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Sample records for artery vascular smooth

  1. Isolation of Endothelial Cells and Vascular Smooth Muscle Cells from Internal Mammary Artery Tissue

    PubMed Central

    Moss, Stephanie C.; Bates, Michael; Parrino, Patrick E.; Woods, T. Cooper

    2007-01-01

    Analyses of vascular smooth muscle cell and endothelial cell function through tissue culture techniques are often employed to investigate the underlying mechanisms regulating cardiovascular disease. As diseases such as diabetes mellitus and chronic kidney disease increase a patient's risk of cardiovascular disease, the development of methods for examining the effects of these diseases on vascular smooth muscle cells and endothelial cells is needed. Commercial sources of endothelial cells and vascular smooth muscle cells generally provide minimal donor information and are in limited supply. This study was designed to determine if vascular smooth muscle cells and endothelial cells could be isolated from human internal mammary arteries obtained from donors undergoing coronary artery bypass graft surgery. As coronary artery bypass graft surgery is a commonly performed procedure, this method would provide a new source for these cells that when combined with the donor's medical history will greatly enhance our studies of the effects of complicating diseases on vascular biology. Internal mammary artery tissue was obtained from patients undergoing coronary artery bypass graft surgery. Through a simple method employing two separate tissue digestions, vascular smooth muscle cells and endothelial cells were isolated and characterized. The isolated vascular smooth muscle cells and endothelial cells exhibited the expected morphology and were able to be passaged for further analysis. The vascular smooth muscle cells exhibited positive staining for α-smooth muscle actin and the endothelial cells exhibited positive staining for CD31. The overall purity of the isolations was > 95%. This method allows for the isolation of endothelial cells and vascular smooth muscle cells from internal mammary arteries, providing a new tool for investigations into the interplay of vascular diseases and complicating diseases such as diabetes and kidney disease. PMID:21603530

  2. 3D Reconstruction of Coronary Artery Vascular Smooth Muscle Cells

    PubMed Central

    Luo, Tong; Chen, Huan; Kassab, Ghassan S.

    2016-01-01

    Aims 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. Methods and Results 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±14.9μm, 4.6±0.6μm and 6.2±1.8μm (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°. Conclusions 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. PMID:26882342

  3. [Vascular smooth muscle cells from human umbilical artery undergo osteoblast differentiation and calcification in vitro].

    PubMed

    Guo, Yong Ping; Sun, Ming Shu; Qian, Jia Qi; Ni, Zhao Hui

    2008-04-01

    To research if the vascular smooth muscle cells (VSMCs) from human umbilical artery undergo osteoblast differentiation spontaneously in vitro. The growth curve of vascular smooth muscle cells from human umbilical artery was obtained by MTT method. The course of multicell nodule formation spontaneously by VSMCs was observed morphologically. The apoptosis of VSMCs in the nodules was detected by Hoechst 33258 and TUNEL methods respectively. The expression of alkaline phosphotase in the nodules was detected by immunohistochemical method. And the calcification was studied with transmission electron microscope and by alizarin red S respectively. We found that the umbilical artery smooth muscle cells confluenced after 7 days of passage and exhibited typical "hill and valley" pattern under light microscope. The cells grew into aggregation and formed nodules at the "hill" region with culture-time prolongation. After 4-5 weeks culture, these nodules built up and calcified spontaneously. We also found alkaline phosphotase expression and apoptosis of VSMCs in these nodules at the same time. We conclude that the vascular smooth muscle cells from human umbilical artery just like from aortic artery can undergo osteoblast differentiation spontaneously in vitro, and apoptosis participate this procedure probably.

  4. Vascular smooth cell proliferation in perfusion culture of porcine carotid arteries

    SciTech Connect

    Liao, Dan; Lin, Peter H.; Yao Qizhi; Chen Changyi

    2008-08-08

    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.

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

    NASA Astrophysics Data System (ADS)

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

    2007-11-01

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

  6. Cullin-3 mutation causes arterial stiffness and hypertension through a vascular smooth muscle mechanism

    PubMed Central

    Agbor, Larry N.; Ibeawuchi, Stella-Rita C.; Hu, Chunyan; Davis, Deborah R.; Keen, Henry L.; Quelle, Frederick W.; Sigmund, Curt D.

    2016-01-01

    Cullin-3 (CUL3) mutations (CUL3Δ9) were previously identified in hypertensive patients with pseudohypoaldosteronism type-II (PHAII), but the mechanism causing hypertension and whether this is driven by renal tubular or extratubular mechanisms remains unknown. We report that selective expression of CUL3Δ9 in smooth muscle acts by interfering with expression and function of endogenous CUL3, resulting in impaired turnover of the CUL3 substrate RhoA, increased RhoA activity, and augmented RhoA/Rho kinase signaling. This caused vascular dysfunction and increased arterial pressure under baseline conditions and a marked increase in arterial pressure, collagen deposition, and vascular stiffness in response to a subpressor dose of angiotensin II, which did not cause hypertension in control mice. Inhibition of total cullin activity increased the level of CUL3 substrates cyclin E and RhoA, and expression of CUL3Δ9 decreased the level of the active form of endogenous CUL3 in human aortic smooth muscle cells. These data indicate that selective expression of the Cul3Δ9 mutation in vascular smooth muscle phenocopies the hypertension observed in Cul3Δ9 human subjects and suggest that mutations in CUL3 cause human hypertension in part through a mechanism involving smooth muscle dysfunction initiated by a loss of CUL3-mediated degradation of RhoA. PMID:27882355

  7. Proteomic analysis of vascular smooth muscle cells in physiological condition and in pulmonary arterial hypertension: Toward contractile versus synthetic phenotypes.

    PubMed

    Régent, Alexis; Ly, Kim Heang; Lofek, Sébastien; Clary, Guilhem; Tamby, Mathieu; Tamas, Nicolas; Federici, Christian; Broussard, Cédric; Chafey, Philippe; Liaudet-Coopman, Emmanuelle; Humbert, Marc; Perros, Frédéric; Mouthon, Luc

    2016-10-01

    Vascular smooth muscle cells (VSMCs) are highly specialized cells that regulate vascular tone and participate in vessel remodeling in physiological and pathological conditions. It is unclear why certain vascular pathologies involve one type of vessel and spare others. Our objective was to compare the proteomes of normal human VSMC from aorta (human aortic smooth muscle cells, HAoSMC), umbilical artery (human umbilical artery smooth muscle cells, HUASMC), pulmonary artery (HPASMC), or pulmonary artery VSMC from patients with pulmonary arterial hypertension (PAH-SMC). Proteomes of VSMC were compared by 2D DIGE and MS. Only 19 proteins were differentially expressed between HAoSMC and HPASMC while 132 and 124 were differentially expressed between HUASMC and HAoSMC or HPASMC, respectively (fold change 1.5≤ or -1.5≥, p < 0.05). As much as 336 proteins were differentially expressed between HPASMC and PAH-SMC (fold change 1.5≤ or -1.5≥, p < 0.05). HUASMC expressed increased amount of α-smooth muscle actin compared to either HPASMC or HAoSMC (although not statistically significant). In addition, PAH-SMC expressed decreased amount of smooth muscle myosin heavy chain and proliferation rate was increased compared to HPASMC thus supporting that PAH-SMC have a more synthetic phenotype. Analysis with Ingenuity identified paxillin and (embryonic lethal, abnormal vision, drosophila) like 1 (ELAVL1) as molecules linked with a lot of proteins differentially expressed between HPASMC and PAH-SMC. There was a trend toward reduced proliferation of PAH-SMC with paxillin-si-RNA and increased proliferation with ELAVL1-siRNA. Thus, VSMCs have very diverse protein content depending on their origin and this is in link with phenotypic differentiation. Paxillin targeting may be a promising treatment of PAH. ELAVL1 also participate in the regulation of PAH-SMC proliferation.

  8. Pathophysiological role of vascular smooth muscle alkaline phosphatase in medial artery calcification†

    PubMed Central

    Sheen, Campbell R.; Kuss, Pia; Narisawa, Sonoko; Yadav, Manisha C.; Nigro, Jessica; Wang, Wei; Chhea, T. Nicole; Sergienko, Eduard A.; Kapoor, Kapil; Jackson, Michael R.; Hoylaerts, Marc. F.; Pinkerton, Anthony B.; O'Neill, W. Charles; Millán, Jose Luis

    2015-01-01

    Medial vascular calcification (MVC) is a pathological phenomenon common to a variety of conditions, including aging, chronic kidney disease, diabetes, obesity, and a variety of rare genetic diseases, that causes vascular stiffening and can lead to heart failure. These conditions share the common feature of tissue-nonspecific alkaline phosphatase (TNAP) upregulation in the vasculature. To evaluate the role of TNAP in MVC, we developed a mouse model that overexpresses human TNAP in vascular smooth muscle cells in an X-linked manner. Hemizygous overexpressor male mice (Tagln-Cre+/-; HprtALPL/Y, or TNAP-OE) show extensive vascular calcification, high blood pressure, cardiac hypertrophy and have a median age of death of 44 days, whereas the cardiovascular phenotype is much less pronounced and life expectancy is longer in heterozygous (Tagln-Cre+/-; HprtALPL/-) female TNAP-OE mice. Gene expression analysis showed upregulation of osteoblast and chondrocyte markers and decreased expression of vascular smooth muscle markers in the aortas of TNAP-OE mice. Through medicinal chemistry efforts, we developed inhibitors of TNAP with drug-like pharmacokinetic characteristics. TNAP-OE mice were treated with the prototypical TNAP inhibitor SBI-425 or vehicle to evaluate the feasibility of TNAP inhibition in vivo. Treatment with this inhibitor significantly reduced aortic calcification and cardiac hypertrophy, and extended lifespan over vehicle-treated controls, in the absence of secondary effects on the skeleton. This study shows that TNAP in the vasculature contributes to the pathology of MVC and that it is a druggable target. This article is protected by copyright. All rights reserved PMID:25428889

  9. A method for three-dimensional quantification of vascular smooth muscle orientation: application in viable murine carotid arteries.

    PubMed

    Spronck, Bart; Megens, Remco T A; Reesink, Koen D; Delhaas, Tammo

    2016-04-01

    When studying in vivo arterial mechanical behaviour using constitutive models, smooth muscle cells (SMCs) should be considered, while they play an important role in regulating arterial vessel tone. Current constitutive models assume a strictly circumferential SMC orientation, without any dispersion. We hypothesised that SMC orientation would show considerable dispersion in three dimensions and that helical dispersion would be greater than transversal dispersion. To test these hypotheses, we developed a method to quantify the 3D orientation of arterial SMCs. Fluorescently labelled SMC nuclei of left and right carotid arteries of ten mice were imaged using two-photon laser scanning microscopy. Arteries were imaged at a range of luminal pressures. 3D image processing was used to identify individual nuclei and their orientations. SMCs showed to be arranged in two distinct layers. Orientations were quantified by fitting a Bingham distribution to the observed orientations. As hypothesised, orientation dispersion was much larger helically than transversally. With increasing luminal pressure, transversal dispersion decreased significantly, whereas helical dispersion remained unaltered. Additionally, SMC orientations showed a statistically significant (p < 0.05) mean right-handed helix angle in both left and right arteries and in both layers, which is a relevant finding from a developmental biology perspective. In conclusion, vascular SMC orientation (1) can be quantified in 3D; (2) shows considerable dispersion, predominantly in the helical direction; and (3) has a distinct right-handed helical component in both left and right carotid arteries. The obtained quantitative distribution data are instrumental for constitutive modelling of the artery wall and illustrate the merit of our method.

  10. Assays for in vitro monitoring of human airway smooth muscle (ASM) and human pulmonary arterial vascular smooth muscle (VSM) cell migration.

    PubMed

    Goncharova, Elena A; Goncharov, Dmitry A; Krymskaya, Vera P

    2006-01-01

    Migration of human pulmonary vascular smooth muscle (VSM) cells contributes to vascular remodeling in pulmonary arterial hypertension and atherosclerosis. Evidence also indicates that, in part, migration of airway smooth muscle (ASM) cells may contribute to airway remodeling associated with asthma. Here we describe migration of VSM and ASM cells in vitro using Transwell or Boyden chamber assays. Because dissecting signaling mechanisms regulating cell migration requires molecular approaches, our protocol also describes how to assess migration of transfected VSM and ASM cells. Transwell or Boyden chamber assays can be completed in approximately 8 h and include plating of serum-deprived VSM or ASM cell suspension on membrane precoated with collagen, migration of cells toward chemotactic gradient and visual (Transwell) or digital (Boyden chamber) analysis of membrane. Although the Transwell assay is easy, the Boyden chamber assay requires hands-on experience; however, both assays are reliable cell-based approaches providing valuable information on how chemotactic and inflammatory factors modulate VSM and ASM migration.

  11. A novel inhibitory effect of oxazol-5-one compounds on ROCKII signaling in human coronary artery vascular smooth muscle cells

    PubMed Central

    Al-Ghabkari, Abdulhameed; Deng, Jing-Ti; McDonald, Paul C.; Dedhar, Shoukat; Alshehri, Mana; Walsh, Michael P.; MacDonald, Justin A.

    2016-01-01

    The selectivity of (4Z)-2-(4-chloro-3-nitrophenyl)-4-(pyridin-3-ylmethylidene)-1,3-oxazol-5-one (DI) for zipper-interacting protein kinase (ZIPK) was previously described by in silico computational modeling, screening a large panel of kinases, and determining the inhibition efficacy. Our assessment of DI revealed another target, the Rho-associated coiled-coil-containing protein kinase 2 (ROCKII). In vitro studies showed DI to be a competitive inhibitor of ROCKII (Ki, 132 nM with respect to ATP). This finding was supported by in silico molecular surface docking of DI with the ROCKII ATP-binding pocket. Time course analysis of myosin regulatory light chain (LC20) phosphorylation catalyzed by ROCKII in vitro revealed a significant decrease upon treatment with DI. ROCKII signaling was investigated in situ in human coronary artery vascular smooth muscle cells (CASMCs). ROCKII down-regulation using siRNA revealed several potential substrates involved in smooth muscle contraction (e.g., LC20, Par-4, MYPT1) and actin cytoskeletal dynamics (cofilin). The application of DI to CASMCs attenuated LC20, Par-4, LIMK, and cofilin phosphorylations. Notably, cofilin phosphorylation was not significantly decreased with a novel ZIPK selective inhibitor (HS-38). In addition, CASMCs treated with DI underwent cytoskeletal changes that were associated with diminution of cofilin phosphorylation. We conclude that DI is not selective for ZIPK and is a potent inhibitor of ROCKII. PMID:27573465

  12. MicroRNA-32 promotes calcification in vascular smooth muscle cells: Implications as a novel marker for coronary artery calcification

    PubMed Central

    Shen, Yingying; Chen, Ling; Xu, Canxin; Zhao, Heng; Wu, Ying; Zhang, Qinghai; Zhong, Jing; Tang, Zhenwang; Liu, Changhui; Zhao, Qiang; Zheng, Yi; Cao, Renxian; Zu, Xuyu

    2017-01-01

    Cardiovascular calcification is one of the most severe outcomes associated with cardiovascular disease and often results in significant morbidity and mortality. Previous reports indicated that epigenomic regulation of microRNAs (miRNAs) might play important roles in vascular smooth muscle cell (VSMC) calcification. Here, we identified potential key miRNAs involved in vascular calcification in vivo and investigated the role of miR-32-5p (miR-32). According to microarray analysis, we observed increased expression of miR-125b, miR-30a, and miR-32 and decreased expression of miR-29a, miR-210, and miR-320 during the progression of vascularcalcification. Additionally, gain- and loss-of-function studies of miR-32 confirmed promotion of VSMC calcification in mice through the enhanced expression of bonemorphogenetic protein-2, runt-related transcription factor-2(RUNX2), osteopontin, and the bone-specific phosphoprotein matrix GLA protein in vitro. Moreover, miR-32 modulated vascularcalcification progression by activating phosphoinositide 3-kinase (PI3K)signaling and increasing RUNX2 expression and phosphorylation by targeting the 3′-untranslated region of phosphatase and tensin homolog Mrna (PTEN) in mouse VSMCs. Furthermore, we detected higher miR-32 levels in plasmafrom patients with coronary artery disease with coronary artery calcification (CAC) as compared with levels observed in non-CAC patients (P = 0.016), further confirming miR-32 as a critical modulator and potential diagnostic marker for CAC. PMID:28319142

  13. 12S-lipoxygenase protein associates with {alpha}-actin fibers in human umbilical artery vascular smooth muscle cells

    SciTech Connect

    Weisinger, Gary . E-mail: gary_w@tasmc.health.gov.il; Limor, Rona; Marcus-Perlman, Yonit; Knoll, Esther; Kohen, Fortune; Schinder, Vera; Firer, Michael; Stern, Naftali

    2007-05-11

    The current study sets out to characterize the intracellular localization of the platelet-type 12S-lipoxygenase (12-LO), an enzyme involved in angiotensin-II induced signaling in vascular smooth muscle cells (VSMC). Immunohistochemical analysis of VSMC in vitro or human umbilical arteries in vivo showed a clear cytoplasmic localization. On immunogold electron microscopy, 12-LO was found primarily associated with cytoplasmic VSMC muscle fibrils. Upon angiotensin-II treatment of cultured VSMC, immunoprecipitated 12-LO was found bound to {alpha}-actin, a component of the cytoplasmic myofilaments. 12-LO/{alpha}-actin binding was blocked by VSMC pretreatment with the 12-LO inhibitors, baicalien or esculetine and the protein synthesis inhibitor, cycloheximide. Moreover, the binding of 12-LO to {alpha}-actin was not associated with 12-LO serine or tyrosine phosphorylation. These observations suggest a previously unrecognized angiotensin-II dependent protein interaction in VSMC through which 12-LO protein may be trafficked, for yet undiscovered purposes towards the much more abundantly expressed cytoskeletal protein {alpha}-actin.

  14. Oxygen-Sensitive Calcium Channels in Vascular Smooth Muscle and Their Possible Role in Hypoxic Arterial Relaxation

    NASA Astrophysics Data System (ADS)

    Franco-Obregon, A.; Urena, J.; Lopez-Barneo, J.

    1995-05-01

    We have investigated the modifications of cytosolic [Ca2+] and the activity of Ca2+ channels in freshly dispersed arterial myocytes to test whether lowering O_2 tension (PO_2) directly influences Ca2+ homeostasis in these cells. Unclamped cells loaded with fura-2 AM exhibit oscillations of cytosolic Ca2+ whose frequency depends on extracellular Ca2+ influx. Switching from a PO_2 of 150 to 20 mmHg leads to a reversible attenuation of the Ca2+ oscillations. In voltage-clamped cells, hypoxia reversibly reduces the influx of Ca2+ through voltage-dependent channels, which can account for the inhibition of the Ca2+ oscillations. Low PO_2 selectively inhibits L-type Ca2+ channel activity, whereas the current mediated by T-type channels is unaltered by hypoxia. The effect of low PO_2 on the L-type channels is markedly voltage dependent, being more apparent with moderate depolarizations. These findings demonstrate the existence of O_2-sensitive, voltage-dependent, Ca2+ channels in vascular smooth muscle that may critically contribute to the local regulation of circulation.

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

    PubMed Central

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

    2015-01-01

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

  16. Phenotypic modifications of vascular smooth muscle cells could be responsible for vascular hyporeactivity to contracting agent in mechanically injured rat carotid artery.

    PubMed

    Popolo, A; Marzocco, S; Nasti, C; Lippolis, L; di Villa Bianca, R d'Emmanuele; Sorrentino, R; Autore, G; Pinto, A

    2005-12-01

    Vascular smooth muscle cells (VSMCs) that accumulate in neointima after angioplastic injury show different phenotypic characteristics from those of medial layer and an impaired reactivity to contracting agents. The aim of the study was to correlate the vascular hyporesponsiveness to the changes in intracellular calcium concentration [Ca(2+)](i) and the expression of proteins necessary for its utilization in mechanically injured rat carotid arteries (IC) at 14 and 28 days after angioplastic balloon. IC showed a significant reduction (P<0.01) to PE- or KCl-induced contraction as compared to uninjured carotid (UC). Fura-2AM-loaded VSMCs isolated from IC revealed that this hyporeactivity to PE or KCl was accompanied by the impairment of the increase in [Ca(2+)](i) induced by contracting agents in both Ca(2+)-free or -containing medium. Similar results were observed following the ryanodine challenge in VSMC. Western blot analysis showed a significant (P<0.05) reduction in myosin heavy chain (MHC) and IP(3)-type III receptor expression in IC isolated at 14 days from injury compared to UC, while an improvement of these proteins expression was observed at 28 days after damage. On the other hand, in IC tissue, SERCA2 and alpha-actin expression, compared to UC was significantly higher at 14 days than at 28 days. These data indicate that vascular hyporeactivity induced by mechanical injury may be due to alterations of either [Ca(2+)](i) or contractile proteins. These modifications could be related to the changes of VSMC phenotypic characteristics, as supported by the observed modifications in MHC, SERCA2 and alpha-actin expression, proteins considered as biological markers of cellular differentiation.

  17. Notch Signaling in Vascular Smooth Muscle Cells.

    PubMed

    Baeten, J T; Lilly, B

    2017-01-01

    The Notch signaling pathway is a highly conserved pathway involved in cell fate determination in embryonic development and also functions in the regulation of physiological processes in several systems. It plays an especially important role in vascular development and physiology by influencing angiogenesis, vessel patterning, arterial/venous specification, and vascular smooth muscle biology. Aberrant or dysregulated Notch signaling is the cause of or a contributing factor to many vascular disorders, including inherited vascular diseases, such as cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy, associated with degeneration of the smooth muscle layer in cerebral arteries. Like most signaling pathways, the Notch signaling axis is influenced by complex interactions with mediators of other signaling pathways. This complexity is also compounded by different members of the Notch family having both overlapping and unique functions. Thus, it is vital to fully understand the roles and interactions of each Notch family member in order to effectively and specifically target their exact contributions to vascular disease. In this chapter, we will review the Notch signaling pathway in vascular smooth muscle cells as it relates to vascular development and human disease.

  18. Role of platelets in smooth muscle cell proliferation and migration after vascular injury in rat carotid artery.

    PubMed Central

    Fingerle, J; Johnson, R; Clowes, A W; Majesky, M W; Reidy, M A

    1989-01-01

    Intimal lesion formation was investigated in rats made thrombocytopenic by a single i.p. injection of a polyclonal antibody made against rat platelets that reduced circulating platelet counts to less than 1% of normal. The carotid artery was then denuded of endothelium with a 2 French balloon catheter, after which no platelets were found adhering to the exposed subendothelium. In control animals, platelets adhered instantly to the denuded artery. Six hours after denudation mRNA for ornithine decarboxylase, a marker for early G1 events, was found to be elevated in both thrombocytopenic and control arteries. Two days after injury the smooth muscle cell replication rate in thrombocytopenic rats was found to be significantly elevated as compared with that in uninjured carotids (13.7% +/- 8.4% vs. 0.65% +/- 0.23%) but was similar to the replication rate observed in denuded carotid arteries from animals treated with nonimmune IgG. One important difference between these animals was that no intimal thickening was observed in thrombocytopenic animals at day 4, and by day 7 the intimas were still significantly smaller than those from control rats. In a separate group of animals which were thrombocytopenic for the entire experiment, no intimal lesions were observed 7 days after injury by balloon catheter. From these results, we conclude that platelets do not play a role in the initiation of smooth muscle cell proliferation after injury by balloon catheter but may regulate their movement into the intima. Images PMID:2813399

  19. Stat3-dependent acute Rantes production in vascular smooth muscle cells modulates inflammation following arterial injury in mice

    PubMed Central

    Kovacic, Jason C.; Gupta, Rohit; Lee, Angela C.; Ma, Mingchao; Fang, Fang; Tolbert, Claire N.; Walts, Avram D.; Beltran, Leilani E.; San, Hong; Chen, Guibin; St. Hilaire, Cynthia; Boehm, Manfred

    2009-01-01

    Inflammation is a key component of arterial injury, with VSMC proliferation and neointimal formation serving as the final outcomes of this process. However, the acute events transpiring immediately after arterial injury that establish the blueprint for this inflammatory program are largely unknown. We therefore studied these events in mice and found that immediately following arterial injury, medial VSMCs upregulated Rantes in an acute manner dependent on Stat3 and NF-κB (p65 subunit). This led to early T cell and macrophage recruitment, processes also under the regulation of the cyclin-dependent kinase inhibitor p21Cip1. Unique to VSMCs, Rantes production was initiated by Tnf-α, but not by Il-6/gp130. This Rantes production was dependent on the binding of a p65/Stat3 complex to NF-κB–binding sites within the Rantes promoter, with shRNA knockdown of either Stat3 or p65 markedly attenuating Rantes production. In vivo, acute NF-κB and Stat3 activation in medial VSMCs was identified, with acute Rantes production after injury substantially reduced in Tnfa–/– mice compared with controls. Finally, we generated mice with SMC-specific conditional Stat3 deficiency and confirmed the Stat3 dependence of acute Rantes production by VSMCs. Together, these observations unify inflammatory events after vascular injury, demonstrating that VSMCs orchestrate the arterial inflammatory response program via acute Rantes production and subsequent inflammatory cell recruitment. PMID:20038813

  20. Feline immunodeficiency virus and retrovirus-mediated adventitial ex vivo gene transfer to rabbit carotid artery using autologous vascular smooth muscle cells.

    PubMed

    Kankkonen, Hanna M; Turunen, Mikko P; Hiltunen, Mikko O; Lehtolainen, Pauliina; Koponen, Jonna; Leppänen, Pia; Turunen, Anna-Mari; Ylä-Herttuala, Seppo

    2004-03-01

    We have developed an ex vivo gene transfer technique to rabbit arterial wall using autologous smooth muscle cells (SMCs). SMCs were harvested from rabbit ear artery, transduced in vitro with vesicular stomatitis virus G-glycoprotein pseudotyped retrovirus or feline immunodeficiency virus (FIV) and returned to the adventitial surface of the carotid artery using a periadventitial silicone collar or collagen sheet placed around the artery. Beta-galactosidase (lacZ) and human apolipoprotein E3 (apoE3) cDNAs were used as transgenes. After retrovirus-mediated gene transfer of lacZ the selected cells implanted with high efficiency and expressed lacZ marker gene at a very high level 7 and 14 days after the operation. The level of lacZ expression decreased thereafter but was still detectable 12 weeks after the gene transfer, and was exclusively localized to the site of cell implantation inside the collar. Utilizing FIV vector expressing apoE3, low levels of apoE were measured from serum collected from a low-density lipoprotein receptor deficient Watanabe heritable hyperlipidemic rabbits 1 month after the gene transfer. The physiological effect of apoE expression was detected as transiently elevated serum cholesterol levels. The results indicate that the model can be used for high efficiency local gene transfer in arteries, e.g. during vascular surgery. The model is also valuable for studying expression, stability and safety of new gene transfer vectors and their expression products in vivo.

  1. Interference of IP-10 expression inhibits vascular smooth muscle cell proliferation and intimal hyperplasia in carotid artery: a new insight in the prevention of restenosis.

    PubMed

    Zuojun, Hu; Lingyu, Hu; Wei, He; Henghui, Yin; Chonggang, Zhang; Jingsong, Wang; Mian, Wang; Yong, Liu; Shenming, Wang

    2012-01-01

    After vascular angioplasty, vascular smooth muscle cell (VSMC) proliferation causes atherosclerosis and intimal hyperplasia leading to restenosis. Interferon-γ-inducible protein (IP)-10 plays a role in atherogenesis, but the mechanism remains unclear. We evaluated the role of IP-10 in intimal hyperplasia and restenosis. IP-10 expression was determined in arterial specimens from 20 arteriosclerotic obliteration patients and 6 healthy individuals. VSMCs were stimulated in vitro with IFN-γ and transfected with IP-10 siRNA. Silencing was verified with RT-PCR/Western blot; cell proliferation rate was detected by methyl-thiazol-tetrazolium. The carotid artery model of atherosclerosis injury was established with IP-10 siRNA. IP-10 expression was detected at 1 and 4 weeks using RT-PCR and immunohistochemistry. Artery morphology was assessed with hematoxylin-and-eosin staining, and intimal hyperplasia was evaluated by electron microscopy. IP-10 was overexpressed in arteriosclerotic obliteration group compared with control group (P < 0.05). IP-10 expression in transfected group was significantly lower than in untransfected group. The intima-to-media ratio of transfected group at 4 weeks was lower than that of untransfected group (P < 0.01). The transfected group exhibited more regular intimal structure and less hyperplasia under electron microscopy. We, therefore, concluded that IP-10 played an important role in intimal hyperplasia as siRNA-mediated IP-10 silencing inhibited aberrant VSMCs hyperplasia and reduced restenosis.

  2. Sympathetic innervation promotes vascular smooth muscle differentiation.

    PubMed

    Damon, Deborah H

    2005-06-01

    The sympathetic nervous system (SNS) is an important modulator of vascular smooth muscle (VSM) growth and function. Several lines of evidence suggest that the SNS also promotes VSM differentiation. The present study tests this hypothesis. Expression of smooth muscle myosin (SM2) and alpha-actin were assessed by Western analysis as indexes of VSM differentiation. SM2 expression (normalized to alpha-actin) in adult innervated rat femoral and tail arteries was 479 +/- 115% of that in noninnervated carotid arteries. Expression of alpha-actin (normalized to GAPDH or total protein) in 30-day-innervated rat femoral arteries was greater than in corresponding noninnervated femoral arteries from guanethidine-sympathectomized rats. SM2 expression (normalized to alpha-actin) in neonatal femoral arteries grown in vitro for 7 days in the presence of sympathetic ganglia was greater than SM2 expression in corresponding arteries grown in the absence of sympathetic ganglia. In VSM-endothelial cell cultures grown in the presence of dissociated sympathetic neurons, alpha-actin (normalized to GAPDH) was 300 +/- 66% of that in corresponding cultures grown in the absence of neurons. This effect was inhibited by an antibody that neutralized the activity of transforming growth factor-beta2. All of these data indicate that sympathetic innervation increased VSM contractile protein expression and thereby suggest that the SNS promotes and/or maintains VSM differentiation.

  3. Vascular Protective Effect of an Ethanol Extract of Camellia japonica Fruit: Endothelium-Dependent Relaxation of Coronary Artery and Reduction of Smooth Muscle Cell Migration.

    PubMed

    Park, Sin-Hee; Shim, Bong-Sup; Yoon, Jun-Seong; Lee, Hyun-Ho; Lee, Hye-Won; Yoo, Seok-Bong; Wi, An-Jin; Park, Whoa-Shig; Kim, Hyun-Jung; Kim, Dong-Wok; Oak, Min-Ho

    2015-01-01

    Camellia japonica is a popular garden plant in Asia and widely used as cosmetic sources and traditional medicine. However, the possibility that C. japonica affects cardiovascular system remains unclear. The aim of the present study was to evaluate vascular effects of an extract of C. japonica. Vascular reactivity was assessed in organ baths using porcine coronary arteries and inhibition of proliferation and migration were assessed using human vascular smooth muscle cells (VSMCs). All four different parts, leaf, stem, flower, and fruits, caused concentration-dependent relaxations and C. japonica fruit (CJF) extract showed the strongest vasorelaxation and its effect was endothelium dependent. Relaxations to CJF were markedly reduced by inhibitor of endothelial nitric oxide synthase (eNOS) and inhibitor of PI3-kinase, but not affected by inhibitor of cyclooxygenase and endothelium-derived hyperpolarizing factor-mediated response. CJF induced activated a time- and concentration-dependent phosphorylation of eNOS in endothelial cells. Altogether, these studies have demonstrated that CJF is a potent endothelium-dependent vasodilator and this effect was involved in, at least in part, PI3K-eNOS-NO pathway. Moreover, CJF attenuated TNF-α induced proliferation and PDGF-BB induced migration of VSMCs. The present findings indicate that CJF could be a valuable candidate of herbal medicine for cardiovascular diseases associated with endothelial dysfunction and atherosclerosis.

  4. Pharmacological characterization of the dopamine receptor coupled to cyclic AMP formation expressed by rat mesenteric artery vascular smooth muscle cells in culture.

    PubMed Central

    Hall, A. S.; Bryson, S. E.; Vaughan, P. F.; Ball, S. G.; Balmforth, A. J.

    1993-01-01

    1. Mesenteric artery vascular smooth muscle cells derived from male Wistar rats and grown in culture were prelabelled with [3H]-adenine and exposed to a range of dopamine receptor agonists and antagonists. Resultant [3H]-cyclic AMP formation was determined and concentration-effect curves constructed, in the presence of propranolol (10-6) M) and the phosphodiesterase inhibitor IBMX (5 x 10(-4) M). 2. Ka apparent values for D1/DA1 dopamine receptor agonists SKF 38393, fenoldopam, 6,7-ADTN, and dopamine were 0.06, 0.59, 4.06 and 5.77 x 10(-6) M respectively. Although fenoldopam and SKF 38393 were more potent than dopamine, they were partial agonists with efficacies, relative to dopamine of approximately 48% and 24% respectively. 6,7-ADTN, in contrast, behaved as a full agonist. 3. Dopamine-stimulated cyclic AMP formation was inhibited in a concentration-dependent manner by the D1/DA1 dopamine receptor selective antagonists, SCH 23390 and cis-flupenthixol (Ki values 0.53 and 36.1 x 10(-1) M respectively). In contrast, the D2/DA2 dopamine receptor selective antagonists, domperidone and (-)-sulpiride, were less potent (Ki values 2.06 and 5.82 x 10(-6) M respectively). Furthermore, the stereoisomers of SCH 23390 and cis-flupenthixol, SCH 23388 and trans-flupenthixol, were at least two orders of magnitude less potent (Ki values 0.14 and 13.2 x 10(-6) M respectively) indicating the stereoselective nature of this receptor. 4. Our results indicate that rat mesenteric artery vascular smooth muscle cells in culture express a dopamine receptor coupled to cyclic AMP formation, which has the pharmacological profile, characteristic of the D1 dopamine receptor subfamily. PMID:7902178

  5. Elastin Degradation and Vascular Smooth Muscle Cell Phenotype Change Precede Cell Loss and Arterial Medial Calcification in a Uremic Mouse Model of Chronic Kidney Disease

    PubMed Central

    Pai, Ashwini; Leaf, Elizabeth M.; El-Abbadi, Mohga; Giachelli, Cecilia M.

    2011-01-01

    Arterial medial calcification (AMC), a hallmark of vascular disease in uremic patients, is highly correlated with serum phosphate levels and cardiovascular mortality. To determine the mechanisms of AMC, mice were made uremic by partial right-side renal ablation (week 0), followed by left-side nephrectomy at week 2. At 3 weeks, mice were switched to a high-phosphate diet, and various parameters of disease progression were examined over time. Serum phosphate, calcium, and fibroblast growth factor 23 (FGF-23) were up-regulated as early as week 4. Whereas serum phosphate and calcium levels declined to normal by 10 weeks, FGF-23 levels remained elevated through 16 weeks, consistent with an increased phosphate load. Elastin turnover and vascular smooth muscle cell (VSMC) phenotype change were early events, detected by week 4 and before AMC. Both AMC and VSMC loss were significantly elevated by week 8. Matrix metalloprotease 2 (MMP-2) and cathepsin S were present at baseline and were significantly elevated at weeks 8 and 12. In contrast, MMP-9 was not up-regulated until week 12. These findings over time suggest that VSMC phenotype change and VSMC loss (early phosphate-dependent events) may be necessary and sufficient to promote AMC in uremic mice fed a high-phosphate diet, whereas elastin degradation might be necessary but is not sufficient to induce AMC (because elastin degradation occurred also in uremic mice on a normal-phosphate diet, but they did not develop AMC). PMID:21281809

  6. Mechanics of Vascular Smooth Muscle.

    PubMed

    Ratz, Paul H

    2015-12-15

    Vascular smooth muscle (VSM; see Table 1 for a list of abbreviations) is a heterogeneous biomaterial comprised of cells and extracellular matrix. By surrounding tubes of endothelial cells, VSM forms a regulated network, the vasculature, through which oxygenated blood supplies specialized organs, permitting the development of large multicellular organisms. VSM cells, the engine of the vasculature, house a set of regulated nanomotors that permit rapid stress-development, sustained stress-maintenance and vessel constriction. Viscoelastic materials within, surrounding and attached to VSM cells, comprised largely of polymeric proteins with complex mechanical characteristics, assist the engine with countering loads imposed by the heart pump, and with control of relengthening after constriction. The complexity of this smart material can be reduced by classical mechanical studies combined with circuit modeling using spring and dashpot elements. Evaluation of the mechanical characteristics of VSM requires a more complete understanding of the mechanics and regulation of its biochemical parts, and ultimately, an understanding of how these parts work together to form the machinery of the vascular tree. Current molecular studies provide detailed mechanical data about single polymeric molecules, revealing viscoelasticity and plasticity at the protein domain level, the unique biological slip-catch bond, and a regulated two-step actomyosin power stroke. At the tissue level, new insight into acutely dynamic stress-strain behavior reveals smooth muscle to exhibit adaptive plasticity. At its core, physiology aims to describe the complex interactions of molecular systems, clarifying structure-function relationships and regulation of biological machines. The intent of this review is to provide a comprehensive presentation of one biomachine, VSM.

  7. Postsynaptic alpha-adrenoceptors, calcium mobilization and (/sup 3/H), 4-dihydropyridine binding in vascular smooth muscle of rat tail artery

    SciTech Connect

    Su, C.M.

    1985-01-01

    Pharmacologic characterization of post-synaptic ..cap alpha..-adrenoceptors in rat tail artery was examined by using selective agonists and antagonists. In this tissue, the ..cap alpha..-adrenoceptor agonists employed all produced concentration-dependent mechanical responses with rank order of potency, clonidine > norepinephrine > norepinephrine > phenylephrine > UK > 14304 > B-HT 920. This order of agonists activities not consistent with a simple classification into ..cap alpha../sub 1/- and ..cap alpha../sub 2/-adrenoceptors in the rat tail artery. Antagonism by prazosin and yohimbine of phenylephrine, norepinephrine and clonidine responses did not reveal the anticipated discrimination between ..cap alpha../sub 1/- and ..cap alpha../sub 2/-adrenoceptors. Potassium depolarization-induced responses were very sensitive to antagonism by the Ca/sup 2 +/ antagonists nifedipine and D 600. The sensitivity sequence of ..cap alpha..-adrenoceptor agonist induced responses to nifedipine and D 600 is H-HT 920 (> clonidine) > phenylephrine > norepinephrine. This disagrees with the thesis that ..cap alpha../sub 2/-adrenoceptor mediated responses in vascular smooth muscle are more sensitive than are ..cap alpha../sub 1/-adrenoceptor mediated responses to Ca/sup 2 +/ channel antagonists. Radioligand binding studies of (/sup 3/H)nitrendipine and (/sup 3/H)Bay K 8644 to microsomal preparations of tail artery membrane a single set of high affinity binding sites and there is a good correlation between the pharmacological potencies and binding affinities of these agents. In addition, study of the displacement of (/sup 3/H)nitrendipine by Bay K 8644 revealed IC/sub 50/ and K/sub l/ values which are in approximate accord with those determined for pharmacologic experiments.

  8. Vascular smooth muscle phenotypic diversity and function

    PubMed Central

    2010-01-01

    The control of force production in vascular smooth muscle is critical to the normal regulation of blood flow and pressure, and altered regulation is common to diseases such as hypertension, heart failure, and ischemia. A great deal has been learned about imbalances in vasoconstrictor and vasodilator signals, e.g., angiotensin, endothelin, norepinephrine, and nitric oxide, that regulate vascular tone in normal and disease contexts. In contrast there has been limited study of how the phenotypic state of the vascular smooth muscle cell may influence the contractile response to these signaling pathways dependent upon the developmental, tissue-specific (vascular bed) or disease context. Smooth, skeletal, and cardiac muscle lineages are traditionally classified into fast or slow sublineages based on rates of contraction and relaxation, recognizing that this simple dichotomy vastly underrepresents muscle phenotypic diversity. A great deal has been learned about developmental specification of the striated muscle sublineages and their phenotypic interconversions in the mature animal under the control of mechanical load, neural input, and hormones. In contrast there has been relatively limited study of smooth muscle contractile phenotypic diversity. This is surprising given the number of diseases in which smooth muscle contractile dysfunction plays a key role. This review focuses on smooth muscle contractile phenotypic diversity in the vascular system, how it is generated, and how it may determine vascular function in developmental and disease contexts. PMID:20736412

  9. Airway epithelial-derived factor relaxes pulmonary vascular smooth muscle.

    PubMed

    Farah, Omar R; Li, Dongge; McIntyre, Brendan A S; Pan, Jingyi; Belik, Jaques

    2009-01-01

    The factors controlling the pulmonary vascular resistance under physiological conditions are poorly understood. We have previously reported on an apparent cross talk between the airway and adjacent pulmonary arterial bed where a factor likely derived from the bronchial epithelial cells reduced the magnitude of agonist-stimulated force in the vascular smooth muscle. The main purpose of this investigation was to evaluate whether bronchial epithelial cells release a pulmonary arterial smooth muscle relaxant factor. Conditioned media from SPOC-1 or BEAS-2B, a rat- and a human-derived bronchial epithelial cell line, respectively, were utilized. This media significantly relaxed precontracted adult but not fetal pulmonary arterial muscle in an oxygen tension-dependent manner. This response was mediated via soluble guanylate cyclase, involving AKT/PI3-kinase and neuronal nitric oxide synthase. Airway epithelial cell-conditioned media increased AKT phosphorylation in pulmonary smooth muscle cells (SMC) and reduced intracellular calcium change following ATP stimulation to a significantly greater extent than observed for bronchial SMC. The present data strongly support the evidence for bronchial epithelial cells releasing a stable and soluble factor capable of inducing pulmonary arterial SMC relaxation. We speculate that under physiological conditions, the maintenance of a low pulmonary vascular resistance, postnatally, is in part modulated by the airway epithelium.

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

    PubMed

    Kurabayashi, Masahiko

    2015-05-01

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

  11. Smooth muscle-selective CPI-17 expression increases vascular smooth muscle contraction and blood pressure

    PubMed Central

    Su, Wen; Xie, Zhongwen; Liu, Shu; Calderon, Lindsay E.; Guo, Zhenheng

    2013-01-01

    Recent data revealed that protein kinase C-potentiated myosin phosphatase inhibitor of 17 kDa (CPI-17), a myosin phosphatase inhibitory protein preferentially expressed in smooth muscle, is upregulated/activated in several diseases but whether this CPI-17 increase plays a causal role in pathologically enhanced vascular smooth muscle contractility and blood pressure remains unclear. To address this possibility, we generated a smooth muscle-specific CPI-17 transgenic mouse model (CPI-17-Tg) and demonstrated that the CPI-17 transgene was selectively expressed in smooth muscle-enriched tissues, including mesenteric arteries. The isometric contractions in the isolated second-order branch of mesenteric artery helical strips from CPI-17-Tg mice were significantly enhanced compared with controls in response to phenylephrine, U-46619, serotonin, ANG II, high potassium, and calcium. The perfusion pressure increases in isolated perfused mesenteric vascular beds in response to norepinephrine were also enhanced in CPI-17-Tg mice. The hypercontractility was associated with increased phosphorylation of CPI-17 and 20-kDa myosin light chain under basal and stimulated conditions. Surprisingly, the protein levels of rho kinase 2 and protein kinase Cα/δ were significantly increased in CPI-17-Tg mouse mesenteric arteries. Radiotelemetry measurements demonstrated that blood pressure was significantly increased in CPI-17-Tg mice. However, no vascular remodeling was detected by morphometric analysis. Taken together, our results demonstrate that increased CPI-17 expression in smooth muscle promotes vascular smooth muscle contractility and increases blood pressure, implicating a pathological significant role of CPI-17 upregulation. PMID:23604714

  12. Vascular Calcification: Mechanisms of Vascular Smooth Muscle Cell Calcification

    PubMed Central

    Leopold, Jane A.

    2014-01-01

    Vascular calcification is highly prevalent and, when present, is associated with major adverse cardiovascular events. Vascular smooth muscle cells play an integral role in mediating vessel calcification by undergoing differentiation to osteoblast-like cells and generating matrix vesicles that serve as a nidus for calcium-phosphate deposition in the vessel wall. Once believed to be a passive process, it is now recognized that vascular calcification is a complex and highly regulated process that involves activation of cellular signaling pathways, circulating inhibitors of calcification, genetic factors, and hormones. This review will examine several of the key mechanisms linking vascular smooth muscle cells to vessel calcification that may be targeted to reduce vessel wall mineralization and, thereby, reduce cardiovascular risk. PMID:25435520

  13. The Smooth Muscle of the Artery

    DTIC Science & Technology

    1975-01-01

    to keep up with inter- national standards. The German Cancer Research Center as well as the Hax-Planck Inntitutes of Heidelberg are well equipped to...SOMLYO: I plan to briefly review some of the aspects of normal function of vascular smooth muscle with particular ---- I SMOOTH MUSCLE STRUCTURE 35...schematic review of the data on catabolism in connective tissue cells smooth muscle cells. The increasing number of electron microscopic studies of

  14. Vascular Extracellular Matrix and Arterial Mechanics

    PubMed Central

    WAGENSEIL, JESSICA E.; MECHAM, ROBERT P.

    2009-01-01

    An important factor in the transition from an open to a closed circulatory system was a change in vessel wall structure and composition that enabled the large arteries to store and release energy during the cardiac cycle. The component of the arterial wall in vertebrates that accounts for these properties is the elastic fiber network organized by medial smooth muscle. Beginning with the onset of pulsatile blood flow in the developing aorta, smooth muscle cells in the vessel wall produce a complex extracellular matrix (ECM) that will ultimately define the mechanical properties that are critical for proper function of the adult vascular system. This review discusses the structural ECM proteins in the vertebrate aortic wall and will explore how the choice of ECM components has changed through evolution as the cardiovascular system became more advanced and pulse pressure increased. By correlating vessel mechanics with physiological blood pressure across animal species and in mice with altered vessel compliance, we show that cardiac and vascular development are physiologically coupled, and we provide evidence for a universal elastic modulus that controls the parameters of ECM deposition in vessel wall development. We also discuss mechanical models that can be used to design better tissue-engineered vessels and to test the efficacy of clinical treatments. PMID:19584318

  15. Vascular leiomyoma of the lung arising from pulmonary artery.

    PubMed

    Terada, Tadashi

    2013-01-01

    Leiomyoma of the lung is extremely rare. The entity is not described in WHO blue book. Less than 100 cases of leiomyoma of the lung have been reported in the literature. However, vascular leiomyoma has not been reported in the literature, to the author's best knowledge. Herein reported is the first case of vascular leiomyoma of the lung arising from smooth muscles of the pulmonary artery. A 62-year-old woman (non-smoker) was found to have a small tumor in the upper lobe in the right lung in routine check. Imaging modalities including CT demonstrated no metastatic lesions. Although clinical cytology and biopsy revealed no malignant cell, right upper lobectomy was performed under the clinical diagnosis of lung carcinoma. Grossly, a white tumor of 1 x 0.8 cm was recognized in the lung. Microscopically, the tumor was connected to the pulmonary arteries. The tumor was composed of mature smooth muscles. Small pulmonary arteries are embedded in the tumor. No lymphatics were seen. Immunohistochemically, the tumor cells were poisitive for alpha-smooth muscle actin, vimentin and Ki-67 (labeling 2%). However, they were negative for cytokeratin (CK) AE1/3, CK CAM5.2, desmin, S100 protein, p53, CD34, KIT, HMB45, estrogen receptor, progesterone receptor, and myoglobin. A pathological diagnosis of primary vascular leiomyoma arising from the smooth muscle of pulmonary artery was made. The patient is now free from tumor, and is now alive 10 year after the operation.

  16. Cobalt contraction of vascular smooth muscle

    SciTech Connect

    Dominiczak, A.; Clyde, E.; Bohr, D. )

    1991-03-11

    Although it has been reported that cobalt causes contraction of vascular smooth muscle, the mechanism responsible for this contraction has not been defined. The authors studied these contractions in rat aortic rings. Concentration-response studies indicated that the threshold for contraction was 10{sup {minus}8}M, maximum contraction occurred at 3 {times} 10{sup 7}M and relaxation began at 10{sup {minus}6}M. No contraction occurred in a calcium-free physiological salt solution and the contraction was not inhibited by H-7, a protein kinase C inhibitor. The authors conclude the cobalt in low concentrations causes contraction by activating calcium channels and that in high concentrations it causes relaxation by inactivating these same channels.

