 3-nitrotyrosine (3-NO₂Tyr) in SCD livers. These data support the hypothesis that SCD increases release of MPO, which generates oxidants that impair EC function and injure livers. Inhibiting MPO is an effective strategy for decreasing oxidative stress and liver injury and restoring EC function in SCD.

  7. Platform technologies for decellularization, tunic-specific cell seeding, and in vitro conditioning of extended length, small diameter vascular grafts.

    Science.gov (United States)

    Fercana, George; Bowser, Devon; Portilla, Margarita; Langan, Eugene M; Carsten, Christopher G; Cull, David L; Sierad, Leslie N; Simionescu, Dan T

    2014-12-01

    The aim of this study was to generate extended length, small diameter vascular scaffolds that could serve as potential grafts for treatment of acute ischemia. Biological tissues are considered excellent scaffolds, which exhibit adequate biological, mechanical, and handling properties; however, they tend to degenerate, dilate, and calcify after implantation. We hypothesized that chemically stabilized acellular arteries would be ideal scaffolds for development of vascular grafts for peripheral surgery applications. Based on promising historical data from our laboratory and others, we chose to decellularize bovine mammary and femoral arteries and test them as scaffolds for vascular grafting. Decellularization of such long structures required development of a novel "bioprocessing" system and a sequence of detergents and enzymes that generated completely acellular, galactose-(α1,3)-galactose (α-Gal) xenoantigen-free scaffolds with preserved collagen, elastin, and basement membrane components. Acellular arteries exhibited excellent mechanical properties, including burst pressure, suture holding strength, and elastic recoil. To reduce elastin degeneration, we treated the scaffolds with penta-galloyl glucose and then revitalized them in vitro using a tunic-specific cell approach. A novel atraumatic endothelialization protocol using an external stent was also developed for the long grafts and cell-seeded constructs were conditioned in a flow bioreactor. Both decellularization and revitalization are feasible but cell retention in vitro continues to pose challenges. These studies support further efforts toward clinical use of small diameter acellular arteries as vascular grafts.

  8. Adhesion and growth of vascular endothelial and smooth muscle cells on artificial vascular prostheses and protein-coated surfaces

    Czech Academy of Sciences Publication Activity Database

    Chlupáč, Jaroslav; Filová, Elena; Brynda, Eduard; Remy-Zolghadri, M.; Bačáková, Lucie

    2005-01-01

    Roč. 3, č. S1 (2005), S8-S9 ISSN 1214-021X. [Cells /6./ - Biological days /18./. 24.10.2005-26.10.2005, České Budějovice] R&D Projects: GA MŠk(CZ) 1M0510; GA AV ČR(CZ) IAA5011301; GA AV ČR(CZ) IAA4050202 Institutional research plan: CEZ:AV0Z50110509 Keywords : tissue engineering * bioartificial vessel * dynamic culture system * hemodynamic bench * fibronectin * collagen * endothelialization * sex-related differences Subject RIV: EI - Biotechnology ; Bionics

  9. Vascular Endothelial Growth Factor, Irradiation, and Axitinib Have Diverse Effects on Motility and Proliferation of Glioblastoma Multiforme Cells

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    Reinhardt Krcek

    2017-08-01

    Full Text Available Glioblastoma multiforme (GBM is the most common primary brain tumor. It is highly aggressive with an unfavorable prognosis for the patients despite therapies including surgery, irradiation, and chemotherapy. One important characteristic of highly vascularized GBM is the strong expression of vascular endothelial growth factor (VEGF. VEGF has become a new target in the treatment of GBM, and targeted therapies such as the VEGF-receptor blocker axitinib are in clinical trials. Most studies focus on VEGF-induced angiogenesis, but only very few investigations analyze autocrine or paracrine effects of VEGF on the tumor cells. In this study, we examined the impact of VEGF, irradiation, and axitinib on cell proliferation and cell motility in human GBM cell lines U-251 and U-373. VEGF receptor 2 was shown to be expressed within both cell lines by using PCR and immunochemistry. Moreover, we performed 24-h videography to analyze motility, and a viability assay for cell proliferation. We observed increasing effects of VEGF and irradiation on cell motility in both cell lines, as well as strong inhibiting effects on cellular motility by VEGF-receptor blockade using axitinib. Moreover, axitinib diminished irradiation induced accelerating effects. While VEGF stimulation or irradiation did not affect cell proliferation, axitinib significantly decreased cell proliferation in both cell lines. Therefore, the impairment of VEGF signaling might have a crucial role in the treatment of GBM.