  17. Adult Vascular Wall Resident Multipotent Vascular Stem Cells, Matrix Metalloproteinases, and Arterial Aneurysms

    PubMed Central

    Amato, Bruno; Compagna, Rita; Amato, Maurizio; Grande, Raffaele; Butrico, Lucia; Rossi, Alessio; Naso, Agostino; Ruggiero, Michele; de Franciscis, Stefano

    2015-01-01

    Evidences have shown the presence of multipotent stem cells (SCs) at sites of arterial aneurysms: they can differentiate into smooth muscle cells (SMCs) and are activated after residing in a quiescent state in the vascular wall. Recent studies have implicated the role of matrix metalloproteinases in the pathogenesis of arterial aneurysms: in fact the increased synthesis of MMPs by arterial SMCs is thought to be a pivotal mechanism in aneurysm formation. The factors and signaling pathways involved in regulating wall resident SC recruitment, survival, proliferation, growth factor production, and differentiation may be also related to selective expression of different MMPs. This review explores the relationship between adult vascular wall resident multipotent vascular SCs, MMPs, and arterial aneurysms. PMID:25866513

  18. Biophysical Induction of Vascular Smooth Muscle Cell Podosomes

    PubMed Central

    Kim, Na Young; Kohn, Julie C.; Huynh, John; Carey, Shawn P.; Mason, Brooke N.; Vouyouka, Ageliki G.; Reinhart-King, Cynthia A.

    2015-01-01

    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. PMID:25785437

  19. Versican accumulates in vascular lesions in pulmonary arterial hypertension

    PubMed Central

    Chan, Christina K.; Eriksson, Inger; Johnson, Pamela Y.; Cao, Xiaofang; Westöö, Christian; Norvik, Christian; Andersson-Sjöland, Annika; Westergren-Thorsson, Gunilla; Johansson, Staffan; Hedin, Ulf; Kjellén, Lena; Wight, Thomas N.; Tran-Lundmark, Karin

    2016-01-01

    Abstract Pulmonary arterial hypertension (PAH) is a lethal condition for which there is no effective curative pharmacotherapy. PAH is characterized by vasoconstriction, wall thickening of pulmonary arteries, and increased vascular resistance. Versican is a chondroitin sulfate proteoglycan in the vascular extracellular matrix that accumulates following vascular injury and promotes smooth-muscle cell proliferation in systemic arteries. Here, we investigated whether versican may play a similar role in PAH. Paraffin-embedded lung sections from patients who underwent lung transplantation to treat PAH were used for immunohistochemistry. The etiologies of PAH in the subjects involved in this study were idiopathic PAH, scleroderma, and congenital heart disease (atrial septal defect) with left-to-right shunt. Independent of the underlying etiology, increased versican immunostaining was observed in areas of medial thickening, in neointima, and in plexiform lesions. Western blot of lung tissue lysates confirmed accumulation of versican in patients with PAH. Double staining for versican and CD45 showed only occasional colocalization in neointima of high-grade lesions and plexiform lesions. In vitro, metabolic labeling with [35S]sulfate showed that human pulmonary artery smooth-muscle cells (hPASMCs) produce mainly chondroitin sulfate glycosaminoglycans. In addition, hypoxia, but not cyclic stretch, was demonstrated to increase both versican messenger RNA expression and protein synthesis by hPASMCs. Versican accumulates in vascular lesions of PAH, and the amount of versican correlates more with lesion severity than with underlying etiology or inflammation. Hypoxia is a possible regulator of versican accumulation, which may promote proliferation of pulmonary smooth-muscle cells and vascular remodeling in PAH. PMID:27683612

  20. Arterial Myogenic Activation through Smooth Muscle Filamin A.

    PubMed

    Retailleau, Kevin; Arhatte, Malika; Demolombe, Sophie; Peyronnet, Rémi; Baudrie, Véronique; Jodar, Martine; Bourreau, Jennifer; Henrion, Daniel; Offermanns, Stefan; Nakamura, Fumihiko; Feng, Yuanyi; Patel, Amanda; Duprat, Fabrice; Honoré, Eric

    2016-03-08

    Mutations in the filamin A (FlnA) gene are frequently associated with severe arterial abnormalities, although the physiological role for this cytoskeletal element remains poorly understood in vascular cells. We used a conditional mouse model to selectively delete FlnA in smooth muscle (sm) cells at the adult stage, thus avoiding the developmental effects of the knockout. Basal blood pressure was significantly reduced in conscious smFlnA knockout mice. Remarkably, pressure-dependent tone of the resistance caudal artery was lost, whereas reactivity to vasoconstrictors was preserved. Impairment of the myogenic behavior was correlated with a lack of calcium influx in arterial myocytes upon an increase in intraluminal pressure. Notably, the stretch activation of CaV1.2 was blunted in the absence of smFlnA. In conclusion, FlnA is a critical upstream element of the signaling cascade underlying the myogenic tone. These findings allow a better understanding of the molecular basis of arterial autoregulation and associated disease states.

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

    PubMed Central

    Chen, Xiu-Juan; Wang, Na; Yi, Peng-Fei; Song, Min; Zhang, Bo; Wang, Yu-Zhong; Liang, Qiu-Hua

    2016-01-01

    Vascular calcification is an active process of osteoblastic differentiation of vascular smooth muscle cells; however, its definite mechanism remains unknown. Vinpocetine, a derivative of the alkaloid vincamine, has been demonstrated to inhibit the high glucose-induced proliferation of vascular smooth muscle cells; however, it remains unknown whether vinpocetine can affect the osteoblastic differentiation of vascular smooth muscle cells. We hereby investigated the effect of vinpocetine on vascular calcification using a beta-glycerophosphate-induced cell model. Our results showed that vinpocetine significantly reduced the osteoblast-like phenotypes of vascular smooth muscle cells including ALP activity, osteocalcin, collagen type I, Runx2 and BMP-2 expression as well as the formation of mineralized nodule. Vinpocetine, binding to translocation protein, induced phosphorylation of extracellular signal-related kinase and Akt and thus inhibited the translocation of nuclear factor-kappa B into the nucleus. Silencing of translocator protein significantly attenuated the inhibitory effect of vinpocetine on osteoblastic differentiation of vascular smooth muscle cells. Taken together, vinpocetine may be a promising candidate for the clinical therapy of vascular calcification. PMID:27589055

  2. Vascular Balloon Injury and Intraluminal Administration in Rat Carotid Artery

    PubMed Central

    Zhang, Wei; Trebak, Mohamed

    2014-01-01

    The carotid artery balloon injury model in rats has been well established for over two decades. It remains an important method to study the molecular and cellular mechanisms involved in vascular smooth muscle dedifferentiation, neointima formation and vascular remodeling. Male Sprague-Dawley rats are the most frequently employed animals for this model. Female rats are not preferred as female hormones are protective against vascular diseases and thus introduce a variation into this procedure. The left carotid is typically injured with the right carotid serving as a negative control. Left carotid injury is caused by the inflated balloon that denudes the endothelium and distends the vessel wall. Following injury, potential therapeutic strategies such as the use of pharmacological compounds and either gene or shRNA transfer can be evaluated. Typically for gene or shRNA transfer, the injured section of the vessel lumen is locally transduced for 30 min with viral particles encoding either a protein or shRNA for delivery and expression in the injured vessel wall. Neointimal thickening representing proliferative vascular smooth muscle cells usually peaks at 2 weeks after injury. Vessels are mostly harvested at this time point for cellular and molecular analysis of cell signaling pathways as well as gene and protein expression. Vessels can also be harvested at earlier time points to determine the onset of expression and/or activation of a specific protein or pathway, depending on the experimental aims intended. Vessels can be characterized and evaluated using histological staining, immunohistochemistry, protein/mRNA assays, and activity assays. The intact right carotid artery from the same animal is an ideal internal control. Injury-induced changes in molecular and cellular parameters can be evaluated by comparing the injured artery to the internal right control artery. Likewise, therapeutic modalities can be evaluated by comparing the injured and treated artery to the

  3. Smooth muscle cell contraction increases the critical buckling pressure of arteries.

    PubMed

    Hayman, Danika M; Zhang, Jinzhou; Liu, Qin; Xiao, Yangming; Han, Hai-Chao

    2013-02-22

    Recent in vitro experiments demonstrated that arteries under increased internal pressure or decreased axial stretch may buckle into the tortuous pattern that is commonly observed in aging or diseased arteries in vivo. It suggests that buckling is a possible mechanism for the development of artery tortuosity. Vascular tone has significant effects on arterial mechanical properties but its effect on artery buckling is unknown. The objective of this study was to determine the effects of smooth muscle cell contraction on the critical buckling pressure of arteries. Porcine common carotid arteries were perfused in an ex vivo organ culture system overnight under physiological flow and pressure. The perfusion pressure was adjusted to determine the critical buckling pressure of these arteries at in vivo and reduced axial stretch ratios (1.5 and 1.3) at baseline and after smooth muscle contraction and relaxation stimulated by norepinephrine and sodium nitroprusside, respectively. Our results demonstrated that the critical buckling pressure was significantly higher when the smooth muscle was contracted compared with relaxed condition (97.3mmHg vs 72.9mmHg at axial stretch ratio of 1.3 and 93.7mmHg vs 58.6mmHg at 1.5, p<0.05). These results indicate that arterial smooth muscle cell contraction increased artery stability.

  4. Smooth Muscle Cell Contraction Increases the Critical Buckling Pressure of Arteries

    PubMed Central

    Hayman, Danika M.; Zhang, Jinzhou; Liu, Qin; Xiao, Yangming; Han, Hai-Chao

    2012-01-01

    Recent in vitro experiments demonstrated that arteries under increased internal pressure or decreased axial stretch may buckle into the tortuous pattern that is commonly observed in aging or diseased arteries in vivo. It suggests that buckling is a possible mechanism for the development of artery tortuosity. Vascular tone has significant effects on arterial mechanical properties but its effect on artery buckling is unknown. The objective of this study was to determine the effects of smooth muscle cell contraction on the critical buckling pressure of arteries. Porcine common carotid arteries were perfused in an ex vivo organ culture system overnight under physiological flow and pressure. The perfusion pressure was adjusted to determine the critical buckling pressure of these arteries at in vivo and reduced axial stretch ratios (1.5 and 1.3) at baseline and after smooth muscle contraction and relaxation stimulated by norepinephrine and sodium nitroprusside, respectively. Our results demonstrated that the critical buckling pressure was significantly higher when the smooth muscle was contracted compared with relaxed condition (97.3mmHg versus 72.9mmHg at axial stretch ratio of 1.3 and 93.7mmHg vs 58.6mmHg at 1.5, p<0.05). These results indicate that arterial smooth muscle cell contraction increased artery stability. PMID:23261241

  5. Long-term expression of human adenosine deaminase in vascular smooth muscle cells of rats: A model for gene therapy

    SciTech Connect

    Lynch, C.M.; Miller, A.D. ); Clowes, M.M.; Osborne, W.R.A.; Clowes, A.W. )

    1992-02-01

    Gene transfer into vascular smooth muscle cells in animals was examined by using recombinant retroviral vectors containing an Escherichia coli {beta}-galactosidase gene or a human adenosine deaminase gene. Direct gene transfer by infusion of virus into rat carotid arteries was not observed. However, gene transfer by infection of smooth muscle cells in culture and seeding of the transduced cells onto arteries that had been denuded of endothelial cells was successful. Potentially therapeutic levels of human adenosine deaminase activity were detected over 6 months of observation, indicating the utility of vascular smooth muscle cells for gene therapy in humans.

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

    PubMed

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

    2017-02-01

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

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

    PubMed Central

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

    2017-01-01

    The role of smooth muscle endothelinB (ETB) receptors in regulating vascular function, blood pressure (BP), and neointimal remodeling has not been established. Selective knockout mice were generated to address the hypothesis that loss of smooth muscle ETB receptors would reduce BP, alter vascular contractility, and inhibit neointimal remodeling. ETB receptors were selectively deleted from smooth muscle by crossing floxed ETB mice with those expressing cre-recombinase controlled by the transgelin promoter. Functional consequences of ETB deletion were assessed using myography. BP was measured by telemetry, and neointimal lesion formation induced by femoral artery injury. Lesion size and composition (day 28) were analyzed using optical projection tomography, histology, and immunohistochemistry. Selective deletion of ETB was confirmed by genotyping, autoradiography, polymerase chain reaction, and immunohistochemistry. ETB-mediated contraction was reduced in trachea, but abolished from mesenteric veins, of knockout mice. Induction of ETB-mediated contraction in mesenteric arteries was also abolished in these mice. Femoral artery function was unaltered, and baseline BP modestly elevated in smooth muscle ETB knockout compared with controls (+4.2±0.2 mm Hg; P<0.0001), but salt-induced and ETB blockade–mediated hypertension were unaltered. Circulating endothelin-1 was not altered in knockout mice. ETB-mediated contraction was not induced in femoral arteries by incubation in culture medium or lesion formation, and lesion size was not altered in smooth muscle ETB knockout mice. In the absence of other pathology, ETB receptors in vascular smooth muscle make a small but significant contribution to ETB-dependent regulation of BP. These ETB receptors have no effect on vascular contraction or neointimal remodeling. PMID:28028193

  8. Smooth muscle cell-extrinsic vascular spasm arises from cardiomyocyte degeneration in sarcoglycan-deficient cardiomyopathy.

    PubMed

    Wheeler, Matthew T; Allikian, Michael J; Heydemann, Ahlke; Hadhazy, Michele; Zarnegar, Sara; McNally, Elizabeth M

    2004-03-01

    Vascular spasm is a poorly understood but critical biomedical process because it can acutely reduce blood supply and tissue oxygenation. Cardiomyopathy in mice lacking gamma-sarcoglycan or delta-sarcoglycan is characterized by focal damage. In the heart, sarcoglycan gene mutations produce regional defects in membrane permeability and focal degeneration, and it was hypothesized that vascular spasm was responsible for this focal necrosis. Supporting this notion, vascular spasm was noted in coronary arteries, and disruption of the sarcoglycan complex was observed in vascular smooth muscle providing a molecular mechanism for spasm. Using a transgene rescue strategy in the background of sarcoglycan-null mice, we replaced cardiomyocyte sarcoglycan expression. Cardiomyocyte-specific sarcoglycan expression was sufficient to correct cardiac focal degeneration. Intriguingly, successful restoration of the cardiomyocyte sarcoglycan complex also eliminated coronary artery vascular spasm, while restoration of smooth muscle sarcoglycan in the background of sarcoglycan-null alleles did not. This mechanism, whereby tissue damage leads to vascular spasm, can be partially corrected by NO synthase inhibitors. Therefore, we propose that cytokine release from damaged cardiomyocytes can feed back to produce vascular spasm. Moreover, vascular spasm feeds forward to produce additional cardiac damage.

  9. Functional preservation of vascular smooth muscle tissue

    NASA Technical Reports Server (NTRS)

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

    1973-01-01

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

  10. Vinpocetine Suppresses Pathological Vascular Remodeling by Inhibiting Vascular Smooth Muscle Cell Proliferation and Migration

    PubMed Central

    Cai, Yujun; Knight, Walter E.; Guo, Shujie; Li, Jian-Dong; Knight, Peter A.

    2012-01-01

    Abnormal vascular smooth muscle cell (SMC) activation is associated with various vascular disorders such as atherosclerosis, in-stent restenosis, vein graft disease, and transplantation-associated vasculopathy. Vinpocetine, a derivative of the alkaloid vincamine, has long been used as a cerebral blood flow enhancer for treating cognitive impairment. However, its role in pathological vascular remodeling remains unexplored. Herein, we show that systemic administration of vinpocetine significantly reduced neointimal formation in carotid arteries after ligation injury. Vinpocetine also markedly decreased spontaneous remodeling of human saphenous vein explants in ex vivo culture. In cultured SMCs, vinpocetine dose-dependently suppressed cell proliferation and caused G1-phase cell cycle arrest, which is associated with a decrease in cyclin D1 and an increase in p27Kip1 levels. In addition, vinpocetine dose-dependently inhibited platelet-derived growth factor (PDGF)-stimulated SMC migration as determined by the two-dimensional migration assays and three-dimensional aortic medial explant invasive assay. Moreover, vinpocetine significantly reduced PDGF-induced type I collagen and fibronectin expression. It is noteworthy that PDGF-stimulated phosphorylation of extracellular signal-regulated kinases 1/2 (ERK1/2), but not protein kinase B, was specifically inhibited by vinpocetine. Vinpocetine powerfully attenuated intracellular reactive oxidative species (ROS) production, which largely mediates the inhibitory effects of vinpocetine on ERK1/2 activation and SMC growth. Taken together, our results reveal a novel function of vinpocetine in attenuating neointimal hyperplasia and pathological vascular remodeling, at least partially through suppressing ROS production and ERK1/2 activation in SMCs. Given the safety profile of vinpocetine, this study provides insight into the therapeutic potential of vinpocetine in proliferative vascular disorders. PMID:22915768

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

    SciTech Connect

    Gao, Fu; Chambon, Pierre; Tellides, George; Kong, Wei; Zhang, Xiaoming; Li, Wei

    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.

  12. Atorvastatin inhibits myocardin expression in vascular smooth muscle cells.

    PubMed

    Li, Jingjing; Jiang, Jixin; Yin, Hao; Wang, Lifeng; Tian, Ruijuan; Li, Haijie; Wang, Zengyong; Li, Dong; Wang, Yuebing; Gui, Yu; Walsh, Michael P; Zheng, Xi-Long

    2012-07-01

    Atorvastatin (ATV), an inhibitor of 3-hydroxy-3-methylglutaryl-coenzyme A reductase, is widely prescribed as a lipid-lowering drug. It also inhibits the RhoA-Rho-associated kinase pathway in vascular smooth muscle (SM) cells and critically inhibits SM function. Myocardin is a coactivator of serum response factor, which upregulates SM contractile proteins. The RhoA-Rho-associated kinase pathway, which directly triggers SM contraction, also increases myocardin gene expression. Therefore, we investigated whether ATV inhibits myocardin gene expression in SM cells. In mice injected with ATV (IP 20 μg/g per day) for 5 days, myocardin gene expression was significantly downregulated in aortic and carotid arterial tissues with decreased expression of myocardin target genes SM α-actin and SM22. Correspondingly, the contractility of aortic rings in mice treated with ATV or the Rho-associated kinase inhibitor Y-27632 was reduced in response to treatment with either KCl or phenylephrine. In cultured mouse and human aortic SM cells, KCl treatment stimulated the expression of myocardin, SM α-actin, and SM22. These stimulatory effects were prevented by ATV treatment. ATV-induced inhibition of myocardin expression was prevented by pretreatment with either mevalonate or geranylgeranylpyrophosphate but not farnesylpyrophosphate. Treatment with Y-27632 mimicked ATV effects on the gene expression of myocardin, SM α-actin, and SM22, further suggesting a role for the RhoA-Rho-associated kinase pathway in ATV effects. Furthermore, ATV treatment inhibited RhoA membrane translocation and activation; these effects were prevented by pretreatment with mevalonate. We conclude that ATV inhibits myocardin gene expression in vivo and in vitro, suggesting a novel mechanism for ATV inhibition of vascular contraction.

  13. Mechanisms of Vascular Smooth Muscle Contraction and the Basis for Pharmacologic Treatment of Smooth Muscle Disorders

    PubMed Central

    Brozovich, F.V.; Nicholson, C.J.; Degen, C.V.; Gao, Yuan Z.; Aggarwal, M.

    2016-01-01

    The smooth muscle cell directly drives the contraction of the vascular wall and hence regulates the size of the blood vessel lumen. We review here the current understanding of the molecular mechanisms by which agonists, therapeutics, and diseases regulate contractility of the vascular smooth muscle cell and we place this within the context of whole body function. We also discuss the implications for personalized medicine and highlight specific potential target molecules that may provide opportunities for the future development of new therapeutics to regulate vascular function. PMID:27037223

  14. Troglitazone inhibits vascular smooth muscle cell growth and intimal hyperplasia.

    PubMed Central

    Law, R E; Meehan, W P; Xi, X P; Graf, K; Wuthrich, D A; Coats, W; Faxon, D; Hsueh, W A

    1996-01-01

    Vascular smooth muscle cell (VSMC) proliferation and migration are responses to arterial injury that are highly important to the processes of restenosis and atherosclerosis. In the arterial balloon injury model in the rat, platelet-derived growth factor (PDGF) and basic fibroblast growth factor (bFGF) are induced in the vessel wall and regulate these VSMC activities. Novel insulin sensitizing agents, thiazolidinediones, have been demonstrated to inhibit insulin and epidermal growth factor-induced growth of VSMCs. We hypothesized that these agents might also inhibit the effect of PDGF and bFGF on cultured VSMCs and intimal hyperplasia in vivo. Troglitazone (1 microM), a member of the thiazolidinedione class, produced a near complete inhibition of both bFGF-induced DNA synthesis as measured by bromodeoxyuridine incorporation (6.5+/-3.9 vs. 17.6+/-4.3% cells labeled, P < 0.05) and c-fos induction. This effect was associated with an inhibition (by 73+/-4%, P < 0.01) by troglitazone of the transactivation of the serum response element, which regulates c-fos expression. Inhibition of c-fos induction by troglitazone appeared to occur via a blockade of the MAP kinase pathway at a point downstream of MAP kinase activation by MAP kinase kinase. At this dose, troglitazone also inhibited PDGF-BB-directed migration of VSMC (by 70+/-6%, P < 0.01). These in vitro effects were operative in vivo. Quantitative image analysis revealed that troglitazone-treated rats had 62% (P < 0.001) less neointima/media area ratio 14 d after balloon injury of the aorta compared with injured rats that received no troglitazone. These results suggest troglitazone is a potent inhibitor of VSMC proliferation and migration and, thus, may be a useful agent to prevent restenosis and possibly atherosclerosis. PMID:8878442

  15. Mitochondrial Fission of Smooth Muscle Cells Is Involved in Artery Constriction.

    PubMed

    Liu, Ming-Yu; Jin, Jing; Li, Shan-Liang; Yan, Jie; Zhen, Chang-Lin; Gao, Jin-Lai; Zhang, Yong-Hui; Zhang, Yan-Qiu; Shen, Xin; Zhang, Liang-Shuan; Wei, Yuan-Yuan; Zhao, Yu; Wang, Chen-Guang; Bai, Yun-Long; Dong, De-Li

    2016-11-01

    Mitochondria are dynamic organelles and continuously undergo fission and fusion processes. Mitochondrial fission is involved in multiple physiological or pathological processes, but the role of mitochondrial fission of smooth muscle cells in artery constriction is unknown. The role of mitochondrial fission of smooth muscle cells in arterial function was investigated by measuring the tension of rat mesenteric arteries and thoracic aorta and by evaluating mitochondrial fission, mitochondrial reactive oxygen species, and cytosolic [Ca(2+)]i in rat vascular smooth muscle cells. Mitochondrial fission inhibitors mdivi-1 and dynasore antagonized phenylephrine- and high K(+)-induced constriction of rat mesenteric arteries. Mdivi-1 relaxed phenylephrine-induced constriction, and mdivi-1 pretreatment prevented phenylephrine-induced constriction in mice, rat aorta, and human mesenteric arteries. Phenylephrine- and high K(+)-induced increase of mitochondrial fission in smooth muscle cells of rat aorta and the increase was inhibited by mdivi-1. Mdivi-1 inhibited high K(+)-induced increases of mitochondrial fission, mitochondrial reactive oxygen species, and cytosolic [Ca(2+)]i in rat vascular smooth muscle cells. Prechelation of cytosolic Ca(2+) prevented high K(+)-induced cytosolic [Ca(2+)]i increase, mitochondrial fission, and mitochondrial reactive oxygen species overproduction. Mitochondria-targeted antioxidant mito-TEMPO antagonized phenylephrine- and high K(+)-induced constriction of rat mesenteric arteries. Nitroglycerin and ROCK (Rho-associated protein kinase) inhibitor Y27632, the 2 vasodilators with different vasorelaxant mechanisms, relaxed high K(+)-induced vasoconstriction and inhibited high K(+)-induced mitochondrial fission. In conclusion, the mitochondrial fission of smooth muscle cells is involved in artery constriction.

  16. Effects of sodium selenite on vascular smooth muscle reactivity.

    PubMed

    Togna, G; Russo, P; Pierconti, F; Caprino, L

    2000-02-01

    The effects of sodium selenite (Na(2)SeO(3)) on the vascular smooth muscle reactivity of rabbit aorta were studied. In isolated rabbit aorta, Na(2)SeO(3) inhibited contractile response to phenylephrine and developed a lasting contracture in the vascular tissue. Relaxation in phenylephrine-precontracted aortic rings induced by sodium nitroprusside and 8-bromo-guanosine 3':5'-cyclic-monophosphate was also inhibited. Preliminary data obtained with ascorbic acid suggested a partial involvement of an oxidative mechanism. Excluding the possibility that Se damages actin or modifies its distribution (immunohistochemical evaluation), results indicate that Se alters vascular smooth muscle reactivity by inhibiting both its contracting and relaxing properties. Calcium-dependent mechanisms appear to be primarily involved and an interference with calcium re-uptake by sarcoplasmic reticulum as a possible site of Se vascular action could be hypothesized.

  17. Cannabinoid CB{sub 1} receptor inhibition decreases vascular smooth muscle migration and proliferation

    SciTech Connect

    Rajesh, Mohanraj; Mukhopadhyay, Partha; Hasko, Gyoergy; Pacher, Pal

    2008-12-26

    Vascular smooth muscle proliferation and migration triggered by inflammatory stimuli and chemoattractants such as platelet-derived growth factor (PDGF) are key events in the development and progression of atherosclerosis and restenosis. Cannabinoids may modulate cell proliferation and migration in various cell types through cannabinoid receptors. Here we investigated the effects of CB{sub 1} receptor antagonist rimonabant (SR141716A), which has recently been shown to have anti-atherosclerotic effects both in mice and humans, on PDGF-induced proliferation, migration, and signal transduction of human coronary artery smooth muscle cells (HCASMCs). PDGF induced Ras and ERK 1/2 activation, while increasing proliferation and migration of HCASMCs, which were dose dependently attenuated by CB{sub 1} antagonist, rimonabant. These findings suggest that in addition to improving plasma lipid alterations and decreasing inflammatory cell migration and inflammatory response, CB{sub 1} antagonists may exert beneficial effects in atherosclerosis and restenosis by decreasing vascular smooth muscle proliferation and migration.

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

    PubMed

    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.

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

    PubMed Central

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

    2015-01-01

    Background Hypertension is a public health problem and increases the incidence of cardiovascular diseases. Objective To evaluate the effects of a resistance exercise session on the contractile and relaxing mechanisms of vascular smooth muscle in mesenteric arteries of NG-nitro L-arginine methyl ester (L-NAME)-induced hypertensive rats. Methods Wistar rats were divided into three groups: control (C), hypertensive (H), and exercised hypertensive (EH). Hypertension was induced by administration of 20 mg/kg of L-NAME for 7 days prior to experimental protocols. The resistance exercise protocol consisted of 10 sets of 10 repetitions and intensity of 40% of one repetition maximum. The reactivity of vascular smooth muscle was evaluated by concentration‑response curves to phenylephrine (PHEN), potassium chloride (KCl) and sodium nitroprusside (SNP). Results Rats treated with L-NAME showed an increase (p < 0.001) in systolic blood pressure (SBP), diastolic blood pressure (DBP) and mean arterial pressure (MAP) compared to the initial period of induction. No difference in PHEN sensitivity was observed between groups H and EH. Acute resistance exercise reduced (p < 0.001) the contractile response induced by KCl at concentrations of 40 and 60 mM in group EH. Greater (p < 0.01) smooth muscle sensitivity to NPS was observed in group EH as compared to group H. Conclusion One resistance exercise session reduces the contractile response induced by KCl in addition to increasing the sensitivity of smooth muscle to NO in mesenteric arteries of hypertensive rats. PMID:26107814

  20. Maintenance of GLUT4 expression in smooth muscle prevents hypertension-induced changes in vascular reactivity.

    PubMed

    Atkins, Kevin B; Seki, Yoshinori; Saha, Jharna; Eichinger, Felix; Charron, Maureen J; Brosius, Frank C

    2015-02-01

    Previous studies have shown that expression of GLUT4 is decreased in arterial smooth muscle of hypertensive rats and mice and that total body overexpression of GLUT4 in mice prevents enhanced arterial reactivity in hypertension. To demonstrate that the effect of GLUT4 overexpression on vascular responses is dependent on vascular smooth muscle GLUT4 rather than on some systemic effect we developed and tested smooth-muscle-specific GLUT4 transgenic mice (SMG4). When made hypertensive with angiotensin II, both wild-type and SMG4 mice exhibited similarly increased systolic blood pressure. Responsiveness to phenylephrine, serotonin, and prostaglandin F2α was significantly increased in endothelium-intact aortic rings from hypertensive wild-type mice but not in aortae of SMG4 mice. Inhibition of Rho-kinase equally reduced serotonin-stimulated contractility in aortae of hypertensive wild-type and SMG4-mice. In addition, acetylcholine-stimulated relaxation was significantly decreased in aortic rings of hypertensive wild-type mice, but not in rings of SMG4 mice. Inhibition of either prostacylin receptors or cyclooxygenase-2 reduced relaxation in rings of hypertensive SMG4 mice. Inhibition of cyclooxygenase-2 had no effect on relaxation in rings of hypertensive wild-type mice. Cyclooxygenase-2 protein expression was decreased in hypertensive wild-type aortae but not in hypertensive SMG4 aortae compared to nonhypertensive controls. Our results demonstrate that smooth muscle expression of GLUT4 exerts a major effect on smooth muscle contractile responses and endothelium-dependent vasorelaxation and that normal expression of GLUT4 in vascular smooth muscle is required for appropriate smooth muscle and endothelial responses.

  1. Perfusion of veins at arterial pressure increases the expression of KLF5 and cell cycle genes in smooth muscle cells

    SciTech Connect

    Amirak, Emre; Zakkar, Mustafa; Evans, Paul C.; Kemp, Paul R.

    2010-01-01

    Vascular smooth muscle cell (VSMC) proliferation remains a major cause of veno-arterial graft failure. We hypothesised that exposure of venous SMCs to arterial pressure would increase KLF5 expression and that of cell cycle genes. Porcine jugular veins were perfused at arterial or venous pressure in the absence of growth factors. The KLF5, c-myc, cyclin-D and cyclin-E expression were elevated within 24 h of perfusion at arterial pressure but not at venous pressure. Arterial pressure also reduced the decline in SM-myosin heavy chain expression. These data suggest a role for KLF5 in initiating venous SMCs proliferation in response to arterial pressure.

  2. [Mechanism of losartan suppressing vascular calcification in rat aortic artery].

    PubMed

    Shao, Juan; Wu, Panfeng; Wu, Jiliang; Li, Mincai

    2016-08-01

    Objective To investigate the effect of the angiotensin II receptor 1 (AT1R) blocker losartan on vascular calcification in rat aortic artery and explore the underlying mechanisms. Methods SD rats were divided randomly into control group, vascular calcification model group and treatment group. Vascular calcification models were made by subcutaneous injection of warfarin plus vitamin K1 for two weeks. Rats in the treatment group were subcutaneously injected with losartan (10 mg/kg) at the end of the first week and consecutively for one week. We observed the morphological changes by HE staining and the calcium deposition by Alizarin red staining in the artery vascular wall. The mRNA expressions of bone morphogenetic protein 2 (BMP2) and Runt-related transcription factor 2 (RUNX2) were analyzed by reverse transcription PCR. The BMP2 and RUNX2 protein expressions were determined by Western blotting. The apoptosis of smooth muscle cells (SMCs) were detected by TUNEL. The AT1R expression was tested by fluorescent immunohistochemistry. Results The aortic vascular calcification was induced by warfarin and vitamin K1. Compared with the vascular calcification model group, the mRNA and protein expressions of BMP2 and RUNX2 were significantly downregulated in the aorta in the losartan treatment group. Furthermore, the apoptosis of SMCs and the AT1R expression obviously decreased. Conclusion AT1R blocker losartan inhibits the apoptosis of SMCs and reduces AT1R expression; it downregulates the BMP2 and RUNX2 expressions in the vascular calcification process.

  3. Tobacco constituents are mitogenic for arterial smooth-muscle cells

    SciTech Connect

    Becker, C.G.; Hajjar, D.P.; Hefton, J.M.

    1985-07-01

    Tobacco glycoprotein (TGP) purified from flue-cured tobacco leaves, tar-derived material (TAR), the water soluble, nondialyzable, delipidized extract of cigarette smoke condensate, rutin-bovine serum albumin conjugates, quercetin, and chlorogenic acid are mitogenic for bovine aortic smooth-muscle cells, but not adventitial fibroblasts. The mitogenicity appears to depend on polyphenol epitopes on carrier molecules. Ellagic acid, another plant polyphenol, inhibited arterial smooth-muscle proliferation. These results suggest that a number of ubiquitous, plant-derived substances may influence smooth-muscle cell proliferation in the arterial wall.

  4. Vascular mechanics of the coronary artery

    NASA Technical Reports Server (NTRS)

    Veress, A. I.; Vince, D. G.; Anderson, P. M.; Cornhill, J. F.; Herderick, E. E.; Klingensmith, J. D.; Kuban, B. D.; Greenberg, N. L.; Thomas, J. D.

    2000-01-01

    This paper describes our research into the vascular mechanics of the coronary artery and plaque. The three sections describe the determination of arterial mechanical properties using intravascular ultrasound (IVUS), a constitutive relation for the arterial wall, and finite element method (FEM) models of the arterial wall and atheroma. METHODS: Inflation testing of porcine left anterior descending coronary arteries was conducted. The changes in the vessel geometry were monitored using IVUS, and intracoronary pressure was recorded using a pressure transducer. The creep and quasistatic stress/strain responses were determined. A Standard Linear Solid (SLS) was modified to reproduce the non-linear elastic behavior of the arterial wall. This Standard Non-linear Solid (SNS) was implemented into an axisymetric thick-walled cylinder numerical model. Finite element analysis models were created for five age groups and four levels of stenosis using the Pathobiological Determinants of Atherosclerosis Youth (PDAY) database. RESULTS: The arteries exhibited non-linear elastic behavior. The total tissue creep strain was epsilon creep = 0.082 +/- 0.018 mm/mm. The numerical model could reproduce both the non-linearity of the porcine data and time dependent behavior of the arterial wall found in the literature with a correlation coefficient of 0.985. Increasing age had a strong positive correlation with the shoulder stress level, (r = 0.95). The 30% stenosis had the highest shoulder stress due to the combination of a fully formed lipid pool and a thin cap. CONCLUSIONS: Studying the solid mechanics of the arterial wall and the atheroma provide important insights into the mechanisms involved in plaque rupture.

  5. Piezo1 in Smooth Muscle Cells Is Involved in Hypertension-Dependent Arterial Remodeling.

    PubMed

    Retailleau, Kevin; Duprat, Fabrice; Arhatte, Malika; Ranade, Sanjeev Sumant; Peyronnet, Rémi; Martins, Joana Raquel; Jodar, Martine; Moro, Céline; Offermanns, Stefan; Feng, Yuanyi; Demolombe, Sophie; Patel, Amanda; Honoré, Eric

    2015-11-10

    The mechanically activated non-selective cation channel Piezo1 is a determinant of vascular architecture during early development. Piezo1-deficient embryos die at midgestation with disorganized blood vessels. However, the role of stretch-activated ion channels (SACs) in arterial smooth muscle cells in the adult remains unknown. Here, we show that Piezo1 is highly expressed in myocytes of small-diameter arteries and that smooth-muscle-specific Piezo1 deletion fully impairs SAC activity. While Piezo1 is dispensable for the arterial myogenic tone, it is involved in the structural remodeling of small arteries. Increased Piezo1 opening has a trophic effect on resistance arteries, influencing both diameter and wall thickness in hypertension. Piezo1 mediates a rise in cytosolic calcium and stimulates activity of transglutaminases, cross-linking enzymes required for the remodeling of small arteries. In conclusion, we have established the connection between an early mechanosensitive process, involving Piezo1 in smooth muscle cells, and a clinically relevant arterial remodeling.

  6. Smooth muscle BK channel activity influences blood pressure independent of vascular tone in mice

    PubMed Central

    Sachse, Gregor; Faulhaber, Jörg; Seniuk, Anika; Ehmke, Heimo; Pongs, Olaf

    2014-01-01

    The large conductance voltage- and Ca2+-activated K+ (BK) channel is an important determinant of vascular tone and contributes to blood pressure regulation. Both activities depend on the ancillary BKβ1 subunit. To determine the significance of smooth muscle BK channel activity for blood pressure regulation, we investigated the potential link between changes in arterial tone and altered blood pressure in BKβ1 knockout (BKβ1−/−) mice from three different genetically defined strains. While vascular tone was consistently increased in all BKβ1−/− mice independent of genetic background, BKβ1−/− strains exhibited increased (strain A), unaltered (strain B) or decreased (strain C) mean arterial blood pressures compared to their corresponding BKβ1+/+ controls. In agreement with previous data on aldosterone regulation by renal/adrenal BK channel function, BKβ1−/− strain A mice have increased plasma aldosterone and increased blood pressure. Consistently, blockade of mineralocorticoid receptors by spironolactone treatment reversibly restored the elevated blood pressure to the BKβ1+/+ strain A level. In contrast, loss of BKβ1 did not affect plasma aldosterone in strain C mice. Smooth muscle-restricted restoration of BKβ1 expression increased blood pressure in BKβ1−/− strain C mice, implying that impaired smooth muscle BK channel activity lowers blood pressure in these animals. We conclude that BK channel activity directly affects vascular tone but influences blood pressure independent of this effect via different pathways. PMID:24687584

  7. Caveolin-1 regulates contractility in differentiated vascular smooth muscle.

    PubMed

    Je, Hyun-Dong; Gallant, Cynthia; Leavis, Paul C; Morgan, Kathleen G

    2004-01-01

    Caveolin is a principal component of caveolar membranes. In the present study, we utilized a decoy peptide approach to define the degree of involvement of caveolin in PKC-dependent regulation of contractility of differentiated vascular smooth muscle. The primary isoform of caveolin in ferret aorta vascular smooth muscle is caveolin-1. Chemical loading of contractile vascular smooth muscle tissue with a synthetic caveolin-1 scaffolding domain peptide inhibited PKC-dependent increases in contractility induced by a phorbol ester or an alpha agonist. Peptide loading also resulted in a significant inhibition of phorbol ester-induced adducin Ser662 phosphorylation, an intracellular monitor of PKC kinase activity, ERK1/2 activation, and Ser789 phosphorylation of the actin binding protein caldesmon. alpha-Agonist-induced ERK1-1/2 activation was also inhibited by the caveolin-1 peptide. Scrambled peptide-loaded tissues or sham-loaded tissues were unaffected with respect to both contractility and signaling. Depolarization-induced activation of contraction was not affected by caveolin peptide loading. Similar results with respect to contractility and ERK1/2 activation during exposure to the phorbol ester or the alpha-agonist were obtained with the cholesterol-depleting agent methyl-beta-cyclodextrin. These results are consistent with a role for caveolin-1 in the coordination of signaling leading to the regulation of contractility of smooth muscle.

  8. Nuclear reprogramming and its role in vascular smooth muscle cells.

    PubMed

    Zaina, Silvio; del Pilar Valencia-Morales, Maria; Tristán-Flores, Fabiola E; Lund, Gertrud

    2013-09-01

    In general terms, "nuclear reprogramming" refers to a change in gene expression profile that results in a significant switch in cellular phenotype. Nuclear reprogramming was first addressed by pioneering studies of cell differentiation during embryonic development. In recent years, nuclear reprogramming has been studied in great detail in the context of experimentally controlled dedifferentiation and transdifferentiation of mammalian cells for therapeutic purposes. In this review, we present a perspective on nuclear reprogramming in the context of spontaneous, pathophysiological phenotypic switch of vascular cells occurring in the atherosclerotic lesion. In particular, we focus on the current knowledge of epigenetic mechanisms participating in the extraordinary flexibility of the gene expression profile of vascular smooth muscle cells and other cell types participating in atherogenesis. Understanding how epigenetic changes participate in vascular cell plasticity may lead to effective therapies based on the remodelling of the vascular architecture.

  9. SGLT inhibitors attenuate NO-dependent vascular relaxation in the pulmonary artery but not in the coronary artery

    PubMed Central

    Han, Ying; Cho, Young-Eun; Ayon, Ramon; Guo, Rui; Youssef, Katia D.; Pan, Minglin; Dai, Anzhi; Yuan, Jason X.-J.

    2015-01-01

    Inhibitors of sodium-glucose cotransporter (SGLT)2 are a new class of oral drugs for type 2 diabetic patients that reduce plasma glucose levels by inhibiting renal glucose reabsorption. There is increasing evidence showing the beneficial effect of SGLT2 inhibitors on glucose control; however, less information is available regarding the impact of SGLT2 inhibitors on cardiovascular outcomes. The present study was designed to determine whether SGLT inhibitors regulate vascular relaxation in mouse pulmonary and coronary arteries. Phlorizin (a nonspecific SGLT inhibitor) and canagliflozin (a SGLT2-specific inhibitor) relaxed pulmonary arteries in a dose-dependent manner, but they had little or no effect on coronary arteries. Pretreatment with phlorizin or canagliflozin significantly inhibited sodium nitroprusside (SNP; a nitric oxide donor)-induced vascular relaxation in pulmonary arteries but not in coronary arteries. Phlorizin had no effect on cGMP-dependent relaxation in pulmonary arteries. SNP induced membrane hyperpolarization in human pulmonary artery smooth muscle cells, and pretreatment of cells with phlorizin and canagliflozin attenuated SNP-induced membrane hyperpolarization by decreasing K+ activities induced by SNP. Contrary to the result observed in ex vivo experiments with SGLT inhibitors, SNP-dependent relaxation in pulmonary arteries was not altered by chronic administration of canagliflozin. On the other hand, canagliflozin administration significantly enhanced SNP-dependent relaxation in coronary arteries in diabetic mice. These data suggest that SGLT inhibitors differentially regulate vascular relaxation depending on the type of arteries, duration of the treatment, and health condition, such as diabetes. PMID:26361875

  10. Smoking and Female Sex: Independent Predictors of Human Vascular Smooth Muscle Cells Stiffening

    PubMed Central

    Dinardo, Carla Luana; Santos, Hadassa Campos; Vaquero, André Ramos; Martelini, André Ricardo; Dallan, Luis Alberto Oliveira; Alencar, Adriano Mesquita; Krieger, José Eduardo; Pereira, Alexandre Costa

    2015-01-01

    Aims Recent evidence shows the rigidity of vascular smooth muscle cells (VSMC) contributes to vascular mechanics. Arterial rigidity is an independent cardiovascular risk factor whose associated modifications in VSMC viscoelasticity have never been investigated. This study’s objective was to evaluate if the arterial rigidity risk factors aging, African ancestry, female sex, smoking and diabetes mellitus are associated with VMSC stiffening in an experimental model using a human derived vascular smooth muscle primary cell line repository. Methods Eighty patients subjected to coronary artery bypass surgery were enrolled. VSMCs were extracted from internal thoracic artery fragments and mechanically evaluated using Optical Magnetic Twisting Cytometry assay. The obtained mechanical variables were correlated with the clinical variables: age, gender, African ancestry, smoking and diabetes mellitus. Results The mechanical variables Gr, G’r and G”r had a normal distribution, demonstrating an inter-individual variability of VSMC viscoelasticity, which has never been reported before. Female sex and smoking were independently associated with VSMC stiffening: Gr (apparent cell stiffness) p = 0.022 and p = 0.018, R2 0.164; G’r (elastic modulus) p = 0.019 and p = 0.009, R2 0.184 and G”r (dissipative modulus) p = 0.011 and p = 0.66, R2 0.141. Conclusion Female sex and smoking are independent predictors of VSMC stiffening. This pro-rigidity effect represents an important element for understanding the vascular rigidity observed in post-menopausal females and smokers, as well as a potential therapeutic target to be explored in the future. There is a significant inter-individual variation of VSMC viscoelasticity, which is slightly modulated by clinical variables and probably relies on molecular factors. PMID:26661469

  11. [Arterial vascular injuries in fractures and dislocations].

    PubMed

    Piatek, S; Bürger, T; Halloul, Z; Westphal, T; Holmenschlager, F; Winckler, S

    2001-05-01

    We analyzed reasons, numbers and results of arterial lesions accompanying fractures (n = 21) and luxations (n = 6) in a 6-year-period (1993-1998) retrospectively. Traffic accidents were in nearly 50% responsible for the injuries. 8 patients had suffered multiple injuries. In 17 patients the lower, and in 10 patients the upper extremities were affected. The vascular wall was completely disrupted or severed in 74%. In 7 cases (26%), patients had suffered blunt or indirect arterial trauma with intima- and media-lacerations due to subcapital fracture of the humerus (n = 2), fractured femoral bone (n = 1), luxation of the knee joint (n = 3) or the elbow (n = 1). The mean preoperative time period was 6 hours and 20 minutes (2 to 16 hours) in patients with complete ischaemia. Vascular reconstruction was performed by interposition of an autologous vein graft or an autologous venous bypass (n = 20), by direct reconstruction and primary suturing (n = 2), by use of a venous patch plasty (n = 2) and, in a single case, by autologous bypass procedure. In one case, a crural artery was ligated, in another case with a Mangled Extremity Severity Score (MESS) of 7 points a primary amputation of the lower leg was necessary. In 5 patients (19%) secondary amputations were performed. No patient died. The final outcome is mostly influenced by the preoperative period of ischaemia.

  12. Statistical modeling of the arterial vascular tree

    NASA Astrophysics Data System (ADS)

    Beck, Thomas; Godenschwager, Christian; Bauer, Miriam; Bernhardt, Dominik; Dillmann, Rüdiger

    2011-03-01

    Automatic examination of medical images becomes increasingly important due to the rising amount of data. Therefore automated methods are required which combine anatomical knowledge and robust segmentation to examine the structure of interest. We propose a statistical model of the vascular tree based on vascular landmarks and unbranched vessel sections. An undirected graph provides anatomical topology, semantics, existing landmarks and attached vessel sections. The atlas was built using semi-automatically generated geometric models of various body regions ranging from carotid arteries to the lower legs. Geometric models contain vessel centerlines as well as orthogonal cross-sections in equidistant intervals with the vessel contour having the form of a polygon path. The geometric vascular model is supplemented by anatomical landmarks which are not necessarily related to the vascular system. These anatomical landmarks define point correspondences which are used for registration with a Thin-Plate-Spline interpolation. After the registration process, the models were merged to form the statistical model which can be mapped to unseen images based on a subset of anatomical landmarks. This approach provides probability distributions for the location of landmarks, vessel-specific geometric properties including shape, expected radii and branching points and vascular topology. The applications of this statistical model include model-based extraction of the vascular tree which greatly benefits from vessel-specific geometry description and variation ranges. Furthermore, the statistical model can be applied as a basis for computer aided diagnosis systems as indicator for pathologically deformed vessels and the interaction with the geometric model is significantly more user friendly for physicians through anatomical names.

  13. Role of blood and vascular smooth muscle in the vasoactivity of nitrite.

    PubMed

    Liu, Taiming; Schroeder, Hobe J; Barcelo, Lisa; Bragg, Shannon L; Terry, Michael H; Wilson, Sean M; Power, Gordon G; Blood, Arlin B

    2014-10-01

    Recent evidence from humans and rats indicates that nitrite is a vasodilator under hypoxic conditions by reacting with metal-containing proteins to produce nitric oxide (NO). We tested the hypothesis that near-physiological concentrations of nitrite would produce vasodilation in a hypoxia- and concentration-dependent manner in the hind limb of sheep. Anesthetized sheep were instrumented to measure arterial blood pressure and femoral blood flows continuously in both hind limbs. Nitrite was infused into one femoral artery to raise the nitrite concentration in the femoral vein by 10 to 15-fold while the sheep breathed 50%, 14% or 12% oxygen in inspired air. In contrast to reports in humans and rats, the nitrite infusion had no measurable effect on mean femoral blood flows or vascular conductances, regardless of inspired O2 levels. In vitro experiments showed no significant difference in the release of NO from nitrite in sheep and human red blood cells. Further experiments demonstrated nitrite is converted to NO in rat artery homogenates faster than sheep arteries, and that this source of NO production is attenuated in the presence of a heme oxidizer. Finally, western blots indicate that concentrations of the heme-containing protein cytoglobin, but not myoglobin, are markedly lower in sheep arteries compared with rats. Overall, the results demonstrate that nitrite is not a physiological vasodilator in sheep. This is likely due to a lack of conversion of nitrite to NO within the vascular smooth muscle, perhaps due to deficient amounts of the heme-containing protein cytoglobin.