  10. Protein kinase A-alpha directly phosphorylates FoxO1 in vascular endothelial cells to regulate expression of vascular cellular adhesion molecule-1 mRNA.

    Science.gov (United States)

    Lee, Ji-Won; Chen, Hui; Pullikotil, Philomena; Quon, Michael J

    2011-02-25

    FoxO1, a forkhead box O class transcription factor, is abundant in insulin-responsive tissues. Akt, downstream from phosphatidylinositol 3-kinase in insulin signaling, phosphorylates FoxO1 at Thr(24), Ser(256), and Ser(319), negatively regulating its function. We previously reported that dehydroepiandrosterone-stimulated phosphorylation of FoxO1 in endothelial cells requires cAMP-dependent protein kinase α (PKA-α). Therefore, we hypothesized that FoxO1 is a novel direct substrate for PKA-α. Using an immune complex kinase assay with [γ-(32)P]ATP, purified PKA-α directly phosphorylated wild-type FoxO1 but not FoxO1-AAA (mutant with alanine substitutions at known Akt phosphorylation sites). Phosphorylation of wild-type FoxO1 (but not FoxO1-AAA) was detectable using phospho-specific antibodies. Similar results were obtained using purified GST-FoxO1 protein as the substrate. Thus, FoxO1 is a direct substrate for PKA-α in vitro. In bovine aortic endothelial cells, interaction between endogenous PKA-α and endogenous FoxO1 was detected by co-immunoprecipitation. In human aortic endothelial cells (HAEC), pretreatment with H89 (PKA inhibitor) or siRNA knockdown of PKA-α decreased forskolin- or prostaglandin E(2)-stimulated phosphorylation of FoxO1. In HAEC transfected with a FoxO-promoter luciferase reporter, co-expression of the catalytic domain of PKA-α, catalytically inactive mutant PKA-α, or siRNA against PKA-α caused corresponding increases or decreases in transactivation of the FoxO promoter. Expression of vascular cellular adhesion molecule-1 mRNA, up-regulated by FoxO1 in endothelial cells, was enhanced by siRNA knockdown of PKA-α or treatment of HAEC with the PKA inhibitor H89. Adhesion of monocytes to endothelial cells was enhanced by H89 treatment or overexpression of FoxO1-AAA, similar to effects of TNF-α treatment. We conclude that FoxO1 is a novel physiological substrate for PKA-α in vascular endothelial cells.

  11. Schedule-Dependent Antiangiogenic and Cytotoxic Effects of Chemotherapy on Vascular Endothelial and Retinoblastoma Cells.

    Directory of Open Access Journals (Sweden)