  14. Vascular smooth muscle cells from injured rat aortas display elevated matrix production associated with transforming growth factor-beta activity.

    PubMed Central

    Rasmussen, L. M.; Wolf, Y. G.; Ruoslahti, E.

    1995-01-01

    The arterial response to injury is characterized by a short period of increased proliferation and migration of vascular smooth muscle cells, followed by an extended period of extracellular matrix accumulation in the intima. Transforming growth factor-beta (TGF-beta) has been implicated as a causative factor in the formation of extracellular matrix in this process, which leads to progressive thickening of the intima, known as intimal hyperplasia. In vitro analysis of vascular smooth muscle cells harvested from normal rat aortas and from aortas injured 14 days earlier showed that both types of cells attached equally well to culture dishes but that the initial spreading of the cells was increased in cells derived from injured vessels. Cells from the injured arteries produced more fibronectin and proteoglycans into the culture medium than the cells from normal arteries and contained more TGF-beta 1 mRNA. TGF-beta 1 increased proteoglycan synthesis by normal smooth muscle cells, and the presence of a neutralizing anti-TGF-beta 1 antibody reduced proteoglycan synthesis by the cells from injured arteries in culture. Fibronectin synthesis was not altered by these treatments. These results indicate that the accumulation of extracellular matrix components in neointimal lesions is at least partially caused by autocrine TGF-beta activity in vascular smooth muscle cells. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 PMID:7573349

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

    PubMed Central

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

    2015-01-01

    Arterial stiffness is recognized as a risk factor for many cardiovascular diseases. Aldosterone via its binding to and activation of the mineralocorticoid receptors (MRs) is a main regulator of blood pressure by controlling renal sodium reabsorption. Although both clinical and experimental data indicate that MR activation by aldosterone is involved in arterial stiffening, the molecular mechanism is not known. In addition to the kidney, MR is expressed in both endothelial and vascular smooth muscle cells (VSMCs), but the specific contribution of the VSMC MR to aldosterone-induced vascular stiffness remains to be explored. To address this question, we generated a mouse model with conditional inactivation of the MR in VSMC (MRSMKO). MRSMKO mice show no alteration in renal sodium handling or vascular structure, but they have decreased blood pressure when compared with control littermate mice. In vivo at baseline, large vessels of mutant mice presented with normal elastic properties, whereas carotids displayed a smaller diameter when compared with those of the control group. As expected after aldosterone/salt challenge, the arterial stiffness increased in control mice; however, it remained unchanged in MRSMKO mice, without significant modification in vascular collagen/elastin ratio. Instead, we found that the fibronectin/α5-subunit integrin ratio is profoundly altered in MRSMKO mice because the induction of α5 expression by aldosterone/salt challenge is prevented in mice lacking VSMC MR. Altogether, our data reveal in the aldosterone/salt hypertension model that MR activation specifically in VSMC leads to the arterial stiffening by modulation of cell-matrix attachment proteins independent of major vascular structural changes. PMID:24296280

  16. Smooth muscle cell proliferation in the occluded rat carotid artery: lack of requirement for luminal platelets.

    PubMed Central

    Guyton, J. R.; Karnovsky, M. J.

    1979-01-01

    The relationship of intimal smooth muscle cell proliferation in the permanently occluded rat carotid artery to the presence or absence of luminal platelets was examined. Blood was rinsed from the arterial lumen immediately after occlusion and was replaced by autologous, citrated platelet-rich plasma (PRP, 6 to 20 X 10(5) platelets/microliter) or filtered platelet-poor plasma (PPP, less than 100 platelets/microliter). Occluded arteries were studied after 1 to 28 days by light and electron microscopy. Events occurring within the first 2 days included fibrin clot formation, endothelial degeneration and denudation, transmural migration of polymorphonucelar leukocytes and monocytes, and, in PRP-filled arteries, degranulation and disappearance of platelets. By 7 days a neointima was formed by macrophages and undifferentiated cells. The latter cells had some features of vascular smooth muscle cells and were apparently derived from medial cells which traversed the internal elastic lamina. After 14 days, identifiable smooth muscle cells emerged as the predominant cell type in a rapidly growing intimal plaque. No differences could be discerned between arteries originally filled with PRP or PPP. This experimental model is similar to atherosclerosis in dimensions of avascular area and in coexistence of degenerative, inflammatory, and proliferative processes. Cell proliferation deep within an atherosclerotic plaque could be initiated by factors other than platelets, perhaps by products of inflammatory cells. Images Figure 4 Figure 7 Figure 6 Figure 1 Figure 2 Figure 3 Figure 8 Figure 5 PMID:426040

  17. IP3 receptors regulate vascular smooth muscle contractility and hypertension

    PubMed Central

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

    2016-01-01

    Inositol 1, 4, 5-trisphosphate receptor–mediated (IP3R-mediated) calcium (Ca2+) release has been proposed to play an important role in regulating vascular smooth muscle cell (VSMC) contraction for decades. However, whether and how IP3R regulates blood pressure in vivo remains unclear. To address these questions, we have generated a smooth muscle–specific IP3R triple-knockout (smTKO) mouse model using a tamoxifen-inducible system. In this study, the role of IP3R-mediated Ca2+ release in adult VSMCs on aortic vascular contractility and blood pressure was assessed following tamoxifen induction. We demonstrated that deletion of IP3Rs significantly reduced aortic contractile responses to vasoconstrictors, including phenylephrine, U46619, serotonin, and endothelin 1. Deletion of IP3Rs also dramatically reduced the phosphorylation of MLC20 and MYPT1 induced by U46619. Furthermore, although the basal blood pressure of smTKO mice remained similar to that of wild-type controls, the increase in systolic blood pressure upon chronic infusion of angiotensin II was significantly attenuated in smTKO mice. Taken together, our results demonstrate an important role for IP3R-mediated Ca2+ release in VSMCs in regulating vascular contractility and hypertension. PMID:27777977

  18. Oncostatin M Promotes Osteoblastic Differentiation of Human Vascular Smooth Muscle Cells Through JAK3-STAT3 Pathway.

    PubMed

    Kakutani, Yoshinori; Shioi, Atsushi; Shoji, Tetsuo; Okazaki, Hirokazu; Koyama, Hidenori; Emoto, Masanori; Inaba, Masaaki

    2015-07-01

    Vascular calcification is a clinically significant component of atherosclerosis and arises from chronic vascular inflammation. Oncostatin M (OSM) derived from plaque macrophages may contribute to the development of atherosclerotic calcification. Here, we investigated the stimulatory effects of OSM on osteoblastic differentiation of human vascular smooth muscle cells (HVSMC) derived from various arteries including umbilical artery, aorta, and coronary artery and its signaling pathway. Osteoblastic differentiation was induced by exposure of HVSMC to osteogenic differentiation medium (ODM) (10% fetal bovine serum, 0.1 μM dexamethasone, 10 mM β-glycerophosphate and 50 μg/ml ascorbic acid 2-phosphate in Dulbecco's modified Eagle's medium [DMEM]). OSM significantly increased alkaline phosphate (ALP) activity and matrix mineralization in HVSMC from all sources. Osteoblast marker genes such as ALP and Runx2 were also up-regulated by OSM in these cells. OSM treatment induced activation of STAT3 in HVSMC from umbilical artery as evidenced by immunoblot. Moreover, not only a JAK3 inhibitor, WHI-P154, but also knockdown of JAK3 by siRNA prevented the OSM-induced ALP activity and matrix mineralization in umbilical artery HVSMC. On the other hand, silencing of STAT3 almost completely suppressed OSM-induced ALP expression and matrix mineralization in HVSMC from all sources. These data suggest that OSM promotes osteoblastic differentiation of vascular smooth muscle cells through JAK3/STAT3 pathway and may contribute to the development of atherosclerotic calcification.

  19. Slug Is Increased in Vascular Remodeling and Induces a Smooth Muscle Cell Proliferative Phenotype

    PubMed Central

    Coll-Bonfill, Núria; Peinado, Victor I.; Pisano, María V.; Párrizas, Marcelina; Blanco, Isabel; Evers, Maurits; Engelmann, Julia C.; García-Lucio, Jessica; Tura-Ceide, Olga; Meister, Gunter

    2016-01-01

    Objective Previous studies have confirmed Slug as a key player in regulating phenotypic changes in several cell models, however, its role in smooth muscle cells (SMC) has never been assessed. The purpose of this study was to evaluate the expression of Slug during the phenotypic switch of SMC in vitro and throughout the development of vascular remodeling. Methods and Results Slug expression was decreased during both cell-to-cell contact and TGFβ1 induced SMC differentiation. Tumor necrosis factor-α (TNFα), a known inductor of a proliferative/dedifferentiated SMC phenotype, induces the expression of Slug in SMC. Slug knockdown blocked TNFα-induced SMC phenotypic change and significantly reduced both SMC proliferation and migration, while its overexpression blocked the TGFβ1-induced SMC differentiation and induced proliferation and migration. Genome-wide transcriptomic analysis showed that in SMC, Slug knockdown induced changes mainly in genes related to proliferation and migration, indicating that Slug controls these processes in SMC. Notably, Slug expression was significantly up-regulated in lungs of mice using a model of pulmonary hypertension-related vascular remodeling. Highly remodeled human pulmonary arteries also showed an increase of Slug expression compared to less remodeled arteries. Conclusions Slug emerges as a key transcription factor driving SMC towards a proliferative phenotype. The increased Slug expression observed in vivo in highly remodeled arteries of mice and human suggests a role of Slug in the pathogenesis of pulmonary vascular diseases. PMID:27441378

  20. Adult vascular smooth muscle cells in culture express neural stem cell markers typical of resident multipotent vascular stem cells.

    PubMed

    Kennedy, Eimear; Mooney, Ciaran J; Hakimjavadi, Roya; Fitzpatrick, Emma; Guha, Shaunta; Collins, Laura E; Loscher, Christine E; Morrow, David; Redmond, Eileen M; Cahill, Paul A

    2014-10-01

    Differentiation of resident multipotent vascular stem cells (MVSCs) or de-differentiation of vascular smooth muscle cells (vSMCs) might be responsible for the SMC phenotype that plays a major role in vascular diseases such as arteriosclerosis and restenosis. We examined vSMCs from three different species (rat, murine and bovine) to establish whether they exhibit neural stem cell characteristics typical of MVSCs. We determined their SMC differentiation, neural stem cell marker expression and multipotency following induction in vitro by using immunocytochemistry, confocal microscopy, fluorescence-activated cell sorting analysis and quantitative real-time polymerase chain reaction. MVSCs isolated from rat aortic explants, enzymatically dispersed rat SMCs and rat bone-marrow-derived mesenchymal stem cells served as controls. Murine carotid artery lysates and primary rat aortic vSMCs were both myosin-heavy-chain-positive but weakly expressed the neural crest stem cell marker, Sox10. Each vSMC line examined expressed SMC differentiation markers (smooth muscle α-actin, myosin heavy chain and calponin), neural crest stem cell markers (Sox10(+), Sox17(+)) and a glia marker (S100β(+)). Serum deprivation significantly increased calponin and myosin heavy chain expression and decreased stem cell marker expression, when compared with serum-rich conditions. vSMCs did not differentiate to adipocytes or osteoblasts following adipogenic or osteogenic inductive stimulation, respectively, or respond to transforming growth factor-β1 or Notch following γ-secretase inhibition. Thus, vascular SMCs in culture express neural stem cell markers typical of MVSCs, concomitant with SMC differentiation markers, but do not retain their multipotency. The ultimate origin of these cells might have important implications for their use in investigations of vascular proliferative disease in vitro.

  1. Interaction of Vascular Smooth Muscle Cells Under Low Shear Stress

    NASA Technical Reports Server (NTRS)

    Seidel, Charles L.

    1998-01-01

    The blood vessel wall consists of three cellular layers, an outer adventitial, a middle medial and an inner intimal layer. When the blood vessel forms in the embryo it begins as a tube composed of a single cell type called endothelial cells. Over time, other cells are recruited from the surrounding tissue to form additional layers on the outer surface of the endothelial tube. The cells that are recruited are called mesenchymal cells. Mesenchymal cells are responsible for the production of connective tissue that holds the blood vessel together and for developing into vascular smooth muscle cells that are responsible for regulating the diameter of the vessel (1) and therefore, blood flow. In a fully developed blood vessel, the endothelial cells make- up the majority of cells in the intimal layer while the mesenchymal cells make-up the majority of cells in the medial and adventitial layers. Within the medial layer of a mature vessel, cells are organized into multiple circular layers of alternating bands of connective tissue and cells. The cell layer is composed of a mixture of mesenchymal cells that have not developed into smooth muscle cells and fully developed smooth muscle cells (2). The assembly and organization of complex tissues is directed in part by a signaling system composed of proteins on the cell surface called adhesion molecules. Adhesion molecules enable cells to recognize each other as well as the composition of the connective tissue in which they reside (3). It was hypothesized that the different cell types that compose the vascular wall possess different adhesion molecules that enable them to recognize each other and through this recognition system, form the complex layered organization of the vascular wall. In other words, the layered organization is an intrinsic property of the cells. If this hypothesis is correct then the different cells that make up the vessel wall, when mixed together, should organize themselves into a layered structure

  2. Vascular smooth muscle cell culture in microfluidic devices

    PubMed Central

    Wei, Y. C.; Chen, F.; Zhang, T.; Chen, D. Y.; Jia, X.; Wang, J. B.; Guo, W.; Chen, J.

    2014-01-01

    This paper presents a microfluidic device enabling culture of vascular smooth muscle cells (VSMCs) where extracellular matrix coating, VSMC seeding, culture, and immunostaining are demonstrated in a tubing-free manner. By optimizing droplet volume differences between inlets and outlets of micro channels, VSMCs were evenly seeded into microfluidic devices. Furthermore, the effects of extracellular matrix (e.g., collagen, poly-l-Lysine (PLL), and fibronectin) on VSMC proliferation and phenotype expression were explored. As a platform technology, this microfluidic device may function as a new VSMC culture model enabling VSMC studies. PMID:25379109

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

    SciTech Connect

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

    2008-08-08

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

  4. Ablation of the androgen receptor from vascular smooth muscle cells demonstrates a role for testosterone in vascular calcification

    PubMed Central

    Zhu, Dongxing; Hadoke, Patrick W. F.; Wu, Junxi; Vesey, Alex T.; Lerman, Daniel. A.; Dweck, Marc R.; Newby, David E.; Smith, Lee B.; MacRae, Vicky E.

    2016-01-01

    Vascular calcification powerfully predicts mortality and morbidity from cardiovascular disease. Men have a greater risk of cardiovascular disease, compared to women of a similar age. These gender disparities suggest an influence of sex hormones. Testosterone is the primary and most well-recognised androgen in men. Therefore, we addressed the hypothesis that exogenous androgen treatment induces vascular calcification. Immunohistochemical analysis revealed expression of androgen receptor (AR) in the calcified media of human femoral artery tissue and calcified human valves. Furthermore, in vitro studies revealed increased phosphate (Pi)-induced mouse vascular smooth muscle cell (VSMC) calcification following either testosterone or dihydrotestosterone (DHT) treatment for 9 days. Testosterone and DHT treatment increased tissue non-specific alkaline phosphatase (Alpl) mRNA expression. Testosterone-induced calcification was blunted in VSMC-specific AR-ablated (SM-ARKO) VSMCs compared to WT. Consistent with these data, SM-ARKO VSMCs showed a reduction in Osterix mRNA expression. However, intriguingly, a counter-intuitive increase in Alpl was observed. These novel data demonstrate that androgens play a role in inducing vascular calcification through the AR. Androgen signalling may represent a novel potential therapeutic target for clinical intervention. PMID:27095121

  5. Insulin-like growth factor I stimulates elastin synthesis by bovine pulmonary arterial smooth muscle cells.

    PubMed

    Badesch, D B; Lee, P D; Parks, W C; Stenmark, K R

    1989-04-14

    Insulin-like growth factor I stimulates mitogenesis in smooth muscle cells, and upregulates elastin synthesis in embryonic aortic tissue. Increased smooth muscle elastin synthesis may play an important role in vascular remodeling in chronic pulmonary hypertension. Therefore, we studied the effect of IGF-I on elastin and total protein synthesis by pulmonary arterial smooth muscle cells in vitro. Tropoelastin synthesis was measured by enzyme immunoassay, and total protein synthesis was measured by [3H]-leucine incorporation. In addition, the steady-state levels of tropoelastin mRNA were determined by slot blot hybridization. Incubation of confluent cultures with various concentrations of IGF-I resulted in a dose-dependent stimulation of elastin synthesis, with a 2.4-fold increase over control levels at 1000 ng/ml of IGF. The increase in elastin synthesis was reflected by a stimulation of the steady-state levels of tropoelastin mRNA. We conclude that IGF-I has potent elastogenic effects on vascular smooth muscle cells, and speculate that it may contribute to vascular wall remodeling in chronic hypertension.

  6. Proliferation of pulmonary artery smooth muscle cells in the development of ascites syndrome in broilers induced by low ambient temperature.

    PubMed

    Wang, J; Qiao, J; Zhao, L H; Li, K; Wang, H; Xu, T; Tian, Y; Gao, M; Wang, X

    2007-12-01

    Pulmonary vascular remodelling, mainly characterized by arterial medial thickening, is an important pathological feature of broiler ascites syndrome (AS). Since vascular smooth muscle cells (VSMC) form the major cellular component of arterial medial layer, we speculate that VSMC proliferation is one of the causes of pulmonary arterial medial thickening in ascitic broilers. Hence, the present study was designed to investigate the role of VSMC proliferation in pulmonary vascular remodelling in development of AS induced by low ambient temperature. Broilers in control group (22 +/- 1.5 degrees C) and low temperature group (11 +/- 2 degrees C) were sampled every week at 15-50 days of age. Proliferative indexes of VSMC in pulmonary arteries were assessed with proliferating cell nuclear antigen, and the relative medial thickness (RMT) and relative wall area (RWA), as indexes of pulmonary vascular remodelling, were examined by computer-image analysing system. The results showed that the high incidence (18.75%) of AS was induced by low temperature, and a significantly increased VSMC proliferation was observed in pulmonary arteries in the low temperature group at 22-50 days of age (P < 0.05). In addition, RMT and RWA in pulmonary arteries were significantly elevated in the low temperature group from 36 days of age (P < 0.05), indicating that pulmonary vascular remodelling occurred following VSMC proliferation in AS. Our data suggest that proliferation of VSMC may facilitate pulmonary vascular remodelling and have a pivotal role in AS induced by low ambient temperature.

  7. Loss of smooth muscle cell hypoxia inducible factor-1α underlies increased vascular contractility in pulmonary hypertension.

    PubMed

    Barnes, Elizabeth A; Chen, Chih-Hsin; Sedan, Oshra; Cornfield, David N

    2017-02-01

    Pulmonary arterial hypertension (PAH) is an often fatal disease with limited treatment options. Whereas current data support the notion that, in pulmonary artery endothelial cells (PAECs), expression of transcription factor hypoxia inducible factor-1α (HIF-1α) is increased, the role of HIF-1α in pulmonary artery smooth muscle cells (PASMCs) remains controversial. This study investigates the hypothesis that, in PASMCs from patients with PAH, decreases in HIF-1α expression and activity underlie augmented pulmonary vascular contractility. PASMCs and tissues were isolated from nonhypertensive control patients and patients with PAH. Compared with controls, HIF-1α and Kv1.5 protein expression were decreased in PAH smooth muscle cells (primary culture). Myosin light chain (MLC) phosphorylation and MLC kinase (MLCK) activity-major determinants of vascular tone-were increased in patients with PAH. Cofactors involved in prolyl hydroxylase domain activity were increased in PAH smooth muscle cells. Functionally, PASMC contractility was inversely correlated with HIF-1α activity. In PASMCs derived from patients with PAH, HIF-1α expression is decreased, and MLCK activity, MLC phosphorylation, and cell contraction are increased. We conclude that compromised PASMC HIF-1α expression may contribute to the increased tone that characterizes pulmonary hypertension.-Barnes, E. A., Chen, C.-H., Sedan, O., Cornfield, D. N. Loss of smooth muscle cell hypoxia inducible factor-1α underlies increased vascular contractility in pulmonary hypertension.

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

    SciTech Connect

    Nawrath, H.; Raschack, M.

    1987-09-01

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

  9. Diffuse and uncontrolled vascular smooth muscle cell proliferation in rapidly progressing pediatric moyamoya disease.

    PubMed

    Reid, Amy J; Bhattacharjee, Meenakshi B; Regalado, Ellen S; Milewicz, Allen L; El-Hakam, Lisa M; Dauser, Robert C; Milewicz, Dianna M

    2010-09-01

    Moyamoya disease is a rare stroke syndrome of unknown etiology resulting from stenosis or occlusion of the supraclinoid internal carotid artery (ICA) in association with an abnormal vascular network in the basal ganglia. Although the highest incidence of moyamoya disease is in pediatric patients, pathology reports have been primarily limited to adult samples and describe occlusive fibrocellular lesions in the intimae of affected arteries. We describe the case of a young girl with primary moyamoya disease who presented at 18 months of age with right hemiparesis following an ischemic stroke. Angiography showed stenosis of the distal left ICA, left middle cerebral artery, and right ICA. An emergent left-sided dural inversion was performed. Recurrent strokes and alternating hemiplegia necessitated a right dural inversion 6 months later. Nonetheless, her aggressive disease proved uniquely refractory to surgical revascularization, and she succumbed to recurrent strokes and neurological deterioration at 2.5 years of age. Pathological specimens revealed a striking bilateral occlusion of the anterior carotid circulation resulting from intimal proliferation of smooth muscle cells (SMCs). Most strikingly, the ascending aorta and the superior mesenteric artery demonstrated similar intimal proliferation, along with SMC proliferation in the media. The systemic pathology involving multiple arteries in this extremely young child, the first case of its kind available for autopsy, suggests that globally uncontrolled SMC proliferation, in the absence of environmental risk factors and likely resulting from an underlying genetic alteration, may be a primary etiologic event leading to moyamoya disease.

  10. Arterial vascularization patterns of the splenium: An anatomical study.

    PubMed

    Kahilogullari, G; Comert, A; Ozdemir, M; Brohi, R A; Ozgural, O; Esmer, A F; Egemen, N; Karahan, S T

    2013-09-01

    The aim of this study was to provide detailed information about the arterial vascularization of the splenium of the corpus callosum (CC). The splenium is unique in that it is part of the largest commissural tract in the brain and a region in which pathologies are seen frequently. An exact description of the arterial vascularization of this part of the CC remains under debate. Thirty adult human brains (60 hemispheres) were obtained from routine autopsies. Cerebral arteries were separately cannulated and injected with colored latex. Then, the brains were fixed in formaldehyde, and dissections were performed using a surgical microscope. The diameter of the arterial branches supplying the splenium of the CC at their origin was investigated, and the vascularization patterns of these branches were observed. Vascular supply to the splenium was provided by the anterior pericallosal artery (40%) from the anterior circulation and by the posterior pericallosal artery (88%) and posterior accessory pericallosal artery (50%) from the posterior circulation. The vascularization pattern of the splenium differs in each hemisphere and is usually supplied by multiple branches. The arterial vascularization of the splenium of the CC was studied comprehensively considering the ongoing debate and the inadequacy of the studies on this issue currently available in the literature. This anatomical knowledge is essential during the treatment of pathologies in this region and especially for splenial arteriovenous malformations.

  11. Pericytes are progenitors for coronary artery smooth muscle

    PubMed Central

    Volz, Katharina S; Jacobs, Andrew H; Chen, Heidi I; Poduri, Aruna; McKay, Andrew S; Riordan, Daniel P; Kofler, Natalie; Kitajewski, Jan; Weissman, Irving; Red-Horse, Kristy

    2015-01-01

    Epicardial cells on the heart’s surface give rise to coronary artery smooth muscle cells (caSMCs) located deep in the myocardium. However, the differentiation steps between epicardial cells and caSMCs are unknown as are the final maturation signals at coronary arteries. Here, we use clonal analysis and lineage tracing to show that caSMCs derive from pericytes, mural cells associated with microvessels, and that these cells are present in adults. During development following the onset of blood flow, pericytes at arterial remodeling sites upregulate Notch3 while endothelial cells express Jagged-1. Deletion of Notch3 disrupts caSMC differentiation. Our data support a model wherein epicardial-derived pericytes populate the entire coronary microvasculature, but differentiate into caSMCs at arterial remodeling zones in response to Notch signaling. Our data are the first demonstration that pericytes are progenitors for smooth muscle, and their presence in adult hearts reveals a new potential cell type for targeting during cardiovascular disease. DOI: http://dx.doi.org/10.7554/eLife.10036.001 PMID:26479710

  12. Coronary endothelial function and vascular smooth muscle proliferation are programmed by early-gestation dexamethasone exposure in sheep

    PubMed Central

    Volk, Kenneth A.; Roghair, Robert D.; Jung, Felicia; Scholz, Thomas D.; Lamb, Fred S.

    2010-01-01

    Exposure of the early-gestation ovine fetus to exogenous glucocorticoids induces changes in postnatal cardiovascular physiology. We sought to characterize coronary artery vascular function in this model by elucidating the contribution of nitric oxide and reactive oxygen species to altered coronary vascular reactivity and examining the proliferative potential of coronary artery vascular smooth muscle cells. Dexamethasone (dex, 0.28 mg·kg−1·day−1 for 48 h) was administered to pregnant ewes at 27–28-day gestation (term 145 days). Coronary arteries were isolated from 1- to 2-wk-old dex-exposed offspring and aged-matched controls. Compared with controls, coronary arteries from dex-exposed lambs demonstrated enhanced vasoconstriction to endothelin-1 and ACh that was abolished by endothelial removal or preincubation with the nitric oxide synthase inhibitor l-NNA, membrane-permeable superoxide dismutase + catalase, or apamin + charybdotoxin, but not indomethacin. The rate of coronary vascular smooth muscle cell (VSMC) proliferation was also significantly greater in dex-exposed lambs. Protein levels of the proliferating cell nuclear antigen were increased and α-smooth muscle actin decreased in dex-exposed coronary VSMC, consistent with a proliferative state. Finally, expression of the NADPH oxidase Nox 4, but not Nox 1, mRNA was also decreased in coronary VSMC from dex-exposed lambs. These findings suggest an important interaction exists between early-gestation glucocorticoid exposure and reactive oxygen species that is associated with alterations in endothelial function and coronary VSMC proliferation. These changes in coronary physiology are consistent with those associated with the development of atherosclerosis and may provide an important link between an adverse intrauterine environment and increased risk for coronary artery disease. PMID:20335378

  13. Role of blood and vascular smooth muscle in the vasoactivity of nitrite

    PubMed Central

    Liu, Taiming; Schroeder, Hobe J.; Barcelo, Lisa; Bragg, Shannon L.; Terry, Michael H.; Wilson, Sean M.; Power, Gordon G.

    2014-01-01

    Recent evidence from humans and rats indicates that nitrite is a vasodilator under hypoxic conditions by reacting with metal-containing proteins to produce nitric oxide (NO). We tested the hypothesis that near-physiological concentrations of nitrite would produce vasodilation in a hypoxia- and concentration-dependent manner in the hind limb of sheep. Anesthetized sheep were instrumented to measure arterial blood pressure and femoral blood flows continuously in both hind limbs. Nitrite was infused into one femoral artery to raise the nitrite concentration in the femoral vein by 10 to 15-fold while the sheep breathed 50%, 14% or 12% oxygen in inspired air. In contrast to reports in humans and rats, the nitrite infusion had no measurable effect on mean femoral blood flows or vascular conductances, regardless of inspired O2 levels. In vitro experiments showed no significant difference in the release of NO from nitrite in sheep and human red blood cells. Further experiments demonstrated nitrite is converted to NO in rat artery homogenates faster than sheep arteries, and that this source of NO production is attenuated in the presence of a heme oxidizer. Finally, western blots indicate that concentrations of the heme-containing protein cytoglobin, but not myoglobin, are markedly lower in sheep arteries compared with rats. Overall, the results demonstrate that nitrite is not a physiological vasodilator in sheep. This is likely due to a lack of conversion of nitrite to NO within the vascular smooth muscle, perhaps due to deficient amounts of the heme-containing protein cytoglobin. PMID:25108012

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

    NASA Technical Reports Server (NTRS)

    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.

  15. Vascular smooth muscle cell response on thin films of collagen.

    PubMed

    Elliott, John T; Woodward, John T; Langenbach, Kurt J; Tona, Alex; Jones, Peter L; Plant, Anne L

    2005-10-01

    Vascular smooth muscle cells (vSMC) cultured on gels of fibrillar type I collagen or denatured collagen (gelatin) comprise a model system that has been widely used for studying the role of the extracellular matrix in vascular diseases such as hypertension, restenosis and athrosclerosis. Despite the wide use of this model system, there are several disadvantages to using collagen gels for cellular studies. These include poor optical characteristics for microscopy, difficulty in verifying that the properties of the preparations are identical from experiment to experiment, heterogeneity within the gels, and difficulty in handling the gels because they are fragile. Previously, we developed an alternative collagen matrix by forming thin films of native fibrillar collagen or denatured collagen on self-assembled monolayers of alkanethiols [Elliott, J.T., Tona, A., Woodward, J., Jones,P., Plant, A., 2003a. Thin films of collagen affect smooth muscle cell morphology. Langmuir 19, 1506-1514.]. These substrates are robust and can be characterized by surface analytical techniques that allow both verification of the reproducibility of the preparation and high-resolution analysis of collagen structure. In addition, they have excellent optical properties that allow more details of the cell-matrix interactions to be observed by microscopy. In this study, we performed a side-by-side structural and functional comparison of collagen gels with thin films of collagen. Our results indicate that vSMC on thin films of collagen are nearly identical to vSMC on thick gels as determined by morphology, proliferation rate, integrin ligation, tenascin-C expression and intracellular signaling events. These results suggest that the features of collagen gels that direct the observed vSMC responses are adequately reconstituted in the thin films of collagen. These thin films will be useful for elucidating the features of the collagen matrix that regulate vSMC response and may be applicable to high

  16. Carvacrol induces the apoptosis of pulmonary artery smooth muscle cells under hypoxia.

    PubMed

    Zhang, Qianlong; Fan, Kai; Wang, Peng; Yu, Juan; Liu, Ruxia; Qi, Hanping; Sun, Hongli; Cao, Yonggang

    2016-01-05

    The abnormal apoptosis of pulmonary artery smooth muscle cells (PASMCs) is an important pathophysiological process in pulmonary vascular remodeling and pulmonary arterial hypertension (PAH). Carvacrol, an essential oil compound from oregano and thyme, has displayed antimicrobial, antitumor, and antioxidant properties. Although carvacrol has pro-apoptosis properties in tumor cells, the underlying mechanisms of carvacrol in PASMC apoptosis remain unclear. Thus, in this study, we aim to investigate the role of carvacrol in pulmonary vascular remodeling and PASMC apoptosis in hypoxia. Right Ventricular Hypertrophy Measurements and pulmonary pathomorphology data show that the ratio of the heart weight/tibia length (HW/TL), the right ventricle/left ventricle plus septum (RV/LV+S) and the medial width of the pulmonary artery increased in chronic hypoxia and were reversed by carvacrol treatment under hypoxia. Additionally, carvacrol inhibited PASMC viability, attenuated oxidative stress, induced mitochondria membrane depolarization, increased the percentage of apoptotic cells, suppressed Bcl-2 expression, decreased procaspase-3 expression, promoted caspase-3 activation, and inhibited the ERK1/2 and PI3K/Akt pathway. Taken together, these findings suggest that carvacrol attenuates the pulmonary vascular remodeling and promotes PASMC apoptosis by acting on, at least in part, the intrinsic apoptotic pathway. This process might provide us new insight into the development of hypoxic pulmonary hypertension.

  17. Effect of sinomenine on vascular smooth muscle cell dedifferentiation and neointima formation after vascular injury in mice.

    PubMed

    Zhu, Lihua; Hao, Yarong; Guan, Hongjing; Cui, Changping; Tian, Song; Yang, Da; Wang, Xinan; Zhang, Shuming; Wang, Lang; Jiang, Hong

    2013-01-01

    Sinomenine, a pure alkaloid extract from Sinomenium acutum, has anti-inflammatory and immunoregulatory functions. This study investigated the efficiency and the signalling pathways involved in the effect of sinomenine on vascular smooth muscle cell (VSMC) dedifferentiation in response to platelet-derived growth factor (PDGF)-BB stimulation and vascular injury. VSMCs were isolated from rat aorta and preincubated with sinomenine before being stimulated with PDGF-BB. WST and BrdU incorporation assays were used to evaluate VSMC proliferation. Flow cytometric analysis was performed for testing the cell cycle progression. The cell migration of VSMCs were analysed using a Transwell system. The expression of VSMC specific genes and signalling proteins were tested by Western blot. For the animal study, C57/BL6 mice were fed either normal rodent chow diets or sinomenine chow diets that supplemented with 0.09 % sinomenine (w/w) in the normal chows for 14 days before carotid artery wire injury. PDGF-BB activated the dedifferentiation of VSMCs characterised by decreased expression of SMA, Smoothelin and SM22α. However, sinomenine treatment preserved the dedifferentiation in response to PDGF-BB. The activations of mitogen-activated protein kinase extracellular signal-regulated kinases, Akt, GSK3β and STAT3 induced by PDGF-BB were also inhibited in sinomenine-treated VSMCs. In vivo evidence with wire-injured mice exhibited a reduction in neointimal area and an increase in smooth muscle-specific gene expression in the sinomenine-treated group. In this study, we found that sinomenine-suppressed VSMC phenotype switching induced by PDGF-BB in vitro and neointimal formation in vivo. Therefore, sinomenine is a potential candidate to be used in the treatment of vascular proliferative disease.

  18. Monocyte-expressed urokinase regulates human vascular smooth muscle cell migration in a coculture model.

    PubMed

    Kusch, Angelika; Tkachuk, Sergey; Lutter, Steffen; Haller, Hermann; Dietz, Rainer; Lipp, Martin; Dumler, Inna

    2002-01-01

    Interactions of vascular smooth muscle cells (VSMC) with monocytes recruited to the arterial wall at a site of injury, with resultant modulation of VSMC growth and migration, are central to the development of vascular intimal thickening. Urokinase-type plasminogen activator (uPA) expressed by monocytes is a potent chemotactic factor for VSMC and might serve for the acceleration of vascular remodeling. In this report, we demonstrate that coculture of human VSMC with freshly isolated peripheral blood-derived human monocytes results in significant VSMC migration that increases during the coculture period. Accordingly, VSMC adhesion was inhibited with similar kinetics. VSMC proliferation, however, was not affected and remained at the same basal level during the whole period of coculture. The increase of VSMC migration in coculture was equivalent to the uPA-induced migration of monocultured VSMC and was blocked by addition into coculture of soluble uPAR (suPAR). Analysis of uPA and uPAR expression in cocultured cells demonstrated that monocytes are a major source of uPA, whose expression increases in coculture five-fold, whereas VSMC display an increased expression of cell surface-associated uPAR. These findings indicate that upregulated uPA production by monocytes following vascular injury acts most likely as an endogenous activator of VSMC migration contributing to the remodeling of vessel walls.

  19. Vascular smooth muscle G(q) signaling is involved in high blood pressure in both induced renal and genetic vascular smooth muscle-derived models of hypertension.

    PubMed

    Harris, David M; Cohn, Heather I; Pesant, Stéphanie; Zhou, Rui-Hai; Eckhart, Andrea D

    2007-11-01

    More than 30% of the US population has high blood pressure (BP), and less than a third of people treated for hypertension have it controlled. In addition, the etiology of most high BP is not known. Having a better understanding of the mechanisms underlying hypertension could potentially increase the effectiveness of treatment. Because G(q) signaling mediates vasoconstriction and vascular function can cause BP abnormalities, we were interested in determining the role of vascular smooth muscle (VSM) G(q) signaling in two divergent models of hypertension: a renovascular model of hypertension through renal artery stenosis and a genetic model of hypertension using mice with VSM-derived high BP. Inhibition of VSM G(q) signaling attenuated BP increases induced by renal artery stenosis to a similar extent as losartan, an ANG II receptor blocker and current antihypertensive therapy. Inhibition of G(q) signaling also attenuated high BP in our genetic VSM-derived hypertensive model. In contrast, BP remained elevated 25% following treatment with losartan, and prazosin, an alpha(1)-adrenergic receptor antagonist, only decreased BP by 35%. Inhibition of G(q) signaling attenuated VSM reactivity to ANG II and resulted in a 2.4-fold rightward shift in EC(50). We also determined that inhibition of G(q) signaling was able to reverse VSM hypertrophy in the genetic VSM-derived hypertensive model. These results suggest that G(q) signaling is an important signaling pathway in two divergent models of hypertension and, perhaps, optimization of antihypertensive therapy could occur with the identification of particular G(q)-coupled receptors involved.

  20. Calcifying nanoparticles promote mineralization in vascular smooth muscle cells: implications for atherosclerosis

    PubMed Central

    Hunter, Larry W; Charlesworth, Jon E; Yu, Sam; Lieske, John C; Miller, Virginia M

    2014-01-01

    Background Nano-sized complexes of calcium phosphate mineral and proteins (calcifying nanoparticles [CNPs]) serve as mineral chaperones. Thus, CNPs may be both a result and cause of soft tissue calcification processes. This study determined if CNPs could augment calcification of arterial vascular smooth muscle cells in vitro. Methods CNPs 210 nm in diameter were propagated in vitro from human serum. Porcine aortic smooth muscle cells were cultured for up to 28 days in medium in the absence (control) or presence of 2 mM phosphate ([P] positive calcification control) or after a single 3-day exposure to CNPs. Transmission electron-microscopy was used to characterize CNPs and to examine their cellular uptake. Calcium deposits were visualized by light microscopy and von Kossa staining and were quantified by colorimetry. Cell viability was quantified by confocal microscopy of live-/dead-stained cells and apoptosis was examined concurrently by fluorescent labeling of exposed phosphatidylserine. Results CNPs, as well as smaller calcium crystals, were observed by transmission electron-microscopy on day 3 in CNP-treated but not P-treated cells. By day 28, calcium deposits were visible in similar amounts within multicellular nodules of both CNP- and P-treated cells. Apoptosis increased with cell density under all treatments. CNP treatment augmented the density of apoptotic bodies and cellular debris in association with mineralized multicellular nodules. Conclusion Exogenous CNPs are taken up by aortic smooth muscle cells in vitro and potentiate accumulation of smooth-muscle-derived apoptotic bodies at sites of mineralization. Thus, CNPs may accelerate vascular calcification. PMID:24920905

  1. Vascular smooth muscle cell functional contractility depends on extracellular mechanical properties

    PubMed Central

    Steucke, Kerianne E.; Tracy, Paige V.; Hald, Eric S.; Hall, Jennifer L.; Alford, Patrick W.

    2015-01-01

    Vascular smooth muscle cells’ primary function is to maintain vascular homeostasis through active contraction and relaxation. In diseases such as hypertension and atherosclerosis, this function is inhibited concurrent to changes in the mechanical environment surrounding vascular smooth muscle cells. It is well established that cell function and extracellular mechanics are interconnected; variations in substrate modulus affect cell migration, proliferation, and differentiation. To date, it is unknown how the evolving extracellular mechanical environment of vascular smooth muscle cells affects their contractile function. Here, we have built upon previous vascular muscular thin film technology to develop a variable-modulus vascular muscular thin film that measures vascular tissue functional contractility on substrates with a range of pathological and physiological moduli. Using this modified vascular muscular thin film, we found that vascular smooth muscle cells generated greater stress on substrates with higher moduli compared to substrates with lower moduli. We then measured protein markers typically thought to indicate a contractile phenotype in vascular smooth muscle cells and found that phenotype is unaffected by substrate modulus. These data suggest that mechanical properties of vascular smooth muscle cells’ extracellular environment directly influence their functional behavior and do so without inducing phenotype switching. PMID:26283412

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

    NASA Astrophysics Data System (ADS)

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

    1986-08-01

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

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

    PubMed Central

    Koike, Sayo; Yano, Shozo; Tanaka, Sayuri; Sheikh, Abdullah M.; Nagai, Atsushi; Sugimoto, Toshitsugu

    2016-01-01

    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(P)H 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(P)H 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. PMID:27649164

  4. Vasoconstrictor effect of endothelin-1 on hypertensive pulmonary arterial smooth muscle involves Rho-kinase and protein kinase C.

    PubMed

    Barman, Scott A

    2007-08-01

    Although one of the common characteristics of pulmonary hypertension is abnormal sustained vasoconstriction, the signaling pathways that mediate this heightened pulmonary vascular response are still not well defined. Protein kinase C (PKC) and Rho-kinase are regulators of smooth muscle contraction induced by G protein-coupled receptor agonists including endothelin-1 (ET-1), which has been implicated as a signaling pathway in pulmonary hypertension. Toward this end, it was hypothesized that both Rho-kinase and PKC mediate the pulmonary vascular response to ET-1 in hypertensive pulmonary arterial smooth muscle, and therefore, the purpose of this study was to determine the role of PKC and Rho-kinase signaling in ET-1-induced vasoconstriction in both normotensive (Sprague-Dawley) and hypertensive (Fawn-Hooded) rat pulmonary arterial smooth muscle. Results indicate that ET-1 caused greater vasoconstriction in hypertensive pulmonary arteries compared with the normal vessels, and treatment with the PKC antagonists chelerythrine, rottlerin, and Gö 6983 inhibited the vasoconstrictor response to ET-1 in the hypertensive vessels. In addition, the specific Rho-kinase inhibitor Y-27632 significantly attenuated the effect of ET-1 in both normotensive and hypertensive phenotypes, with greater inhibition occurring in the hypertensive arteries. Furthermore, Western blot analysis revealed that ET-1 increased RhoA expression in both normotensive and hypertensive pulmonary arteries, with expression being greater in the hypertensive state. These results suggest that both PKC and Rho/Rho-kinase mediate the heightened pulmonary vascular response to ET-1 in hypertensive pulmonary arterial smooth muscle.

  5. IL-22 activates oxidant signaling in pulmonary vascular smooth muscle cells.

    PubMed

    Bansal, Geetanjali; Das, Dividutta; Hsieh, Cheng-Ying; Wang, Yi-Hsuan; Gilmore, Brent A; Wong, Chi-Ming; Suzuki, Yuichiro J

    2013-12-01

    Reactive oxygen species (ROS) mediate cell-signaling processes in response to various ligands and play important roles in the pathogenesis of cardiovascular diseases. The present study reports that interleukin-22 (IL-22) elicits signal transduction in vascular smooth muscle cells (SMCs) through a ROS-dependent mechanism. We find that pulmonary artery SMCs express IL-22 receptor alpha 1 and that IL-22 activates STAT3 through this receptor. IL-22-induced signaling is found to be mediated by NADPH oxidase, as indicated by the observations that the inhibition and siRNA knock-down of this enzyme inhibit IL-22 signaling. IL-22 triggers the oxidative modifications of proteins through protein carbonylation and protein glutathionylation. Mass spectrometry identified some proteins that are carbonylated in response to IL-22 stimulation, including α-enolase, heat shock cognate 71kDa protein, mitochondrial 60kDa heat shock protein, and cytoplasmic 2 actin and determined that α-tubulin is glutathionylated. Protein glutathionylation and STAT3 phosphorylation are enhanced by the siRNA knock-down of glutaredoxin, while IL-22-mediated STAT3 phosphorylation is suppressed by knocking down thioredoxin interacting protein, an inhibitor of thioredoxin. IL-22 is also found to promote the growth of SMCs via NADPH oxidase. In rats, pulmonary hypertension is found to be associated with increased smooth muscle IL-22 expression. These results show that IL-22 promotes the growth of pulmonary vascular SMCs via a signaling mechanism that involves NADPH oxidase-dependent oxidation.

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

    PubMed

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

    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(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.

  7. Mice lacking hypertension candidate gene ATP2B1 in vascular smooth muscle cells show significant blood pressure elevation.

    PubMed

    Kobayashi, Yusuke; Hirawa, Nobuhito; Tabara, Yasuharu; Muraoka, Hidenori; Fujita, Megumi; Miyazaki, Nobuko; Fujiwara, Akira; Ichikawa, Yasuhiro; Yamamoto, Yuichiro; Ichihara, Naoaki; Saka, Sanae; Wakui, Hiromichi; Yoshida, Shin-ichiro; Yatsu, Keisuke; Toya, Yoshiyuki; Yasuda, Gen; Kohara, Katsuhiko; Kita, Yoshikuni; Takei, Kohtaro; Goshima, Yoshio; Ishikawa, Yoshihiro; Ueshima, Hirotsugu; Miki, Tetsuro; Umemura, Satoshi

    2012-04-01

    We reported previously that ATP2B1 was one of the genes for hypertension receptivity in a large-scale Japanese population, which has been replicated recently in Europeans and Koreans. ATP2B1 encodes the plasma membrane calcium ATPase isoform 1, which plays a critical role in intracellular calcium homeostasis. In addition, it is suggested that ATP2B1 plays a major role in vascular smooth muscle contraction. Because the ATP2B1 knockout (KO) mouse is embryo-lethal, we generated mice with vascular smooth muscle cell-specific KO of ATP2B1 using the Cre-loxP system to clarify the relationship between ATP2B1 and hypertension. The KO mice expressed significantly lower levels of ATP2B1 mRNA and protein in the aorta compared with control mice. KO mice showed significantly higher systolic blood pressure as measured by tail-cuff method and radiotelemetric method. Similar to ATP2B1, the expression of the Na(+)-Ca(2+) exchanger isoform 1 mRNA was decreased in vascular smooth muscle cells of KO mice. However, ATP2B4 expression was increased in KO mice. The cultured vascular smooth muscle cells of KO mice showed increased intracellular calcium concentration not only in basal condition but also in phenylephrine-stimulated condition. Furthermore, phenylephrine-induced vasoconstriction was significantly increased in vascular rings of the femoral artery of KO mice. These results suggest that ATP2B1 plays important roles in the regulation of blood pressure through alteration of calcium handling and vasoconstriction in vascular smooth muscle cells.

  8. Influence of constriction, wall tension, smooth muscle activation and cellular deformation on rat resistance artery vasodilator reactivity.

    PubMed

    Colton, Ilsley; Mandalà, Maurizio; Morton, Jude; Davidge, Sandra T; Osol, George

    2012-01-01

    This study investigated how vasoconstriction (tone), wall tension, smooth muscle activation, and vascular wall deformation influence resistance artery vasodilator reactivity. Resistance arteries, from two different regional circulations (splanchnic, uterine) and from pregnant and non-pregnant rats, were cannulated and pressurized, or mounted on a wire myograph under isometric conditions prior to being exposed to both endothelium-dependent (acetylcholine, ACh) and -independent (sodium nitroprusside, SNP) vasodilator agonists. A consistent pattern of reduced vasodilator sensitivity was noted as a function of extent of preconstriction for both agonists noted in pressurized arteries. A similar pattern regarding activation was noted in wire-mounted arteries in response to SNP but not ACh. Wall tension proved to be a major determinant of vascular smooth muscle vasodilator reactivity and its normalization reversed this pattern, as more constricted vessels were more sensitive to ACh relaxation without any change in SNP sensitivity, suggesting that endothelial deformation secondary to vasoconstriction augments its vasodilator output. To our knowledge, this is the first study to dissect out the complex interplay between biophysical forces impinging on VSM (pressure, wall tension), the ambient level of tone (vasoconstriction, smooth muscle cell activation), and consequences of cellular (particularly endothelial) deformation secondary to constriction in determining resistance artery vasodilatory reactivity.

  9. Interaction between human monocytes and vascular smooth muscle cells induces vascular endothelial growth factor expression.

    PubMed

    Hojo, Y; Ikeda, U; Maeda, Y; Takahashi, M; Takizawa, T; Okada, M; Funayama, H; Shimada, K

    2000-05-01

    The objective of this study was to investigate whether synthesis of vascular endothelial growth factor (VEGF), a major mitogen for vascular endothelial cells, was induced by a cell-to-cell interaction between monocytes and vascular smooth muscle cells (VSMCs). Human VSMCs and THP-1 cells (human monocytoid cell) were cocultured. VEGF levels in the coculture medium were determined by enzyme-linked immunosorbent assay. Northern blot analysis of VEGF mRNA was performed using a specific cDNA probe. Immunohistochemistry was performed to determine which types of cell produce VEGF. Adding THP-1 cells to VSMCs for 24 h increased VEGF levels of the culture media, 8- and 10-fold relative to those of THP-1 cells and VSMCs alone, respectively. Northern blot analysis showed that VEGF mRNA expression was induced in the cocultured cells and peaked after 12 h. Immunohistochemistry disclosed that both types of cell in the coculture produced VEGF. Separate coculture experiments revealed that both direct contact and a soluble factor(s) contributed to VEGF production. Neutralizing anti-interleukin (IL)-6 antibody inhibited VEGF production by the coculture of THP-1 cells and VSMCs. A cell-to-cell interaction between monocytes and VSMCs induced VEGF synthesis in both types of cell. An IL-6 mediated mechanism is at least partially involved in VEGF production by the cocultures. Local VEGF production induced by a monocyte-VSMC interaction may play an important role in atherosclerosis and vascular remodeling.

  10. CADASIL mutations and shRNA silencing of NOTCH3 affect actin organization in cultured vascular smooth muscle cells.

    PubMed

    Tikka, Saara; Ng, Yan Peng; Di Maio, Giuseppe; Mykkänen, Kati; Siitonen, Maija; Lepikhova, Tatiana; Pöyhönen, Minna; Viitanen, Matti; Virtanen, Ismo; Kalimo, Hannu; Baumann, Marc

    2012-12-01

    Cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL) is the most common hereditary vascular dementia caused by mutations in NOTCH3 gene. Pathology is manifested in small- and middle-sized arteries throughout the body, though primarily in cerebral white matter. Hemodynamics is altered in CADASIL and NOTCH3 is suggested to regulate actin filament polymerization and thereby vascular tone. We analyzed NOTCH3 expression and morphology of actin cytoskeleton in genetically genuine cultured human CADASIL vascular smooth muscle cells (VSMCs) (including a cell line homozygous for p.Arg133Cys mutation) derived from different organs, and in control VSMCs with short hairpin RNA (shRNA)-silenced NOTCH3. NOTCH3 protein level was higher in VSMCs derived from adult than newborn arteries in both CADASIL and control VSMCs. CADASIL VSMCs showed altered actin cytoskeleton including increased branching and node formation, and more numerous and smaller adhesion sites than control VSMCs. Alterations in actin cytoskeleton in shRNA-silenced VSMCs were similar as in CADASIL VSMCs. Severity of the alterations in actin filaments corresponded to NOTCH3 expression level being most severe in VSMCs derived from adult cerebral arteries. These observations suggest that hypomorphic NOTCH3 activity causes alterations in actin organization in CADASIL. Furthermore, arteries from different organs have specific characteristics, which modify the effects of the NOTCH3 mutation and which is one explanation for the exceptional susceptibility of cerebral white matter arteries.

  11. CCN1 suppresses pulmonary vascular smooth muscle contraction in response to hypoxia.

    PubMed

    Lee, Seon-Jin; Zhang, Meng; Hu, Kebin; Lin, Ling; Zhang, Duo; Jin, Yang

    2015-12-01

    Pulmonary vasoconstriction and increased vascular resistance are common features in pulmonary hypertension (PH). One of the contributing factors in the development of pulmonary vasoconstriction is increased pulmonary artery smooth muscle cell (PASMC) contraction. Here we report that CCN1, an extracellular matrix molecule, suppressed PASMC contraction in response to hypoxia. CCN1 (Cyr61), discovered in past decade, belongs to the Cyr61-CTGF-Nov (CCN) family. It carries a variety of cellular functions, including angiogenesis and cell adhesion, death, and proliferation. Hypoxia robustly upregulated the expression of CCN1 in the pulmonary vessels and lung parenchyma. Given that CCN1 is a secreted protein and functions in a paracine manner, we examined the potential effects of CCN1 on the adjacent smooth muscle cells. Interestingly, bioactive recombinant CCN1 significantly suppressed hypoxia-induced contraction in human PASMCs in vitro. Consistently, in the in vivo functional studies, administration of bioactive CCN1 protein significantly decreased right ventricular pressure in three different PH animal models. Mechanistically, protein kinase A-pathway inhibitors abolished the effects of CCN1 in suppressing PASMC contraction. Furthermore, CCN1-inhibited smooth muscle contraction was independent of the known vasodilators, such as nitric oxide. Taken together, our studies indicated a novel cellular function of CCN1, potentially regulating the pathogenesis of PH.

  12. Protein kinase C–independent inhibition of arterial smooth muscle K+ channels by a diacylglycerol analogue

    PubMed Central

    Rainbow, RD; Parker, AM; Davies, NW

    2011-01-01

    BACKGROUND AND PURPOSE Analogues of the endogenous diacylglycerols have been used extensively as pharmacological activators of protein kinase C (PKC). Several reports show that some of these compounds have additional effects that are independent of PKC activation, including direct block of K+ and Ca2+ channels. We investigated whether dioctanoyl-sn-glycerol (DiC8), a commonly used diacylglycerol analogue, blocks K+ currents of rat mesenteric arterial smooth muscle in a PKC-independent manner. EXPERIMENTAL APPROACH Conventional whole-cell and inside-out patch clamp was used to measure the inhibition of K+ currents of rat isolated mesenteric smooth muscle cells by DiC8 in the absence and presence of PKC inhibitor peptide. KEY RESULTS Mesenteric artery smooth muscle Kv currents inactivated very slowly with a time constant of about 2 s following pulses from −65 to +40 mV. Application of 1 µM DiC8 produced an approximate 40-fold increase in the apparent rate of inactivation. Pretreatment of the cells with PKC inhibitor peptide had a minimal effect on the action of DiC8, and substantial inactivation still occurred, indicating that this effect was mainly independent of PKC. We also found that DiC8 blocked BK and KATP currents, and again a significant proportion of these blocks occurred independently of PKC activation. CONCLUSIONS AND IMPLICATIONS These results show that DiC8 has a direct effect on arterial smooth muscle K+ channels, and this precludes its use as a PKC activator when investigating PKC-mediated effects on vascular K+ channels. PMID:21323899

  13. Dihydrotestosterone alters cyclooxygenase-2 levels in human coronary artery smooth muscle cells

    PubMed Central

    Osterlund, Kristen L.; Handa, Robert J.

    2010-01-01

    Both protective and nonprotective effects of androgens on the cardiovascular system have been reported. Our previous studies show that the potent androgen receptor (AR) agonist dihydrotestosterone (DHT) increases levels of the vascular inflammatory mediator cyclooxygenase (COX)-2 in rodent cerebral arteries independent of an inflammatory stimulus. Little is known about the effects of androgens on inflammation in human vascular tissues. Therefore, we tested the hypothesis that DHT alters COX-2 levels in the absence and presence of induced inflammation in primary human coronary artery smooth muscle cells (HCASMC). Furthermore, we tested the ancillary hypothesis that DHT's effects on COX-2 levels are AR-dependent. Cells were treated with DHT (10 nM) or vehicle for 6 h in the presence or absence of LPS or IL-1β. Similar to previous observations in rodent arteries, in HCASMC, DHT alone increased COX-2 levels compared with vehicle. This effect of DHT was attenuated in the presence of the AR antagonist bicalutamide. Conversely, in the presence of LPS or IL-1β, increases in COX-2 were attenuated by cotreatment with DHT. Bicalutamide did not affect this response, suggesting that DHT-induced decreases in COX-2 levels occur independent of AR stimulation. Thus we conclude that DHT differentially influences COX-2 levels under physiological and pathophysiological conditions in HCASMC. This effect of DHT on COX-2 involves AR-dependent and- independent mechanisms, depending on the physiological state of the cell. PMID:20103743

  14. Extracellular calcium sensing in rat aortic vascular smooth muscle cells

    SciTech Connect

    Smajilovic, Sanela; Hansen, Jakob Lerche; Christoffersen, Tue E.H.

    2006-10-06

    Extracellular calcium (Ca2+o) can act as a first messenger in many cell types through a G protein-coupled receptor, calcium-sensing receptor (CaR). It is still debated whether the CaR is expressed in vascular smooth muscle cells (VSMCs). Here, we report the expression of CaR mRNA and protein in rat aortic VSMCs and show that Ca2+o stimulates proliferation of the cells. The effects of Ca2+o were attenuated by pre-treatment with MAPK kinase 1 (MEK1) inhibitor, as well as an allosteric modulator, NPS 2390. Furthermore, stimulation of the VSMCs with Ca2+o-induced phosphorylation of ERK1/2, but surprisingly did not cause inositol phosphate accumulation. We were not able to conclusively state that the CaR mediates Ca2+o-induced cell proliferation. Rather, an additional calcium-sensing mechanism may exist. Our findings may be of importance with regard to atherosclerosis, an inflammatory disease characterized by abnormal proliferation of VSMCs and high local levels of calcium.

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

    PubMed

    Uryga, Anna K; Bennett, Martin R

    2016-04-15

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

  16. Heterogeneity in vascular smooth muscle cell embryonic origin in relation to adult structure, physiology, and disease

    PubMed Central

    Pfaltzgraff, Elise R.; Bader, David M.

    2015-01-01

    Regional differences in vascular physiology and disease response exist throughout the vascular tree. While these differences in physiology and disease correspond to regional vascular environmental conditions, there is also compelling evidence that the embryonic origins of the smooth muscle inherent to the vessels may play a role. Here we review what is known regarding the role of embryonic origin of vascular smooth muscle cells during vascular development. The focus of this review is to highlight the heterogeneity in the origins of vascular smooth muscle cells and the resulting regional physiologies of the vessels. Our goal is to stimulate future investigation into this area and provide a better understanding of vascular organogenesis and disease. PMID:25546231

  17. Carvedilol inhibits proliferation of cultured pulmonary artery smooth muscle cells of patients with idiopathic pulmonary arterial hypertension.

    PubMed

    Fujio, Hideki; Nakamura, Kazufumi; Matsubara, Hiromi; Kusano, Kengo Fukushima; Miyaji, Katsumasa; Nagase, Satoshi; Ikeda, Tetsuya; Ogawa, Aiko; Ohta-Ogo, Keiko; Miura, Daiji; Miura, Aya; Miyazaki, Masahiro; Date, Hiroshi; Ohe, Tohru

    2006-02-01

    Idiopathic pulmonary arterial hypertension (IPAH) is associated with proliferation of smooth muscle cells (SMCs) in small pulmonary arteries. Inhibition of proliferation of pulmonary artery smooth muscle cells (PASMCs) may be an effective treatment of patients with idiopathic pulmonary arterial hypertension. Recent studies have shown that carvedilol, an alpha- and beta-blocker with antioxidant and calcium channel blocking properties, inhibits the proliferation of cultured normal human pulmonary artery smooth muscle cells. In this study, we tested the hypothesis that carvedilol has antiproliferative effects on pulmonary artery smooth muscle cells of patients with idiopathic pulmonary arterial hypertension. Pulmonary artery smooth muscle cells from six idiopathic pulmonary arterial hypertension patients who had undergone lung transplantation were cultured. To determine cell proliferation, H-thymidine incorporation was measured. Platelet-derived growth factor-induced proliferation of IPAH-PASMCs was significantly greater than that of normal control pulmonary artery smooth muscle cells. Carvedilol (0.1 microM to 10 microM) inhibited the proliferation of idiopathic pulmonary arterial hypertension-pulmonary artery smooth muscle cells in a concentration-dependent manner. Prazosin (an alpha-blocker) and N-acetyl L cysteine (an antioxidant agent) (0.1 microM to 10 microM) did not inhibit their proliferation, but the high concentration of propranolol (a beta-blocker) and nifedipine (a calcium channel blocker) (10 microM) inhibited the proliferation. The combination of propranolol and nifedipine inhibited the proliferation but only at a high concentration (10 microM) combination. Cell cycle analysis revealed that carvedilol (10 microM) significantly decreased the number of cells in S and G2/M phases. These results indicate that carvedilol inhibits the exaggerated proliferation of pulmonary artery smooth muscle cells of patients with idiopathic pulmonary arterial hypertension

  18. Factors influencing acute thrombus formation on carotid artery vascular grafts

    SciTech Connect

    Torem, S.; Schneider, P.A.; Paxton, L.D.; Yasuda, H.; Hanson, S.R.

    1988-10-01

    Scintillation camera imaging of 111Indium-labeled platelets has been used to measure acute thrombus formation on modified expanded Teflon (ePTFE) vascular grafts placed in the carotid arteries of normal baboons. Platelet deposition plateaued over 2 hr postoperatively and occurred primarily at the graft-vessel anastomoses. A positive correlation was found between the circulating platelet count in individual animals and the extent of early platelet thrombus deposition. Unmodified ePTFE grafts accumulated 4.6 +/- 1.2 x 10(9) platelets per graft, or 2.3 +/- 0.71 x 10(9) platelets per anastomosis. Acutely, platelet accumulation was reduced versus control graft results by coating the graft lumenal surfaces with a smooth layer of silicone rubber polymer (0.60 +/- 0.19 x 10(9) platelets per anastomosis; P less than 0.02) but not by coating the grafts using a plasma polymer based on methane, which did not modify graft texture (8.2 +/- 1.7 x 10(9) platelets per graft; P greater than 0.10). The benefit of the silicone rubber coating persisted for at least 48 hr. However, longer term patency was not preserved because 10 of 12 grafts placed had failed within 1 to 2 months.

  19. Vascular C-reactive protein in the pathogenesis of coronary artery disease: role of vascular inflammation and oxidative stress.

    PubMed

    Inoue, Nobutaka

    2006-12-01

    Atherosclerosis is considered to be a chronic inflammatory disease. Vascular inflammation occurs in response to injury induced by various stimuli, such as oxidative stress, shear stress, infection, and so on. This concept is supported by the recent clinical findings that C-reactive protein (CRP) is an independent risk factor for coronary heart disease. CRP, which was originally identified as a protein that could precipitate the C-polysaccharide of pneumococcal cell walls, has been widely used as a clinical marker of the state of inflammation, since its production by hepatocytes increases during the acute phase of the inflammatory response. Recent investigations have provided two new concepts for the research field of CRP, namely, its extra-hepatic production and its potent biological activities such as the induction of adhesion molecules and chemokines. Recently, we demonstrated that smooth muscle cells and macrophages in coronary arteries expressed CRP protein and mRNA, as evaluated using coronary specimens of coronary artery disease (CAD) patients obtained by atherectomy. The expression of vascular CRP was closely associated with NAD(P)H oxidase, an important enzymatic origin of reactive oxygen species (ROS) in vessel walls. Furthermore, CRP directly up-regulated NAD(P)H oxidase p22(phox) and enhanced ROS generation in cultured coronary artery smooth muscle cells. Thus, vascular CRP is likely to be a direct participant in vascular inflammation and lesion formation via its potent biological effects. Since lysophosphatidylcholine, a major atherogenic lipid of oxidized LDL, was reported to activate vascular NAD(P)H oxidase, we speculate that there is a vicious circle consisting of vascular NAD(P)H oxidase, ROS and oxidized LDL. Since phagocytic NAD(P)H oxidase is at the first line of the host defense system, it is important to selectively suppress vascular NAD(P)H oxidase in the localized inflammatory lesions in therapeutic strategies for CAD. In this review, we

  20. Myosin light chain kinase controls voltage-dependent calcium channels in vascular smooth muscle.

    PubMed

    Martinsen, A; Schakman, O; Yerna, X; Dessy, C; Morel, N

    2014-07-01

    The Ca(2+)-dependent kinase myosin light chain kinase (MLCK) is the activator of smooth muscle contraction. In addition, it has been reported to be involved in Ca(2+) channel regulation in cultured cells, and we previously showed that the MLCK inhibitor ML-7 decreases arginine vasopressin (AVP)-induced Ca(2+) influx in rat aorta. This study was designed to investigate whether MLCK is involved in Ca(2+) regulation in resistance artery smooth muscle cell, which plays a major role in the control of blood pressure. As ML compounds were shown to have off-target effects, MLCK was downregulated by transfection with a small interfering RNA targeting MLCK (MLCK-siRNA) in rat small resistance mesenteric artery (RMA) and in the rat embryonic aortic cell line A7r5. Noradrenaline-induced contraction and Ca(2+) signal were significantly depressed in MLCK-siRNA compared to scramble-siRNA-transfected RMA. Contraction and Ca(2+) signal induced by high KCl and voltage-activated Ca(2+) current were also significantly decreased in MLCK-siRNA-transfected RMA, suggesting that MLCK depletion modifies voltage-operated Ca(2+) channels. KCl- and AVP-induced Ca(2+) signals and voltage-activated Ca(2+) current were decreased in MLCK-depleted A7r5 cells. Eventually, real-time quantitative PCR analysis indicated that in A7r5, MLCK controlled mRNA expression of CaV1.2 (L-type) and CaV3.1 (T-type) voltage-dependent Ca(2+) channels. Our results suggest that MLCK controls the transcription of voltage-dependent Ca(2+) channels in vascular smooth muscle cells.

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

    SciTech Connect

    Viitanen, Matti; Sundström, Erik; Baumann, Marc; Tikka, Saara

    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.

  2. Inducible expression of vascular cell adhesion molecule-1 by vascular smooth muscle cells in vitro and within rabbit atheroma.

    PubMed Central

    Li, H.; Cybulsky, M. I.; Gimbrone, M. A.; Libby, P.

    1993-01-01

    Vascular cell adhesion molecule-1 (VCAM-1), a mononuclear leukocyte adhesion molecule, is expressed in cultured vascular endothelial cells activated by cytokines and is induced in rabbit aortic endothelium in vivo within 1 week after initiation of an atherogenic diet. We now demonstrate that vascular smooth muscle cells can also express VCAM-1 in rabbit atherosclerotic lesions in vivo and in response to cytokines in vitro. Immunohistochemical staining of aortas from rabbits fed a 0.3% cholesterol-containing diet revealed that a portion of smooth muscle cells within intimal foam cell-rich lesions expressed VCAM-1. The intimal VCAM-1-expressing cells localized predominantly in regions above the internal elastic lamina. These VCAM-1-positive cells had the typical spindle shape of smooth muscle cells but had reduced alpha-actin expression in comparison to normal medial smooth muscle cells, and did not bear markers for endothelium, macrophages, and T cells. In culture, rabbit aortic smooth muscle cells expressed VCAM-1 mRNA and protein in a time- and concentration-dependent fashion when exposed to interferon-gamma or Gram-negative bacterial lipopolysaccharide. Cultured human vascular smooth muscle cells also expressed VCAM-1 mRNA and protein in response to lipopolysaccharide, interferon-gamma, and interleukin-4. The monokines interleukin-1 alpha and tumor necrosis factor-alpha did not induce VCAM-1 expression in either rabbit or human vascular smooth muscle cells. Inducible VCAM-1 expression by vascular smooth muscle cells in vivo during hypercholesterolemia and in vitro in response to certain cytokines suggests a broader range of VCAM-1 functions in vascular biology than heretofore appreciated. Images Figure 1 Figure 2 Figure 3 Figure 4 PMID:7504883

  3. The properties and distribution of inward rectifier potassium currents in pig coronary arterial smooth muscle.

    PubMed Central

    Quayle, J M; Dart, C; Standen, N B

    1996-01-01

    1. Whole-cell potassium currents were studied in single smooth muscle cells enzymatically isolated from pig coronary arteries. 2. In cells isolated from small diameter branches of the left anterior descending coronary artery (LAD), an inward rectifier potassium current (IK(IR)) was identified, which was inhibited by extracellular barium ions, suggesting the presence of inward rectifier potassium (KIR) channels. 3. The conductance for IK(IR) measured in 6, 12, 60 and 140 mM extracellular potassium was a function of membrane potential and the extracellular potassium concentration. 4. On hyperpolarization, IK(IR) activated along an exponential time course with a time constant that was voltage dependent. 5. Inward rectifier current was compared in cells isolated from coronary vessels taken from different points along the vascular tree. Current density was greater in cells isolated from small diameter coronary arteries; at -140 mV it was -20.5 +/- 4.4 pA pF-1 (n = 23) in 4th order branches of the LAD, but -0.8 +/- 0.2 pA pF-1 (n = 11) in the LAD itself. 6. In contrast to IK(IR), there was little effect of arterial diameter on the density of voltage-dependent potassium current; densities at +30 mV were 12.8 +/- 1.3 pA pF-1 (n = 19) in 4th order branches and 17.4 +/- 3.1 pA pF-1 (n = 11) in the LAD. 7. We conclude that KIR channels are present in pig coronary arteries, and that they are expressed at a higher density in small diameter arteries. The presence of an enhanced IK(IR) may have functional consequences for the regulation of cell membrane potential and tone in small coronary arteries. PMID:8865069

  4. Role of KCNQ channels in skeletal muscle arteries and periadventitial vascular dysfunction.

    PubMed

    Zavaritskaya, Olga; Zhuravleva, Nadezda; Schleifenbaum, Johanna; Gloe, Torsten; Devermann, Lena; Kluge, Reinhart; Mladenov, Mitko; Frey, Manfred; Gagov, Hristo; Fésüs, Gabor; Gollasch, Maik; Schubert, Rudolf

    2013-01-01

    KCNQ channels have been identified in arterial smooth muscle. However, their role in vasoregulation and chronic vascular diseases remains elusive. We tested the hypothesis that KCNQ channels contribute to periadventitial vasoregulation in peripheral skeletal muscle arteries by perivascular adipose tissue and that they represent novel targets to rescue periadventitial vascular dysfunction. Two models, spontaneously hypertensive rats and New Zealand obese mice, were studied using quantitative polymerase chain reaction, the patch-clamp technique, membrane potential measurements, myography of isolated vessels, and blood pressure telemetry. In rat Gracilis muscle arteries, anticontractile effects of perivascular fat were inhibited by the KCNQ channel blockers XE991 and linopirdine but not by other selective K(+) channel inhibitors. Accordingly, XE991 and linopirdine blocked noninactivating K(+) currents in freshly isolated Gracilis artery smooth muscle cells. mRNAs of several KCNQ channel subtypes were detected in those arteries, with KCNQ4 channels being dominant. In spontaneously hypertensive rats, the anticontractile effect of perivascular fat in Gracilis muscle arteries was largely reduced compared with Wistar rats. However, the vasodilator effects of KCNQ channel openers and mRNA expression of KCNQ channels were normal. Furthermore, KCNQ channel openers restored the diminished anticontractile effects of perivascular fat in spontaneously hypertensive rats. Moreover, KCNQ channel openers reduced arterial blood pressure in both models of hypertension independent of ganglionic blockade. Thus, our data suggest that KCNQ channels play a pivotal role in periadventitial vasoregulation of peripheral skeletal muscle arteries, and KCNQ channel opening may be an effective mechanism to improve impaired periadventitial vasoregulation and associated hypertension.

  5. Quantification of Adventitial Vasa Vasorum Vascularization in Double-injury Restenotic Arteries

    PubMed Central

    Ye, Meng; Zhang, Bai-Gen; Zhang, Lan; Xie, Hui; Zhang, Hao

    2015-01-01

    Background: Accumulating evidence indicates a potential role of adventitial vasa vasorum (VV) dysfunction in the pathophysiology of restenosis. However, characterization of VV vascularization in restenotic arteries with primary lesions is still missing. In this study, we quantitatively evaluated the response of adventitial VV to vascular injury resulting from balloon angioplasty in diseased arteries. Methods: Primary atherosclerotic-like lesions were induced by the placement of an absorbable thread surrounding the carotid artery of New Zealand rabbits. Four weeks following double-injury induced that was induced by secondary balloon dilation, three-dimensional patterns of adventitial VV were reconstructed; the number, density, and endothelial surface of VV were quantified using micro-computed tomography. Histology and immunohistochemistry were performed in order to examine the development of intimal hyperplasia. Results: Results from our study suggest that double injured arteries have a greater number of VV, increased luminal surface, and an elevation in the intima/media ratio (I/M), along with an accumulation of macrophages and smooth muscle cells in the intima, as compared to sham or single injury arteries. I/M and the number of VV were positively correlated (R2 = 0.82, P < 0.001). Conclusions: Extensive adventitial VV neovascularization occurs in injured arteries after balloon angioplasty, which is associated with intimal hyperplasia. Quantitative assessment of adventitial VV response may provide insight into the basic biological process of postangioplasty restenosis. PMID:26228224

  6. Reconstruction of small diameter arteries using decellularized vascular scaffolds.

    PubMed

    Nagaoka, Yuki; Yamada, Hiroshi; Kimura, Tsuyoshi; Kishida, Akio; Fujisato, Toshia; Takakuda, Kazuo

    2014-03-19

    Although artificial vessels are available for large diameter arteries, there are no artificial vessels for small diameter arteries of < 4 mm. We created a decellularized vascular scaffold (length, 10 mm; outer diameter, 1.5 mm; inner diameter, 1.3 mm) from rat abdominal arteries. We measured the biomechanical characteristics of the scaffolds, implanted them to defects made in rat carotid arteries, and evaluated their patency and the endothelial cell linings. Silastic grafts were implanted as controls. The decellularized scaffolds demonstrated similar mechanical characteristics to normal arteries. All of the control grafts were occluded. Fibroblast-like cells were discovered in the thrombus, and fibrous organization was apparent. In contrast, patency of the grafts in 10 of 12 animals was observed 4 weeks after implantation. The internal cavity of the patent scaffold was completely lined by endotheliallike cells. Thus, the possibility of small artery reconstruction using decellularized scaffolds was demonstrated.

  7. Hypoxia Does neither Stimulate Pulmonary Artery Endothelial Cell Proliferation in Mice and Rats with Pulmonary Hypertension and Vascular Remodeling nor in Human Pulmonary Artery Endothelial Cells

    PubMed Central

    Yu, Lunyin; Hales, Charles A.

    2011-01-01

    Background Hypoxia results in pulmonary hypertension and vascular remodeling due to induction of pulmonary artery cell proliferation. Besides pulmonary artery smooth muscle cells, pulmonary artery endothelial cells (PAECs) are also involved in the development of pulmonary hypertension, but the effect of hypoxia on PAEC proliferation has not been completely understood. Methods We investigated PAEC proliferation in mice and rats with hypoxia-induced pulmonary hypertension and vascular remodeling as well as in human PAECs under hypoxia. Results and Conclusion We did not find significant PAEC proliferation in chronically hypoxic rats or mice. There was a slight decrease in proliferation in mice and rats with pulmonary hypertension and vascular remodeling. We also did not find significant human PAEC proliferation and cell cycle progression under different levels of oxygen (1, 2, 3, 5 and 10%) for one day, although the same conditions of hypoxia induced significant proliferation and cell cycle progression in pulmonary artery smooth muscle cells and pulmonary artery fibroblasts. Exposure to hypoxia for 7 days also did not increase PAEC proliferation. These results demonstrated that hypoxia alone is not a stimulus to PAEC proliferation in vivo and in vitro. The present study provides a novel role for PAECs in hypoxia-induced pulmonary hypertension and vascular remodeling. PMID:21691120

  8. [Arterial vascularization of the triceps sural muscle].

    PubMed

    Mairesse, J L; Mestdagh, H; Procyk, S; Depreux, R

    1984-01-01

    The triceps surae muscle, the dorsal and medial leg skin constitute a very important reserve of muscular and myocutaneous flaps. The material on which the study was carried out consisted of 20 legs from standard cadavers. The superficialis femoral artery was injected with terebenthene and minimum mixture. The medial head of gastrocnemius is 23.3 em long, 6.9 cm wide, 1.25 mm thick at distal third. Its dominant blood supply is carried by the medialis gastrocnemius artery. It rises from popliteal artery 1.2 cm above the femoral tibial articulation with 1.9 mm diameter. It runs 3 cm down before entering muscle where it provides 2 or 3 mean branches. These branches give musculocutaneous arteries to the skin of the dorsal leg. The same study was performed for the lateral head of gastrocnemius and soleus. We studied also arteries of dorsomedial leg skin. The characteristics of long saphenous and short saphenous arteries were described. These muscles and dorsomedial leg skin can be used as muscular or myocutaneous flap for covering defects between the lower leg and the lower thigh.

  9. Molecular Mechanisms of Pulmonary Vascular Remodeling in Pulmonary Arterial Hypertension

    PubMed Central

    Leopold, Jane A.; Maron, Bradley A.

    2016-01-01

    Pulmonary arterial hypertension (PAH) is a devastating disease that is precipitated by hypertrophic pulmonary vascular remodeling of distal arterioles to increase pulmonary artery pressure and pulmonary vascular resistance in the absence of left heart, lung parenchymal, or thromboembolic disease. Despite available medical therapy, pulmonary artery remodeling and its attendant hemodynamic consequences result in right ventricular dysfunction, failure, and early death. To limit morbidity and mortality, attention has focused on identifying the cellular and molecular mechanisms underlying aberrant pulmonary artery remodeling to identify pathways for intervention. While there is a well-recognized heritable genetic component to PAH, there is also evidence of other genetic perturbations, including pulmonary vascular cell DNA damage, activation of the DNA damage response, and variations in microRNA expression. These findings likely contribute, in part, to dysregulation of proliferation and apoptosis signaling pathways akin to what is observed in cancer; changes in cellular metabolism, metabolic flux, and mitochondrial function; and endothelial-to-mesenchymal transition as key signaling pathways that promote pulmonary vascular remodeling. This review will highlight recent advances in the field with an emphasis on the aforementioned molecular mechanisms as contributors to the pulmonary vascular disease pathophenotype. PMID:27213345

  10. PPARβ/δ, a Novel Regulator for Vascular Smooth Muscle Cells Phenotypic Modulation and Vascular Remodeling after Subarachnoid Hemorrhage in Rats.

    PubMed

    Zhang, Hongrong; Jiang, Li; Guo, Zongduo; Zhong, Jianjun; Wu, Jingchuan; He, Junchi; Liu, Han; He, Zhaohui; Wu, Haitao; Cheng, Chongjie; Sun, Xiaochuan

    2017-03-22

    Cerebral vascular smooth muscle cell (VSMC) phenotypic switch is involved in the pathophysiology of vascular injury after aneurysmal subarachnoid hemorrhage (aSAH), whereas the molecular mechanism underlying it remains largely speculative. Peroxisome proliferator-activated receptor β/δ (PPARβ/δ) has been implicated to modulate the vascular cells proliferation and vascular homeostasis. In the present study, we investigated the potential role of PPARβ/δ in VSMC phenotypic switch following SAH. Activation of PPARβ/δ by GW0742 and adenoviruses PPARβ/δ (Ad-PPARβ/δ) significantly inhibited hemoglobin-induced VSMC phenotypic switch. However, the effects of PPARβ/δ on VSMC phenotypic switch were partly obstacled in the presence of LY294002, a potent inhibitor of Phosphatidyl-Inositol-3 Kinase-AKT (PI3K/AKT). Furthermore, following study demonstrated that PPARβ/δ-induced PI3K/AKT activation can also contribute to Serum Response Factor (SRF) nucleus localization and Myocardin expression, which was highly associated with VSMC phenotypic switch. Finally, we found that Ad-PPARβ/δ positively modulated vascular remodeling in SAH rats, i.e. the diameter of basilar artery and the thickness of vessel wall. In addition, overexpression of PPARβ/δ by adenoviruses significantly improved neurological outcome. Taken together, this study identified PPARβ/δ as a useful regulator for VSMC phenotypic switch and vascular remodeling following SAH, providing novel insights into the therapeutic strategies of delayed cerebral ischemia.

  11. PPARβ/δ, a Novel Regulator for Vascular Smooth Muscle Cells Phenotypic Modulation and Vascular Remodeling after Subarachnoid Hemorrhage in Rats

    PubMed Central

    Zhang, Hongrong; Jiang, Li; Guo, Zongduo; Zhong, Jianjun; Wu, Jingchuan; He, Junchi; Liu, Han; He, Zhaohui; Wu, Haitao; Cheng, Chongjie; Sun, Xiaochuan

    2017-01-01

    Cerebral vascular smooth muscle cell (VSMC) phenotypic switch is involved in the pathophysiology of vascular injury after aneurysmal subarachnoid hemorrhage (aSAH), whereas the molecular mechanism underlying it remains largely speculative. Peroxisome proliferator-activated receptor β/δ (PPARβ/δ) has been implicated to modulate the vascular cells proliferation and vascular homeostasis. In the present study, we investigated the potential role of PPARβ/δ in VSMC phenotypic switch following SAH. Activation of PPARβ/δ by GW0742 and adenoviruses PPARβ/δ (Ad-PPARβ/δ) significantly inhibited hemoglobin-induced VSMC phenotypic switch. However, the effects of PPARβ/δ on VSMC phenotypic switch were partly obstacled in the presence of LY294002, a potent inhibitor of Phosphatidyl-Inositol-3 Kinase-AKT (PI3K/AKT). Furthermore, following study demonstrated that PPARβ/δ-induced PI3K/AKT activation can also contribute to Serum Response Factor (SRF) nucleus localization and Myocardin expression, which was highly associated with VSMC phenotypic switch. Finally, we found that Ad-PPARβ/δ positively modulated vascular remodeling in SAH rats, i.e. the diameter of basilar artery and the thickness of vessel wall. In addition, overexpression of PPARβ/δ by adenoviruses significantly improved neurological outcome. Taken together, this study identified PPARβ/δ as a useful regulator for VSMC phenotypic switch and vascular remodeling following SAH, providing novel insights into the therapeutic strategies of delayed cerebral ischemia. PMID:28327554

  12. Evidence for a role of collagen synthesis in arterial smooth muscle cell migration.

    PubMed Central

    Rocnik, E F; Chan, B M; Pickering, J G

    1998-01-01

    Migration of smooth muscle cells (SMCs) and collagen synthesis by SMCs are central to the pathophysiology of vascular disease. Both processes can be induced shortly after vascular injury; however, a functional relationship between them has not been established. In this study, we determined if collagen synthesis was required for SMC migration, using ethyl-3,4-dihydroxybenzoate (EDHB), an inhibitor of prolyl-4-hydroxylase, and 3,4-DL-dehydroproline (DHP), a proline analogue, which we demonstrate inhibit collagen elaboration by porcine arterial SMCs. SMCs exposed to EDHB or DHP attached normally to collagen- and vitronectin-coated substrates; however, spreading on collagen but not vitronectin was inhibited. SMC migration speed, quantified by digital time-lapse video microscopy, was significantly and reversibly reduced by EDHB and DHP. Flow cytometry revealed that expression of beta1 integrins, through which SMCs interact with collagen, was unaffected by EDHB or DHP. However, both inhibitors prevented normal clustering of beta1 integrins on the surface of SMCs, consistent with a lack of appropriate matrix ligands for integrin engagement. Moreover, there was impaired recruitment of vinculin into focal adhesion complexes of spreading SMCs and disassembly of the smooth muscle alpha-actin-containing cytoskeleton. These findings suggest that de novo collagen synthesis plays a role in SMC migration and implicates a mechanism whereby newly synthesized collagen may be necessary to maintain the transcellular traction system required for effective locomotion. PMID:9576753

  13. Inhibition of Rho protein stimulates iNOS expression in rat vascular smooth muscle cells.

    PubMed

    Muniyappa, R; Xu, R; Ram, J L; Sowers, J R

    2000-06-01

    Inducible nitric oxide synthase (iNOS) in vascular smooth muscle cells (VSMCs) is upregulated in arterial injury and plays a role in regulating VSMC proliferation and restenosis. Inflammatory cytokines [e.g., interleukin-1beta (IL-1beta)] released during vascular injury induce iNOS. Small GTP-binding proteins of the Ras superfamily play a major role in IL-1beta-dependent signaling pathways. In this study, we examined the role of Rho GTPases in regulating iNOS expression in VSMCs. Treatment of VSMCs with mevastatin, which inhibits isoprenylation of Rho and other small GTP-binding proteins, produced significantly higher amounts of IL-1beta-evoked NO and iNOS protein compared with control. Similarly, bacterial toxins [Toxin B from Clostridium difficile and C3 ADP-ribosyl transferase (C3) toxin from Clostridium botulinium] that specifically inactivate Rho proteins increased NOS products (NO and citrulline) and iNOS expression. Toxin B increased the activity of iNOS promoter-reporter construct in VSMCs. Both toxins enhanced IL-1beta-stimulated iNOS expression and NO production. These data demonstrate for the first time that inhibition of Rho induces iNOS and suggest a role for Rho protein in IL-1beta-stimulated NO production in VSMCs.

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

    SciTech Connect

    Perez-Vizcaino, Francisco . E-mail: fperez@med.ucm.es; Bishop-Bailley, David; Lodi, Federica; Duarte, Juan; Cogolludo, Angel; Moreno, Laura; Bosca, Lisardo; Mitchell, Jane A.; Warner, Timothy D.

    2006-08-04

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

  15. PDT-induced apoptosis in arterial smooth muscles cells

    NASA Astrophysics Data System (ADS)

    Nyamekye, Isaac; Renick, R.; Gilbert, C.; McEwan, Jean R.; Evan, G.; Bishop, Christopher C. R.; Bown, Stephen G.

    1995-03-01

    PDT kills smooth muscle cells (SMC) in vivo and thus prevents intimal hyperplasia after angioplasty. It causes little inflammation and structural integrity of the artery is not compromised. We have studied the process of the SMC death in vitro. Cultured rat SMC (cell line sv40 ATCC) were sensitized with aluminum disulphonated phthalocyanine (AlS2Pc), and then irradiated with 675 nm laser light (2.5 J/cm2). Controls were studied using only sensitizer or laser for treatment. The cells were incubated and the dying process observed with a time lapse video and microscope system. PDT caused a characteristic pattern of death. Cells lost contact with neighbors, shrank, and showed hyperactivity and membrane ruffling. The cells imploded into active and condensed membrane bound vesicles which were terminally reduced to residual bodies. These are the morphological changes of apoptosis. The control cells which were given AlS2Pc alone or laser alone showed no death. PDT induced cultured arterial SMC death by apoptosis rather than necrosis. An apoptotic mechanism of cell death in vivo would explain the relative lack of inflammation and local tissue destruction in the face of massive death.

  16. [Primary culture and functional identification of distal pulmonary artery smooth muscle cells in mice].

    PubMed

    Li, M C; Chen, Y Q; Zhang, C T; Jiang, Q; Lu, W J; Wang, J

    2017-02-12

    Objective: To establish a method of isolation and primary culture of mice distal pulmonary artery smooth muscle cells (PASMCs) and identify the functional properties. Methods: PASMCs were harvested from the distal pulmonary artery (PA) tissue of mice by enzymatic digestion of collagenaseⅠand papain; and the growth characteristics were observed under inverted microscope and identified by Immunofluorescence technique. Effects on the intracellular calcium ion concentration of distal PASMCs were detected by Fura-2-AM fluorescent probe tracer under a fluorescence microscope in Krebs solution containing clopiazonic acid (CPA) and nifedipin (Nif). Results: PASMCs density reached approximately to 80% in a typical valley-peak-like shape after 6 days. Cell α-smooth muscle actin (α-SMA) immunofluorescence identified that 95% of the cultured cells were PASMCs. More than 95% PASMCs responded well to calcium-potassium Krebs solution (potassium ion concentration of 60 mmol/L) and showed a rapid increase in basal [Ca(2+) ](i) after 1 minute's perfusion (Δ[Ca(2+) ](i)>50), which demonstrated that the voltage-dependent calcium channels (VDCC) of distal PASMCs were in good function; after the perfusion of calcium Krebs, calcium-free/calcium-Krebs containing CPA and Nif, distal PASMCs showed two typical peaks, indicated the full function of store-operated calcium channel (SOCC) in distal PASMCs. Conclusion: This experiment successfully established a stable and reliable mice distal PASMCs model and the study of pulmonary vascular diseases could benefit from its higher purity and better functional condition.

  17. Transdifferentiation of endothelial cells to smooth muscle cells play an important role in vascular remodelling

    PubMed Central

    Coll-Bonfill, Núria; Musri, Melina Mara; Ivo, Victor; Barberà, Joan Albert; Tura-Ceide, Olga

    2015-01-01

    Pulmonary artery remodelling it is a major feature of pulmonary hypertension (PH). It is characterised by cellular and structural changes of the pulmonary arteries causing higher pulmonar vascular resistance and right ventricular failure. Abnormal deposition of smooth muscle-like (SM-like) cells in normally non-muscular, small diameter vessels and a deregulated control of endothelial cells are considered pathological features of PH. The origin of the SM-like cells and the mechanisms underlying the development and progression of this remodelling process are not understood. Endothelial cells within the intima may migrate from their organised layer of cells and transition to mesenchymal or SM-like phenotype in a process called endothelial-mesenchymal transition (EnMT). Traditionally, Waddington’s epigenetic landscape illustrates that fates of somatic cells are progressively determined to compulsorily follow a downhill differentiation pathway. EnMT induces the transformation of cells with stem cell traits, therefore contrasting Waddington’s theory and confirming that cell fate seems to be far more flexible than previously thought. The prospect of therapeutic inhibition of EnMT to delay or prevent PH may represent a promising new treatment modality. PMID:25973327

  18. Transdifferentiation of endothelial cells to smooth muscle cells play an important role in vascular remodelling.

    PubMed

    Coll-Bonfill, Núria; Musri, Melina Mara; Ivo, Victor; Barberà, Joan Albert; Tura-Ceide, Olga

    2015-01-01

    Pulmonary artery remodelling it is a major feature of pulmonary hypertension (PH). It is characterised by cellular and structural changes of the pulmonary arteries causing higher pulmonar vascular resistance and right ventricular failure. Abnormal deposition of smooth muscle-like (SM-like) cells in normally non-muscular, small diameter vessels and a deregulated control of endothelial cells are considered pathological features of PH. The origin of the SM-like cells and the mechanisms underlying the development and progression of this remodelling process are not understood. Endothelial cells within the intima may migrate from their organised layer of cells and transition to mesenchymal or SM-like phenotype in a process called endothelial-mesenchymal transition (EnMT). Traditionally, Waddington's epigenetic landscape illustrates that fates of somatic cells are progressively determined to compulsorily follow a downhill differentiation pathway. EnMT induces the transformation of cells with stem cell traits, therefore contrasting Waddington's theory and confirming that cell fate seems to be far more flexible than previously thought. The prospect of therapeutic inhibition of EnMT to delay or prevent PH may represent a promising new treatment modality.

  19. Increased endothelial and vascular smooth muscle cell adhesion on nanostructured titanium and CoCrMo

    PubMed Central

    Choudhary, Saba; Berhe, Mikal; Haberstroh, Karen M; Webster, Thomas J

    2006-01-01

    In the body, vascular cells continuously interact with tissues that possess nanostructured surface features due to the presence of proteins (such as collagen and elastin) embedded in the vascular wall. Despite this fact, vascular stents intended to restore blood flow do not have nanoscale surface features but rather are smooth at the nanoscale. As the first step towards creating the next generation of vascular stent materials, the objective of this in vitro study was to investigate vascular cell (specifically, endothelial, and vascular smooth muscle cell) adhesion on nanostructured compared with conventional commercially pure (cp) Ti and CoCrMo. Nanostructured cp Ti and CoCrMo compacts were created by separately utilizing either constituent cp Ti or CoCrMo nanoparticles as opposed to conventional micronsized particles. Results of this study showed for the first time increased endothelial and vascular smooth muscle cell adhesion on nanostructured compared with conventional cp Ti and CoCrMo after 4 hours’ adhesion. Moreover, compared with their respective conventional counterparts, the ratio of endothelial to vascular smooth muscle cells increased on nanostructured cp Ti and CoCrMo. In addition, endothelial and vascular smooth muscle cells had a better spread morphology on the nanostructured metals compared with conventional metals. Overall, vascular cell adhesion was better on CoCrMo than on cp Ti. Results of surface characterization studies demonstrated similar chemistry but significantly greater root-mean-square (rms) surface roughness as measured by atomic force microscopy (AFM) for nanostructured compared with respective conventional metals. For these reasons, results from the present in vitro study provided evidence that vascular stents composed of nanometer compared with micron-sized metal particles (specifically, either cp Ti or CoCrMo) may invoke cellular responses promising for improved vascular stent applications. PMID:17722261

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

    PubMed

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

    2016-05-10

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

  1. Magnolol inhibits migration of vascular smooth muscle cells via cytoskeletal remodeling pathway to attenuate neointima formation

    SciTech Connect

    Karki, Rajendra; Kim, Seong-Bin; Kim, Dong-Wook

    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

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

    PubMed Central

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

    2016-01-01

    Cyclic stretch is an important inducer of vascular smooth muscle cell (VSMC) proliferation, which is crucial in vascular remodeling during hypertension. However, the molecular mechanism remains unclear. We studied the effects of emerin and lamin A/C, two important nuclear envelope proteins, on VSMC proliferation in hypertension and the underlying mechano-mechanisms. In common carotid artery of hypertensive rats in vivo and in cultured cells subjected to high (15%) cyclic stretch in vitro, VSMC proliferation was increased significantly, and the expression of emerin and lamin A/C was repressed compared with normotensive or normal (5%) cyclic stretch controls. Using targeted siRNA to mimic the repressed expression of emerin or lamin A/C induced by 15% stretch, we found that VSMC proliferation was enhanced under static and 5%-stretch conditions. Overexpression of emerin or lamin A/C reversed VSMC proliferation induced by 15% stretch. Hence, emerin and lamin A/C play critical roles in suppressing VSMC hyperproliferation induced by hyperstretch. ChIP-on-chip and MOTIF analyses showed that the DNAs binding with emerin contain three transcription factor motifs: CCNGGA, CCMGCC, and ABTTCCG; DNAs binding with lamin A/C contain the motifs CVGGAA, GCCGCYGC, and DAAGAAA. Protein/DNA array proved that altered emerin or lamin A/C expression modulated the activation of various transcription factors. Furthermore, accelerating local expression of emerin or lamin A/C reversed cell proliferation in the carotid artery of hypertensive rats in vivo. Our findings establish the pathogenetic role of emerin and lamin A/C repression in stretch-induced VSMC proliferation and suggest mechanobiological mechanism underlying this process that involves the sequence-specific binding of emerin and lamin A/C to specific transcription factor motifs. PMID:27114541

  3. ECM-mimetic heparin glycosamioglycan-functionalized surface favors constructing functional vascular smooth muscle tissue in vitro.

    PubMed

    Zhang, Jimin; Wang, Jianing; Wei, Yongzhen; Gao, Cheng; Chen, Xuejiao; Kong, Wei; Kong, Deling; Zhao, Qiang

    2016-10-01

    Contractile vascular smooth muscle accounts for the normal physiological function of artery. Heparin, as a native glycosaminoglycan, has been well known for its important function in promoting or maintaining the contractile phenotype of vascular smooth muscle cells (VSMCs). In this study, heparin-functionalized non-woven poly(ε-caprolactone) (PCL) mat was fabricated by a facile and efficient surface modification protocol, which enables the control of surface heparin density within a broad range. Surface heparization remarkably increased the hydrophilicity of PCL, and reduced platelet adhesion. MTT assay showed that VSMC proliferation was evidently inhibited on the heparin-functionalized PCL surface in a dose-dependent manner. Gene analysis confirmed that surface heparization also promoted the transition of VSMCs from synthetic phenotype to contractile one. Furthermore, with a proper surface density of heparin, it allowed VSMCs to grow in a certain rate, while exhibiting contractile phenotype. Culture of VSMCs on a modified PCL mat with moderate heparin density (PCL-Hep-20) for 2 days resulted in a confluent layer of contractile smooth muscle cells. These data suggest that the heparin-modified PCL scaffolds may be a promising candidate to generate functional vascular tissues in vitro.