    Ursula Winter

    Full Text Available Current treatment of retinoblastoma involves using the maximum dose of chemotherapy that induces tumor control and is tolerated by patients. The impact of dose and schedule on the cytotoxicity of chemotherapy has not been studied. Our aim was to gain insight into the cytotoxic and antiangiogenic effect of the treatment scheme of chemotherapy used in retinoblastoma by means of different in vitro models and to evaluate potential effects on multi-drug resistance proteins. Two commercial and two patient-derived retinoblastoma cell types and two human vascular endothelial cell types were exposed to increasing concentrations of melphalan or topotecan in a conventional (single exposure or metronomic (7-day continuous exposure treatment scheme. The concentration of chemotherapy causing a 50% decrease in cell proliferation (IC50 was determined by MTT and induction of apoptosis was evaluated by flow cytometry. Expression of ABCB1, ABCG2 and ABCC1 after conventional or metronomic treatments was assessed by RT-qPCR. We also evaluated the in vivo response to conventional (0.6 mg/kg once a week for 2 weeks and metronomic (5 days a week for 2 weeks topotecan in a retinoblastoma xenograft model. Melphalan and topotecan were cytotoxic to both retinoblastoma and endothelial cells after conventional and metronomic treatments. A significant decrease in the IC50 (median, 13-fold; range: 3-23 was observed following metronomic chemotherapy treatment in retinoblastoma and endothelial cell types compared to conventional treatment (p0.05. In mice, continuous topotecan lead to significantly lower tumor volumes compared to conventional treatment after 14 days of treatment (p<0.05. Continuous exposure to melphalan or topotecan increased the chemosensitivity of retinoblastoma and endothelial cells to both chemotherapy agents with lower IC50 values compared to short-term treatment. These findings were validated in an in vivo model. None of the dosing modalities induced

  12. Gut flora-dependent metabolite Trimethylamine-N-oxide accelerates endothelial cell senescence and vascular aging through oxidative stress.

    Science.gov (United States)

    Ke, Yilang; Li, Dang; Zhao, Mingming; Liu, Changjie; Liu, Jia; Zeng, Aiping; Shi, Xiaoyun; Cheng, Si; Pan, Bing; Zheng, Lemin; Hong, Huashan

    2018-02-20

    Trimethylamine-N-oxide (TMAO), gut microbiota-dependent metabolites, has been shown to be associated with cardiovascular diseases. However, little is known about the relationship between TMAO and vascular aging. Here, we observed a change in TMAO during the aging process and the effects of TMAO on vascular aging and endothelial cell (EC) senescence. We analyzed age-related plasma levels of TMAO in young adults (18-44 years old), older adults (≥ 65 years old), and 1-month-old, 3-month-old, 6-month-old and 10-month-old senescence-accelerated mouse prone 8 (SAMP8) and age-matched senescence-accelerated mouse resistance 1 (SAMR1) models. We found that circulating TMAO increased with age both in humans and mice. Next, we observed that a TMAO treatment for 16 weeks induced vascular aging in SAMR1 mice and accelerated the process in SAMP8 mice, as measured by an upregulation of senescence markers including senescence-associated β-galactosidase (SA-β-gal), p53, and p21, vascular dysfunction and remodeling. In vitro, we demonstrated that prolonged TMAO treatment induced senescence in human umbilical vein endothelial cells (HUVECs), characterized by reduced cell proliferation, increased expressions of senescence markers, stagnate G0/G1, and impaired cell migration. Furthermore, TMAO suppressed sirtuin 1 (SIRT1) expression and increased oxidative stress both in vivo and in vitro and then activated the p53/p21/Rb pathway resulting in increased p53, acetylation of p53, p21, and decreased CDK2, cyclinE1, and phosphorylation of Rb. In summary, these data suggest that elevated circulating TMAO during the aging process may deteriorate EC senescence and vascular aging, which is probably associated with repression of SIRT1 expression and increased oxidative stress, and, thus, the activation of the p53/p21/Rb pathway. Copyright © 2018 Elsevier Inc. All rights reserved.

  13. 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.

  14. Extracellular Vesicles from Vascular Endothelial Cells Promote Survival, Proliferation and Motility of Oligodendrocyte Precursor Cells.