  4. Attenuation of endothelin-1-induced calcium response by tyrosine kinase inhibitors in vascular smooth muscle cells.

    PubMed

    Liu, C Y; Sturek, M

    1996-06-01

    Although tyrosine kinases play an important role in cell growth and have been implicated in regulation of smooth muscle contraction, their role in agonist-induced myoplasmic Ca2+ responses is unclear. We examined effects of the tyrosine kinase inhibitors genistein and methyl 2,5-dihydroxycinnamate (MDHC) on the endothelin-1 (ET-1)-induced Ca2+ response and determined underlying mechanisms for the effects. Freshly isolated smooth muscle cells from porcine coronary arteries were loaded with fura 2 ester, and myoplasmic free Ca2+ (Ca2+ (m)) concentration was estimated with fura 2 microfluorometry. Both genistein and MDHC inhibited the initial transient Cam2+ response to ET by 54 and 81%, respectively (P < 0.05), in the presence of extracellular Ca2+. Genistein also significantly delayed the Cam2+ response, with the latent period from ET-1 application to the beginning of the Cam2+ response being increased from 1.08 +/- 0.17 to 2.65 +/- 0.52 min (P < 0.05). In the absence of extracellular Ca2+, genistein inhibited the ET-1-induced Cam2+ response by 93% (P < 0.05). The Cam2+ responses to caffeine (5 mM) or inositol trisphosphate (IP3) applied intracellularly via a patch-clamp pipette were not affected by genistein. Both genistein and MDHC also abolished the sustained Cam2+ response to ET-1. However, the Cam2+ response to depolarization by 80 mM K+ was not inhibited by MDHC and only inhibited 22% by genistein (P < 0.05). These results indicate that 1) activation of tyrosine kinases is an important regulatory mechanism for the ET-1-induced Cam2+ response in vascular smooth muscle and 2) tyrosine kinases mediate ET-1-induced Ca2+ release with no direct effect on IP3-mediated Ca2+ release. Thus ET-1-mediated signaling upstream of IP3 interaction with the Ca2+ stores is regulated by tyrosine kinases.

  5. Vanin-1 pantetheinase drives smooth muscle cell activation in post-arterial injury neointimal hyperplasia.

    PubMed

    Dammanahalli, K Jagadeesha; Stevens, Stephanie; Terkeltaub, Robert

    2012-01-01

    The pantetheinase vanin-1 generates cysteamine, which inhibits reduced glutathione (GSH) synthesis. Vanin-1 promotes inflammation and tissue injury partly by inducing oxidative stress, and partly by peroxisome proliferator-activated receptor gamma (PPARγ) expression. Vascular smooth muscle cells (SMCs) contribute to neointimal hyperplasia in response to injury, by multiple mechanisms including modulation of oxidative stress and PPARγ. Therefore, we tested the hypothesis that vanin-1 drives SMC activation and neointimal hyperplasia. We studied reactive oxygen species (ROS) generation and functional responses to platelet-derived growth factor (PDGF) and the pro-oxidant diamide in cultured mouse aortic SMCs, and also assessed neointima formation after carotid artery ligation in vanin-1 deficiency. Vnn1(-/-) SMCs demonstrated decreased oxidative stress, proliferation, migration, and matrix metalloproteinase 9 (MMP-9) activity in response to PDGF and/or diamide, with the effects on proliferation linked, in these studies, to both increased GSH levels and PPARγ expression. Vnn1(-/-) mice displayed markedly decreased neointima formation in response to carotid artery ligation, including decreased intima:media ratio and cross-sectional area of the neointima. We conclude that vanin-1, via dual modulation of GSH and PPARγ, critically regulates the activation of cultured SMCs and development of neointimal hyperplasia in response to carotid artery ligation. Vanin-1 is a novel potential therapeutic target for neointimal hyperplasia following revascularization.

  6. Laminar shear stress stimulates vascular smooth muscle cell apoptosis via the Akt pathway.

    PubMed

    Fitzgerald, Tamara N; Shepherd, Benjamin R; Asada, Hidenori; Teso, Desarom; Muto, Akihito; Fancher, Tiffany; Pimiento, Jose M; Maloney, Stephen P; Dardik, Alan

    2008-08-01

    Vascular smooth muscle cells (SMC) may be directly exposed to blood flow after an endothelial-denuding injury. It is not known whether direct exposure of SMC to shear stress reduces SMC turnover and contributes to the low rate of restenosis after most vascular interventions. This study examines if laminar shear stress inhibits SMC proliferation or stimulates apoptosis. Bovine aortic SMC were exposed to arterial magnitudes of laminar shear stress (11 dynes/cm(2)) for up to 24 h and compared to control SMC (0 dynes/cm(2)). SMC density was assessed by cell counting, DNA synthesis by (3)[H]-thymidine incorporation, and apoptosis by TUNEL staining. Akt, caspase, bax, and bcl-2 phosphorylation were assessed by Western blotting; caspase activity was also measured with an in vitro assay. Analysis of variance was used to compare groups. SMC exposed to laminar shear stress had a 38% decrease in cell number (n = 4, P = 0.03), 54% reduction in (3)[H]-thymidine incorporation (n = 3, P = 0.003), and 15-fold increase in TUNEL staining (n = 4, P < 0.0001). Akt phosphorylation was reduced by 67% (n = 3, P < 0.0001), whereas bax/bcl-2 phosphorylation was increased by 1.8-fold (n = 3, P = 0.01). Caspase-3 activity was increased threefold (n = 5, P = 0.03). Pretreatment of cells with ZVAD-fmk or wortmannin resulted in 42% increased cell retention (n = 3, P < 0.01) and a fourfold increase in apoptosis (n = 3, P < 0.04), respectively. Cells transduced with constitutively-active Akt had twofold decreased apoptosis (n = 3, P < 0.002). SMC exposed to laminar shear stress have decreased proliferation and increased apoptosis, mediated by the Akt pathway. These results suggest that augmentation of SMC apoptosis may be an alternative strategy to inhibit restenosis after vascular injury.

  7. Nitric Oxide and the Mechanism of Rat Vascular Smooth Muscle Photorelaxation

    PubMed Central

    Flitney, Frederick Werner; Megson, Ian L

    2003-01-01

    Photorelaxation of vascular smooth muscle (VSM) was studied using segments of tail artery from normotensive rats (NTR) or spontaneously hypertensive rats (SHR). Isolated vessels with intact endothelium were perfused with Krebs solution containing phenylephrine. Perfusion pressures were recorded while arteries were irradiated with either visible (VIS; λ = 514.5 nm) or long wavelength ultra-violet (UVA; λ = 366 nm) light. VIS light produced a transient vasodilator response: a rapid decrease of pressure that recovered fully during the period (6 min) of illumination. An irradiated artery was refractory to a second period of illumination delivered immediately after the first, but its photosensitivity recovered slowly in the dark, a process called ‘repriming’. Photorelaxations generated by UVA light were qualitatively different and consisted of two components: a phasic (or p-) component superimposed on a sustained (or s-) component. The p-component is similar to the VIS light-induced response in that both exhibit refractoriness and repriming depends upon endothelium-derived NO. In contrast, the s-component persists throughout the period of illumination and does not show refractoriness. We conclude that VIS light-induced photorelaxations and the p-component of UVA light-induced responses are mediated by the photochemical release of NO from a finite molecular ‘store’ that can be reconstituted afterwards in the dark. The s-component of the UVA light-induced response does not depend directly on endothelial NO and may result instead from a stimulatory effect of UVA light on soluble guanylate cyclase. NO-dependent photorelaxation is impaired in vessels from SHR while the s-component is enhanced. PMID:12824453

  8. A comparative study of potassium-induced relaxation in vascular smooth muscle of tiger salamanders and rats.

    PubMed

    Malvin, G M; Webb, R C

    1984-07-01

    This study compares potassium-induced relaxation in vascular tissue of an amphibian (Ambystoma tigrinum) and a mammal (rat). Aortas (salamanders) and tail arteries (rats) were cut into helical strips for isometric force recording. After norepinephrine-induced contraction in potassium-free solution, arteries relaxed in response to added potassium (1-20 mmol/l). Potassium-induced relaxation was greater in rat tail arteries than in salamander aortas. Half-maximal relaxation occurred at a potassium concentration of approximately 3 mmol/l in both species. Ouabain inhibited potassium-induced relaxation; salamanders were more sensitive to the glycoside than rats. Potassium-induced relaxation decreased as the temperature of the bathing medium was lowered; half-maximal inhibition occurred at 19 and 29 degrees C for salamander aortas and rat tail arteries, respectively. Potassium-induced relaxation also varied with the interval in potassium-free solution, the hydrogen ion concentration (rats only), and the magnitude of norepinephrine-induced contraction. It appears that the cellular mechanism causing potassium-induced relaxation is similar in blood vessels of salamanders and rats. The observations are consistent with the hypothesis that stimulated electrogenic sodium transport produced membrane hyperpolarization and relaxation in vascular smooth muscle.

  9. Antagonism of Nav channels and α1-adrenergic receptors contributes to vascular smooth muscle effects of ranolazine

    PubMed Central

    Virsolvy, Anne; Farah, Charlotte; Pertuit, Nolwenn; Kong, Lingyan; Lacampagne, Alain; Reboul, Cyril; Aimond, Franck; Richard, Sylvain

    2015-01-01

    Ranolazine is a recently developed drug used for the treatment of patients with chronic stable angina. It is a selective inhibitor of the persistent cardiac Na+ current (INa), and is known to reduce the Na+-dependent Ca2+ overload that occurs in cardiomyocytes during ischemia. Vascular effects of ranolazine, such as vasorelaxation,have been reported and may involve multiple pathways. As voltage-gated Na+ channels (Nav) present in arteries play a role in contraction, we hypothesized that ranolazine could target these channels. We studied the effects of ranolazine in vitro on cultured aortic smooth muscle cells (SMC) and ex vivo on rat aortas in conditions known to specifically activate or promote INa. We observed that in the presence of the Nav channel agonist veratridine, ranolazine inhibited INa and intracellular Ca2+ calcium increase in SMC, and arterial vasoconstriction. In arterial SMC, ranolazine inhibited the activity of tetrodotoxin-sensitive voltage-gated Nav channels and thus antagonized contraction promoted by low KCl depolarization. Furthermore, the vasorelaxant effects of ranolazine, also observed in human arteries and independent of the endothelium, involved antagonization of the α1-adrenergic receptor. Combined α1-adrenergic antagonization and inhibition of SMCs Nav channels could be involved in the vascular effects of ranolazine. PMID:26655634

  10. Antagonism of Nav channels and α1-adrenergic receptors contributes to vascular smooth muscle effects of ranolazine.

    PubMed

    Virsolvy, Anne; Farah, Charlotte; Pertuit, Nolwenn; Kong, Lingyan; Lacampagne, Alain; Reboul, Cyril; Aimond, Franck; Richard, Sylvain

    2015-12-10

    Ranolazine is a recently developed drug used for the treatment of patients with chronic stable angina. It is a selective inhibitor of the persistent cardiac Na(+) current (INa), and is known to reduce the Na(+)-dependent Ca(2+) overload that occurs in cardiomyocytes during ischemia. Vascular effects of ranolazine, such as vasorelaxation,have been reported and may involve multiple pathways. As voltage-gated Na(+) channels (Nav) present in arteries play a role in contraction, we hypothesized that ranolazine could target these channels. We studied the effects of ranolazine in vitro on cultured aortic smooth muscle cells (SMC) and ex vivo on rat aortas in conditions known to specifically activate or promote INa. We observed that in the presence of the Nav channel agonist veratridine, ranolazine inhibited INa and intracellular Ca(2+) calcium increase in SMC, and arterial vasoconstriction. In arterial SMC, ranolazine inhibited the activity of tetrodotoxin-sensitive voltage-gated Nav channels and thus antagonized contraction promoted by low KCl depolarization. Furthermore, the vasorelaxant effects of ranolazine, also observed in human arteries and independent of the endothelium, involved antagonization of the α1-adrenergic receptor. Combined α1-adrenergic antagonization and inhibition of SMCs Nav channels could be involved in the vascular effects of ranolazine.

  11. Lanthanum prevents high phosphate-induced vascular calcification by preserving vascular smooth muscle lineage markers.

    PubMed

    Ciceri, Paola; Elli, Francesca; Brenna, Irene; Volpi, Elisa; Romagnoli, Solange; Tosi, Delfina; Braidotti, Paola; Brancaccio, Diego; Cozzolino, Mario

    2013-06-01

    Vascular calcification (VC) represents a major cardiovascular risk factor in chronic kidney disease patients. High phosphate (Pi) levels are strongly associated with VC in this population. Therefore, Pi binders are commonly used to control high Pi levels. The aim of this work was to study the mechanism of action of lanthanum chloride (LaCl3) on the progression of Pi-induced VC through its direct effect on vascular smooth muscle cells (VSMCs) in vitro. High Pi induced VSCM Ca deposition. We evaluated the action of LaCl3, compared to gadolinium chloride (GdCl3), and found different effects on the modulation of VSMC lineage markers, such as α-actin and SM22α. In fact, only LaCl3 preserved the expression of both VSMC lineage markers compared to high Pi-treated cells. Interestingly, both LaCl3 and GdCl3 reduced the high Pi-induced elevations of bone morphogenic protein 2 mRNA expression, with no reduction of the high core binding factor-alpha 1 mRNA levels observed in calcified VSMCs. Furthermore, we also found that only LaCl3 completely prevented the matrix GLA protein mRNA levels and osteonectin protein expression elevations induced by high Pi compared to GdCl3. Finally, LaCl3, in contrast to GdCl3, prevented the high Pi-induced downregulation of Axl, a membrane tyrosine kinase receptor involved in apoptosis. Thus, our results suggest that LaCl3 prevents VC by preserving VSMC lineage markers and by decreasing high Pi-induced osteoblastic differentiation.

  12. Importance of extracellular Ca2+ and intracellular Ca2+ release in ethanol-induced contraction of cerebral arterial smooth muscle.

    PubMed

    Yang, Z; Wang, J; Zheng, T; Altura, B T; Altura, B M

    2001-07-01

    The present study was designed to investigate the roles of extracellular Ca2+ ([Ca2+]0) influx and intracellular free Ca2+ ([Ca2+]i) release in ethanol-induced contractions of isolated canine cerebral arteries and primary cultured, cerebral vascular smooth muscle cells. Ethanol (20-200 mM) produced significant contractions in isolated canine basilar arterial rings in a concentration-dependent manner. Removal of [Ca2+]0 and pretreatment of canine basilar arterial rings with verapamil (an antagonist of voltage-gated Ca2+ channels), thapsigargin (a selective antagonist of the sarcoplasmic reticulum Ca2+ pump), caffeine plus ryanodine (a specific antagonist of ryanodine-sensitive Ca2+ release), or heparin (an inositol 1,4,5,-trisphosphate [InsP3]-mediated Ca2+ release antagonist) markedly attenuated (approximately 50%-80%) ethanol-induced contractions. The absence of [Ca2+]0 and preincubation of primary single smooth muscle cells obtained from canine basilar arteries with verapamil, thapsigargin, heparin, or caffeine plus ryanodine markedly attenuated (approximately 50%-80%) the transient and sustained elevations in [Ca2+]i induced by ethanol. Results of the present study suggest to us that both Ca2+ influx through voltage-gated Ca2+ channels and Ca2+ release from intracellular stores (both InsP3 sensitive and ryanodine sensitive) are required for ethanol-induced contractions of isolated canine basilar arteries.

  13. Arterial structure and function in vascular ageing: are you as old as your arteries?

    PubMed

    Thijssen, Dick H J; Carter, Sophie E; Green, Daniel J

    2016-04-15

    Advancing age may be the most potent independent predictor of future cardiovascular events, a relationship that is not fully explained by time-related changes in traditional cardiovascular risk factors. Since some arteries exhibit differential susceptibility to atherosclerosis, generalisations regarding the impact of ageing in humans may be overly simplistic, whereas in vivo assessment of arterial function and health provide direct insight. Coronary and peripheral (conduit, resistance and skin) arteries demonstrate a gradual, age-related impairment in vascular function that is likely to be related to a reduction in endothelium-derived nitric oxide bioavailability and/or increased production of vasoconstrictors (e.g. endothelin-1). Increased exposure and impaired ability for defence mechanisms to resist oxidative stress and inflammation, but also cellular senescence processes, may contribute to age-related changes in vascular function and health. Arteries also undergo structural changes as they age. Gradual thickening of the arterial wall, changes in wall content (i.e. less elastin, advanced glycation end-products) and increase in conduit artery diameter are observed with older age and occur similarly in central and peripheral arteries. These changes in structure have important interactive effects on artery function, with increases in small and large arterial stiffness representing a characteristic change with older age. Importantly, direct measures of arterial function and structure predict future cardiovascular events, independent of age or other cardiovascular risk factors. Taken together, and given the differential susceptibility of arteries to atherosclerosis in humans, direct measurement of arterial function and health may help to distinguish between biological and chronological age-related change in arterial health in humans.

  14. Cocaine mediated apoptosis of vascular cells as a mechanism for carotid artery dissection leading to ischemic stroke.

    PubMed

    Dabbouseh, Noura M; Ardelt, Agnieszka

    2011-08-01

    In arterial dissection, blood may enter the arterial wall through an intimal tear, splitting the arterial wall and activating the coagulation cascade at the site of endothelial damage. Dissection of extracranial and intracranial vessels may lead to ischemic stroke through thromboembolic or hemodynamic mechanisms. Major blunt trauma or rapid acceleration-deceleration may cause dissection, but in patients with inherent arterial wall weakness, dissection can occur spontaneously or as a result of minor neck movement. Cocaine use has been associated with dissection of the aortic arch and coronary and renal arteries through cocaine-mediated hypertension. Recent preclinical studies have suggested, however, that cocaine may cause apoptosis of cells in the vascular wall. In this article, we postulate that cocaine may cause apoptosis of vascular endothelial and/or smooth muscle cells, thus weakening the vascular wall and resulting in a dissection-prone state. We review the literature and propose a biological basis for vasculopathy, vascular dissection, and ischemic stroke in the setting of cocaine use. Further research studies on vascular cells, as well as focused analysis of human pathological material, will be important in providing evidence for or against our hypotheses.

  15. Endothelial dysfunction impairs vascular neurotransmission in tail arteries.

    PubMed

    Sousa, Joana B; Fresco, Paula; Diniz, Carmen

    2015-01-01

    The present study intends to clarify if endothelium dysfunction impairs vascular sympathetic neurotransmission. Electrically-evoked tritium overflow (100 pulses/5 Hz) was evaluated in arteries (intact and denuded) or exhibiting some degree of endothelium dysfunction (spontaneously hypertensive arteries), pre-incubated with [(3)H]-noradrenaline in the presence of enzymes (nitric oxide synthase (NOS); nicotinamide adenine dinucleotide phosphate (NADPH) oxidase; xanthine oxidase; cyclooxygenase; adenosine kinase) inhibitors and a nucleoside transporter inhibitor. Inhibition of endothelial nitric oxide synthase with L-NIO dihydrochloride reduced tritium overflow in intact arteries whereas inhibition of neuronal nitric oxide synthase with Nω-Propyl-L-arginine hydrochloride was devoid of effect showing that only endothelial nitric oxide synthase is involved in vascular sympathetic neuromodulation. Inhibition of enzymes involved in reactive oxygen species or prostaglandins production with apocynin and allopurinol or indomethacin, respectively, failed to alter tritium overflow. A facilitation or reduction of tritium overflow was observed in the presence of 8-cyclopentyl-1,3-dipropylxanthine (DPCPX) or of 5-iodotubericidin, respectively, but only in intact arteries. These effects can be ascribed to a tonic inhibitory effect mediated by A1 receptors. In denuded and hypertensive arteries, 7-(2-phenylethyl)-5-amino-2-(2-furyl)-pyrazolo-[4,3-e]-1,2,4-triazolo[1,5-c] pyrimidine (SCH 58261) reduced tritium overflow, suggesting the occurrence of a tonic activation of A2A receptors. When endogenous adenosine bioavailability was increased by the nucleoside transporter inhibitor, S-(4-Nitrobenzyl)-6-thioinosine, tritium overflow increased in intact, denuded and hypertensive arteries. Among the endothelium-derived substances studied that could alter vascular sympathetic transmission only adenosine/adenosine receptor mediated mechanisms were clearly impaired by endothelium injury/dysfunction.

  16. Association of T Cell and Macrophage Activation with Arterial Vascular Health in HIV.

    PubMed

    Grome, Heather N; Barnett, Louise; Hagar, Cindy C; Harrison, David G; Kalams, Spyros A; Koethe, John R

    2017-02-01

    HIV-infected individuals are at increased risk of cardiovascular disease (CVD), but the arterial vascular functions affected by persistent innate and cellular immune activation are not well described. We assessed the relationship between immunologic and vascular parameters in 70 HIV-infected adults on efavirenz, tenofovir, and emtricitabine with more than 2 years of virologic suppression and no history of CVD. We measured brachial artery flow-mediated dilation (FMD) using ultrasound and circulating intercellular adhesion molecule 1 (ICAM-1) and vascular cell adhesion molecule 1 (VCAM-1) by multiple immunoassay. We also measured circulating naive (CD45RO(-)CCR7(+)CD27(+)), activated (CD38(+) and CD38(+)DR(+)), exhausted (PD1(+)), senescent (CD57(+)), and memory (CD45RO(+)) CD4(+) and CD8(+) T cell subsets by flow cytometry, and macrophage activation markers by ELISA and multiple immunoassay. Regression models were adjusted for age, sex, smoking, duration of antiretroviral therapy (ART), and body mass index. Median age was 45 years (IQR 39, 50), median CD4(+) count 701 cells/μl (IQR 540, 954), and 43% were female. Lower brachial FMD was associated with a higher percentage of activated CD8(+) T cells (p < .01), but not associated with macrophage activation. In contrast, higher ICAM-1 and VCAM-1 were associated with sCD163 (p < = .01 for both), macrophage inflammatory protein-1α (p < = .02 for both), and sCD14 (p = .01 for ICAM-1 only). These findings are consistent with the hypothesis that circulating CD8(+) T cell activation may impair arterial smooth muscle relaxation, while macrophage activation has a role in the expression of endothelial cell proteins involved in immune cell translocation. Both innate and cellular immune activation appear to promote arterial vascular disease in HIV-infected persons on ART using differing mechanisms.

  17. Acetylcholine-induced K+ currents in smooth muscle cells of intact rat small arteries.

    PubMed Central

    Weidelt, T; Boldt, W; Markwardt, F

    1997-01-01

    1. The mechanism of the sustained acetylcholine-induced endothelium-dependent hyperpolarization (EDH) in intact rat small mesenteric arteries prestimulated with noradrenaline (10(-6) M) was investigated by means of the single microelectrode voltage-clamp method. 2. The vascular smooth muscle cells (VSMCs) in this preparation are poorly or even not coupled for the reasons that: (1) the mean input resistance Rlnp of the clamped vascular smooth muscle increases from 120 M omega under control conditions to 440 M omega after application of K+ channel blocking drugs, (2) the voltage relaxation after injection of hyperpolarizing currents has a monoexponential time course and is linearly dependent on Rlnp, and (3) voltage steps induced by current-clamp steps are not transferred to locations in the vascular musculature 120 microns apart from the current injecting microelectrode. 3. Sustained (> 5 min) application of ACh (10(-5) M) hyperpolarized the VSMCs by induction of a hyperpolarizing current. This effect was completely blocked by the inhibitor of the nitric oxide (NO) synthase L-NAME (10(-3) M) but not by the inhibitor of the soluble guanylate cyclase (sGCl) Methylene Blue (MB, 10(-4) M). 4. Application of the NO donor sodium nitroprusside (SNP, 10(-6) M) for more than 5 min mimicked the induction of the endothelium-dependent hyperpolarizing current in vessels with destroyed endothelium. The reversal potential of this current is dependent on the extracellular K+ concentration. The effect of SNP could also not be blocked by MB. 5. The blockers of ATP-dependent and Ca(2+)-dependent K+ channels, glibenclamide (Glb, 10(-5) M) and charybdotoxin (CTX, 5 x 10(-8) M), respectively, blocked a hyperpolarizing current in the VSMCs similar to the ACh- or SNP-induced current. 6. The isolated application of either Glb or CTX did not block the activation of the hyperpolarizing current by SNP. Only the combined administration of Glb and CTX blocked the SNP-induced current completely

  18. Low levels of the reverse transactivator fail to induce target transgene expression in vascular smooth muscle cells.

    PubMed

    Viceconte, Nikenza; McKenna, Tomás; Eriksson, Maria

    2014-01-01

    Hutchinson-Gilford progeria syndrome (HGPS) is a genetic disease with multiple features that are suggestive of premature aging. Most patients with HGPS carry a mutation on one of their copies of the LMNA gene. The LMNA gene encodes the lamin A and lamin C proteins, which are the major proteins of the nuclear lamina. The organs of the cardiovascular system are amongst those that are most severely affected in HGPS, undergoing a progressive depletion of vascular smooth muscle cells, and most children with HGPS die in their early teens from cardio-vascular disease and other complications from atherosclerosis. In this study, we developed a transgenic mouse model based on the tet-ON system to increase the understanding of the molecular mechanisms leading to the most lethal aspect of HGPS. To induce the expression of the most common HGPS mutation, LMNA c.1824C>T; p.G608G, in the vascular smooth muscle cells of the aortic arch and thoracic aorta, we used the previously described reverse tetracycline-controlled transactivator, sm22α-rtTA. However, the expression of the reverse sm22α-transactivator was barely detectable in the arteries, and this low level of expression was not sufficient to induce the expression of the target human lamin A minigene. The results from this study are important because they suggest caution during the use of previously functional transgenic animal models and emphasize the importance of assessing transgene expression over time.

  19. Low Levels of the Reverse Transactivator Fail to Induce Target Transgene Expression in Vascular Smooth Muscle Cells

    PubMed Central

    Viceconte, Nikenza; McKenna, Tomás; Eriksson, Maria

    2014-01-01

    Hutchinson-Gilford progeria syndrome (HGPS) is a genetic disease with multiple features that are suggestive of premature aging. Most patients with HGPS carry a mutation on one of their copies of the LMNA gene. The LMNA gene encodes the lamin A and lamin C proteins, which are the major proteins of the nuclear lamina. The organs of the cardiovascular system are amongst those that are most severely affected in HGPS, undergoing a progressive depletion of vascular smooth muscle cells, and most children with HGPS die in their early teens from cardio-vascular disease and other complications from atherosclerosis. In this study, we developed a transgenic mouse model based on the tet-ON system to increase the understanding of the molecular mechanisms leading to the most lethal aspect of HGPS. To induce the expression of the most common HGPS mutation, LMNA c.1824C>T; p.G608G, in the vascular smooth muscle cells of the aortic arch and thoracic aorta, we used the previously described reverse tetracycline-controlled transactivator, sm22α-rtTA. However, the expression of the reverse sm22α-transactivator was barely detectable in the arteries, and this low level of expression was not sufficient to induce the expression of the target human lamin A minigene. The results from this study are important because they suggest caution during the use of previously functional transgenic animal models and emphasize the importance of assessing transgene expression over time. PMID:25090270

  20. Internal carotid artery occlusion: association with atherosclerotic disease in other arterial beds and vascular risk factors.

    PubMed

    Paraskevas, Kosmas I; Mikhailidis, Dimitri P; Liapis, Christos D

    2007-01-01

    The aim of this article is to investigate the association between internal carotid artery occlusion (ICAO) and the presence of atherosclerotic disease and vascular risk factors. The clinical characteristics and risk factors of 120 patients presenting with ICAO were retrospectively reviewed. All patients (n = 120) had at least 1 of the 4 vascular risk factor (diabetes, smoking, hypercholesterolemia, and hypertension); 2, 3, or all 4 risk factors were present in 14 to 82 of the patients (11.7% to 68.3%), 10 to 39 of the patients (8.3% to 32.5%), and 9 of the patients (7.5%), respectively. A total of 84 patients (70%) with ICAO had disease in at least 1 additional vascular bed (aorta, coronary or lower limb arteries). In addition to ICAO, vascular disease was present in 2 and all 3 of these arterial beds in 42 (35%) and 9 (7.5%) patients, respectively. Furthermore, stenosis or occlusion of the ipsilateral or contralateral vertebral arteries was recorded in 19 of 120 patients (15.8%). Regarding the contralateral carotid artery, 1 patient had bilateral ICAO. One patient had contralateral common carotid artery occlusion, and 1 patient was excluded from the analysis because of surgery to the contralateral carotid artery. Of the remaining 117 patients, 34 (29.0%) had less than 50% contralateral carotid artery stenosis. Thirty-two patients (27.4%) had 50% to 69%, and 51 (43.6%) had 70% to 99% stenosis. Ultrasonographic imaging of the carotid plaque of the contralateral carotid artery revealed that 52 of the 120 arteries (43.3%) were uniformly or predominantly echolucent (types I and II, respectively). Fifty-nine (49.2%) were predominantly or uniformly echogenic (types III and IV), and 9 (7.5%) could not be classified. A similar distribution of echomorphology was observed on the occluded side. ICAO is associated with widespread atherosclerotic disease and a high prevalence of vascular risk factors. Detection of ICAO should prompt the investigation of other arterial beds and

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

    PubMed

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

    1998-12-01

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

  2. Association of miRNA-145 expression in vascular smooth muscle cells with vascular damages in patients with lupus nephritis.

    PubMed

    Ding, Yan; Liao, Wang; Yi, Zhuwen; Xiang, Wei; He, Xiaojie

    2015-01-01

    miRNAs have been found to contribute to the regulation of multiple cellular processes, including cell apoptosis, differentiation and proliferation. The patients with lupus nephritis (LN) exhibit thickened renal vascular membrane and highly proliferative vascular smooth muscle cells (VSMCs). Of various miRNAs discovered, miR-145 is essential to mediate the proliferation of VSMCs and the formation of atherosclerotic plaques. In this study, we studied the pathological and vascular damage of renal LN, and the correlation between miR-145 expression in VSMCs and the vascular damages. Serum, urine, and renal biopsies were obtained from 41 patients with active LN. The serum and urinary VEGF levels were examined to confirm the renal damage of each patient. Biopsies were stained to observe the glomerular segmental lesions, sclerosis, and to evaluate the vascular damages. The expression of miR-145 was also examined to determine the correlation between its expression and the vascular damages. The expression of miR-145 was mainly detected in the renal VSMCs and the epithelial cells of glomerular proximal convoluted tubule. Nevertheless, the expression of miR-145 reduced as the tunicae media vasorum ratios increased, indicating the development of LN inhibits the expression of miR-145. Furthermore, our studies revealed no significant correlation among renal interstitial vascular damage, glomerular damage and severity classification of LN. Therefore, we suggest the damage of renal interstitial vascular should be considered as one of the factors to evaluate the severity of the LN.

  3. Cytoskeletal remodeling in differentiated vascular smooth muscle is actin isoform dependent and stimulus dependent.

    PubMed

    Kim, Hak Rim; Gallant, Cynthia; Leavis, Paul C; Gunst, Susan J; Morgan, Kathleen G

    2008-09-01

    Dynamic remodeling of the actin cytoskeleton plays an essential role in the migration and proliferation of vascular smooth muscle cells. It has been suggested that actin remodeling may also play an important functional role in nonmigrating, nonproliferating differentiated vascular smooth muscle (dVSM). In the present study, we show that contractile agonists increase the net polymerization of actin in dVSM, as measured by the differential ultracentrifugation of vascular smooth muscle tissue and the costaining of single freshly dissociated cells with fluorescent probes specific for globular and filamentous actin. Furthermore, induced alterations of the actin polymerization state, as well as actin decoy peptides, inhibit contractility in a stimulus-dependent manner. Latrunculin pretreatment or actin decoy peptides significantly inhibit contractility induced by a phorbol ester or an alpha-agonist, but these procedures have no effect on contractions induced by KCl. Aorta dVSM expresses alpha-smooth muscle actin, beta-actin, nonmuscle gamma-actin, and smooth muscle gamma-actin. The incorporation of isoform-specific cell-permeant synthetic actin decoy peptides, as well as isoform-specific probing of cell fractions and two-dimensional gels, demonstrates that actin remodeling during alpha-agonist contractions involves the remodeling of primarily gamma-actin and, to a lesser extent, beta-actin. Taken together, these results show that net isoform- and agonist-dependent increases in actin polymerization regulate vascular contractility.

  4. Thrombospondin-1 limits ischemic tissue survival by inhibiting nitric oxide–mediated vascular smooth muscle relaxation

    PubMed Central

    Isenberg, Jeff S.; Hyodo, Fuminori; Matsumoto, Ken-Ichiro; Romeo, Martin J.; Abu-Asab, Mones; Tsokos, Maria; Kuppusamy, Periannan; Wink, David A.; Krishna, Murali C.

    2007-01-01

    The nitric oxide (NO)/cGMP pathway, by relaxing vascular smooth muscle cells, is a major physiologic regulator of tissue perfusion. We now identify thrombospondin-1 as a potent antagonist of NO for regulating F-actin assembly and myosin light chain phosphorylation in vascular smooth muscle cells. Thrombospondin-1 prevents NO-mediated relaxation of precontracted vascular smooth muscle cells in a collagen matrix. Functional magnetic resonance imaging demonstrated that an NO-mediated increase in skeletal muscle perfusion was enhanced in thrombospondin-1–null relative to wild-type mice, implicating endogenous thrombospondin-1 as a physiologic antagonist of NO-mediated vasodilation. Using a random myocutaneous flap model for ischemic injury, tissue survival was significantly enhanced in thrombospondin-1–null mice. Improved flap survival correlated with increased recovery of oxygen levels in the ischemic tissue of thrombospondin-1–null mice as measured by electron paramagnetic resonance oximetry. These findings demonstrate an important antag-onistic relation between NO/cGMP signaling and thrombospondin-1 in vascular smooth muscle cells to regulate vascular tone and tissue perfusion. PMID:17082319

  5. Dual effect of initial [K] on vascular tone in rat mesenteric arteries.

    PubMed

    Brochet, Didier X P; Langton, Philip D

    2006-10-01

    A slight increase in extracellular concentration of potassium ([K(+)](o)) can act as a vasodilator in rat mesenteric vascular bed. However, in recent years, several groups have failed to consistently observe relaxation of rat mesenteric arteries in these conditions. The aim of the present study was to provide a mechanistic understanding of this discrepancy. In rat small mesenteric arteries, 37 of 40 arteries mounted for measurement of isometric force and pre-contracted with phenylephrine (PE) did not relax when ([K(+)](o) was raised from 5.9 mM (control ([K(+)](o) to 11.2 or 21.2 mM. However, when ([K(+)](o) was briefly lowered to 1.2 mM, increasing ([K(+)](o) to between 5.9 and 41.2 mM evoked relaxation. This relaxation was not reduced by barium or by removal of the endothelium, but was abolished by 0.1 mM ouabain. Raising ([K(+)](o) from concentrations between 0 and 5.9 mM to 13.8 mM elicited a relaxation of PE-induced tone that was inversely proportional to initial ([K(+)](o). Relaxation was associated with a ouabain-sensitive hyperpolarization of smooth muscle cells. In arteries exposed to dihydroouabain (DHO), raising ([K(+)](o) from 5.9 to 13.8 mM and simultaneously washing out DHO resulted in relaxation of PE-induced force. These results suggest that only when the initial ([K(+)](o) is less than approximately 5 mM do small elevations in ([K(+)](o) evoke smooth muscle hyperpolarization and relaxation via activation of Na,K-ATPase, and not inwardly rectifying K(+) channels. Therefore, small differences in the initial ([K(+)](o) (4.6 vs 5.9 mM) can strongly influence the variations of vascular tone to increases in ([K(+)](o).

  6. Vascular nanomedicine: Site specific delivery of elastin stabilizing therapeutics to damaged arteries

    NASA Astrophysics Data System (ADS)

    Sinha, Aditi

    Elastin, a structural protein in the extra-cellular matrix, plays a critical role in the normal functioning of blood vessels. Apart from performing its primary function of providing resilience to arteries, it also plays major role in regulating cell-cell and cell-matrix interactions, response to injury, and morphogenesis. Medial arterial calcification (MAC) and abdominal aortic aneurysm (AAA) are two diseases where the structural and functional integrity of elastin is severely compromised. Although the clinical presentation of MAC and AAA differ, they have one common underlying causative mechanism---pathological degradation of elastin. Hence prevention of elastin degradation in the early stages of MAC and AAA can mitigate, partially if not wholly, the fatal consequences of both the diseases. The work presented here is motivated by the overwhelming statistics of people afflicted by elastin associated cardiovascular diseases and the unavailability of cure for the same. Overall goal of our research is to understand role of elastin degradation in cardiovascular diseases and to develop a targeted vascular drug delivery system that is minimally invasive, biodegradable, and non-toxic, that prevents elastin from degradation. Our hope is that such treatment will also help regenerate elastin, thereby providing a multi-fold treatment option for elasto-degenerative vascular diseases. For this purpose, we have first confirmed the combined role of degraded elastin and hyperglycemia in the pathogenesis of MAC. We have shown that in the absence of degraded elastin and TGF-beta1 (abundantly present in diabetic arteries) vascular smooth muscle cells maintain their homeostatic state, regardless of environmental glucose concentrations. However simultaneous exposure to glucose, elastin peptides and TGF-beta1 causes the pathological transgenesis of vascular cells to osteoblast-like cells. We show that plant derived polyphenols bind to vascular elastin with great affinity resulting in

  7. Adrenal Androgen Dehydroepiandrosterone Sulfate Inhibits Vascular Remodeling Following Arterial Injury

    PubMed Central

    Ii, Masaaki; Hoshiga, Masaaki; Negoro, Nobuyuki; Fukui, Ryosuke; Nakakoji, Takahiro; Kohbayashi, Eiko; Shibata, Nobuhiko; Furutama, Daisuke; Ishihara, Tadashi; Hanafusa, Toshiaki; Losordo, Douglas W.; Ohsawa, Nakaaki

    2009-01-01

    Recent epidemiologic studies have suggested that serum dehydroepiandrosterone sulfate (DHEAS) levels have a significant inverse correlation with the incidence of cardiovascular diseases. However, direct evidence for the association with DHEAS and vascular disorders has not yet been explored. DHEAS significantly reduced neointima formation 28 days after surgery without altering other serum metabolite levels in a rabbit carotid balloon injury model. Immunohistochemical analyses revealed the reduction of proliferating cell nuclear antigen (PCNA) index and increase of TdT-mediated dUTP-biotin Nick End Labeling (TUNEL) index, expressing differentiated vascular smooth muscle cell (VSMC) markers in the media 7 days after surgery. In vitro, DHEAS exhibited inhibitory effects on VSMC proliferation and migration activities, inducing G1 cell cycle arrest with upregulation of one of the cyclin dependent kinase (CDK) inhibitors p16INK4a and apoptosis with activating peroxisome proliferator-activated receptor (PPAR)-α in VSMCs. DHEAS inhibits vascular remodeling reducing neointima formation after vascular injury via its effects on VSMC phenotypic modulation, functions and apoptosis upregulating p16INK4a/activating PPARα. DHEAS may play a pathophysiological role for vascular remodeling in cardiovascular disease. PMID:19298964

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

    SciTech Connect

    Helkin, Alex; Maier, Kristopher G.; Gahtan, Vivian

    2015-09-04

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

  9. Uteroplacental insufficiency and lactational environment separately influence arterial stiffness and vascular function in adult male rats.

    PubMed

    Tare, Marianne; Parkington, Helena C; Bubb, Kristen J; Wlodek, Mary E

    2012-08-01

    Early life environmental influences can have lifelong consequences for health, including the risk of cardiovascular disease. Uteroplacental insufficiency causes fetal undernutrition and impairs fetal growth. Previously we have shown that uteroplacental insufficiency is associated with impaired maternal mammary development, compromising postnatal growth leading to hypertension in male rat offspring. In this study we investigated the roles of prenatal and postnatal nutritional environments on endothelial and smooth muscle reactivity and passive wall stiffness of resistance arteries of male rat offspring. Fetal growth restriction was induced by maternal bilateral uterine vessel ligation (restricted) on day 18 of pregnancy. Control offspring were from mothers that had sham surgery (control) and another group from mothers with their litter size reduced (reduced; litter size reduced to 5 at birth, equivalent to the restricted group). On postnatal day 1, offspring (control, restricted, and reduced) were cross-fostered onto control or restricted mothers. At 6 months, mesenteric and femoral arteries were studied using wire and pressure myography. In restricted-on-restricted rats, wall stiffness was increased, and sensitivity to phenylephrine and relaxation evoked by endothelium-derived hyperpolarizing factor and sodium nitroprusside were impaired in mesenteric arteries. In femoral arteries, relaxation to sodium nitroprusside was reduced, whereas wall stiffness was unaltered. Cross-fostering restricted offspring onto control mothers alleviated deficits in vascular stiffness and reactivity. Control or reduced offspring who suckled a restricted mother had marked vascular stiffening. In conclusion, prenatal and early postnatal environments separately influence vascular function and stiffness. Furthermore, the early postnatal lactational environment is a determinant of later cardiovascular function.

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

    PubMed

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

    2001-11-09

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

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

    SciTech Connect

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

    2013-11-15

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

  12. Cellular mechanism through which parathyroid hormone-related protein induces proliferation in arterial smooth muscle cells: definition of an arterial smooth muscle PTHrP/p27kip1 pathway.

    PubMed

    Fiaschi-Taesch, Nathalie; Sicari, Brian M; Ubriani, Kiran; Bigatel, Todd; Takane, Karen K; Cozar-Castellano, Irene; Bisello, Alessandro; Law, Brian; Stewart, Andrew F

    2006-10-27

    Parathyroid hormone-related protein (PTHrP) is present in vascular smooth muscle (VSM), is markedly upregulated in response to arterial injury, is essential for normal VSM proliferation, and also markedly accentuates neointima formation following rat carotid angioplasty. PTHrP contains a nuclear localization signal (NLS) through which it enters the nucleus and leads to marked increases in retinoblastoma protein (pRb) phosphorylation and cell cycle progression. Our goal was to define key cell cycle molecules upstream of pRb that mediate cell cycle acceleration induced by PTHrP. The cyclin D/cdk-4,-6 system and its upstream regulators, the inhibitory kinases (INKs), are not appreciably influenced by PTHrP. In striking contrast, cyclin E/cdk-2 kinase activity is markedly increased by PTHrP, and this is a result of a specific, marked, PTHrP-induced proteasomal degradation of p27(kip1). Adenoviral restoration of p27(kip1) fully reverses PTHrP-induced cell cycle progression, indicating that PTHrP mediates its cell cycle acceleration in VSM via p27(kip1). In confirmation, adenoviral delivery of PTHrP to murine primary vascular smooth muscle cells (VSMCs) significantly decreases p27(kip1) expression and accelerates cell cycle progression. p27(kip1) is well known to be a central cell cycle regulatory molecule involved in both normal and pathological VSM proliferation and is a target of widely used drug-eluting stents. The current observations define a novel "PTHrP/p27(kip1) pathway" in the arterial wall and suggest that this pathway is important in normal arterial biology and a potential target for therapeutic manipulation of the arterial response to injury.

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

    NASA Technical Reports Server (NTRS)

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

    1998-01-01

    Recent studies have demonstrated that vascular smooth muscle cells are responsive to changes in their local hemodynamic environment. The effects of shear stress on the expression of human protease activated receptor-1 (PAR-1) and tissue plasminogen activator (tPA) mRNA and protein were investigated in human aortic smooth muscle cells (HASMCs). Under conditions of low shear stress (5 dyn/cm2), PAR-1 mRNA expression was increased transiently at 2 hours compared with stationary control values, whereas at high shear stress (25 dyn/cm2), mRNA expression was decreased (to 29% of stationary control; P<0.05) at all examined time points (2 to 24 hours). mRNA half-life studies showed that this response was not due to increased mRNA instability. tPA mRNA expression was decreased (to 10% of stationary control; P<0.05) by low shear stress after 12 hours of exposure and was increased (to 250% of stationary control; P<0.05) after 24 hours at high shear stress. The same trends in PAR-1 mRNA levels were observed in rat smooth muscle cells, indicating that the effects of shear stress on human PAR-1 were not species-specific. Flow cytometry and ELISA techniques using rat smooth muscle cells and HASMCs, respectively, provided evidence that shear stress exerted similar effects on cell surface-associated PAR-1 and tPA protein released into the conditioned media. The decrease in PAR-1 mRNA and protein had functional consequences for HASMCs, such as inhibition of [Ca2+] mobilization in response to thrombin stimulation. These data indicate that human PAR-1 and tPA gene expression are regulated differentially by shear stress, in a pattern consistent with their putative roles in several arterial vascular pathologies.

  14. Vascular Smooth Muscle Sirtuin-1 Protects Against Aortic Dissection During Angiotensin II–Induced Hypertension

    PubMed Central

    Fry, Jessica L; Shiraishi, Yasunaga; Turcotte, Raphaël; Yu, Xunjie; Gao, Yuan Z; Akiki, Rachid; Bachschmid, Markus; Zhang, Yanhang; Morgan, Kathleen G; Cohen, Richard A; Seta, Francesca

    2015-01-01

    Background Sirtuin-1 (SirT1), a nicotinamide adenine dinucleotide+–dependent deacetylase, is a key enzyme in the cellular response to metabolic, inflammatory, and oxidative stresses; however, the role of endogenous SirT1 in the vasculature has not been fully elucidated. Our goal was to evaluate the role of vascular smooth muscle SirT1 in the physiological response of the aortic wall to angiotensin II, a potent hypertrophic, oxidant, and inflammatory stimulus. Methods and Results Mice lacking SirT1 in vascular smooth muscle (ie, smooth muscle SirT1 knockout) had drastically high mortality (70%) caused by aortic dissection after angiotensin II infusion (1 mg/kg per day) but not after an equipotent dose of norepinephrine, despite comparable blood pressure increases. Smooth muscle SirT1 knockout mice did not show any abnormal aortic morphology or blood pressure compared with wild-type littermates. Nonetheless, in response to angiotensin II, aortas from smooth muscle SirT1 knockout mice had severely disorganized elastic lamellae with frequent elastin breaks, increased oxidant production, and aortic stiffness compared with angiotensin II–treated wild-type mice. Matrix metalloproteinase expression and activity were increased in the aortas of angiotensin II–treated smooth muscle SirT1 knockout mice and were prevented in mice overexpressing SirT1 in vascular smooth muscle or with use of the oxidant scavenger tempol. Conclusions Endogenous SirT1 in aortic smooth muscle is required to maintain the structural integrity of the aortic wall in response to oxidant and inflammatory stimuli, at least in part, by suppressing oxidant-induced matrix metalloproteinase activity. SirT1 activators could potentially be a novel therapeutic approach to prevent aortic dissection and rupture in patients at risk, such as those with hypertension or genetic disorders, such as Marfan’s syndrome. PMID:26376991

  15. Real-time vascular mechanosensation through ex vivo artery perfusion

    PubMed Central

    2014-01-01

    Background Cell-based perfusion studies have provided great insight into fluid-sensing mechanisms, such as primary cilia in the renal and vascular systems. However, the intrinsic limitations of in vitro cell culture, such as the inability to reflect cellular organization within tissues, has distanced observed paradigms from possible clinical developments. Here we describe a protocol that applies ex vivo artery perfusion and calcium imaging to observe real-time cellular responses to fluid-shear stress. Results Through our ex vivo artery perfusion method, we were able to simulate physiological flow and initiate distinct fluid shear stress mechanosensory responses, as well as induced acetylcholine responses in mouse aortic tissue. The observed calcium profiles confirm results found through previous in vitro cell culture experiments. The overall procedure, including dissection, sample preparation and perfusion, takes around 3 hours to complete. Conclusion Through our unique method, we are able to induce laminar flow within intact mouse aortic tissue and illicit subsequent cellular responses. This method of ex vivo artery perfusion provides the opportunity to bridge the novel findings of in vitro studies with subsequent physiological models of fluid-shear stress mechanosensation in vascular tissues. PMID:24685068

  16. Signalling pathways activated by 5-HT(1B)/5-HT(1D) receptors in native smooth muscle and primary cultures of rabbit renal artery smooth muscle cells.