    Directory of Open Access Journals (Sweden)

    Masashi Kurachi

    Full Text Available We previously examined the effect of brain microvascular endothelial cell (MVEC transplantation on rat white matter infarction, and found that MVEC transplantation promoted remyelination of demyelinated axons in the infarct region and reduced apoptotic death of oligodendrocyte precursor cells (OPCs. We also found that the conditioned medium (CM from cultured MVECs inhibited apoptosis of cultured OPCs. In this study, we examined contribution of extracellular vesicles (EVs contained in the CM to its inhibitory effect on OPC apoptosis. Removal of EVs from the CM by ultracentrifugation reduced its inhibitory effect on OPC apoptosis. To confirm whether EVs derived from MVECs are taken up by cultured OPCs, we labeled EVs with PKH67, a fluorescent dye, and added them to OPC cultures. Many vesicular structures labeled with PKH67 were found within OPCs immediately after their addition. Next we examined the effect of MVEC-derived EVs on OPC behaviors. After 2 days in culture with EVs, there was significantly less pyknotic and more BrdU-positive OPCs when compared to control. We also examined the effect of EVs on motility of OPCs. OPCs migrated longer in the presence of EVs when compared to control. To examine whether these effects on cultured OPCs are shared by EVs from endothelial cells, we prepared EVs from conditioned media of several types of endothelial cells, and tested their effects on cultured OPCs. EVs from all types of endothelial cells we examined reduced apoptosis of OPCs and promoted their motility. Identification of the molecules contained in EVs from endothelial cells may prove helpful for establishment of effective therapies for demyelinating diseases.

  15. Low toxic and high soluble camptothecin derivative 2–47 effectively induces apoptosis of tumor cells in vitro

    International Nuclear Information System (INIS)

    Zhou, Yao; Zhao, Hong-Ye; Jiang, Du; Wang, Lu-Yao; Xiang, Cen; Wen, Shao-Peng; Fan, Zhen-Chuan; Zhang, Yong-Min; Guo, Na; Teng, Yu-Ou; Yu, Peng

    2016-01-01

    The cytotoxic activity of camptothecin derivatives is so high that these compounds need to be further modified before their successful application as anti-cancer agents clinically. In this study, we reported the synthesis and biological evaluation of a novel camptothecin derivative called compound 2–47. The changes in structure did not reduce its activity to inhibit DNA topoisomerase I. Compound 2–47 induced apoptosis of many tumor cells including leukemia cells K562, Jurkat, HL-60, breast cancer cell BT-549, colon cancer cell HT-29 and liver cancer cell HepG2 with a half maximal inhibitory concentration (IC 50 ) of 2- to 3-fold lower than HCPT as a control. In particular, 2–47 inhibited the proliferation of Jurkat cells with an IC 50 of as low as 40 nM. By making use of Jurkat cell as a model, following treatment of Jurkat cells, compound 2–47 activated caspase-3 and PARP, resulting in a decreased Bcl-2/Bax ratio. These data showed that compound 2–47 induces Jurkat cell death through the mitochondrial apoptotic pathway. In addition, compound 2–47 showed a decreased cytotoxic activity against normal cells and an improved solubility in low-polar solvent. For example, compound 2–47 solutes in CHCl 3 130-fold higher than HCPT. Taken together, our data demonstrated that camptothecin derivative 2–47 notably inhibits the tumor cell proliferation through mitochondrial-mediated apoptosis in vitro. - Highlights: • Compound 2–47 showed a wide inhibitory effect on the tested tumor cell lines with an IC 50 of 3 times lower than that of HCPT in general. • Compound 2–47 inhibited the proliferation of the human leukemia cell Jurkat at an IC 50 of as low as 40 nM. • As compared to HCPT, compound 2–47 showed much reduced cytotoxicity on normal human cells. • As compared to others, compound 2–47 showed a hundreds-fold higher solubility in non-polar organic solution.

  16. Low toxic and high soluble camptothecin derivative 2–47 effectively induces apoptosis of tumor cells in vitro

    Energy Technology Data Exchange (ETDEWEB)