    PubMed

    Hinton, J M; Hill, P; Jeremy, J; Garland, C

    2000-01-01

    The potential of primary cultures of rabbit renal artery vascular smooth muscle cells (VSMCs) was assessed as a means to investigate the signalling pathways linked to 5-hydroxytryptamine (5-HT) 5-HT(1B)/5-HT(1D) receptors in native arteries. In renal artery segments denuded of endothelium, incubated with ketanserin and prazosin (each 1 microM), and prestimulated with 20 mM K(+) Krebs buffer, 5-HT and CP 93,129, a 5-HT(1B) receptor agonist, evoked concentration-dependent contractions. GR 127935, a 5-HT(1B)/5-HT(1D) receptor antagonist, significantly antagonised 5-HT-evoked contractions at nanomolar concentrations. Reverse transcription polymerase chain reaction (RT-PCR) of mRNA from smooth muscle cells from the isolated renal artery and from primary cultures of VSMCs from the same artery expressed mRNA transcripts for the 5-HT(1B) receptor and the 5-HT(1D) receptor in both preparations. The sequence of the PCR fragments corresponded to the known sequence for these receptors. Application of 5-HT evoked a concentration-dependent, pertussis toxin (PTx)-sensitive reduction in cyclic AMP in both cultured cells and intact artery (cyclic AMP concentration reduced by 65.53 +/- 3.33 and 52.65 +/- 5.34% from basal with 10 microM 5-HT, respectively). The effect of 10 microM 5-HT on cAMP was increased in the presence of 20 mM K(+) (reduced by 82.50 +/- 2.50 and 87.54 +/- 3.97%, respectively). In intact arteries, contraction through 5-HT(1B)/5-HT(1D) receptors was significantly attenuated by inhibitors of phosphatidylinositol 3-kinase (wortmannin) and activated mitogen-activated protein kinase (MAPK), MEK (U0126). In the cultured VSMCs, activated MAPK was identified by immunocytochemistry and immunoblotting after stimulation with 5-HT, but only if 20 mM K(+) was present at the onset of stimulation. These data provide the first direct evidence that 5-HT(1B)/5-HT(1B) receptors are linked to the activation of MAPK and indicate that primary cultures of renal VSMCs could provide a

  17. Coronary Artery Disease Associated Transcription Factor TCF21 Regulates Smooth Muscle Precursor Cells That Contribute to the Fibrous Cap

    PubMed Central

    Raiesdana, Azad; Kundu, Ramendra; Miller, Clint L.; Kim, Juyong B.; Arora, Komal; Carcamo-Oribe, Ivan; Xiong, Yiqin; Tellakula, Nikhil; Nanda, Vivek; Murthy, Nikitha; Boisvert, William A.; Hedin, Ulf; Perisic, Ljubica; Aldi, Silvia; Maegdefessel, Lars; Pjanic, Milos; Owens, Gary K.; Tallquist, Michelle D.; Quertermous, Thomas

    2015-01-01

    Recent genome wide association studies have identified a number of genes that contribute to the risk for coronary heart disease. One such gene, TCF21, encodes a basic-helix-loop-helix transcription factor believed to serve a critical role in the development of epicardial progenitor cells that give rise to coronary artery smooth muscle cells (SMC) and cardiac fibroblasts. Using reporter gene and immunolocalization studies with mouse and human tissues we have found that vascular TCF21 expression in the adult is restricted primarily to adventitial cells associated with coronary arteries and also medial SMC in the proximal aorta of mouse. Genome wide RNA-Seq studies in human coronary artery SMC (HCASMC) with siRNA knockdown found a number of putative TCF21 downstream pathways identified by enrichment of terms related to CAD, including “vascular disease,” “disorder of artery,” and “occlusion of artery,” as well as disease-related cellular functions including “cellular movement” and “cellular growth and proliferation.” In vitro studies in HCASMC demonstrated that TCF21 expression promotes proliferation and migration and inhibits SMC lineage marker expression. Detailed in situ expression studies with reporter gene and lineage tracing revealed that vascular wall cells expressing Tcf21 before disease initiation migrate into vascular lesions of ApoE-/- and Ldlr-/- mice. While Tcf21 lineage traced cells are distributed throughout the early lesions, in mature lesions they contribute to the formation of a subcapsular layer of cells, and others become associated with the fibrous cap. The lineage traced fibrous cap cells activate expression of SMC markers and growth factor receptor genes. Taken together, these data suggest that TCF21 may have a role regulating the differentiation state of SMC precursor cells that migrate into vascular lesions and contribute to the fibrous cap and more broadly, in view of the association of this gene with human CAD, provide

  18. Statins inhibited erythropoietin-induced proliferation of rat vascular smooth muscle cells.

    PubMed

    Kaneda, Tae; Tsuruoka, Shuichi; Fujimura, Akio

    2010-12-15

    Erythropoietin (EPO) directly stimulates the proliferation of vascular smooth muscle cells, and this is believed to be one of the mechanisms of vascular access failure of hemodialysis patients. However, precise mechanisms of the EPO-induced proliferation of vascular smooth muscle cells are not certain. HMG-CoA reductase inhibitors (statins) are primarily used to reduce cholesterol levels, but also exert other effects, including reno-protective effects. We evaluated the effect of several statins with various hydrophilicities on the EPO-induced proliferation of primary cultured rat vascular smooth muscle cells (VSMCs) in vitro. EPO significantly and concentration-dependently increased DNA synthesis as assessed by [³H]thymidine incorporation, cell proliferation as assessed by WST-1 assay, and activation of the p44/42MAPK pathway. Therapeutic doses of statins (pravastatin, simvastatin, atorvastatin and fluvastatin) in patients with hypercholesterolemia almost completely suppressed all of the EPO-induced effects in a concentration-dependent manner. Co-addition of mevalonic acid almost completely reversed the effects of statins. Statin alone did not affect the basal proliferation capacity of the cells. The effects were almost similar among the statins. We concluded that statins inhibited EPO-induced proliferation in rat VSMCs at least partly through their inhibition of HMG-CoA reductase activity. In the future, statins might prove useful for the treatment of EPO-induced hyperplasia of vascular access. Because the statins all showed comparable effects irrespective of their hydrophilicities, these effects might be a class effect.

  19. Phosphate binders prevent phosphate-induced cellular senescence of vascular smooth muscle cells and vascular calcification in a modified, adenine-based uremic rat model.

    PubMed

    Yamada, S; Tatsumoto, N; Tokumoto, M; Noguchi, H; Ooboshi, H; Kitazono, T; Tsuruya, K

    2015-04-01

    Clinical and experimental studies have reported that phosphate overload plays a central role in the pathogenesis of vascular calcification in chronic kidney disease. However, it remains undetermined whether phosphate induces cellular senescence during vascular calcification. We established a modified uremic rat model induced by a diet containing 0.3% adenine that showed more slowly progressive kidney failure, more robust vascular calcification, and longer survival than the conventional model (0.75% adenine). To determine the effect of phosphate on senescence of vascular smooth muscle cells (VSMCs) and the protective effect of phosphate binders, rats were divided into four groups: (1) normal control rats; (2) rats fed with the modified adenine-based diet (CKD); (3) CKD rats treated with 6% lanthanum carbonate (CKD-LaC); and (4) CKD rats treated with 6% calcium carbonate (CKD-CaC). After 8 weeks, CKD rats showed circumferential arterial medial calcification, which was inhibited in CKD-LaC and CKD-CaC rats. CKD rats showed increased protein expression of senescence-associated β-galactosidase, bone-related proteins, p16 and p21, and increased oxidative stress levels in the calcified area, which were inhibited by both phosphate binders. However, serum levels of oxidative stress and inflammatory markers, serum fibroblast growth factor 23, and aortic calcium content in CKD-CaC rats were higher than those in CKD-LaC rats. In conclusion, phosphate induces cellular senescence of VSMCs in the modified uremic rat model, and phosphate binders can prevent both cellular senescence and calcification of VSMCs via phosphate unloading. Our modified adenine-based uremic rat model is useful for evaluating uremia-related complications, including vascular calcification.

  20. Alteration of Contractile Function and Calcium Ion Movements in Vascular Smooth Muscle by Gentamicin and Other Aminoglycoside Antibiotics

    PubMed Central

    Adams, H. Richard; Goodman, Frank R.; Weiss, George B.

    1974-01-01

    Experiments were conducted to examine the effects of certain aminoglycoside antibiotics on contractile responses and related calcium ion (Ca2+) movements in isolated vascular smooth muscle. Gentamicin, kanamycin, and streptomycin decreased contractile responses produced by norepinephrine, histamine, and high K+ in rabbit aortic strips. The inhibitory action of these antibiotics on mechanical function was more pronounced when the Ca2+ concentration of the bathing solution was decreased from 1.5 mM (normal Ca2+ solution) to 0.05 mM (low Ca2+ solution). The uptake of radiocalcium (45Ca) into the isolated media-intimal layer of rabbit aortae was decreased in a maintained manner by each antibiotic. With gentamicin, the inhibitory effect on 45Ca uptake was shown to be dependent upon the concentration of gentamicin employed and to be more evident in a 0.1 mM Ca2+ solution than in a normal Ca2+ solution. In addition, the rate of 45Ca efflux from the rabbit aortic media-intimal layer was increased in a sustained manner by gentamicin, streptomycin, and kanamycin. Furthermore, contractile responses induced by high K+ and norepinephrine in canine carotid arterial strips were inhibited by gentamicin. Present findings indicate that aminoglycoside antibiotics interfere with Ca2+-linked events leading to activation of the contractile mechanism of vascular smooth muscle. These in vitro findings may partially explain the occurrence of in vivo cardiovascular depression that has occasionally been observed after the administration of chemically related antimicrobial agents. PMID:15825418

  1. Targeted gene transfection from microbubbles into vascular smooth muscle cells using focused, ultrasound-mediated delivery

    PubMed Central

    Phillips, Linsey C.; Klibanov, Alexander L.; Wamhoff, Brian R.; Hossack, John A.

    2010-01-01

    We investigate a method for gene delivery to vascular smooth muscle cells using ultrasound triggered delivery of plasmid DNA from electrostatically coupled cationic microbubbles. Microbubbles carrying reporter plasmid DNA were acoustically ruptured in the vicinity of smooth muscle cells in vitro under a range of acoustic pressures (0–950 kPa) and pulse durations (0–100 cycles). No effect on gene transfection or viability was observed from application of microbubbles, DNA, or ultrasound alone. Microbubbles in combination with ultrasound (500 kPa, 1MHz, 50 cycle bursts at a Pulse Repetition Frequency [PRF] of 100 Hz) significantly reduced viability both with DNA (53 +/− 27%) and without (19 +/− 8%). Maximal gene transfection (~1% of cells) occurred using 50 cycle, 1 MHz pulses at 300 kPa which resulted in 40% viability of cells. We demonstrated that we can locally deliver DNA to vascular smooth muscle cells in vitro using microbubble carriers and focused ultrasound. PMID:20800174

  2. Histone deacetylase inhibitors promote eNOS expression in vascular smooth muscle cells and suppress hypoxia-induced cell growth.

    PubMed

    Tan, Xiaoling; Feng, Lan; Huang, Xiaoyong; Yang, Yidong; Yang, Chengzhong; Gao, Yuqi

    2017-03-07

    Hypoxia stimulates excessive growth of vascular smooth muscle cells (VSMCs) contributing to vascular remodelling. Recent studies have shown that histone deacetylase inhibitors (HDIs) suppress VSMC proliferation and activate eNOS expression. However, the effects of HDI on hypoxia-induced VSMC growth and the role of activated eNOS in VSMCs are unclear. Using an EdU incorporation assay and flow cytometry analysis, we found that the HDIs, butyrate (Bur) and suberoylanilide hydroxamic acid (SAHA) significantly suppressed the proliferation of hypoxic VSMC lines and induced apoptosis. Remarkable induction of cleaved caspase 3, p21 expression and reduction of PCNA expression were also observed. Increased eNOS expression and enhanced NO secretion by hypoxic VSMC lines were detected using Bur or SAHA treatment. Knockdown of eNOS by siRNA transfection or exposure of hypoxic VSMCs to NO scavengers weakened the effects of Bur and SAHA on the growth of hypoxic VSMCs. In animal experiments, administration of Bur to Wistar rats exposed to hypobaric hypoxia for 28 days ameliorated the thickness and collagen deposition in pulmonary artery walls. Although the mean pulmonary arterial pressure (mPAP) was not obviously decreased with Bur in hypoxic rats, right ventricle hypertrophy index (RVHI) was decreased and the oxygen partial pressure of arterial blood was elevated. Furthermore, cell viability was decreased and eNOS and cleaved caspase 3 were induced in HDI-treated rat pulmonary arterial SMCs. These findings imply that HDIs prevent hypoxia-induced VSMC growth, in correlation with activated eNOS expression and activity in hypoxic VSMCs.

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

    PubMed

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

    2014-04-01

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

  4. Endothelial cells are progenitors of cardiac pericytes and vascular smooth muscle cells

    PubMed Central

    Chen, Qi; Zhang, Hui; Liu, Yang; Adams, Susanne; Eilken, Hanna; Stehling, Martin; Corada, Monica; Dejana, Elisabetta; Zhou, Bin; Adams, Ralf H.

    2016-01-01

    Mural cells of the vessel wall, namely pericytes and vascular smooth muscle cells, are essential for vascular integrity. The developmental sources of these cells and molecular mechanisms controlling their progenitors in the heart are only partially understood. Here we show that endocardial endothelial cells are progenitors of pericytes and vascular smooth muscle cells in the murine embryonic heart. Endocardial cells undergo endothelial–mesenchymal transition and convert into primitive mesenchymal progenitors expressing the platelet-derived growth factor receptors, PDGFRα and PDGFRβ. These progenitors migrate into the myocardium, differentiate and assemble the wall of coronary vessels, which requires canonical Wnt signalling involving Frizzled4, β-catenin and endothelial cell-derived Wnt ligands. Our findings identify a novel and unexpected population of progenitors for coronary mural cells with potential relevance for heart function and disease conditions. PMID:27516371

  5. Kindlin-2 siRNA inhibits vascular smooth muscle cell proliferation, migration and intimal hyperplasia via Wnt signaling.

    PubMed

    Wu, Xiaolin; Liu, Wenwei; Jiang, Hong; Chen, Jing; Wang, Jichun; Zhu, Rui; Li, Bin

    2016-02-01

    It is known that vascular smooth muscle cell (VSMC) proliferation and migration leads to intimal hyperplasia in cases of atherosclerosis and restenosis. In the present study, we investigated the effects of kindlin-2 on VSMC proliferation, migration and intimal hyperplasia, and the underlying mechanisms. The left common carotid artery of Sprague‑Dawley rats were subjected to balloon injury in order to induce intimal hyperplasia, and then transfected with kindlin-2 small interfering RNA (siRNA) lentivirus or negative control siRNA lentivirus. We noted that the degree of intimal hyperplasia 4 weeks after balloon injury was significantly reduced in arteries transfected with kindlin-2 siRNA lentivirus (P<0.05). In vitro, kindlin-2 siRNA suppressed VSMC proliferation and migration induced by Wnt3a (100 ng/ml). Western blot analyses and RT-qPCR revealed that kindlin-2 regulated Wnt/β-catenin signaling and thereby modulated the expression of β-catenin target genes, including c-myc and cyclin D1. This study demonstrated that kindlin-2 plays a critical role in VSMC proliferation, migration and intimal hyperplasia via Wnt signaling. Therefore, blocking the activity of kindlin-2 represents a novel therapeutic strategy for vascular injury.

  6. Distribution of a lanthanide (147 Pm) in vascular smooth muscle.

    PubMed

    Weiss, G B; Goodman, F R

    1976-08-01

    In order to ascertain whether trivalent rare earth ions such as lanthanum (La+++) penetrate the cell membrane under physiological conditions, the extracellular and cellular distribution of promethium (147 Pm), a carrier-free rare earth radioisotope, was examined in rabbit aortic smooth muscle. As the duration of incubation was lengthened, uptake of 147Pm continued to increase; it was inhibited by La+++ and other rare earth ions (Nd+++, Lu+++) only when the 147 Pm/rare earth concentration ratio exceeded 1:10(6). However, equally high concentrations of Ca++ had no effect on 147Pm uptake. Efflux of 147Pm was only transiently increased by 1.5 mM La+++, and exposure to 0.05 mM EDTA elicited an increased 147Pm efflux with both transient and maintained components. The magnitude of the EDTA-induced increase in 147 Pm efflux was similar over a 30-fold range of EDTA concentration (0.05-1.5 mM); the limiting factor for 147Pm efflux is the rate of 147Pm desorption from the tissue rather than the extracellular concentration of EDTA. Loss of 147Pm in the presence of 0.05 mM EDTA could be described in terms of two specific washout components (the more rapid of which included 147Pm within the extracellular space and the slower of which had half-times of washout of approximately 7-10 minutes). Uptake of 147Pm was inhibited by lowering the incubation solution temperature to 0 degrees C or by procaine. However, concentrations of metabolic inhibitors (iodoacetate and dinitrophenol) which diminish loss of Ca++ from the cell did not decrease either the uptake or efflux of 147Pm. Thus, significant quantities of 147Pm do not appear to be accumulated within the cell or transported out of the cell; distribution of 147Pm can be most simply described in terms of a binding at and desorption from surface acessible fiber sites.

  7. Metformin inhibits inflammatory response via AMPK-PTEN pathway in vascular smooth muscle cells.

    PubMed

    Kim, Sun Ae; Choi, Hyoung Chul

    2012-09-07

    Atherosclerosis is a chronic inflammation of the coronary arteries. Vascular smooth muscle cells (VSMCs) stimulated by cytokines and chemokines accelerate the inflammatory response and migrate to the injured endothelium during the progression of atherosclerosis. Activation of AMP activated protein kinase (AMPK), a key sensor maintaining metabolic homeostasis, suppresses the inflammatory response. However, how AMPK regulates the inflammatory response is poorly understood. To identify the mechanism of this response, we focused on phosphatase and tensin homolog (PTEN), which is a negative regulator of inflammation. We investigated that activation of AMPK-induced PTEN expression and suppression of the inflammatory response through the AMPK-PTEN pathway in VSMCs. We treated with the well-known AMPK activator metformin to induce PTEN expression. PTEN was induced by metformin (2mM) and inhibited by compound C (10 μM) and AMPK siRNA. Tumor necrosis factor-alpha (TNF-α) was used to induce inflammation. The inflammatory response was confirmed by cyclooxygenase (COX)-2, inducible nitric oxide synthase (iNOS) expression, and activation of nuclear factor (NF)-κB. Metformin suppressed COX-2 and iNOS mRNA and protein expression dose dependently. Treatment with compound C and bpv (pic) in the presence of metformin, iNOS and COX-2 protein expression increased. NF-κB activation decreased in response to metformin and was restored by inhibiting AMPK and PTEN. Inhibiting AMPK and PTEN restored ROS levels stimulated with TNF-α. Taken together, PTEN could be a possible downstream regulator of AMPK, and the AMPK-PTEN pathway might be important in the regulation of the inflammatory response in VSMCs.

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

    PubMed

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

    2016-08-24

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

  9. Cinnamaldehyde inhibits L-type calcium channels in mouse ventricular cardiomyocytes and vascular smooth muscle cells.

    PubMed

    Alvarez-Collazo, Julio; Alonso-Carbajo, Lucía; López-Medina, Ana I; Alpizar, Yeranddy A; Tajada, Sendoa; Nilius, Bernd; Voets, Thomas; López-López, José Ramón; Talavera, Karel; Pérez-García, María Teresa; Alvarez, Julio L

    2014-11-01

    Cinnamaldehyde (CA), a major component of cinnamon, is known to have important actions in the cardiovascular system, including vasorelaxation and decrease in blood pressure. Although CA-induced activation of the chemosensory cation channel TRPA1 seems to be involved in these phenomena, it has been shown that genetic ablation of Trpa1 is insufficient to abolish CA effects. Here, we confirm that CA relaxes rat aortic rings and report that it has negative inotropic and chronotropic effects on isolated mouse hearts. Considering the major role of L-type Ca(2+) channels in the control of the vascular tone and cardiac contraction, we used whole-cell patch-clamp to test whether CA affects L-type Ca(2+) currents in mouse ventricular cardiomyocytes (VCM, with Ca(2+) as charge carrier) and in mesenteric artery smooth muscle cells (VSMC, with Ba(2+) as charge carrier). We found that CA inhibited L-type currents in both cell types in a concentration-dependent manner, with little voltage-dependent effects. However, CA was more potent in VCM than in VSMC and caused opposite effects on the rate of inactivation. We found these divergences to be at least in part due to the use of different charge carriers. We conclude that CA inhibits L-type Ca(2+) channels and that this effect may contribute to its vasorelaxing action. Importantly, our results demonstrate that TRPA1 is not a specific target of CA and indicate that the inhibition of voltage-gated Ca(2+) channels should be taken into account when using CA to probe the pathophysiological roles of TRPA1.

  10. Osteopontin expression in vascular smooth muscle cells in patients with end-stage renal disease.

    PubMed

    Nakamura, Hironori; Honda, Hirokazu; Inada, Yoshifumi; Kato, Noriyuki; Kato, Kenichi; Kitazawa, Kozo; Sugisaki, Tetsuzo

    2006-06-01

    beta-glycerophosphate, a phosphate donor, and uremic sera induce osteopontin (OPN) expression in bovine vascular smooth muscle cells (VSMCs). However, the correlations of serum phosphorus level with OPN expression, and blood urea nitrogen (BUN) level with OPN expression in humans have not previously been reported. The purpose of the current study is to compare the expression of OPN in VSMCs with clinical data in patients with end-stage renal disease (ESRD). The radial arteries of 33 patients (21 male and 12 female patients) were examined to determine the expression of OPN and collagen type I (Col I) by immunohistochemistry. The correlation of the expression of bone matrix proteins with clinical data was analyzed. Between the low-serum phosphorus (<6 mg/dL) group and high-serum phosphorus (> or =6 mg/dL) group, significant differences were detected in the expression of OPN (P = 0.0049) and the levels of BUN (P = 0.0005), serum phosphorus (P < 0.0001) and calcium x phosphorus products (P < 0.0001). Moreover, between the low-BUN (<70 mg/dL, N = 19) group and high-BUN (> or =70 mg/dL) group, significant differences were detected in the expression of OPN (P = 0.0039) and the levels of BUN (P = 0.0002), serum phosphorus (P = 0.0002) and calcium x phosphorus products (P = 0.0003). We have shown that hyperphosphatemia or azotemia is associated with the expression of OPN in VSMCs in patients with ESRD.

  11. Localisation of members of the vascular endothelial growth factor (VEGF) family and their receptors in human atherosclerotic arteries

    PubMed Central

    Belgore, F; Blann, A; Neil, D; Ahmed, A S; Lip, G Y H

    2004-01-01

    Background: Vascular endothelial growth factor (VEGF) mediates endothelial cell mitogenesis and enhances vascular permeability. The existence of single or multiple VEGF isoforms and receptors suggests that these proteins may have overlapping but distinct functions, which may be reflected in their cell expression and distribution. Methods: The localisation of VEGFs A–C and their receptors (VEGFRs 1–3, respectively) in 30 fresh human atherosclerotic arteries, 15 normal uterine arteries, and 15 saphenous veins using immunohistochemistry and western blotting. Results: Saphenous veins showed no staining for VEGF-B or VEGFR-2. Smooth muscle cells (SMCs) showed the strongest staining for VEGF-A, VEGF-B, VEGFR-1, and VEGFR-2 in all specimens. Conversely, VEGFR-3 and VEGF-C were predominately localised to the endothelial vasa vasorum in normal arteries, whereas medial SMCs showed the strongest staining in atherosclerotic arteries. Western blotting showed variations in VEGF protein localisation, with lower amounts of VEGF-B and VEGF-C in saphenous veins, compared with arterial tissue. Amounts of VEGF-C were lower than those of VEGF-A and VEGF-B in all specimens. Conclusion: This study provides direct evidence of the presence of VEGF proteins and receptors in human physiology and pathology, with variations in both the amounts of VEGF proteins expressed and their cellular distribution in normal arteries compared with atherosclerotic arteries. The presence of VEGFs A–C and their receptors in normal arterial tissue implies that VEGF functions may extend beyond endothelial cell proliferation. Reduced VEGFR-2 staining in atherosclerotic arteries may have implications for the atherosclerosis process and the development of vascular disease and its complications. PMID:14990597

  12. Current Trends in Heparin Use During Arterial Vascular Interventional Radiology

    SciTech Connect

    Durran, Alexandra C.; Watts, Christopher

    2012-12-15

    Purpose: This study was designed to assess the current use of heparinized saline and bolus doses of heparin in non-neurological interventional radiology and to determine whether consensus could be reached to produce guidance for heparin use during arterial vascular intervention. Methods: An interactive electronic questionnaire was distributed to members of the British Society of Interventional Radiology regarding their current practice in the use, dosage, and timing of heparin boluses and heparinized flushing solutions.ResultsA total of 108 completed questionnaires were received. More than 80% of respondents used heparinized saline with varying concentrations; the most prevalent was 1,000 IU/l (international units of heparin per liter) and 5,000 IU/l. Fifty-one percent of interventionalists use 3,000 IU as their standard bolus dose; however, the respondents were split regarding the timing of bolus dose with {approx}60% administering it after arterial access is obtained and 40% after crossing the lesion. There was no consensus on altering dose according to body weight, and only 4% monitored clotting parameters. Conclusions: There seems to be some coherence among practicing interventionalists regarding heparin administration. We hypothesize that heparinized saline should be used at a recognized standard concentration of 1,000 IU/l as a flushing concentration in all arterial vascular interventions and that 3,000 IU bolus is considered the standard dose for straightforward therapeutic procedures and 5000 IU for complex, crural, and endovascular aneurysm repair work. The bolus should be given after arterial access is obtained to allow time for optimal anticoagulation to be achieved by the time of active intervention and stenting. Further research into clotting abnormalities following such interventional procedures would be an interesting quantifiable follow-up to this initial survey of opinions and practice.

  13. Fibroblast growth factor stimulates angiotensin converting enzyme expression in vascular smooth muscle cells. Possible mediator of the response to vascular injury.

    PubMed Central

    Fishel, R S; Thourani, V; Eisenberg, S J; Shai, S Y; Corson, M A; Nabel, E G; Bernstein, K E; Berk, B C

    1995-01-01

    Angiotensin converting enzyme (ACE) activity contributes to the vascular response to injury because ACE inhibition limits neointima formation in rat carotid arteries after balloon injury. To investigate the mechanisms by which ACE may contribute to vascular smooth muscle cell (VSMC) proliferation, we studied expression of ACE in vivo after injury and in vitro after growth factor stimulation. ACE activity 14 d after injury was increased 3.6-fold in the injured vessel. ACE expression, measured by immunohistochemistry, became apparent at 7 d in the neointima and at 14 d was primarily in the most luminal neointimal cells. To characterize hormones that induce ACE in vivo, cultured VSMC were exposed to steroids and growth factors. Among steroids, only glucocorticoids stimulated ACE expression with an 8.0 +/- 2.1-fold increase in activity and a 6.5-fold increase in mRNA (30 nM dexamethasone for 72 h). Among growth factors tested, only fibroblast growth factor (FGF) stimulated ACE expression (4.2 +/- 0.7-fold increase in activity and 1.6-fold increase in mRNA in response to 10 ng/ml FGF for 24 h). Dexamethasone and FGF were synergistic at the indicated concentrations inducing 50.6 +/- 12.4-fold and 32.5-fold increases in activity and mRNA expression, respectively. In addition, when porcine iliac arteries were transfected with recombinant FGF-1 (in the absence of injury), ACE expression increased in neointimal VSMC, to the same extent as injured, nontransfected arteries. The data suggest a temporal sequence for the response to injury in which FGF induces ACE, ACE generates angiotensin II, and angiotensin II stimulates VSMC growth in concert with FGF. Images PMID:7814638

  14. Morphology and Ploidy of Smooth Muscle Cells in Chorionic Arteries under Different Hemodynamic Conditions.

    PubMed

    Gansburgskii, A N; Yal'tsev, A V

    2017-02-01

    Smooth muscle cells from the arterial wall of placental chorion were studied at 39-40-week gestation. The content of mono- and binuclear tetraploid myocytes was higher in sites of arterial branching and turns (27.3% vs. 4.4% straight parts of the arteries; DNA cytophotometry data). Mitoses were found only in these arterial regions (0.18%). Regional changes in the sizes of diploid and polyploid myocytes were detected, associated with the blood flow pattern in the chorion; myocyte hypertrophy was 17-fold more incident in sites of arterial turns and branching than in straight arteries. Possible causes of changes in the proliferative characteristics and subsequent growth of the chorionic arterial wall myocytes are discussed.

  15. Decreased vascular smooth muscle cell density in medial degeneration of human abdominal aortic aneurysms.

    PubMed Central

    López-Candales, A.; Holmes, D. R.; Liao, S.; Scott, M. J.; Wickline, S. A.; Thompson, R. W.

    1997-01-01

    Abdominal aortic aneurysms (AAAs) are characterized by structural deterioration of the aortic wall leading to progressive aortic dilatation and eventual rupture. The histopathological changes in AAAs are particularly evident within the elastic media, which is normally dominated by vascular smooth muscle cells (SMCs). To determine whether a decrease in vascular SMCs contributes to medial degeneration, we measured SMC density in 21 normal and pathological human abdominal aortic tissue specimens using immunohistochemistry for alpha-SMC actin and direct cell counts (medial SMCs per high-power field (HPF)). Medial SMC density was not significantly different between normal aorta (n = 5; 199.5 +/- 14.9 SMCs/HPF) and atherosclerotic occlusive disease (n = 6; 176.4 +/- 13.9 SMCs/HPF), but it was reduced by 74% in AAA (n = 10; 50.9 +/- 6.1 SMCs/HPF; P < 0.01 versus normal aorta). Light and electron microscopy revealed no evidence of overt cellular necrosis, but SMCs in AAAs exhibited ultrastructural changes consistent with apoptosis. Using in situ end-labeling (ISEL) of fragmented DNA to detect apoptotic cells, up to 30% of aortic wall cells were ISEL positive in AAAs. By double-labeling techniques, many of these cells were alpha-actin-positive SMCs distributed throughout the degenerative media. In contrast, ISEL-positive cells were observed only within the intimal plaque in atherosclerotic occlusive disease. The amount of p53 protein detected by immunoblotting was increased nearly fourfold in AAA compared with normal aorta and atherosclerotic occlusive disease (P < 0.01), and immunoreactive p53 was localized to lymphocytes and residual SMCs in the aneurysm wall. Using reverse transcription polymerase chain reaction assays a substantial amount of p53 mRNA expression was observed in AAAs. These results demonstrate that medial SMC density is significantly decreased in human AAA tissues associated with evidence of SMC apoptosis and increased production of p53, a potential

  16. Silencing of osterix expression by siRNA inhibits aldosterone‑induced calcification of vascular smooth muscle cells in mice.

    PubMed

    Gong, Yan-Chun; He, Yue; Wang, Hao; Niu, Wen-Quan; Ji, Kai-Da; Li, Hua

    2016-09-01

    The process of vascular calcification shares numerous similarities with that of skeletal mineralization and involves the deposition of hydroxyapatite crystals in arteries and cardiac valves. However, the underlying cellular mechanism remains to be fully elucidated. Microarray analysis in the present study demonstrated that greater than 2,000 genes were upregulated during the calcification of murine vascular smooth muscle cells (VSMCs), of which osterix (OSX) and integrin‑binding sialoprotein (IBSP) were the most significantly differentially expressed genes. Following the validation of increased OSX and IBSP expression by reverse transcription‑quantitative polymerase chain reaction in calcifying murine VSMCs induced by aldosterone. Subsequent to transfection with siRNA‑OSX, results indicated that OSX may inhibit calcification of VSMCs via IBSP. It was suggested that the increased OSX expression in calcifying VSMCs may reflect the well‑established prenatal role of OSX. A full understanding of the importance of OSX in this pathological process would improve understanding of the pathogenesis of vascular calcification.

  17. G-Protein-Coupled Receptor 35 Mediates Human Saphenous Vein Vascular Smooth Muscle Cell Migration and Endothelial Cell Proliferation

    PubMed Central

    McCallum, Jennifer E.; Mackenzie, Amanda E.; Divorty, Nina; Clarke, Carolyn; Delles, Christian; Milligan, Graeme; Nicklin, Stuart A.

    2016-01-01

    Vascular smooth muscle cell (VSMC) migration and proliferation is central to neointima formation in vein graft failure following coronary artery bypass. However, there are currently no pharmacological interventions that prevent vein graft failure through intimal occlusion. It is hence a therapeutic target. Here, we investigated the contribution of GPR35 to human VSMC and endothelial cell (EC) migration, using a scratch-wound assay, and also the contribution to proliferation, using MTS and BrdU assays, in in vitro models using recently characterized human GPR35 ortholog-selective small-molecule agonists and antagonists. Real-time PCR studies showed GPR35 to be robustly expressed in human VSMCs and ECs. Stimulation of GPR35, with either the human-selective agonist pamoic acid or the reference agonist zaprinast, promoted VSMC migration in the scratch-wound assay. These effects were blocked by coincubation with either of the human GPR35-specific antagonists, CID-2745687 or ML-145. These GPR35-mediated effects were produced by inducing alterations in the actin cytoskeleton via the Rho A/Rho kinase signaling axis. Additionally, the agonist ligands stimulated a proliferative response in ECs. These studies highlight the potential that small molecules that stimulate or block GPR35 activity can modulate vascular proliferation and migration. These data propose GPR35 as a translational therapeutic target in vascular remodeling. PMID:27064272

  18. Heparin modulates the composition of the extracellular matrix domain surrounding arterial smooth muscle cells.

    PubMed Central

    Snow, A. D.; Bolender, R. P.; Wight, T. N.; Clowes, A. W.

    1990-01-01

    Heparin and related molecules influence vascular wall structure by their ability to inhibit smooth muscle cell (smc) proliferation and migration. However, little is known as to whether heparin has an effect on the extracellular matrix. In the present study, the effect of heparin on the content and regional distribution of elastin, collagen, and proteoglycans (PGs) in blood vessels following experimental injury was determined. Two groups of rats were subjected to left common carotid balloon injury and were infused with either 0.9% saline or heparin in a saline solution, for 2 weeks. Using a new morphometric method of analysis, the authors determined changes in volumes of elastin, collagen, and PGs contained within an 'extracellular matrix domain (ECM domain),' the average envelope of connective tissue surrounding each smc. Heparin treatment inhibited intimal thickening and decreased the elastin content in the ECM domain in the upper and lower arterial intima. Collagen also was found to be significantly decreased 5.0-fold and 7.6-fold in the ECM domains of upper and lower intima, respectively, of heparin-treated animals. The decrease in both elastin and collagen was balanced by a significant increase in amorphous and filamentous electron-dense material. Heparin also caused a significant 1.8-fold and 1.9-fold increase in the PG content in the ECM domain in the upper and lower intima, respectively. Immunohistochemical analysis, using antibodies to elastin and PG subclasses, supported the morphometric observations. This study has shown that heparin administered in vivo can alter the accumulation and distribution of each of the major vascular ECM components in a specific and differential manner. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 PMID:2386199

  19. Phospholipase D signaling in serotonin-induced mitogenesis of pulmonary artery smooth muscle cells.

    PubMed

    Liu, Y; Fanburg, B L

    2008-09-01

    We have previously reported the participation of mitogen-activated protein, Rho, and phosphoinositide-3 (PI3) kinases in separate pathways in serotonin (5-HT)-induced proliferation of pulmonary artery smooth muscle cells (SMCs). In this study, we investigated the possible participation of phospholipase D (PLD) and phosphatidic acid (PA) in this growth process. 5-HT stimulated a time-dependent increase in [(3)H]phosphatidylbutanol and PA generation. Exposure of SMCs to 1-butanol or overexpression of an inactive mutant of human PLD1R898R blocked 5-HT-induced proliferation. Furthermore, 1-butanol inhibited 5-HT activation of S6K1 and S6 protein, downstream effectors of mammalian target of rapamycin (mTOR), by 80 and 72%, respectively, and partially blocked activation of extracellular signal-regulated kinase (ERK) by 30% but had no effect on other associated signaling pathways. Exogenous PA caused cellular proliferation and revitalized cyclin D1 expression by 5-HT of the 1-butanol-treated cells. PA also reproduced activations by 5-HT of mTOR, S6K1, and ERK. Transfection with inactive human PLD1 reduced 5-HT-induced activation of S6K1 by approximately 50%. Inhibition of 5-HT receptor 2A (R 2A) with ketaserin blocked PLD activation by 5-HT. Inhibition with PI3-kinase inhibitor failed to block either activation of PLD by 5-HT or PA-dependent S6K1 phosphorylation. Taken together, these results indicate that ligation of the 5-HTR 2A by 5-HT initiates PLD activation in SMCs, and that its product, PA, is an early signaling molecule in 5-HT-induced pulmonary artery SMC proliferation. Signaling by PA produces its downstream effects primarily through the mTOR/S6K1 pathway and to a lesser extent through the ERK pathway. Hydrolysis of cell membrane lipid may be important in vascular effects of 5-HT.

  20. Orai channel-mediated Ca2+ signals in vascular and airway smooth muscle

    PubMed Central

    Spinelli, Amy M.

    2016-01-01

    Orai (Orai1, Orai2, and Orai3) proteins form a family of highly Ca2+-selective plasma membrane channels that are regulated by stromal-interacting molecules (STIM1 and STIM2); STIM proteins are Ca2+ sensors located in the membrane of the endoplasmic reticulum. STIM and Orai proteins are expressed in vascular and airway smooth muscle and constitute the molecular components of the ubiquitous store-operated Ca2+ entry pathway that mediate the Ca2+ release-activated Ca2+ current. STIM/Orai proteins also encode store-independent Ca2+ entry pathways in smooth muscle. Altered expression and function of STIM/Orai proteins have been linked to vascular and airway pathologies, including restenosis, hypertension, and atopic asthma. In this review we discuss our current understanding of Orai proteins and the store-dependent and -independent signaling pathways mediated by these proteins in vascular and airway smooth muscle. We also discuss the current studies linking altered expression and function of Orai proteins with smooth muscle-related pathologies. PMID:26718630

  1. MicroRNA-182 prevents vascular smooth muscle cell dedifferentiation via FGF9/PDGFRβ signaling

    PubMed Central

    Dong, Nana; Wang, Wei; Tian, Jinwei; Xie, Zulong; Lv, Bo; Dai, Jiannan; Jiang, Rui; Huang, Dan; Fang, Shaohong; Tian, Jiangtian; Li, Hulun; Yu, Bo

    2017-01-01

    The abnormal phenotypic transformation of vascular smooth muscle cells (SMCs) causes various proliferative vascular diseases. MicroRNAs (miRNAs or miRs) have been established to play important roles in SMC biology and phenotypic modulation. This study revealed that the expression of miR-182 was markedly altered during rat vascular SMC phenotypic transformation in vitro. We aimed to investigate the role of miR-182 in the vascular SMC phenotypic switch and to determine the potential molecular mechanisms involved. The expression of miR-182 gene was significantly downregulated in cultured SMCs during dedifferentiation from a contractile to a synthetic phenotype. Conversely, the upregulation of miR-182 increased the expression of SMC-specific contractile genes, such as α-smooth muscle actin, smooth muscle 22α and calponin. Additionally, miR-182 overexpression potently inhibited SMC proliferation and migration under both basal conditions and under platelet-derived growth factor-BB stimulation. Furthermore, we identified fibroblast growth factor 9 (FGF9) as the target gene of miR-182 for the phenotypic modulation of SMCs mediated through platelet-derived growth factor receptor β (PDGFRβ) signaling. These data suggest that miR-182 may be a novel SMC phenotypic marker and a modulator that may be used to prevent SMC dedifferentiation via FGF9/PDGFRβ signaling. PMID:28259995

  2. Mutations in Smooth Muscle Alpha-Actin (ACTA2) Cause Coronary Artery Disease, Stroke, and Moyamoya Disease, Along with Thoracic Aortic Disease

    PubMed Central

    Guo, Dong-Chuan; Papke, Christina L.; Tran-Fadulu, Van; Regalado, Ellen S.; Avidan, Nili; Johnson, Ralph Jay; Kim, Dong H.; Pannu, Hariyadarshi; Willing, Marcia C.; Sparks, Elizabeth; Pyeritz, Reed E.; Singh, Michael N.; Dalman, Ronald L.; Grotta, James C.; Marian, Ali J.; Boerwinkle, Eric A.; Frazier, Lorraine Q.; LeMaire, Scott A.; Coselli, Joseph S.; Estrera, Anthony L.; Safi, Hazim J.; Veeraraghavan, Sudha; Muzny, Donna M.; Wheeler, David A.; Willerson, James T.; Yu, Robert K.; Shete, Sanjay S.; Scherer, Steven E.; Raman, C.S.; Buja, L. Maximilian; Milewicz, Dianna M.

    2009-01-01

    The vascular smooth muscle cell (SMC)-specific isoform of α-actin (ACTA2) is a major component of the contractile apparatus in SMCs located throughout the arterial system. Heterozygous ACTA2 mutations cause familial thoracic aortic aneurysms and dissections (TAAD), but only half of mutation carriers have aortic disease. Linkage analysis and association studies of individuals in 20 families with ACTA2 mutations indicate that mutation carriers can have a diversity of vascular diseases, including premature onset of coronary artery disease (CAD) and premature ischemic strokes (including Moyamoya disease [MMD]), as well as previously defined TAAD. Sequencing of DNA from patients with nonfamilial TAAD and from premature-onset CAD patients independently identified ACTA2 mutations in these patients and premature onset strokes in family members with ACTA2 mutations. Vascular pathology and analysis of explanted SMCs and myofibroblasts from patients harboring ACTA2 suggested that increased proliferation of SMCs contributed to occlusive diseases. These results indicate that heterozygous ACTA2 mutations predispose patients to a variety of diffuse and diverse vascular diseases, including TAAD, premature CAD, ischemic strokes, and MMD. These data demonstrate that diffuse vascular diseases resulting from either occluded or enlarged arteries can be caused by mutations in a single gene and have direct implications for clinical management and research on familial vascular diseases. PMID:19409525

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

    SciTech Connect

    Sahni, Abha; Wang, Nadan; Alexis, Jeffrey

    2013-02-15

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

  4. Secreted Protein Acidic and Rich in Cysteine Modulates Molecular Arterial Homeostasis of Human Arterial Smooth Muscle Cells In Vitro.

    PubMed

    Ye, Geng-Fan; Zhu, Shao-Wei; Zhu, Shu-Gan; Li, Feng; Wang, Yun-Yan

    2016-12-01

    Secreted protein acidic and rich in cysteine (SPARC) is widely expressed in the vascular smooth muscle cells (VSMCs) of human intracranial aneurysms (IAs), but the effect and underlying mechanism of SPARC on VSMCs during the formation and progression of IAs needs to be probed. Human umbilical arterial smooth muscle cells (HUASMCs) were treated with a gradient concentrations of SPARC in vitro for different time. Cell counting kit-8 (CCK-8) assay, cell cycle, and cell apoptosis were used to investigate the effect of SPARC on HUASMCs. After exposure to 2 and 4 μg/ml SPARC, cell viability were 89.3 ± 2.00 %, and 87.57 ± 2.17 % (P < 0.05 vs. control), respectively. Induced by 2 μg/ml SPARC, the proportion of cells in G0/G1 phase was 74.77 ± 1.33 % (P < 0.05 vs. control), and the early and late apoptosis ratio were 7.38 ± 1.25 % and 4.86 ± 0.81 % (P < 0.01 vs. control), respectively. After exposure to 2 μg/ml SPARC for 2, 6, 12, 24, and 48 h, Western blot analysis showed that the protein level of p21 was upregulated significantly at 2-12 h (P < 0.05 vs. control), while the expression of p53 remained stable within 48 h. The expression of Bax protein increased markedly and peaked at 24 (P < 0.01 vs. control), while Bcl2 protein decreased significantly at 48 h (P < 0.01 vs. control). Cleaved caspase3 was also upregulated dramatically and peaked at 24 h (P < 0.05 vs. control). The protein level of MMP2 increased significantly and peaked at 24 h (P < 0.01 vs. control), while TIMP2 remained stable and even reduced at 48 h (P < 0.05 vs. control). Taken together, SPARC could arrest HUASMCs in G0/G1 phase by overexpression of p21 and induce mitochondria-mediated apoptosis in vitro, which could result in the decreased cell viability. Besides, SPARC might also lead to the activation of MMP2 instead of MMP9. These results indicated SPARC could reduce the self-repair capability and increase injury of media layer and internal elastic

  5. Preliminary Experience with Vascular Plugs for Parent Artery Occlusion of the Carotid or Vertebral Arteries

    PubMed Central

    Lee, Woosung; Shin, Yong Sam; Kim, Kyung Hyun; Kim, Yong Bae; Hong, Chang-Ki

    2016-01-01

    Objective The purpose of this study was to report the authors' preliminary experience using the Amplatzer Vascular Plug (AVP) (St. Jude Medical, Plymouth, MN, USA) for parent artery occlusion of the internal carotid artery (ICA) or vertebral artery (VA). Materials and Methods Between September 2008 and December 2015, we performed 52 therapeutic parent artery occlusions (PAOs) by an endovascular technique. Among them, 10 patients underwent PAO of the carotid or vertebral arteries using AVPs. Clinical and radiographic data of these patients were retrospectively reviewed. Results The devices were used for VA dissection that presented with subarachnoid hemorrhage (SAH) in five patients, traumatic arteriovenous fistula (AVF) in two patients, spontaneous AVF in one patient, recurrence of carotid-cavernous fistula (CCF) in one patient, and symptomatic unruptured giant ICA aneurysm in one patient. The devices were used in conjunction with detachable and/or pushable coils and in the extracranial segments of the ICA or VA. Complete occlusion of the parent artery was achieved in all patients. There was one intra-procedural rupture of the VA dissection during coiling prior to using the device. Conclusion Results from the current series suggest that the AVP might be used for therapeutic PAO in the extracranial segments of the ICA or VA. PMID:27847763

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

    SciTech Connect

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

    2007-11-16

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

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

    PubMed

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

    2016-03-04

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

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

    NASA Astrophysics Data System (ADS)

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

    2012-02-01

    We investigated the influences on smooth muscle cells after our novel short-duration thermal angioplasty, Photo-thermo Dynamic Balloon Angioplasty (PTDBA), to reveal the mechanism that can suppress neo-intimal hyperplasia after PTDBA. We obtained the sufficient arterial dilatations by short-duration heating (<=15 s, <70°C) and low dilatation pressure (<0.4 MPa) without arterial injuries in our previous in vivo studies. Smooth muscle cells, which play most important role in chronic treatment effects, were heated during PTDBA and stretch-fixed after PTDBA. The dead cell rate by heating, estimated by Arrhenius equation with A=2.5x1016 s-1 and Ea=1.17×105 J mol-1, was 15.7+/-2.2% after PTDBA. The measured deformation rate of smooth muscle cells' nuclei was 1.6+/-0.1 after PTDBA in vivo. We found that the expression of smooth muscle cells' growth factor after PTDBA was inhibited 0.52 fold compared to that after the conventional balloon angioplasty in vivo. The measured neo-intimal hyperplasia occupancy rate was less than 20% after PTDBA in vivo. We prospect that the inhibition of the growth factor's expression by stretch-fixing may result to suppress the neo-intimal hyperplasia. In addition, the decrease of smooth muscle cells' density in the vessel media by heating might be another reason for the neo-intimal hyperplasia suppression.

  9. Vascular narrowing in pulmonary arterial hypertension is heterogeneous: rethinking resistance.

    PubMed

    Rol, Nina; Timmer, Esther M; Faes, Theo J C; Noordegraaf, Anton Vonk; Grünberg, Katrien; Bogaard, Harm-Jan; Westerhof, Nico

    2017-03-01

    In idiopathic pulmonary arterial hypertension (PAH), increased pulmonary vascular resistance is associated with structural narrowing of small (resistance) vessels and increased vascular tone. Current information on pulmonary vascular remodeling is mostly limited to averaged increases in wall thickness, but information on number of vessels affected and internal diameter decreases for vessels of different sizes is limited. Our aim was to quantify numbers of affected vessels and their internal diameter decrease for differently sized vessels in PAH in comparison with non-PAH patients. Internal and external diameters of transversally cut vessels were measured in five control subjects and six PAH patients. Resistance vessels were classified in Strahler orders, internal diameters 13 μm (order 1) to 500 μm (order 8). The number fraction, that is, percentage of affected vessels, and the internal diameter fraction, that is, percentage diameter of normal diameter, were calculated. In PAH, not all resistance vessels are affected. The number fraction is about 30%, that is, 70% of vessels have diameters not different from vessels of control subjects. Within each order, the decrease in diameter of affected vessels is variable with an averaged diameter fraction of 50-70%. Narrowing of resistance vessels is heterogeneous: not all vessels are narrowed, and the decrease in internal diameters, even within a single order, vary largely. This heterogeneous narrowing alone cannot explain the large resistance increase in PAH We suggest that rarefaction could be an important contributor to the hemodynamic changes.

  10. Emerging roles for vascular smooth muscle cell exosomes in calcification and coagulation.

    PubMed

    Kapustin, A N; Shanahan, C M

    2016-06-01

    Vascular smooth muscle cell (VSMC) phenotypic conversion from a contractile to 'synthetic' state contributes to vascular pathologies including restenosis, atherosclerosis and vascular calcification. We have recently found that the secretion of exosomes is a feature of 'synthetic' VSMCs and that exosomes are novel players in vascular repair processes as well as pathological vascular thrombosis and calcification. Pro-inflammatory cytokines and growth factors as well as mineral imbalance stimulate exosome secretion by VSMCs, most likely by the activation of sphingomyelin phosphodiesterase 3 (SMPD3) and cytoskeletal remodelling. Calcium stress induces dramatic changes in VSMC exosome composition and accumulation of phosphatidylserine (PS), annexin A6 and matrix metalloproteinase-2, which converts exosomes into a nidus for calcification. In addition, by presenting PS, VSMC exosomes can also provide the catalytic surface for the activation of coagulation factors. Recent data showing that VSMC exosomes are loaded with proteins and miRNA regulating cell adhesion and migration highlight VSMC exosomes as potentially important communication messengers in vascular repair. Thus, the identification of signalling pathways regulating VSMC exosome secretion, including activation of SMPD3 and cytoskeletal rearrangements, opens up novel avenues for a deeper understanding of vascular remodelling processes.

  11. Attenuation of Chondrogenic Transformation in Vascular Smooth Muscle by Dietary Quercetin in the MGP-Deficient Mouse Model

    PubMed Central

    Borras, Teresa; Nurminskaya, Maria

    2013-01-01

    Rationale Cartilaginous metaplasia of vascular smooth muscle (VSM) is characteristic for arterial calcification in diabetes and uremia and in the background of genetic alterations in matrix Gla protein (MGP). A better understanding of the molecular details of this process is critical for the development of novel therapeutic approaches to VSM transformation and arterial calcification. Objective This study aimed to identify the effects of bioflavonoid quercetin on chondrogenic transformation and calcification of VSM in the MGP-null mouse model and upon TGF-β3 stimulation in vitro, and to characterize the associated alterations in cell signaling. Methods and Results Molecular analysis revealed activation of β-catenin signaling in cartilaginous metaplasia in Mgp-/- aortae in vivo and during chondrogenic transformation of VSMCs in vitro. Quercetin intercepted chondrogenic transformation of VSM and blocked activation of β-catenin both in vivo and in vitro. Although dietary quercetin drastically attenuated calcifying cartilaginous metaplasia in Mgp-/- animals, approximately one-half of total vascular calcium mineral remained as depositions along elastic lamellae. Conclusion Quercetin is potent in preventing VSM chondrogenic transformation caused by diverse stimuli. Combined with the demonstrated efficiency of dietary quercetin in preventing ectopic chondrogenesis in the MGP-null vasculature, these findings indicate a potentially broad therapeutic applicability of this safe for human consumption bioflavonoid in the therapy of cardiovascular conditions linked to cartilaginous metaplasia of VSM. Elastocalcinosis is a major component of MGP-null vascular disease and is controlled by a mechanism different from chondrogenic transformation of VSM and not sensitive to quercetin. PMID:24098781

  12. Gene Expressions Underlying Mishandled Calcium Clearance and Elevated Generation of Reactive Oxygen Species in the Coronary Artery Smooth Muscle Cells of Chronic Heart Failure Rats

    PubMed Central

    Ding, Liang; Su, Xian-Xiu; Zhang, Wen-Hui; Xu, Yu-Xiang; Pan, Xue-Feng

    2017-01-01

    Background: The calcium clearance and reactive oxygen species (ROS) generations in the coronary artery smooth muscle cells in chronic heart failure (HF) have not been fully investigated. Therefore, we attempted to understand the gene expressions underlying the mishandling of calcium clearance and the accumulations of ROS. Methods: We initially established an animal model of chronic HF by making the left anterior descending coronary artery ligation (CAL) in rats, and then isolated the coronary artery vascular smooth muscle cells from the ischemic and the nonischemic parts of the coronary artery vessels in 12 weeks after CAL operation. The intracellular calcium concentration and ROS level were measured using flow cytometry, and the gene expressions of sarco/endoplasmic reticulum Ca2+-ATPase (SERCA2a), encoding sarcoplasmic reticulum Ca2+-ATPase 2a, encoding sodium-calcium exchanger (NCX), and p47phox encoding a subunit of the nicotinamide adenine dinucleotide phosphate (NADPH) oxidase were examined using real-time quantitative reverse transcription polymerase chain reaction and Western blotting, respectively. Results: We found that the calcium accumulation and ROS generation in the coronary artery smooth muscle cells isolated from either the ischemic or the nonischemic part of the CAL coronary artery vessel were significantly increased irrespective of blood supply (all P < 0.01). Moreover, these were accompanied by the increased expressions of NCX and p47phox, the decreased expression of SERCA2a, and the increased amount of phosphorylated forms of p47phox in NADPH oxidase (all P < 0.05). Conclusions: Our results demonstrated that the disordered calcium clearance and the increased ROS generation occurred in the coronary artery smooth muscle cells in rats with chronic HF produced by ligation of the left anterior descending coronary artery (CAL), and which was found to be disassociated from blood supply, and the increased generation of ROS in the cells was found to make

  13. Loss of prolyl hydroxylase domain protein 2 in vascular endothelium increases pericyte coverage and promotes pulmonary arterial remodeling

    PubMed Central

    Xie, Xue-Jiao; Tao, Yong-Kang; He, Xiaochen; Roman, Richard J.; Aschner, Judy L.; Chen, Jian-Xiong

    2016-01-01

    Pulmonary arterial hypertension (PAH) is a leading cause of heart failure. Although pulmonary endothelial dysfunction plays a crucial role in the progression of the PAH, the underlying mechanisms are poorly understood. The HIF-α hydroxylase system is a key player in the regulation of vascular remodeling. Knockout of HIF-2α has been reported to cause pulmonary hypertension. The present study examined the role of endothelial cell specific prolyl hydroxylase-2 (PHD2) in the development of PAH and pulmonary vascular remodeling. The PHD2f/f mouse was crossbred with VE-Cadherin-Cre promoter mouse to generate an endothelial specific PHD2 knockout (Cdh5-Cre-PHD2ECKO) mouse. Pulmonary arterial pressure and the size of the right ventricle was significantly elevated in the PHD2ECKO mice relative to the PHD2f/f controls. Knockout of PHD2 in EC was associated with vascular remodeling, as evidenced by an increase in pulmonary arterial media to lumen ratio and number of muscularized arterioles. The pericyte coverage and vascular smooth muscle cells were also significantly increased in the PA. The increase in vascular pericytes was associated with elevated expression of fibroblast specific protein-1 (FSP-1). Moreover, perivascular interstitial fibrosis of pulmonary arteries was significantly increased in the PHD2ECKO mice. Mechanistically, knockout of PHD2 in EC increased the expression of Notch3 and transforming growth factor (TGF-β) in the lung tissue. We conclude that the expression of PHD2 in endothelial cells plays a critical role in preventing pulmonary arterial remodeling in mice. Increased Notch3/TGF-β signaling and excessive pericyte coverage may be contributing to the development of PAH following deletion of endothelial PHD2. PMID:27613846

  14. Pharmacological evidence for a novel cysteinyl-leukotriene receptor subtype in human pulmonary artery smooth muscle

    PubMed Central

    Walch, Laurence; Norel, Xavier; Bäck, Magnus; Gascard, Jean-Pierre; Dahlén, Sven-Erik; Brink, Charles

    2002-01-01

    To characterize the cysteinyl-leukotriene receptors (CysLT receptors) in isolated human pulmonary arteries, ring preparations were contracted with leukotriene C4 (LTC4) and leukotriene D4 (LTD4) in either the absence or presence of the selective CysLT1 receptor antagonists, ICI 198615, MK 571 or the dual CysLT1/CysLT2 receptor antagonist, BAY u9773. Since the contractions induced by the cysteinyl-leukotrienes (cysLTs) in intact preparations failed to attain a plateau response over the concentration range studied, the endothelium was removed and the tissue treated continuously with indomethacin (Rubbed+INDO). In these latter preparations, the pEC50 for LTC4 and LTD4 were not significantly different (7.61±0.07, n=20 and 7.96±0.09, n=22, respectively). However, the LTC4 and LTD4 contractions were markedly potentiated when compared with data from intact tissues. Leukotriene E4 (LTE4) did not contract human isolated pulmonary arterial preparations. In addition, treatment of preparations with LTE4 (1 μM; 30 min) did not modify either the LTC4 or LTD4 contractions. Treatment of preparations with the S-conjugated glutathione (S-hexyl-GSH; 100 μM, 30 min), an inhibitor of the metabolism of LTC4 to LTD4, did not modify LTC4 contractions. The pEC50 values for LTC4 were significantly reduced by treatment of the preparations with either ICI 198615, MK 571 or BAY u9773 and the pKB values were: 7.20, 7.02 and 6.26, respectively. In contrast, these antagonists did not modify the LTD4 pEC50 values. These findings suggest the presence of two CysLT receptors on human pulmonary arterial vascular smooth muscle. A CysLT1 receptor with a low affinity for CysLT1 antagonists and a novel CysLT receptor subtype, both responsible for vasoconstriction. Activation of this latter receptor by LTC4 and LTD4 induced a contractile response which was resistant to the selective CysLT1 antagonists (ICI 198615 and MK 571) as well as the non-selective (CysLT1/CysLT2) antagonist, BAY u9773. PMID

  15. Design and utilization of macrophage and vascular smooth muscle cell co-culture systems in atherosclerotic cardiovascular disease investigation.

    PubMed

    Zuniga, Mary C; White, Sharla L Powell; Zhou, Wei

    2014-10-01

    Atherosclerotic cardiovascular disease has been acknowledged as a chronic inflammatory condition. Monocytes and macrophages lead the inflammatory pathology of atherosclerosis whereas changes in atheromatous plaque thickness and matrix composition are attributed to vascular smooth muscle cells. Because these cell types are key players in atherosclerosis progression, it is crucial to utilize a reliable system to investigate their interaction. In vitro co-culture systems are useful platforms to study specific molecular mechanisms between cells. This review aims to summarize the various co-culture models that have been developed to investigate vascular smooth muscle cell and monocyte/macrophage interactions, focusing on the monocyte/macrophage effects on vascular smooth muscle cell function.

  16. Voltage-dependent effects of barnidipine in rat vascular smooth muscle.

    PubMed

    Wegener, J W; Korstanje, C; Nawrath, H

    2003-08-01

    The effects of the dihydropyridine nifedipine and its more lipophilic congener, barnidipine, were investigated in smooth muscle preparations from the rat in resting and depolarizing conditions. Both drugs relaxed precontracted aortic rings more potently in depolarizing conditions, barnidipine being more potent than nifedipine. Currents through Ca2+ channels in rat vascular smooth muscle cells (A7r5) and in isolated rat cardiomyocytes were reduced more potently by both drugs at a holding potential of -40 mV than at -80 mV. However, barnidipine and nifedipine were more effective in reducing the current in A7r5 cells than in cardiomyocytes. The IC(50) obtained in aortic rings and in A7r5 cells were similar for barnidipine but an order of magnitude different for nifedipine. The results show that, in depolarizing conditions, barnidipine was more effective than nifedipine. It is suggested that the higher potency of barnidipine acting in vascular smooth muscle is related to both a higher affinity to the inactivated state of vascular Ca2+ channels and to a more lipophilic property as compared with nifedipine.

  17. Role of Na+-K+ ATPase in cyclic GMP-mediated relaxation of canine pulmonary artery smooth muscle cells

    PubMed Central

    Tamaoki, J; Tagaya, E; Nishimura, K; Isono, K; Nagai, A

    1997-01-01

    Sodium-potassium adenosine triphosphatase (Na+-K+ ATPase) plays a role in the regulation of vascular tone, but contribution of this enzyme to nitrovasodilator-induced pulmonary vasodilatation remains uncertain. We thus studied the interaction between guanosine 3′:5′-cyclic monophosphate (cyclic GMP) and Na+-K+ ATPase in smooth muscle cells isolated from canine pulmonary artery. To assess the contractile properties, changes in smooth muscle cell length were determined microscopically. Application of potassium chloride (KCl) shortened the cell length, an effect which was reduced by sodium nitroprusside and 8-bromo-cyclic GMP in a concentration-dependent manner. Pretreatment of cells with the cyclic GMP-dependent kinase inhibitor KT 5823 (2 μM) abolished the effects of sodium nitroprusside and 8-bromo-cyclic GMP. Ouabain (0.3 μM) did not alter the KCl-induced muscle shortening, but inhibited the relaxant responses to sodium nitroprusside and 8-bromo-cyclic GMP. Incubation of smooth muscle cells with sodium nitroprusside concentration-dependently increased intracellular cyclic GMP levels and ouabain-sensitive 86Rb uptake, and these values were significantly correlated. In the presence of KT 5823, sodium nitroprusside increased cyclic GMP levels but did not alter ouabain-sensitive 86Rb uptake. These results suggest that there is a link between accumulation of intracellular cyclic GMP and activation of sarcolemmal Na+-K+ ATPase in pulmonary artery smooth muscle cells and that this link may be involved in the sodium nitroprusside-induced pulmonary vasodilatation. PMID:9298536

  18. Deletion of mineralocorticoid receptors in smooth muscle cells blunts renal vascular resistance following acute cyclosporine administration

    PubMed Central

    Amador, Cristian A.; Bertocchio, Jean-Philippe; Andre-Gregoire, Gwennan; Placier, Sandrine; Van Huyen, Jean-Paul Duong; El Moghrabi, Soumaya; Berger, Stefan; Warnock, David G.; Chatziantoniou, Christos; Jaffe, Iris Z.; Rieu, Philippe; Jaisser, Frederic

    2016-01-01

    Calcineurin inhibitors such as cyclosporine A (CsA) are still commonly used after renal transplantation, despite CsA–induced nephrotoxicity (CIN), which is partly related to vasoactive mechanisms. The mineralocorticoid receptor (MR) is now recognized as a key player in the control of vascular tone, and both endothelial cell- and vascular smooth muscle cell (SMC)-MR modulate the vasoactive responses to vasodilators and vasoconstrictors. Here we tested whether vascular MR is involved in renal hemodynamic changes induced by CsA. The relative contribution of vascular MR in acute CsA treatment was evaluated using mouse models with targeted deletion of MR in endothelial cell or SMC. Results indicate that MR expressed in SMC, but not in endothelium, contributes to the increase of plasma urea and creatinine, the appearance of isometric tubular vacuolization, and overexpression of a kidney injury biomarker (neutrophil gelatinase–associated lipocalin) after CsA treatment. Inactivation of MR in SMC blunted CsA–induced phosphorylation of contractile proteins. Finally, the in vivo increase of renal vascular resistance induced by CsA was blunted when MR was deleted from SMC cells, and this was associated with decreased L-type Ca2+ channel activity. Thus, our study provides new insights into the role of vascular MR in renal hemodynamics during acute CIN, and provides rationale for clinical studies of MR antagonism to manage the side effects of calcineurin inhibitors. PMID:26422501

  19. Procontractile G protein–mediated signaling pathways antagonistically regulate smooth muscle differentiation in vascular remodeling

    PubMed Central

    Althoff, Till F.; Juárez, Julián Albarrán; Troidl, Kerstin; Tang, Cong; Wang, Shengpeng; Wirth, Angela; Takefuji, Mikito; Wettschureck, Nina

    2012-01-01

    Vascular smooth muscle (Sm) cells (VSMCs) are highly plastic. Their differentiation state can be regulated by serum response factor (SRF), which activates genes involved in Sm differentiation and proliferation by recruiting cofactors, such as members of the myocardin family and ternary complex factors (TCFs), respectively. However, the extracellular cues and upstream signaling mechanisms regulating SRF-dependent VSMC differentiation under in vivo conditions are poorly understood. In this study, we show that the procontractile signaling pathways mediated by the G proteins G12/G13 and Gq/G11 antagonistically regulate VSMC plasticity in different models of vascular remodeling. In mice lacking Gα12/Gα13 or their effector, the RhoGEF protein LARG, RhoA-dependent SRF-regulation was blocked and down-regulation of VSMC differentiation marker genes was enhanced. This was accompanied by an excessive vascular remodeling and exacerbation of atherosclerosis. In contrast, Sm-specific Gαq/Gα11 deficiency blocked activation of extracellular signal-regulated kinase 1/2 and the TCF Elk-1, resulting in a reduced VSMC dedifferentiation in response to flow cessation or vascular injury. These data show that the balanced activity of both G protein–mediated pathways in VSMCs is required for an appropriate vessel remodeling response in vascular diseases and suggest new approaches to modulate Sm differentiation in vascular pathologies. PMID:23129751

  20. The effect of deuterium oxide (D sub 2 O) on in vitro vascular smooth muscle contraction

    SciTech Connect

    McWilliam, T.M.; Liepins, A.; Rankin, A.J. )

    1990-02-26

    Deuterium oxide (D{sub 2}O), a stable nonradioactive isotope of water, has been demonstrated to reduce L-type calcium channel conductance in isolated myocytes. Since the concentration of intracellular free calcium has been implicated in the mechanism of vascular smooth muscle contraction, the authors investigated whether it inhibits contraction of vascular smooth muscle. Phenylephrine concentration-contraction curves were carried out in the rat aortic ring preparation to determine whether D{sub 2}O inhibits contraction of rat aorta induced through activation of receptor-operated calcium channels. D{sub 2}O depressed these response curves in a concentration dependent manner with 50% inhibition of maximum contraction observed with 60% D{sub 2}O; this effect proved to be reversible and non-toxic. D{sub 2}O also depressed potassium chloride curves, demonstrating an effect on voltage-operated calcium channels. Since vascular endothelium releases endothelium-derived relaxing factor (EDRF) when stimulated by a range of pharmacological agents, it was examined whether the endothelium has a role in these actions of D{sub 2}O on vascular contraction. Mechanical disruption of the endothelium had no effect.

  1. Function and regulation of large conductance Ca(2+)-activated K+ channel in vascular smooth muscle cells.

    PubMed

    Hu, Xiang-Qun; Zhang, Lubo

    2012-09-01

    Large conductance Ca(2+)-activated K(+) (BK(Ca)) channels are abundantly expressed in vascular smooth muscle cells. Activation of BK(Ca) channels leads to hyperpolarization of cell membrane, which in turn counteracts vasoconstriction. Therefore, BK(Ca) channels have an important role in regulation of vascular tone and blood pressure. The activity of BK(Ca) channels is subject to modulation by various factors. Furthermore, the function of BK(Ca) channels are altered in both physiological and pathophysiological conditions, such as pregnancy, hypertension and diabetes, which has dramatic impacts on vascular tone and hemodynamics. Consequently, compounds and genetic manipulation that alter activity and expression of the channel might be of therapeutic interest.

  2. Monocyte-expressed urokinase inhibits vascular smooth muscle cell growth by activating Stat1.

    PubMed

    Kunigal, Sateesh; Kusch, Angelika; Tkachuk, Natalia; Tkachuk, Sergey; Jerke, Uwe; Haller, Hermann; Dumler, Inna

    2003-12-15

    After vascular injury, a remodeling process occurs that features leukocyte migration and infiltration. Loss of endothelial integrity allows the leukocytes to interact with vascular smooth muscle cells (VSMCs) and to elicit "marching orders"; however, the signaling processes are poorly understood. We found that human monocytes inhibit VSMC proliferation and induce a migratory potential. The monocytes signal the VSMCs through the urokinase-type plasminogen activator (uPA). The VSMC uPA receptor (uPAR) receives the signal and activates the transcription factor Stat1 that, in turn, mediates the antiproliferative effects. These results provide the first evidence that monocytes signal VSMCs by mechanisms involving the fibrinolytic system, and they imply an important link between the uPA/uPAR-related signaling machinery and human vascular disease.

  3. Dependence of Golgi apparatus integrity on nitric oxide in vascular cells: implications in pulmonary arterial hypertension

    PubMed Central

    Lee, Jason E.; Patel, Kirit; Almodóvar, Sharilyn; Tuder, Rubin M.; Flores, Sonia C.

    2011-01-01

    Although reduced bioavailability of nitric oxide (NO) has been implicated in the pathogenesis of pulmonary arterial hypertension (PAH), its consequences on organellar structure and function within vascular cells is largely unexplored. We investigated the effect of reduced NO on the structure of the Golgi apparatus as assayed by giantin or GM130 immunofluorescence in human pulmonary arterial endothelial (HPAECs) and smooth muscle (HPASMCs) cells, bovine PAECs, and human EA.hy926 endothelial cells. Golgi structure was also investigated in cells in tissue sections of pulmonary vascular lesions in idiopathic PAH (IPAH) and in macaques infected with a chimeric simian immunodeficiency virus containing the human immunodeficiency virus (HIV)-nef gene (SHIV-nef) with subcellular three-dimensional (3D) immunoimaging. Compounds with NO scavenging activity including 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide (c-PTIO), methylene blue, N-acetylcysteine, and hemoglobin markedly fragmented the Golgi in all cell types evaluated as did monocrotaline pyrrole, while LY-83583, sildenafil, fasudil, Y-27632, Tiron, Tempol, or H2O2 did not. Golgi fragmentation by NO scavengers was inhibited by diethylamine NONOate, was evident in HPAECs after selective knockdown of endothelial nitric oxide synthase using small interfering RNA (siRNA), was independent of microtubule organization, required the GTPase dynamin 2, and was accompanied by depletion of α-soluble N-ethylmaleimide-sensitive factor (NSF) acceptor protein (α-SNAP) from Golgi membranes and codispersal of the SNAP receptor (SNARE) Vti1a with giantin. Golgi fragmentation was confirmed in endothelial and smooth muscle cells in pulmonary arterial lesions in IPAH and the SHIV-nef-infected macaque with subcellular 3D immunoimaging. In SHIV-nef-infected macaques Golgi fragmentation was observed in cells containing HIV-nef-bearing endosomes. The observed Golgi fragmentation suggests that NO plays a significant role in

  4. Metformin inhibits inflammatory response via AMPK-PTEN pathway in vascular smooth muscle cells

    SciTech Connect

    Kim, Sun Ae; Choi, Hyoung Chul

    2012-09-07

    Highlights: Black-Right-Pointing-Pointer PTEN was induced by metformin and inhibited by compound C and AMPK siRNA. Black-Right-Pointing-Pointer Metformin suppressed TNF-{alpha}-induced COX-2 and iNOS mRNA expression. Black-Right-Pointing-Pointer Compound C and bpv (pic) increased iNOS and COX-2 protein expression. Black-Right-Pointing-Pointer NF-{kappa}B activation was restored by inhibiting AMPK and PTEN. Black-Right-Pointing-Pointer AMPK and PTEN regulated TNF-{alpha}-induced ROS production in VSMCs. -- Abstract: Atherosclerosis is a chronic inflammation of the coronary arteries. Vascular smooth muscle cells (VSMCs) stimulated by cytokines and chemokines accelerate the inflammatory response and migrate to the injured endothelium during the progression of atherosclerosis. Activation of AMP activated protein kinase (AMPK), a key sensor maintaining metabolic homeostasis, suppresses the inflammatory response. However, how AMPK regulates the inflammatory response is poorly understood. To identify the mechanism of this response, we focused on phosphatase and tensin homolog (PTEN), which is a negative regulator of inflammation. We investigated that activation of AMPK-induced PTEN expression and suppression of the inflammatory response through the AMPK-PTEN pathway in VSMCs. We treated with the well-known AMPK activator metformin to induce PTEN expression. PTEN was induced by metformin (2 mM) and inhibited by compound C (10 {mu}M) and AMPK siRNA. Tumor necrosis factor-alpha (TNF-{alpha}) was used to induce inflammation. The inflammatory response was confirmed by cyclooxygenase (COX)-2, inducible nitric oxide synthase (iNOS) expression, and activation of nuclear factor (NF)-{kappa}B. Metformin suppressed COX-2 and iNOS mRNA and protein expression dose dependently. Treatment with compound C and bpv (pic) in the presence of metformin, iNOS and COX-2 protein expression increased. NF-{kappa}B activation decreased in response to metformin and was restored by inhibiting AMPK

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

    PubMed

    Wiciński, Michał; Malinowski, Bartosz; Węclewicz, Mateusz M; Grześk, Elżbieta; Grześk, Grzegorz

    2017-01-01

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

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

    PubMed Central

    Malinowski, Bartosz; Grześk, Elżbieta; Grześk, Grzegorz

    2017-01-01

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

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

    SciTech Connect

    Nakahara, Takehiro; Sato, Hiroko; Shimizu, Takehisa; Tanaka, Toru; Matsui, Hiroki; Kawai-Kowase, Keiko; Sato, Mahito; Iso, Tatsuya; Arai, Masashi; Kurabayashi, Masahiko

    2010-04-02

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

  8. Assessment of vascular autonomic function using peripheral arterial tonometry.

    PubMed

    Hamada, Satomi; Oono, Ai; Ishihara, Yuri; Hasegawa, Yuki; Akaza, Miho; Sumi, Yuki; Inoue, Yoshinori; Izumiyama, Hajime; Hirao, Kenzo; Isobe, Mitsuaki; Sasano, Tetsuo

    2017-03-01

    Peripheral autonomic function is impaired in diabetic polyneuropathy. However, it is difficult to evaluate it due to the lack of non-invasive quantitative assessment. We aimed to establish a novel index to evaluate vascular autonomic function using reactive hyperemia peripheral arterial tonometry (RH-PAT), a widely performed endothelial function test. Sixty-five subjects were enrolled, including healthy subjects, cases with sympathetic nerve blockers, and diabetic patients. RH-PAT was performed with 5-min blood flow occlusion in unilateral arm. We calculated the reduction ratio of the post-occlusion pulse amplitude to the baseline in the non-occluded arm (RPN), with 1-min sliding window. In healthy subjects, RPN gradually increased with time-dependent manner. However, this phenomenon was eliminated in cases with sympathetic nerve blockers. Plasma concentration of norepinephrine was measured before and after the blood flow occlusion, which showed a significant increase. We then compared RPNs with the change in heart rate variability (HRV) parameters. RPN calculated at 5 min after the reperfusion had the highest correlation with the change in sympathetic HRV parameter, and thus, we named sympathetic hypoemia index (SHI). Finally, we studied the relationship between SHI and diabetes. SHI was significantly lower in diabetic patients than matched controls. SHI, a novel index derived from RH-PAT, represented the peripheral sympathetic activity. SHI may be useful for assessing the vascular autonomic activity in diabetic patients.

  9. The bestrophin- and TMEM16A-associated Ca2+-activated Cl– channels in vascular smooth muscles

    PubMed Central

    Dam, Vibeke Secher; Boedtkjer, Donna MB; Aalkjaer, Christian; Matchkov, Vladimir

    2014-01-01

    The presence of Ca2+-activated Cl– currents (ICl(Ca)) in vascular smooth muscle cells (VSMCs) is well established. ICl(Ca) are supposedly important for arterial contraction by linking changes in [Ca2+]i and membrane depolarization. Bestrophins and some members of the TMEM16 protein family were recently associated with ICl(Ca). Two distinct ICl(Ca) are characterized in VSMCs; the cGMP-dependent ICl(Ca) dependent upon bestrophin expression and the ‘classical’ Ca2+-activated Cl– current, which is bestrophin-independent. Interestingly, TMEM16A is essential for both the cGMP-dependent and the classical ICl(Ca). Furthermore, TMEM16A has a role in arterial contraction while bestrophins do not. TMEM16A’s role in the contractile response cannot be explained however only by a simple suppression of the depolarization by Cl– channels. It is suggested that TMEM16A expression modulates voltage-gated Ca2+ influx in a voltage-independent manner and recent studies also demonstrate a complex role of TMEM16A in modulating other membrane proteins. PMID:25478625

  10. Reactive oxygen species and RhoA signaling in vascular smooth muscle: role in chronic hypoxia-induced pulmonary hypertension.

    PubMed

    Resta, Thomas C; Broughton, Brad R S; Jernigan, Nikki L

    2010-01-01

    Increases in myofilament Ca2+ sensitivity resulting from stimulation of RhoA and Rho kinase represent a primary mechanism of vasoconstriction and associated pulmonary hypertension resulting from chronic hypoxia (CH). This chapter summarizes recent advances in the understanding of RhoA/Rho kinase signaling mechanisms in pulmonary vascular smooth muscle (VSM) that increase the sensitivity of the contractile apparatus to Ca2+ and contribute to vasoconstriction in this setting. Such advances include the discovery of myogenic tone in small pulmonary arteries from CH rats that contributes to vasoconstriction through a mechanism inherent to the VSM, dependent on Rho kinase-induced Ca2+ sensitization but independent of L-type voltage-gated Ca2+ channels. Additional studies have revealed an important contribution of superoxide anion (O2-)-induced RhoA activation to both receptor-mediated and membrane depolarization-induced myofilament Ca2+ sensitization in hypertensive pulmonary arteries. Xanthine oxidase and NADPH oxidase isoforms are potential sources of O2- that mediate RhoA-dependent vasoconstriction and associated pulmonary hypertension.

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

    NASA Technical Reports Server (NTRS)

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

    2001-01-01

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

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

    NASA Astrophysics Data System (ADS)

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

    2011-11-01

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

  13. Subclavian artery aneurysm in a patient with vascular Ehlers-Danlos syndrome.

    PubMed

    Yasuda, Shota; Imoto, Kiyotaka; Uchida, Keiji; Uranaka, Yasuko; Kurosawa, Kenji; Masuda, Munetaka

    2016-02-01

    We describe our experience of surgical treatment in a 28-year-old woman with vascular Ehlers-Danlos syndrome. A right subclavian artery aneurysm was detected. The right vertebral artery arose from the aneurysm. Digital subtraction angiography showed interruption of the left vertebral artery. The aneurysm was excised and the right vertebral artery was anastomosed end-to-side to the right common carotid artery under deep hypothermia and circulatory arrest. The patient remained very well 4 years after surgery, with no late vascular complication.

  14. Arterial vascularization and morphological characteristics of adrenal glands in the Pampas deer (Ozotoceros bezoarticus, Linnaeus 1758).

    PubMed

    Erdoğan, S; Pérez, W

    2014-10-01

    This research presents morphological characteristics of adrenal glands and a demonstration of arterial vascularization in the Pampas deer, which is considered to be in extreme danger of extinction. A total of ten deer constituted the material of the study. Vascularization of organs was investigated by using latex injection technique. Left adrenal glands were basically supplied by coeliac, cranial mesenteric, renal and lumbal arteries. The arterial vascularization of the left adrenal glands was very complex in comparison with right adrenal glands. In two examples, branch of the lumbal artery was divided into phrenic caudal artery and cranial adrenal artery. In six examples, it was observed that the caudomedial and ventral regions of the left adrenal glands were also supplied by thinner branches that stemmed from second left lumbal artery. Besides, coeliac and cranial mesenteric arteries also gave off shorter branches supplying the cranial region of the left adrenal glands in five examples. It was determined that two branches originated from abdominal aorta directly for supplying left adrenal glands in only two examples. In four examples, two caudal adrenal arteries stemmed separately from left renal artery in a short distance. Arterial vascularization of right adrenal glands was more constant and supplied by lumbal and renal arteries. The adrenal glands were generally oval or round shaped. In only two examples, left adrenal glands were 'V-' or heart-shaped. There was no significant difference (P > 0.05) in sizes between right and left adrenal glands.

  15. c-Ski inhibits the proliferation of vascular smooth muscle cells via suppressing Smad3 signaling but stimulating p38 pathway.

    PubMed

    Li, Jun; Li, Ping; Zhang, Yan; Li, Gong-Bo; Zhou, Yuan-Guo; Yang, Kang; Dai, Shuang-Shuang

    2013-01-01

    Proliferation of vascular smooth muscle cells (VSMCs) plays key roles in the progression of intimal hyperplasia, but the molecular mechanisms that trigger VSMC proliferation after vascular injury remain unclear. c-Ski, a co-repressor of transforming growth factor β (TGF-β)/Smad signaling, was detected to express in VSMC of rat artery. During the course of arterial VSMC proliferation induced by balloon injury in rat, the endogenous protein expressions of c-Ski decreased markedly in a time-dependent manner. In vivo c-Ski gene delivery was found to significantly suppress balloon injury-induced VSMC proliferation and neointima formation. Further investigation in A10 rat aortic smooth muscle cells demonstrated that overexpression of c-Ski gene inhibited TGF-β1 (1 ng/ml)-induced A10 cell proliferation while knockdown of c-Ski by RNAi enhanced the stimulatory effect of TGF-β1 on A10 cell growth. Western blot for signaling detection showed that suppression of Smad3 phosphorylation while stimulating p38 signaling associated with upregulation of cyclin-dependent kinase inhibitors p21 and p27 was responsible for the inhibitory effect of c-Ski on TGF-β1-induced VSMC proliferation. These data suggest that the decrease of endogenous c-Ski expression is implicated in the progression of VSMC proliferation after arterial injury and c-Ski administration represents a promising role for treating intimal hyperplasia via inhibiting the proliferation of VSMC.

  16. Suppression of Akt1 phosphorylation by adenoviral transfer of the PTEN gene inhibits hypoxia-induced proliferation of rat pulmonary arterial smooth muscle cells

    SciTech Connect

    Luo, Chunxia; Yi, Bin; Bai, Li; Xia, Yongzhi; Wang, Guansong; Qian, Guisheng; Feng, Hua

    2010-07-02

    Recent findings identify the role of proliferation of pulmonary artery smooth muscle cells (PASMCs) in pulmonary vascular remodeling. Phosphoinositide 3 kinase (PI3K) and serine/threonine kinase (Akt) proteins are expressed in vascular smooth muscle cells. In addition, phosphatase and tensin homolog deleted on chromosome 10 (PTEN) has been identified as a negative regulator of cytokine signaling that inhibits the PI3K-Akt pathway. However, little is known about the role of PTEN/Akt signaling in hypoxia-associated vascular remodeling. In this study, we found that hypoxia-induced the expression of Akt1 mRNA and phosphorylated protein by at least twofold in rat PASMCs. Phospho-PTEN significantly decreased in the nuclei of PASMCs after hypoxic stimulation. After forcing over-expression of PTEN by adenovirus-mediated PTEN (Ad-PTEN) transfection, the expression of phospho-Akt1 was significantly suppressed in PASMCs at all time-points measured. Additionally, we showed here that hypoxia increased proliferation of PASMCs by nearly twofold and over-expression of PTEN significantly inhibited hypoxia-induced PASMCs proliferation. These findings suggest that phospho-PTEN loss in the nuclei of PASMCs under hypoxic conditions may be the major cause of aberrant activation of Akt1 and may, therefore, play an important role in hypoxia-associated pulmonary arterial remodeling. Finally, the fact that transfection with Ad-PTEN inhibits the phosphorylation of Akt1 in PASMCs suggests a potential therapeutic effect on hypoxia-associated pulmonary arterial remodeling.

  17. Efficient transduction of vascular smooth muscle cells with a translational AAV2.5 vector: a new perspective for in-stent restenosis gene therapy.

    PubMed

    Lompré, A-M; Hadri, L; Merlet, E; Keuylian, Z; Mougenot, N; Karakikes, I; Chen, J; Atassi, F; Marchand, A; Blaise, R; Limon, I; McPhee, S W J; Samulski, R J; Hajjar, R J; Lipskaia, L

    2013-09-01

    Coronary artery disease represents the leading cause of mortality in the developed world. Percutaneous coronary intervention involving stent placement remains disadvantaged by restenosis or thrombosis. Vascular gene therapy-based methods may be approached, but lack a vascular gene delivery vector. We report a safe and efficient long-term transduction of rat carotid vessels after balloon injury intervention with a translational optimized AAV2.5 vector. Compared with other known adeno-associated virus (AAV) serotypes, AAV2.5 demonstrated the highest transduction efficiency of human coronary artery vascular smooth muscle cells (VSMCs) in vitro. Local delivery of AAV2.5-driven transgenes in injured carotid arteries resulted in transduction as soon as day 2 after surgery and persisted for at least 30 days. In contrast to adenovirus 5 vector, inflammation was not detected in AAV2.5-transduced vessels. The functional effects of AAV2.5-mediated gene transfer on neointimal thickening were assessed using the sarco/endoplasmic reticulum Ca(2+) ATPase isoform 2a (SERCA2a) human gene, known to inhibit VSMC proliferation. At 30 days, human SERCA2a messenger RNA was detected in transduced arteries. Morphometric analysis revealed a significant decrease in neointimal hyperplasia in AAV2.5-SERCA2a-transduced arteries: 28.36±11.30 (n=8) vs 77.96±24.60 (n=10) μm(2), in AAV2.5-green fluorescent protein-infected, P<0.05. In conclusion, AAV2.5 vector can be considered as a promising safe and effective vector for vascular gene therapy.

  18. Neutrophil Elastase Is Produced by Pulmonary Artery Smooth Muscle Cells and Is Linked to Neointimal Lesions

    PubMed Central

    Kim, Yu-Mee; Haghighat, Leila; Spiekerkoetter, Edda; Sawada, Hirofumi; Alvira, Cristina M.; Wang, Lingli; Acharya, Swati; Rodriguez-Colon, Gabriela; Orton, Andrew; Zhao, Mingming; Rabinovitch, Marlene

    2011-01-01

    Previously, we reported that murine gammaherpesvirus-68 (M1-MHV-68) induces pulmonary artery (PA) neointimal lesions in S100A4-overexpressing, but not in wild-type (C57), mice. Lesions were associated with heightened lung elastase activity and PA elastin degradation. We now investigate a direct relationship between elastase and PA neointimal lesions, the nature and source of the enzyme, and its presence in clinical disease. We found an association exists between the percentage of PAs with neointimal lesions and elastin fragmentation in S100A4 mice 6 months after viral infection. Confocal microscopy documented the heightened susceptibility of S100A4 versus C57 PA elastin to degradation by elastase. A transient increase in lung elastase activity occurs in S100A4 mice, 7 days after M1-MHV-68, unrelated to inflammation or viral load and before neointimal lesions. Administration of recombinant elafin, an elastase-specific inhibitor, ameliorates early increases in serine elastase and attenuates later development of neointimal lesions. Neutrophils are the source of elevated elastase (NE) in the S100A4 lung, and NE mRNA and protein levels are greater in PA smooth muscle cells (SMC) from S100A4 mice than from C57 mice. Furthermore, elevated NE is observed in cultured PA SMC from idiopathic PA hypertension versus that in control lungs and localizes to neointimal lesions. Thus, PA SMC produce NE, and heightened production and activity of NE is linked to experimental and clinical pulmonary vascular disease. PMID:21763677

  19. Vascular smooth muscle dysfunction induced by monomethylarsonous acid (MMA III): a contributing factor to arsenic-associated cardiovascular diseases.

    PubMed

    Bae, Ok-Nam; Lim, Eun-Kyung; Lim, Kyung-Min; Noh, Ji-Yoon; Chung, Seung-Min; Lee, Moo-Yeol; Yun, Yeo-Pyo; Kwon, Seong-Chun; Lee, Jun-Ho; Nah, Seung-Yeol; Chung, Jin-Ho

    2008-11-01

    While arsenic in drinking water is known to cause various cardiovascular diseases in human, exact mechanism still remains elusive. Recently, trivalent-methylated arsenicals, the metabolites of inorganic arsenic, were shown to have higher cytotoxic potential than inorganic arsenic. To study the role of these metabolites in arsenic-induced cardiovascular diseases, we investigated the effect of monomethylarsonous acid (MMA III), a major trivalent-methylated arsenical, on vasomotor tone of blood vessels. In isolated rat thoracic aorta and small mesenteric arteries, MMA III irreversibly suppressed normal vasoconstriction induced by three distinct agonists of phenylephrine (PE), serotonin and endothelin-1. Inhibition of vasoconstriction was retained in aortic rings without endothelium, suggesting that MMA III directly impaired the contractile function of vascular smooth muscle. The effect of MMA III was mediated by inhibition of PE-induced Ca2+ increase as found in confocal microscopy and fluorimeter in-lined organ chamber technique. The attenuation of Ca2+ increase was from concomitant inhibition of release from intracellular store and extracellular Ca2+ influx via L-type Ca2+ channel, which was blocked by MMA III as shown in voltage-clamp assay in Xenopus oocytes. MMA III did not affect downstream process of Ca2+, as shown in permeabilized arterial strips. In in vivo rat model, MMA III attenuated PE-induced blood pressure increase indeed, supporting the clinical relevance of these in vitro findings. In conclusion, MMA III-induced smooth muscle dysfunction through disturbance of Ca2+ regulation, which results in impaired vasoconstriction and aberrant blood pressure change. This study will provide a new insight into the role of trivalent-methylated arsenicals in arsenic-associated cardiovascular diseases.

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

    SciTech Connect

    Aguado, Andrea; Galán, María; Zhenyukh, Olha; Wiggers, Giulia A.; Roque, Fernanda R.; Redondo, Santiago; Peçanha, Franck; Martín, Angela; Fortuño, Ana; Cachofeiro, Victoria; Tejerina, Teresa; Salaices, Mercedes; 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

  1. Role played by Prx1-dependent extracellular matrix properties in vascular smooth muscle development in embryonic lungs

    PubMed Central

    Ames, Juliana; Chokshi, Mithil; Aiad, Norman; Sanyal, Sonali; Kawabata, Kimihito C.; Levental, Ilya; Sundararaghavan, Harini G.; Burdick, Jason A.; Janmey, Paul; Miyazono, Kohei; Wells, Rebecca G.; Jones, Peter L.

    2015-01-01

    Abstract Although there are many studies focusing on the molecular pathways underlying lung vascular morphogenesis, the extracellular matrix (ECM)–dependent regulation of mesenchymal cell differentiation in vascular smooth muscle development needs better understanding. In this study, we demonstrate that the paired related homeobox gene transcription factor Prx1 maintains the elastic ECM properties, which are essential for vascular smooth muscle precursor cell differentiation. We have found that Prx1null mouse lungs exhibit defective vascular smooth muscle development, downregulated elastic ECM expression, and compromised transforming growth factor (TGF)–β localization and signaling. Further characterization of ECM properties using decellularized lung ECM scaffolds derived from Prx1 mice demonstrated that Prx1 is required to maintain lung ECM stiffness. The results of cell culture using stiffness-controlled 2-D and 3-D synthetic substrates confirmed that Prx1-dependent ECM stiffness is essential for promotion of smooth muscle precursor differentiation for effective TGF-β stimulation. Supporting these results, both decellularized Prx1null lung ECM and Prx1WT (wild type) ECM scaffolds with blocked TGF-β failed to support mesenchymal cell to 3-D smooth muscle cell differentiation. These results suggest a novel ECM-dependent regulatory pathway of lung vascular development wherein Prx1 regulates lung vascular smooth muscle precursor development by coordinating the ECM biophysical and biochemical properties. PMID:26064466

  2. Calcium homeostasis and sensitization in pulmonary arterial smooth muscle.

    PubMed

    Jernigan, Nikki L; Resta, Thomas C

    2014-04-01

    The pulmonary circulation is a low-pressure, low-resistance vascular bed with little to no resting tone under normal conditions. An increase in the [Ca(2+) ]i in PASMCs is an important determinant of contraction, migration, and proliferation. Both Ca(2+) influx through plasma membrane Ca(2+) channels and Ca(2+) release from the SR contribute to a rise in [Ca(2+) ]i . Additionally important in the pulmonary circulation are several kinase-mediated signaling pathways that act to increase the sensitivity of the contractile apparatus to [Ca(2+) ]i . Similarly, cytoskeletal processes resulting in dynamic remodeling of the actin cytoskeleton can further contribute to contractility in the pulmonary circulation. In addition to endocrine, paracrine, and autocrine factors, alveolar hypoxia is an important stimulus for pulmonary vasoconstriction. However, prolonged hypoxia is a critical pathological stimulus associated with the development of pulmonary hypertension and cor pulmonale. In this review, we will discuss recent advances in our understanding of how Ca(2+) homeostasis and sensitization regulate PASMC contractility under both physiological and pathophysiological conditions.

  3. Induction of cyclin A gene expression by homocysteine in vascular smooth muscle cells.

    PubMed Central

    Tsai, J C; Wang, H; Perrella, M A; Yoshizumi, M; Sibinga, N E; Tan, L C; Haber, E; Chang, T H; Schlegel, R; Lee, M E

    1996-01-01

    Homocysteine is an important and independent risk factor for arteriosclerosis. We showed previously that homocysteine stimulates vascular smooth muscle cell proliferation, a hallmark of arteriosclerosis. We show here that homocysteine and serum increased DNA synthesis synergistically in both human and rat aortic smooth muscle cells (RASMCs). Treatment of quiescent RASMCs with 1 mM homocysteine or 2% calf serum for 36 h increased cyclin A mRNA levels by 8- and 14-fold, respectively, whereas homocysteine plus serum increased cyclin A mRNA levels by 40-fold, indicating a synergistic induction of cyclin A mRNA. Homocysteine did not increase the half-life of cyclin A mRNA (2.9 h), but it did increase the transcriptional rate of the cyclin A gene in nuclear run-on experiments. The positive effect of homocysteine on cyclin A gene transcription was confirmed by our finding that homocysteine increased cyclin A promoter activity and ATF-binding protein levels in RASMCs. Finally, 1 mM homocysteine increased cyclin A protein levels and cyclin A-associated kinase activity by threefold. This homocysteine-induced expression lesions by promoting proliferation of vascular smooth muscle cells. PMID:8550827

  4. Heparin fragments inhibit human vascular smooth muscle cell proliferation in vitro

    SciTech Connect

    Selden, S.C.; Johnson, W.V.; Maciag, T.

    1986-03-01

    The authors have examined the effect of heparin on human abdominal aortic smooth muscle cell growth. Cell proliferation was inhibited by more than 90% at a concentration of 20 ..mu..g/ml in a 12 day growth assay using heparin from Sigma, Upjohn or Calbiochem. Additionally, 200 ..mu..g/ml Upjohn heparin inhibits /sup 3/H-thymidine incorporation by 50% in short term assays using serum or purified platelet-derived growth factor (25-100ng/ml) to initiate the cell cycle. Homogeneous size classes of heparin fragments were prepared by nitrous acid cleavage and BioGel P-10 filtration chromatography. Deca-, octa-, hexa-, tetra-, and di-saccharides inhibited proliferation by 90% at concentrations of 280, 320, 260, 180 and 100 ..mu..g/ml, respectively, in a 12 day growth assay. These data confirm the work of Castellot et.al. and extend the range of inhibitory fragments down to the tetra- and di-saccharide size. These data suggest, therefore, that di-saccharide subunit of heparin is sufficient to inhibit vascular smooth muscle cell proliferation. The authors are now examining the role of the anhydromannose moiety on the reducing end of the nitrous acid generated fragments as a possible mediator of heparin-induced inhibition of vascular smooth muscle cell proliferation.

  5. Phospholipase D1 is involved in α1-adrenergic contraction of murine vascular smooth muscle.

    PubMed

    Wegener, Jörg W; Loga, Florian; Stegner, David; Nieswandt, Bernhard; Hofmann, Franz

    2014-03-01

    α1-Adrenergic stimulation increases blood vessel tone in mammals. This process involves a number of intracellular signaling pathways that include signaling via phospholipase C, diacylglycerol (DAG), and protein kinase C. So far, it is not certain whether signaling via phospholipase D (PLD) and PLD-derived DAG is involved in this process. We asked whether PLD participates in the α1-adrenergic-mediated signaling in vascular smooth muscle. α1-Adrenergic-induced contraction was assessed by myography of isolated aortic rings and by pressure recordings using the hindlimb perfusion model in mice. The effects of the PLD inhibitor 1-butanol (IC50 0.15 vol%) and the inactive congener 2-butanol were comparatively studied. Inhibition of PLD by 1-butanol reduced specifically the α1-adrenergic-induced contraction and the α1-adrenergic-induced pressure increase by 10 and 40% of the maximum, respectively. 1-Butanol did not influence the aortic contractions induced by high extracellular potassium, by the thromboxane analog U46619, or by a phorbol ester. The effects of 1-butanol were absent in mice that lack PLD1 (Pld1(-/-) mice) or that selectively lack the CaV1.2 channel in smooth muscle (sm-CaV1.2(-/-) mice) but still present in the heterozygous control mice. α1-Adrenergic contraction of vascular smooth muscle involves activation of PLD1, which controls a portion of the α1-adrenergic-induced CaV1.2 channel activity.