    Zhou, Yao; Zhao, Hong-Ye; Jiang, Du; Wang, Lu-Yao; Xiang, Cen; Wen, Shao-Peng [Key Laboratory of Industrial Fermentation Microbiology, Tianjin Key Laboratory of Industrial Microbiology, Sino-French Joint Laboratory of Food Nutrition, Safety and Medicinal Chemistry, Tianjin University of Science and Technology, Tianjin 300457 (China); Fan, Zhen-Chuan [Key Laboratory of Food Nutrition and Safety, Tianjin University of Science & Technology, Ministry of Education, Tianjin, 300457 (China); Obesita & Algaegen LLC, College Station, TX 77845 (United States); Zhang, Yong-Min [Université Pierre et Marie Curie-Paris 6, Institut Parisien de Chimie Moléculaire UMR CNRS 8232, 4 place Jussieu, 75005, Paris (France); Guo, Na [Key Laboratory of Industrial Fermentation Microbiology, Tianjin Key Laboratory of Industrial Microbiology, Sino-French Joint Laboratory of Food Nutrition, Safety and Medicinal Chemistry, Tianjin University of Science and Technology, Tianjin 300457 (China); Teng, Yu-Ou, E-mail: tyo201485@tust.edu.cn [Key Laboratory of Industrial Fermentation Microbiology, Tianjin Key Laboratory of Industrial Microbiology, Sino-French Joint Laboratory of Food Nutrition, Safety and Medicinal Chemistry, Tianjin University of Science and Technology, Tianjin 300457 (China); Yu, Peng, E-mail: yupeng@tust.edu.cn [Key Laboratory of Industrial Fermentation Microbiology, Tianjin Key Laboratory of Industrial Microbiology, Sino-French Joint Laboratory of Food Nutrition, Safety and Medicinal Chemistry, Tianjin University of Science and Technology, Tianjin 300457 (China)

    2016-04-08

    The cytotoxic activity of camptothecin derivatives is so high that these compounds need to be further modified before their successful application as anti-cancer agents clinically. In this study, we reported the synthesis and biological evaluation of a novel camptothecin derivative called compound 2–47. The changes in structure did not reduce its activity to inhibit DNA topoisomerase I. Compound 2–47 induced apoptosis of many tumor cells including leukemia cells K562, Jurkat, HL-60, breast cancer cell BT-549, colon cancer cell HT-29 and liver cancer cell HepG2 with a half maximal inhibitory concentration (IC{sub 50}) of 2- to 3-fold lower than HCPT as a control. In particular, 2–47 inhibited the proliferation of Jurkat cells with an IC{sub 50} of as low as 40 nM. By making use of Jurkat cell as a model, following treatment of Jurkat cells, compound 2–47 activated caspase-3 and PARP, resulting in a decreased Bcl-2/Bax ratio. These data showed that compound 2–47 induces Jurkat cell death through the mitochondrial apoptotic pathway. In addition, compound 2–47 showed a decreased cytotoxic activity against normal cells and an improved solubility in low-polar solvent. For example, compound 2–47 solutes in CHCl{sub 3} 130-fold higher than HCPT. Taken together, our data demonstrated that camptothecin derivative 2–47 notably inhibits the tumor cell proliferation through mitochondrial-mediated apoptosis in vitro. - Highlights: • Compound 2–47 showed a wide inhibitory effect on the tested tumor cell lines with an IC{sub 50} of 3 times lower than that of HCPT in general. • Compound 2–47 inhibited the proliferation of the human leukemia cell Jurkat at an IC{sub 50} of as low as 40 nM. • As compared to HCPT, compound 2–47 showed much reduced cytotoxicity on normal human cells. • As compared to others, compound 2–47 showed a hundreds-fold higher solubility in non-polar organic solution.

  17. Soluble common gamma chain exacerbates COPD progress through the regulation of inflammatory T cell response in mice

    Directory of Open Access Journals (Sweden)

    Lee B

    2017-03-01

    Full Text Available Byunghyuk Lee,1 Eunhee Ko,1 Jiyeon Lee,2 Yuna Jo,1 Hyunju Hwang,1 Tae Sik Goh,1,3 Myungsoo Joo,2 Changwan Hong1 1Department of Anatomy and Cell Biology, Pusan National University School of Medicine, 2Division of Applied Medicine, School of Korean Medicine, Pusan National University, Yangsan, 3Department of Orthopedic Surgery, Medical Research Institute, Pusan National University School of Medicine, Busan, South Korea Abstract: Cigarette smoking (CS is a major cause of considerable morbidity and mortality by inducing lung cancer and COPD. COPD, a smoking-related disorder, is closely related to the alteration of immune system and inflammatory processes that are specifically mediated by T cells. Soluble common gamma chain (sγc has recently been identified as a critical regulator of the development and differentiation of T cells. We examined the effects of sγc in a cigarette smoke extract (CSE mouse model. The sγc level in CSE mice serum is significantly downregulated, and the cellularity of lymph node (LN is systemically reduced in the CSE group. Overexpression of sγc enhances the cellularity and IFNγ production of CD8 T cells in LN and also enhances Th1 and Th17 differentiation of CD4 T cells in the respiratory tract. Mechanistically, the downregulation of sγc expression mediated by CSE is required to prevent excessive inflammatory T cell responses. Therefore, our data suggest that sγc may be one of the target molecules for the control of immunopathogenic progresses in COPD. Keywords: COPD, T cell, soluble common gamma chain, cytokine

  18. Pleiotrophin Regulates the Retention and Self-Renewal of Hematopoietic Stem Cells in the Bone Marrow Vascular Niche

    Directory of Open Access Journals (Sweden)

    Heather A. Himburg

    2012-10-01

    Full Text Available The mechanisms through which the bone marrow (BM microenvironment regulates hematopoietic stem cell (HSC fate remain incompletely understood. We examined the role of the heparin-binding growth factor pleiotrophin (PTN in regulating HSC function in the niche. PTN−/− mice displayed significantly decreased BM HSC content and impaired hematopoietic regeneration following myelosuppression. Conversely, mice lacking protein tyrosine phosphatase receptor zeta, which is inactivated by PTN, displayed significantly increased BM HSC content. Transplant studies revealed that PTN action was not HSC autonomous, but rather was mediated by the BM microenvironment. Interestingly, PTN was differentially expressed and secreted by BM sinusoidal endothelial cells within the vascular niche. Furthermore, systemic administration of anti-PTN antibody in mice substantially impaired both the homing of hematopoietic progenitor cells to the niche and the retention of BM HSCs in the niche. PTN is a secreted component of the BM vascular niche that regulates HSC self-renewal and retention in vivo.

  19. Synergy between thrombin and serotonin in inducing vascular smooth muscle cell proliferation.

    Science.gov (United States)

    Pakala, R; Benedict, C

    1999-12-01

    Previous studies have indicated that apart from playing an important role in hemostasis and thrombosis, thrombin may also contribute to the development of postangioplasty restenosis caused by the stimulation of vascular smooth muscle cell (VSMC) proliferation. Because thrombin generation in vivo is accompanied by platelet activation and release of smooth muscle cell (SMC) growth factors such as serotonin, we examined the possible interaction between these two compounds on VSMC proliferation. Thrombin (0.01 to 100 nmol/L), thrombin receptor-activating peptide (0.1 to 1000 micromol/L), and serotonin (5HT; 0.1 to 1000 micromol/L) increased tritiated thymidine incorporation into the DNA of canine aortic VSMCs in a dose-dependent manner. When thrombin and 5HT were added together at sub-threshold concentrations, they acted synergistically in inducing tritiated thymidine incorporation. These findings were paralleled by a 90%+/-5% increase in the cell number at 48 hours, as compared with a 37%+/-2% increase with 50 micromol/L serotonin and a 13%+/-3% increase with 0.1 nmol/L thrombin. We also demonstrated that a brief exposure to thrombin (1 hour) is sufficient to show its potentiating effect on serotonin. The mitogenic effect of serotonin and its synergistic interaction with thrombin on VSMC proliferation was abolished by serotonin type 2 receptor antagonist LY281067. Similarly, gamma-hirudin--a direct thrombin inhibitor--blocked the mitogenic effect of thrombin and its synergistic interaction with serotonin. When LY281067 and gamma-hirudin were used together, they abolished the mitogenic effects of both the agonists. Because clot-bound active thrombin can escape inactivation by anti-thrombin, this thrombin may potentiate the mitogenic effect of serotonin and keep the SMCs in a proliferative state for a long period of time. These findings support the use of 5HT2 receptor antagonists in combination with thrombin inhibitors in the prevention of SMC proliferation after

  20. Enhanced adhesion of early endothelial progenitor cells to radiation-induced senescence-like vascular endothelial cells in vitro

    International Nuclear Information System (INIS)

    Sermsathanasawadi, N.; Inoue, Yoshinori; Iwai, Takehisa; Ishii, Hideto; Yoshida, Masayuki; Igarashi, Kaori; Miura, Masahiko

    2009-01-01

    The effects of ionizing radiation (IR) on tumor neovascularization are still unclear. We previously reported that vascular endothelial cells (ECs) expressing the IR-induced senescence-like (IRSL) phenotype exhibit a significant decrease in angiogenic activity in vitro. In this study, we examined the effects of the IRSL phenotype on adhesion to early endothelial progenitor cells (early EPCs). Adhesion of human peripheral blood-derived early EPCs to human umbilical vein endothelial cells (HUVECs) expressing the IRSL phenotype was evaluated by an adhesion assay under static conditions. It was revealed that the IRSL HUVECs supported significantly more adhesion of early EPCs than normal HUVECs. Expressions of ICAM-1, VCAM-1 and E-selectin were up-regulated in IRSL HUVECs. Pre-treatment of IRSL HUVECs with adhesion-blocking monoclonal antibodies against E-selectin and VCAM-1 significantly reduced early EPC adhesion to IRSL HUVECs, suggesting a potential role for the E-selectin and VCAM-1 in the adhesion between IRSL ECs and early EPCs. Therefore, the IRSL phenotype expressed in ECs may enhance neovascularization via increased homing of early EPCs. Our findings are first to implicate the complex effects of this phenotype on tumor neovascularization following irradiation. (author)

  1. Selective Deletion of Leptin Signaling in Endothelial Cells Enhances Neointima Formation and Phenocopies the Vascular Effects of Diet-Induced Obesity in Mice.

    Science.gov (United States)

    Hubert, Astrid; Bochenek, Magdalena L; Schütz, Eva; Gogiraju, Rajinikanth; Münzel, Thomas; Schäfer, Katrin

    2017-09-01

    Obesity is associated with elevated circulating leptin levels and hypothalamic leptin resistance. Leptin receptors (LepRs) are expressed on endothelial cells, and leptin promotes neointima formation in a receptor-dependent manner. Our aim was to examine the importance of endothelial LepR (End.LepR) signaling during vascular remodeling and to determine whether the cardiovascular consequences of obesity are because of hyperleptinemia or endothelial leptin resistance. Mice with loxP-flanked LepR alleles were mated with mice expressing Cre recombinase controlled by the inducible endothelial receptor tyrosine kinase promoter. Obesity was induced with high-fat diet. Neointima formation was examined after chemical carotid artery injury. Morphometric quantification revealed significantly greater intimal hyperplasia, neointimal cellularity, and proliferation in End.LepR knockout mice, and similar findings were obtained in obese, hyperleptinemic End.LepR wild-type animals. Analysis of primary endothelial cells confirmed abrogated signal transducer and activator of transcription-3 phosphorylation in response to leptin in LepR knockout and obese LepR wild-type mice. Quantitative PCR, ELISA, and immunofluorescence analyses revealed increased expression and release of endothelin-1 in End.LepR-deficient and LepR-resistant cells, and ET receptor A/B antagonists abrogated their paracrine effects on murine aortic smooth muscle cell proliferation. Reduced expression of peroxisome proliferator-activated receptor-γ and increased nuclear activator protein-1 staining was observed in End.LepR-deficient and LepR-resistant cells, and peroxisome proliferator-activated receptor-γ antagonization increased endothelial endothelin-1 expression. Our findings suggest that intac