  6. Transdifferentiation of pulmonary arteriolar endothelial cells into smooth muscle-like cells regulated by myocardin involved in hypoxia-induced pulmonary vascular remodelling

    PubMed Central

    Zhu, Pengcheng; Huang, Lei; Ge, Xiaona; Yan, Fei; Wu, Renliang; Ao, Qilin

    2006-01-01

    Myocardin gene has been identified as a master regulator of smooth muscle cell differentiation. Smooth muscle cells play a critical role in the pathogenesis of hypoxia-induced pulmonary hypertension (PH) and pulmonary vascular remodelling (PVR). The purpose of this study was to investigate the change of myocardin gene expression in the pulmonary vessels of hypoxia-induced PH affected by Sildenafil treatment and the involvement of endothelial cells transdifferentiation into smooth muscle cells in the process of hypoxia-induced PH and PVR. Myocardin and relative markers were investigated in animal models and cultured endothelial cells. Mean pulmonary artery pressure (mPAP) was measured. Immunohistochemistry and immunofluorescence were used to show the expression of smooth muscle α-actin (SMA), in situ hybridization (ISH) and reverse transcription polymerase chain reaction (RT-PCR) were performed respectively to detect the myocardin and SMA expression at mRNA levels. Small interfering RNA (siRNA) induced suppression of myocardin in cultured cells. We confirmed that hypoxia induced the PH and PVR in rats. Sildenafil could attenuate the hypoxia-induced PH. We found that myocardin mRNA expression is upregulated significantly in the hypoxic pulmonary vessels and cultured cells but downregulated in PH with Sildenafil treatment. The porcine pulmonary artery endothelial cells (PAECs) transdifferentiate into smooth muscle-like cells in hypoxic culture while the transdifferentiation did not occur when SiRNA of myocardin was applied. Our results suggest that myocardin gene, as a marker of smooth muscle cell differentiation, was expressed in the pulmonary vessels in hypoxia-induced PH rats, which could be downregulated by Sildenafil treatment, as well as in hypoxic cultured endothelial cells. Hypoxia induced the transdifferentiation of endothelial cells of vessels into smooth muscle-like cells which was regulated by myocardin. PMID:17222214

  7. Identification of transcription factors and gene clusters in rabbit smooth muscle cells during high flow-induced vascular remodeling via microarray.

    PubMed

    Zhang, Zhaolong; Yang, Pengfei; Yao, Pengfei; Dai, Dongwei; Yu, Ying; Zhou, Yu; Huang, Qinghai; Liu, Jianmin

    2016-01-10

    Sustained blood flow, especially high blood flow causes the remodeling of arteries. The molecular mechanism of vascular remodeling has been mainly investigated in cultured cells. However, the in vivo molecular mechanism is poorly understood. In this study, we performed microarray analysis to explore the gene expression profile of smooth muscle cells (SMCs) during vascular remodeling. Transcriptional profiles indicated that 947 genes were differentially expressed in SMCs responding to high flow compared with the sham control, of which 617 genes were up-regulated and 330 genes were down-regulated. Gene ontology analysis revealed the special participation of extracellular matrix related genes during high flow-induced vascular remodeling. KEGG pathway analysis showed the enrichment of metabolism and immune function associated genes in SMCs exposed to high flow. Besides, we also identified 25 differentially expressed transcription factors potentially impacted by hemodynamic insult. Finally, we revealed FOXN4 as a novel transcription factor that could modulate MMP2 and MMP9 transcriptional activity. Collectively, our results revealed major gene clusters and transcription factors in SMCs during vascular remodeling which may provide an insight into the molecular mechanism of vascular remodeling and facilitate the screening of candidate genes for vascular diseases.

  8. BMP-9 regulates the osteoblastic differentiation and calcification of vascular smooth muscle cells through an ALK1 mediated pathway

    PubMed Central

    Zhu, Dongxing; Mackenzie, Neil Charles Wallace; Shanahan, Catherine M; Shroff, Rukshana C; Farquharson, Colin; MacRae, Vicky Elizabeth

    2015-01-01

    The process of vascular calcification shares many similarities with that of physiological skeletal mineralization, and involves the deposition of hydroxyapatite crystals in arteries. However, the cellular mechanisms responsible have yet to be fully explained. Bone morphogenetic protein (BMP-9) has been shown to exert direct effects on both bone development and vascular function. In the present study, we have investigated the role of BMP-9 in vascular smooth muscle cell (VSMC) calcification. Vessel calcification in chronic kidney disease (CKD) begins pre-dialysis, with factors specific to the dialysis milieu triggering accelerated calcification. Intriguingly, BMP-9 was markedly elevated in serum from CKD children on dialysis. Furthermore, in vitro studies revealed that BMP-9 treatment causes a significant increase in VSMC calcium content, alkaline phosphatase (ALP) activity and mRNA expression of osteogenic markers. BMP-9-induced calcium deposition was significantly reduced following treatment with the ALP inhibitor 2,5-Dimethoxy-N-(quinolin-3-yl) benzenesulfonamide confirming the mediatory role of ALP in this process. The inhibition of ALK1 signalling using a soluble chimeric protein significantly reduced calcium deposition and ALP activity, confirming that BMP-9 is a physiological ALK1 ligand. Signal transduction studies revealed that BMP-9 induced Smad2, Smad3 and Smad1/5/8 phosphorylation. As these Smad proteins directly bind to Smad4 to activate target genes, siRNA studies were subsequently undertaken to examine the functional role of Smad4 in VSMC calcification. Smad4-siRNA transfection induced a significant reduction in ALP activity and calcium deposition. These novel data demonstrate that BMP-9 induces VSMC osteogenic differentiation and calcification via ALK1, Smad and ALP dependent mechanisms. This may identify new potential therapeutic strategies for clinical intervention. PMID:25297851

  9. Control of vascular smooth muscle function by Src-family kinases and reactive oxygen species in health and disease.

    PubMed

    MacKay, Charles E; Knock, Greg A

    2015-09-01

    Reactive oxygen species (ROS) are now recognised as second messenger molecules that regulate cellular function by reversibly oxidising specific amino acid residues of key target proteins. Amongst these are the Src-family kinases (SrcFKs), a multi-functional group of non-receptor tyrosine kinases highly expressed in vascular smooth muscle (VSM). In this review we examine the evidence supporting a role for ROS-induced SrcFK activity in normal VSM contractile function and in vascular remodelling in cardiovascular disease. VSM contractile responses to G-protein-coupled receptor stimulation, as well as hypoxia in pulmonary artery, are shown to be dependent on both ROS and SrcFK activity. Specific phosphorylation targets are identified amongst those that alter intracellular Ca(2+) concentration, including transient receptor potential channels, voltage-gated Ca(2+) channels and various types of K(+) channels, as well as amongst those that regulate actin cytoskeleton dynamics and myosin phosphatase activity, including focal adhesion kinase, protein tyrosine kinase-2, Janus kinase, other focal adhesion-associated proteins, and Rho guanine nucleotide exchange factors. We also examine a growing weight of evidence in favour of a key role for SrcFKs in multiple pro-proliferative and anti-apoptotic signalling pathways relating to oxidative stress and vascular remodelling, with a particular focus on pulmonary hypertension, including growth-factor receptor transactivation and downstream signalling, hypoxia-inducible factors, positive feedback between SrcFK and STAT3 signalling and positive feedback between SrcFK and NADPH oxidase dependent ROS production. We also discuss evidence for and against the potential therapeutic targeting of SrcFKs in the treatment of pulmonary hypertension.

  10. Vascular Smooth Muscle LRP6 Limits Arteriosclerotic Calcification in Diabetic LDLR-/- Mice by Restraining Noncanonical Wnt Signals

    PubMed Central

    Cheng, Su-Li; Ramachandran, Bindu; Behrmann, Abraham; Shao, Jian-Su; Mead, Megan; Smith, Carolyn; Krchma, Karen; Arredondo, Yoanna Bello; Kovacs, Attila; Kapoor, Kapil; Brill, Laurence M.; Perera, Ranjan; Williams, Bart O.; Towler, Dwight A.

    2015-01-01

    Rationale Wnt signaling regulates key aspects of diabetic vascular disease. Objective We generated SM22-Cre;LRP6(fl/fl);LDLR-/- mice to determine contributions of Wnt co-receptor LRP6 in the vascular smooth muscle lineage (VSM) of male LDLR-null mice, a background susceptible to diet (HFD) - induced diabetic arteriosclerosis. Methods and Results As compared to LRP6(fl/fl);LDLR-/- controls, SM22-Cre;LRP6(fl/fl);LDLR-/- (LRP6-VKO) siblings exhibited increased aortic calcification on HFD without changes in fasting glucose, lipids, or body composition. Pulse wave velocity (index of arterial stiffness) was also increased. Vascular calcification paralleled enhanced aortic osteochondrogenic programs and circulating osteopontin (OPN), a matricellular regulator of arteriosclerosis. Survey of ligands and Frizzled (Fzd) receptor profiles in LRP6-VKO revealed upregulation of canonical and noncanonical Wnts alongside Fzd10. Fzd10 stimulated noncanonical signaling and OPN promoter activity via an USF-activated cognate inhibited by LRP6. RNAi revealed that USF1 but not USF2 supports OPN expression in LRP6-VKO VSM, and immunoprecipitation confirmed increased USF1 association with OPN chromatin. ML141, an antagonist of cdc42/Rac1 noncanonical signaling, inhibited USF1 activation, osteochondrogenic programs, alkaline phosphatase, and VSM calcification. Mass spectrometry identified LRP6 binding to protein arginine methyltransferase (PRMT) - 1, and nuclear asymmetric dimethylarginine modification was increased with LRP6-VKO. RNAi demonstrated that PRMT1 inhibits OPN and TNAP while PRMT4 supports expression. USF1 complexes containing the H3R17Me2a signature of PRMT4 are increased with LRP6-VKO. Jmjd6, a demethylase downregulated with LRP6 deficiency, inhibits OPN and TNAP expression, USF1:H3R17Me2a complex formation and transactivation. Conclusions LRP6 restrains VSM noncanonical signals that promote osteochondrogenic differentiation, mediated in part via USF1- and arginine methylation

  11. Gallic acid tailoring surface functionalities of plasma-polymerized allylamine-coated 316L SS to selectively direct vascular endothelial and smooth muscle cell fate for enhanced endothelialization.

    PubMed

    Yang, Zhilu; Xiong, Kaiqin; Qi, Pengkai; Yang, Ying; Tu, Qiufen; Wang, Jin; Huang, Nan

    2014-02-26

    The creation of a platform for enhanced vascular endothelia cell (VEC) growth while suppressing vascular smooth muscle cell (VSMC) proliferation offers possibility for advanced coatings of vascular stents. Gallic acid (GA), a chemically unique phenolic acid with important biological functions, presents benefits to the cardiovascular disease therapy because of its superior antioxidant effect and a selectivity to support the growth of ECs more than SMCs. In this study, GA was explored to tailor such a multifunctional stent surface combined with plasma polymerization technique. On the basis of the chemical coupling reaction, GA was bound to an amine-group-rich plasma-polymerized allylamine (PPAam) coating. The GA-functionalized PPAam (GA-PPAam) surface created a favorable microenvironment to obtain high ECs and SMCs selectivity. The GA-PPAam coating showed remarkable enhancement in the adhesion, viability, proliferation, migration, and release of nitric oxide (NO) of human umbilical vein endothelial cells (HUVECs). The GA-PPAam coating also resulted in remarkable inhibition effect on human umbilical artery smooth muscle cell (HUASMC) adhesion and proliferation. These striking findings may provide a guide for designing the new generation of multifunctional vascular devices.

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

    NASA Astrophysics Data System (ADS)

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

    2010-07-01

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

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

    SciTech Connect

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

    2010-07-23

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

  14. New Insights into Mechanisms and Functions of Chemokine (C-X-C Motif) Receptor 4 Heteromerization in Vascular Smooth Muscle

    PubMed Central

    Evans, Ann E.; Tripathi, Abhishek; LaPorte, Heather M.; Brueggemann, Lioubov I.; Singh, Abhay Kumar; Albee, Lauren J.; Byron, Kenneth L.; Tarasova, Nadya I.; Volkman, Brian F.; Cho, Thomas Yoonsang; Gaponenko, Vadim; Majetschak, Matthias

    2016-01-01

    Recent evidence suggests that C-X-C chemokine receptor type 4 (CXCR4) heteromerizes with α1A/B-adrenoceptors (AR) and atypical chemokine receptor 3 (ACKR3) and that CXCR4:α1A/B-AR heteromers are important for α1-AR function in vascular smooth muscle cells (VSMC). Structural determinants for CXCR4 heteromerization and functional consequences of CXCR4:α1A/B-AR heteromerization in intact arteries, however, remain unknown. Utilizing proximity ligation assays (PLA) to visualize receptor interactions in VSMC, we show that peptide analogs of transmembrane-domain (TM) 2 and TM4 of CXCR4 selectively reduce PLA signals for CXCR4:α1A-AR and CXCR4:ACKR3 interactions, respectively. While both peptides inhibit CXCL12-induced chemotaxis, only the TM2 peptide inhibits phenylephrine-induced Ca2+-fluxes, contraction of VSMC and reduces efficacy of phenylephrine to constrict isolated arteries. In a Cre-loxP mouse model to delete CXCR4 in VSMC, we observed 60% knockdown of CXCR4. PLA signals for CXCR4:α1A/B-AR and CXCR4:ACKR3 interactions in VSMC, however, remained constant. Our observations point towards TM2/4 of CXCR4 as possible contact sites for heteromerization and suggest that TM-derived peptide analogs permit selective targeting of CXCR4 heteromers. A molecular dynamics simulation of a receptor complex in which the CXCR4 homodimer interacts with α1A-AR via TM2 and with ACKR3 via TM4 is presented. Our findings further imply that CXCR4:α1A-AR heteromers are important for intrinsic α1-AR function in intact arteries and provide initial and unexpected insights into the regulation of CXCR4 heteromerization in VSMC. PMID:27331810

  15. Inhibition of SRF/myocardin reduces aortic stiffness by targeting vascular smooth muscle cell stiffening in hypertension

    PubMed Central

    Zhou, Ning; Lee, Jia-Jye; Stoll, Shaunrick; Ma, Ben; Wiener, Robert; Wang, Charles; Costa, Kevin D.; Qiu, Hongyu

    2017-01-01

    Aims Increased aortic stiffness is a fundamental manifestation of hypertension. However, the molecular mechanisms involved remain largely unknown. We tested the hypothesis that abnormal intrinsic vascular smooth muscle cell (VSMC) mechanical properties in large arteries, but not in distal arteries, contribute to the pathogenesis of aortic stiffening in hypertension, mediated by the serum response factor (SRF)/myocardin signalling pathway. Methods and results Four month old male spontaneously hypertensive rats (SHR) and normotensive Wistar-Kyoto (WKY) rats were studied. Using atomic force microscopy, significant VSMC stiffening was observed in the large conducting aorta compared with the distal arteries in SHR (P < 0.001), however, this regional variation was not observed in WKY rats (P > 0.4). The increase of VSMC stiffness was accompanied by a parallel increase in the expression of SRF by 9.8-fold and of myocardin by 10.5-fold in thoracic aortic VSMCs from SHR vs. WKY rats, resulting in a significant increase of downstream stiffness-associated genes (all, P < 0.01 vs. WKY). Inhibition of SRF/myocardin expression selectively attenuated aortic VSMC stiffening, and normalized downstream targets in VSMCs isolated from SHR but not from WKY rats. In vivo, 2 weeks of treatment with SRF/myocardin inhibitor delivered by subcutaneous osmotic minipump significantly reduced aortic stiffness and then blood pressure in SHR but not in WKY rats, although concomitant changes in aortic wall remodelling were not detected during this time frame. Conclusions SRF/myocardin pathway acts as a pivotal mediator of aortic VSMC mechanical properties and plays a central role in the pathological aortic stiffening in hypertension. Attenuation of aortic VSMC stiffening by pharmacological inhibition of SRF/myocardin signalling presents a novel therapeutic strategy for the treatment of hypertension by targeting the cellular contributors to aortic stiffness. PMID:28003268

  16. Agonistic Anti-PDGF Receptor Autoantibodies from Patients with Systemic Sclerosis Impact Human Pulmonary Artery Smooth Muscle Cells Function In Vitro

    PubMed Central

    Svegliati, Silvia; Amico, Donatella; Spadoni, Tatiana; Fischetti, Colomba; Finke, Doreen; Moroncini, Gianluca; Paolini, Chiara; Tonnini, Cecilia; Grieco, Antonella; Rovinelli, Marina; Gabrielli, Armando

    2017-01-01

    One of the earliest events in the pathogenesis of systemic sclerosis (SSc) is microvasculature damage with intimal hyperplasia and accumulation of cells expressing PDGF receptor. Stimulatory autoantibodies targeting PDGF receptor have been detected in SSc patients and demonstrated to induce fibrosis in vivo and convert in vitro normal fibroblasts into SSc-like cells. Since there is no evidence of the role of anti-PDGF receptor autoantibodies in the pathogenesis of SSc vascular lesions, we investigated the biologic effect of agonistic anti-PDGF receptor autoantibodies from SSc patients on human pulmonary artery smooth muscle cells and the signaling pathways involved. The synthetic (proliferation, migration, and type I collagen gene α1 chain expression) and contractile (smooth muscle-myosin heavy chain and smooth muscle-calponin expression) profiles of human pulmonary artery smooth muscle cells were assessed in vitro after incubation with SSc anti-PDGF receptors stimulatory autoantibodies. The role of reactive oxygen species, NOX isoforms, and mammalian target of rapamycin (mTOR) was investigated. Human pulmonary artery smooth muscle cells acquired a synthetic phenotype characterized by higher growth rate, migratory activity, gene expression of type I collagen α1 chain, and less expression of markers characteristic of the contractile phenotype such as smooth muscle-myosin heavy chain and smooth muscle-calponin when stimulated with PDGF and autoantibodies against PDGF receptor, but not with normal IgG. This phenotypic profile is mediated by increased generation of reactive oxygen species and expression of NOX4 and mTORC1. Our data indicate that agonistic anti-PDGF receptor autoantibodies may contribute to the pathogenesis of SSc intimal hyperplasia. PMID:28228756

  17. Agonistic Anti-PDGF Receptor Autoantibodies from Patients with Systemic Sclerosis Impact Human Pulmonary Artery Smooth Muscle Cells Function In Vitro.

    PubMed

    Svegliati, Silvia; Amico, Donatella; Spadoni, Tatiana; Fischetti, Colomba; Finke, Doreen; Moroncini, Gianluca; Paolini, Chiara; Tonnini, Cecilia; Grieco, Antonella; Rovinelli, Marina; Gabrielli, Armando

    2017-01-01

    One of the earliest events in the pathogenesis of systemic sclerosis (SSc) is microvasculature damage with intimal hyperplasia and accumulation of cells expressing PDGF receptor. Stimulatory autoantibodies targeting PDGF receptor have been detected in SSc patients and demonstrated to induce fibrosis in vivo and convert in vitro normal fibroblasts into SSc-like cells. Since there is no evidence of the role of anti-PDGF receptor autoantibodies in the pathogenesis of SSc vascular lesions, we investigated the biologic effect of agonistic anti-PDGF receptor autoantibodies from SSc patients on human pulmonary artery smooth muscle cells and the signaling pathways involved. The synthetic (proliferation, migration, and type I collagen gene α1 chain expression) and contractile (smooth muscle-myosin heavy chain and smooth muscle-calponin expression) profiles of human pulmonary artery smooth muscle cells were assessed in vitro after incubation with SSc anti-PDGF receptors stimulatory autoantibodies. The role of reactive oxygen species, NOX isoforms, and mammalian target of rapamycin (mTOR) was investigated. Human pulmonary artery smooth muscle cells acquired a synthetic phenotype characterized by higher growth rate, migratory activity, gene expression of type I collagen α1 chain, and less expression of markers characteristic of the contractile phenotype such as smooth muscle-myosin heavy chain and smooth muscle-calponin when stimulated with PDGF and autoantibodies against PDGF receptor, but not with normal IgG. This phenotypic profile is mediated by increased generation of reactive oxygen species and expression of NOX4 and mTORC1. Our data indicate that agonistic anti-PDGF receptor autoantibodies may contribute to the pathogenesis of SSc intimal hyperplasia.

  18. Downregulation of L-type Ca2+ channel in rat mesenteric arteries leads to loss of smooth muscle contractile phenotype and inward hypertrophic remodeling.

    PubMed

    Kudryavtseva, Olga; Herum, Kate Møller; Dam, Vibeke Secher; Straarup, Marthe Simonsen; Kamaev, Dmitry; Briggs Boedtkjer, Donna M; Matchkov, Vladimir V; Aalkjær, Christian

    2014-05-01

    L-type Ca(2+) channels (LTCCs) are important for vascular smooth muscle cell (VSMC) contraction, as well as VSMC differentiation, as indicated by loss of LTCCs during VSMC dedifferentiation. However, it is not clear whether loss of LTCCs is a primary event underlying phenotypic modulation or whether loss of LTCCs has significance for vascular structure. We used small interference RNA (siRNA) transfection in vivo to investigate the role of LTCCs in VSMC phenotypic expression and structure of rat mesenteric arteries. siRNA reduced LTCC mRNA and protein expression in rat mesenteric arteries 3 days after siRNA transfection to 12.7 ± 0.7% and 47.3 ± 13%, respectively: this was associated with an increased resting intracellular Ca(2+) concentration ([Ca(2+)]i). Despite the high [Ca(2+)]i, the contractility was reduced (tension development to norepinephrine was 3.5 ± 0.2 N/m and 0.8 ± 0.2 N/m for sham-transfected and downregulated arteries respectively; P < 0.05). Expression of contractile phenotype marker genes was reduced in arteries downregulated for LTCCs. Phenotypic changes were associated with a 45% increase in number of VSMCs and a consequent increase of media thickness and media area. Ten days after siRNA transfection arterial structure was again normalized. The contractile responses of LTCC-siRNA transfected arteries were elevated in comparison with matched controls 10 days after transfection. The study provides strong evidence for causal relationships between LTCC expression and VSMC contractile phenotype, as well as novel data addressing the complex relationship between VSMC contractility, phenotype, and vascular structure. These findings are relevant for understanding diseases, associated with phenotype changes of VSMC and vascular remodeling, such as atherosclerosis and hypertension.

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

    NASA Astrophysics Data System (ADS)

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

    2016-08-01

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

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

    PubMed Central

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

    2016-01-01

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

  1. Circumferential alignment of vascular smooth muscle cells in a circular microfluidic channel.

    PubMed

    Choi, Jong Seob; Piao, Yunxian; Seo, Tae Seok

    2014-01-01

    The circumferential alignment of human aortic smooth muscle cells (HASMCs) in an orthogonally micropatterned circular microfluidic channel is reported to form an in vivo-like smooth muscle cell layer. To construct a biomimetic smooth muscle cell layer which is aligned perpendicular to the axis of blood vessel, a half-circular polydimethylsiloxane (PDMS) microchannel is first fabricated by soft lithography using a convex PDMS mold. Then, the orthogonally microwrinkle patterns are generated inside the half-circular microchannel by a strain responsive wrinkling method. During the UV treatment on a PDMS substrate with uniaxial 40% stretch and a subsequent strain releasing step, the microwrinkle patterns perpendicular to the axial direction of the circular microchannel are generated, which can guide the circumferential alignment of HASMCs during cultivation. The analysis of orientation angle, shape index, and contractile protein marker expression indicates that the cultured HASMCs reveal the in vivo-like cell phenotype. Finally, a fully circular microchannel is produced by bonding two half-circular microchannels, and the HASMCs are cultured circumferentially inside the channels with high alignment and viability for 5 days. These results demonstrated the creation of an in vivo-like 3D smooth muscle cell layer in the circular microfluidic channel which can provide a bioassay platforms for in-depth study of HASMC biology and vascular function.

  2. Non-genomic mechanism of 17 beta-oestradiol-induced inhibition of contraction in mammalian vascular smooth muscle.

    PubMed Central

    Kitazawa, T; Hamada, E; Kitazawa, K; Gaznabi, A K

    1997-01-01

    17 beta-Oestradiol (E2) at 0.1-10 microM directly inhibited various tonic and phasic smooth muscle contractions. The mechanism(s) of oestrogen-induced inhibition of contraction was studied using intact and permeabilized strips and isolated single cells of smooth muscle. 2. In endothelium-denuded vascular smooth muscle, E2 attenuated high K(+)-induced force development and myosin light chain phosphorylation, and produced rapid and reversible relaxation. There were no significant differences in these inhibitory effects between tissue types (femoral artery vs. portal vein), species (rat vs. rabbit) or sexes. 3. The inhibitory potencies of several steroidal and non-steroidal oestrogen analogues were examined and their effects were for the most part stereo-specific. However, two steroids with negligible affinities for the nuclear oestrogen receptor also strongly inhibited high K(+)-induced contraction. 4. Genomic modulators including a protein synthesis inhibitor, an RNA synthesis inhibitor, and oestrogen receptor antagonists did not affect the inhibitory actions of E2. Inhibitors of cyclic nucleotide-dependent protein kinases did not reduce the E2 effect. 5. Ca2+ release from intracellular stores by agonists and by inositol 1,4,5-trisphosphate (IP3) does not appear to be modulated by E2. Neither pretreatment with ryanodine nor with thapsigargin affected the E2-induced inhibition of high K(+)-induced contraction. 6. E2 had no effect on either normal or GTP gamma S-increased Ca2+ sensitivity of the regulatory and contractile apparatus. 7. E2 and its analogues rapidly inhibited voltage-dependent L-type Ca2+ channel currents in isolated smooth muscle cells. Repetitive stimulation was not required for E2-induced inhibition of the currents. 8. This study strongly suggests that at pharmacological concentrations oestrogen primarily reduces Ca2+ influx through inhibition of L-type Ca2+ channels in a non-genomic manner and decreases myosin light chain phosphorylation and

  3. Targeting N-acetylglucosamine-bearing polymer-coated liposomes to vascular smooth muscle cells.

    PubMed

    Ise, Mamiko; Ise, Hirohiko; Shiba, Yuji; Kobayashi, Satoshi; Goto, Mitsuaki; Takahashi, Masafumi; Akaike, Toshihiro; Ikeda, Uichi

    2011-12-01

    The targeted delivery of anti-inflammatory agents has great therapeutic potential for treating restenosis following percutaneous coronary intervention. To develop a drug delivery system targeted to injured blood vessels, we examined whether N-acetylglucosamine (GlcNAc)-bearing polymer-coated liposomes (GlcNAc-Ls) are specifically taken up by vascular smooth muscle cells (VSMCs). Flow cytometric analysis revealed that GlcNAc-Ls were taken up by VSMCs in vitro. Furthermore, GlcNAc-Ls were intravenously administered to mice that had undergone wire-mediated vascular injury. GlcNAc-Ls markedly accumulated at the intramural site of the injured vessel walls but not at the contralateral (uninjured) vessel walls. These results demonstrated that GlcNAc-Ls can be specifically taken up by VSMCs both in vitro and in vivo. We propose a novel strategy of using GlcNAc-Ls that has potential for application in drug delivery targeted to injured blood vessels.

  4. Calcium regulation of vascular smooth muscle cell-derived matrix vesicles.

    PubMed

    Kapustin, Alexander N; Shanahan, Catherine M

    2012-07-01

    Vascular calcification is a pathological process common in patients with disorders of mineral metabolism and mediated by vascular smooth muscle cells (VSMCs). A key event in the initiation of VSMC calcification is the release of mineralization-competent matrix vesicles (MVs), small membrane-bound bodies with structural features enabling them to efficiently nucleate hydroxyapatite. These bodies are similar to MVs secreted by chondrocytes during bone development and their properties include the absence of calcification inhibitors, formation of nucleation sites, and accumulation of matrix metalloproteinases such as MMP-2. The mechanisms of MV biogenesis and loading remain poorly understood; however, emerging data have demonstrated that alterations in cytosolic calcium homeostasis can trigger multiple changes in MV composition that promote their mineralization.

  5. Chlorogenic acid prevents isoproterenol-induced DNA damage in vascular smooth muscle cells

    PubMed Central

    Wang, Jingshuai; Li, Jiyang; Liu, Jie; Xu, Mengjiao; Tong, Xiaowen; Wang, Jianjun

    2016-01-01

    Numerous clinical therapeutic agents have been identified as DNA damaging. The present study revealed that isoproterenol (Iso) resulted in DNA damage in vascular smooth muscle cells (VSMCs) and increased the levels of intracellular oxygen free radicals. Administration of chlorogenic acid (CGA) inhibited this effect. Pretreatment with CGA abrogated the increase in protein expression levels of γ-H2A histone family member X, phosphorylated ataxia telangiectasia mutated, phosphorylated Rad3-related protein, breast cancer 1 and C-terminal Src homologous kinase induced by Iso. In addition, the increase in levels of intracellular reactive oxygen species (ROS) induced by Iso was inhibited by CGA pretreatment in a dose-dependent manner. The results of the present study suggest that CGA may inhibit Iso-induced VSMC damage via the suppression of ROS generation. Therefore, CGA may be a novel agent for the treatment of vascular diseases. PMID:27634104

  6. Identification of ostruthin from Peucedanum ostruthium rhizomes as an inhibitor of vascular smooth muscle cell proliferation.

    PubMed

    Joa, Helge; Vogl, Sylvia; Atanasov, Atanas G; Zehl, Martin; Nakel, Thomas; Fakhrudin, Nanang; Heiss, Elke H; Picker, Paolo; Urban, Ernst; Wawrosch, Christoph; Saukel, Johannes; Reznicek, Gottfried; Kopp, Brigitte; Dirsch, Verena M

    2011-06-24

    Inhibition of vascular smooth muscle cell (VSMC) proliferation is of substantial interest in combating cardiovascular disease. A dichloromethane extract from the rhizomes of Peucedanum ostruthium, a traditionally used Austrian medicinal plant with anti-inflammatory properties, was examined for a putative antiproliferative activity in rat aortic VSMC. This extract inhibited serum (10%)-induced VSMC proliferation concentration dependently. Further identification and biological testing of its major constituents revealed that the coumarin ostruthin (7) is the major antiproliferative substance. In summary, a new bioactivity of P. ostruthium rhizomes is described, and 7 has been identified as the responsible compound.

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

    PubMed

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

    1989-02-01

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

  8. Vascular reactivity of rabbit isolated renal and femoral resistance arteries in renal wrap hypertension.

    PubMed

    Khammy, Makhala M; Angus, James A; Wright, Christine E

    2016-02-15

    In rabbits with cellophane renal wrap hypertension, hindquarter and total vascular resistance changes to pressor and depressor agents are amplified compared to those of normotensive rabbits. The aim of the present study was to evaluate the in vitro pharmacodynamics of hypertensive and normotensive rabbit small artery segments isolated from the renal and hindquarter vascular beds. Using wire myography, the full range (Emax) and sensitivity (EC50) to a range of agonists of segments of renal interlobar (≈ 600 µm i.d.), renal arcuate (≈ 250 µm i.d.) and deep femoral branch (≈ 250 µm i.d.) arteries were assessed under normalised conditions of passive tension. Interlobar arteries from hypertensive rabbits were more sensitive (EC50) than those from normotensive rabbits to noradrenaline (6-fold), methoxamine (3-fold) and angiotensin II (3-fold). Arcuate artery reactivity was largely unaffected by hypertension. Deep femoral arteries from hypertensive rabbits had enhanced sensitivity only to noradrenaline (2-fold) and methoxamine (4-fold). Sensitivity to relaxation by acetylcholine was unaffected by hypertension in all arteries. Deep femoral arteries from hypertensive rabbits were more sensitive to sodium nitroprusside than normotensive counterparts. Adenosine caused little relaxation in renal arteries, but full relaxation in deep femoral arteries, unaltered by hypertension. This study found substantial heterogeneity in the pharmacodynamic profile of vessels isolated from different vascular beds and between arterial segments within the kidney. These profiles were differentially affected by hypertension suggesting that hypertension per se is not a resultant of general vascular dysfunction.

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

    SciTech Connect

    Davies, P.F.

    1986-03-01

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

    PubMed Central

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

    2016-01-01

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

  11. Regulatory mechanism of human vascular smooth muscle cell phenotypic transformation induced by NELIN

    PubMed Central

    PEI, CHANGAN; QIN, SHIYONG; WANG, MINGHAI; ZHANG, SHUGUANG

    2015-01-01

    Vascular disorders, including hypertension, atherosclerosis and restenosis, arise from dysregulation of vascular smooth muscle cell (VSMC) differentiation, which can be controlled by regulatory factors. The present study investigated the regulatory mechanism of the phenotypic transformation of human VSMCs by NELIN in order to evaluate its potential as a preventive and therapeutic of vascular disorders. An in vitro model of NELIN-overexpressing VSMCs was prepared by transfection with a lentiviral (LV) vector (NELIN-VSMCs) and NELIN was slienced using an a lentiviral vector with small interfering (si)RNA in another group (LV-NELIN-siRNA-VSMCs). The effects of NELIN overexpression or knockdown on the phenotypic transformation of human VSMCs were observed, and its regulatory mechanism was studied. Compared with the control group, cells in the NELIN-VSMCs group presented a contractile phenotype with a significant increase of NELIN mRNA, NELIN protein, smooth muscle (SM)α-actin and total Ras homolog gene family member A (RhoA) protein expression. The intra-nuclear translocation of SMα-actin-serum response factor (SMα-actin-SRF) occurred in these cells simultaneously. Following exposure to Rho kinsase inhibitor Y-27632, SRF and SMα-actin expression decreased. However, cells in the LV-NELIN-siRNA-VSMCs group presented a synthetic phenotype, and the expression of NELIN mRNA, NELIN protein, SMα-actin protein and total RhoA protein was decreased. The occurrence of SRF extra-nuclear translocation was observed. In conclusion, the present study suggested that NELIN was able to activate regulatory factors of SMα-actin, RhoA and SRF successively in human VSMCs cultured in vitro. Furthermore, NELIN-induced phenotypic transformation of human VSMCs was regulated via the RhoA/SRF signaling pathway. The results of the present study provide a foundation for the use of NELIN in preventive and therapeutic treatment of vascular remodeling diseases, including varicosity and

  12. Activated mineralocorticoid receptor regulates micro-RNA-29b in vascular smooth muscle cells.

    PubMed

    Bretschneider, Maria; Busch, Bianca; Mueller, Daniel; Nolze, Alexander; Schreier, Barbara; Gekle, Michael; Grossmann, Claudia

    2016-04-01

    Inappropriately activated mineralocorticoid receptor (MR) is a risk factor for vascular remodeling with unclear molecular mechanism. Recent findings suggest that post-transcriptional regulation by micro-RNAs (miRs) may be involved. Our aim was to search for MR-dependent miRs in vascular smooth muscle cells (VSMCs) and to explore the underlying molecular mechanism and the pathologic relevance. We detected that aldosteroneviathe MR reduces miR-29bin vivoin murine aorta and in human primary and cultured VSMCs (ED50= 0.07 nM) but not in endothelial cells [quantitative PCR (qPCR), luciferase assays]. This effect was mediated by an increased decay of miR-29b in the cytoplasm with unchanged miR-29 family member or primary-miR levels. Decreased miR-29b led to an increase in extracellular matrix measured by ELISA and qPCR and enhanced VSMC migration in single cell-tracking experiments. Additionally, cell proliferation and the apoptosis/necrosis ratio (caspase/lactate dehydrogenase assay) was modulated by miR-29b. Enhanced VSMC migration by aldosterone required miR-29b regulation. Control experiments were performed with scrambled RNA and empty plasmids, by comparing aldosterone-stimulated with vehicle-incubated cells. Overall, our findings provide novel insights into the molecular mechanism of aldosterone-mediated vascular pathogenesis by identifying miR-29b as a pathophysiologic relevant target of activated MR in VSMCs and by highlighting the importance of miR processing for miR regulation.-Bretschneider, M., Busch, B., Mueller, D., Nolze, A., Schreier, B., Gekle, M., Grossmann, C. Activated mineralocorticoid receptor regulates micro-RNA-29b in vascular smooth muscle cells.

  13. Phenylephrine Decreases Vascular Tension in Goat Arteries in Specific Circumstances

    PubMed Central

    Raj, Renu R.

    2016-01-01

    Phenylephrine (PE) causes vasoconstriction through alpha adrenergic receptors. PE-induced vasodilatation has also been reported earlier in pre-constricted vessels. Here we demonstrate in spiral strips of goat arteries that addition of PE can decrease tone even from base-line levels (i.e. not pre-constricted) and show that this process requires nitric oxide (NO) and alpha adrenergic stimulation, but is cGMP-independent. Under control conditions, PE caused vasoconstriction, but under conditions where NO levels are higher, as with L-Arginine or sodium nitroprusside, PE decreased vessel tension. L-Arginine/PE combination was not able to decrease tension when alpha adrenoceptors were blocked with Phentolamine or endothelial nitric oxide synthase (eNOS) was blocked with Nω-Nitro-L-arginine (L-NNA). Propranolol, a beta blocker, was unable to prevent the reduction in tension by the L-Arginine/PE combination. Adrenaline and noradrenaline (and not isoproterenol) also reduced vessel tension in the presence of L-Arginine. Even when NO levels were not enhanced, relieving NO from having to stimulate the enzyme soluble guanylyl cyclase (sGC) (either by using sGC blockers, namely ODQ or methylene blue, or by enhancing cGMP levels (with sildenafil) which by negative feedback probably inhibits sGC) led to PE-induced reduction of vascular tension. PMA—phorbol myristate acetate—an agonist which stimulates Protein Kinase C was able to prevent the ability of PE to reduce vascular tension in a high NO environment. Our conclusion is that PE reduces vascular tension through alpha adrenoceptors if there is excess NO availability to activate a putative pathway. Though the reduction of vessel tone by PE is dependent on NO, it is independent of cGMP. Prior treatment with PMA or PE itself can prevent further PE-induced reduction of tension in a high NO environment. The results here suggest, counter-intuitively, that alpha blockers may be of help in the treatment of septic shock where

  14. Phenylephrine Decreases Vascular Tension in Goat Arteries in Specific Circumstances.

    PubMed

    Raj, Renu R; Subramani, Sathya

    2016-01-01

    Phenylephrine (PE) causes vasoconstriction through alpha adrenergic receptors. PE-induced vasodilatation has also been reported earlier in pre-constricted vessels. Here we demonstrate in spiral strips of goat arteries that addition of PE can decrease tone even from base-line levels (i.e. not pre-constricted) and show that this process requires nitric oxide (NO) and alpha adrenergic stimulation, but is cGMP-independent. Under control conditions, PE caused vasoconstriction, but under conditions where NO levels are higher, as with L-Arginine or sodium nitroprusside, PE decreased vessel tension. L-Arginine/PE combination was not able to decrease tension when alpha adrenoceptors were blocked with Phentolamine or endothelial nitric oxide synthase (eNOS) was blocked with Nω-Nitro-L-arginine (L-NNA). Propranolol, a beta blocker, was unable to prevent the reduction in tension by the L-Arginine/PE combination. Adrenaline and noradrenaline (and not isoproterenol) also reduced vessel tension in the presence of L-Arginine. Even when NO levels were not enhanced, relieving NO from having to stimulate the enzyme soluble guanylyl cyclase (sGC) (either by using sGC blockers, namely ODQ or methylene blue, or by enhancing cGMP levels (with sildenafil) which by negative feedback probably inhibits sGC) led to PE-induced reduction of vascular tension. PMA-phorbol myristate acetate-an agonist which stimulates Protein Kinase C was able to prevent the ability of PE to reduce vascular tension in a high NO environment. Our conclusion is that PE reduces vascular tension through alpha adrenoceptors if there is excess NO availability to activate a putative pathway. Though the reduction of vessel tone by PE is dependent on NO, it is independent of cGMP. Prior treatment with PMA or PE itself can prevent further PE-induced reduction of tension in a high NO environment. The results here suggest, counter-intuitively, that alpha blockers may be of help in the treatment of septic shock where nitric

  15. Vascular Access System for Continuous Arterial Infusion of a Protease Inhibitor in Acute Necrotizing Pancreatitis

    SciTech Connect

    Ganaha, Fumikiyo; Yamada, Tetsuhisa; Yorozu, Naoya; Ujita, Masuo; Irie, Takeo; Fukuda, Yasushi; Fukuda, Kunihiko; Tada, Shimpei

    1999-09-15

    We used a vascular access system (VAS) for continuous arterial infusion (CAI) of a protease inhibitor in two patients with acute necrotizing pancreatitis. The infusion catheter was placed into the dorsal pancreatic artery in the first patient and into the gastroduodenal artery in the second, via a femoral artery approach. An implantable port was then connected to the catheter and was secured in a subcutaneous pocket prepared in the right lower abdomen. No complications related to the VAS were encountered. This system provided safe and uncontaminated vascular access for successful CAI for acute pancreatitis.

  16. Anatomy of the medial femoral circumflex artery with respect to the vascularity of the femoral head.

    PubMed

    Zlotorowicz, M; Szczodry, M; Czubak, J; Ciszek, B

    2011-11-01

    We performed a series of 16 anatomical dissections on Caucasian cadaver material to determine the surgical anatomy of the medial femoral circumflex artery (MFCA) and its anastomoses. These confirmed that the femoral head receives its blood supply primarily from the MFCA via a group of posterior superior nutrient arteries and the posterior inferior nutrient artery. In terms of anastomoses that may also contribute to the blood supply, the anastomosis with the inferior gluteal artery, via the piriformis branch, is the most important. These dissections provide a base of knowledge for further radiological studies on the vascularity of the normal femoral head and its vascularity after dislocation of the hip.

  17. Differential Effects of Long Term FTY720 Treatment on Endothelial versus Smooth Muscle Cell Signaling to S1P in Rat Mesenteric Arteries.

    PubMed

    Hamidi Shishavan, Mahdi; Bidadkosh, Arash; Yazdani, Saleh; Lambooy, Sebastiaan; van den Born, Jacob; Buikema, Hendrik; Henning, Robert H; Deelman, Leo E

    2016-01-01

    The sphingosine-1-phosphate (S1P) analog FTY720 exerts pleiotropic effects on the cardiovascular system and causes down-regulation of S1P receptors. Myogenic constriction is an important mechanism regulating resistance vessel function and is known to be modulated by S1P. Here we investigated myogenic constriction and vascular function of mesenteric arteries of rats chronically treated with FTY720. Wistar rats received FTY720 1mg/kg/daily for six weeks. At termination, blood pressure was recorded and small mesenteric arteries collected for vascular studies in a perfusion set up. Myogenic constriction to increased intraluminal pressure was low, but a sub-threshold dose of S1P profoundly augmented myogenic constriction in arteries of both controls and animals chronically treated with FTY720. Interestingly, endothelial denudation blocked the response to S1P in arteries of FTY720-treated animals, but not in control rats. In acute experiments, presence of FTY720 significantly augmented the contractile response to S1P, an effect that was partially abolished after the inhibition of cyclooxygenase (COX-)-derived prostaglandins. FTY720 down regulated S1P1 but not S1P2 in renal resistance arteries and in cultured human endothelial cells. This study therefore demonstrates the endothelium is able to compensate for the complete loss of responsiveness of the smooth muscle layer to S1P after long term FTY720 treatment through a mechanism that most likely involves enhanced production of contractile prostaglandins by the endothelium.

  18. A phosphorus nuclear magnetic resonance study of metabolites and intracellular pH in rabbit vascular smooth muscle.

    PubMed Central

    Spurway, N C; Wray, S

    1987-01-01

    1. 31P nuclear magnetic resonance (n.m.r.) spectroscopy was used to investigate metabolites, intracellular pH (pHi) and the effects of pHi on tone in rabbit blood vessels. The vessels were bathed in mammalian Ringer solution and maintained at 20 degrees C while inside the spectrometer. 2. Vascular spectra showed relatively low phosphocreatine (PCr) concentrations compared to skeletal muscle. The [PCr]/[ATP] ratio was only 1.32 +/- 0.09 (n = 7). There was also a prominent phosphomonoester (PME) peak. Similar features have been reported for other smooth muscles examined by 31P n.m.r. 3. The [PCr] was higher and the inorganic phosphate (Pi) concentration lower than values deduced from chemical analysis of arterial extracts. However, the [PCr] value fell within the range obtained for other smooth muscles when studied by 31P n.m.r. 4. Measurement of pHi under control conditions (external pH 7.25) gave a mean value of 7.19 +/- 0.03 at 20 degrees C (n = 5). Metabolic inhibition brought about by 0.5 mM-cyanide and 0.2 mM-fluoride did not significantly alter pHi. At higher inhibitor concentrations (3 and 1 mM respectively) there was a significant acidosis. 5. The effects of NH4Cl upon pH were investigated in metabolically inhibited preparations. During 10 min applications of 30 mM-NH4Cl (isosmotically substituted for NaCl) the pHi rose; during subsequent NH4Cl removal it fell below control values. In the least inhibited tissues the total pHi excursion between NH4Cl applications and removals was 0.5 unit. 6. Rabbit ear vessels have been found to increase vascular tone during manoeuvres which were expected to decrease pHi. From the direct measurement of pHi reported in this study, it is concluded that the vascular tone changes brought about by NH4Cl application and withdrawal may be attributed to the alteration of pHi. PMID:3446806

  19. [Effect of potassium ions on the contractile activity of renal artery smooth muscle].

    PubMed

    Orlov, R S; Aĭvar, Iu P

    1979-07-01

    Study of isolated segments of renal arteries in rabbits showed that decrease of potassium ion concentration in the bathing fluid was followed by increase in tension, while its increase from 5 meq/l to 10 meq/l was accompanied by gradual relaxation of vessel muscles and increase of their sensitivity to noradrenalin (NA). This relationship was lacking in segments activated with NA. The ability of NA and angiotensin to activate renal arterial muscles by electromechanic and pharmacomechanic coupling mechanismes was proved experimentally. The paper discussed the role of the cell membrane sodium potassium pump in vascular muscles.

  20. Ursolic acid suppresses leptin-induced cell proliferation in rat vascular smooth muscle cells.

    PubMed

    Yu, Ya-Mei; Tsai, Chiang-Chin; Tzeng, Yu-Wen; Chang, Weng-Cheng; Chiang, Su-Yin; Lee, Ming-Fen

    2017-01-29

    Accumulating lines of evidence indicate that high leptin levels are associated with adverse cardiovascular health in obese individuals. Proatherogenic effects of leptin include endothelial cell activation, vascular smooth muscle cell proliferation and migration. Ursolic acid (UA) has been reported to exhibit multiple biological effects including antioxidant and anti-inflammatory properties. In this study, we investigated the effect of UA on leptin-induced biological responses in rat vascular smooth muscle cells (VSMCs). A-10 VSMCs were treated with leptin in the presence or absence of UA. Intracellular reactive oxygen species (ROS) was probed by 2',7'-dichlorofluorescein diacetate. The expression of extracellular signal-regulated kinase (ERK)1/2, phospho-(ERK)1/2, nuclear factor-kappa B (NF-κB) p65 and p50, and matrix metalloproteinase-2 (MMP2) was determined by Western blotting. Immunocytochemistry and confocal laser scanning microscopy were also used for the detection of NF-κB. The secretion of MMP2 was detected by gelatin zymography. UA exhibited antioxidant activities in vitro. In rat VSMCs, UA effectively inhibited cell growth and the activity of MMP2 induced by leptin. These suppressive effects appeared by decreasing the activation of (ERK)1/2, the nuclear expression and translocation of NF-κB, and the production of ROS. UA appeared to inhibit leptin-induced atherosclerosis, which may prevent the development of obesity-induced cardiovascular diseases.

  1. The apoptosis induced by HMME-based photodynamic therapy in rabbit vascular smooth muscle cells

    NASA Astrophysics Data System (ADS)

    Yin, Huijuan; Li, Xiaoyuan; Lin, Hong; Liu, Jianzhong; Yu, Hongkui

    2007-02-01

    Objective To study the effects of HMME-based photodynamic therapy on proliferation and apoptosis of rabbit vascular smooth muscle cells(VSMCs). Method The cytotoxic effect of HMME-PDT on rabbit vascular smooth muscle cells was studied by means of Trypan Blue assay, HMME at 10μg/ml concentration and the light dose at 2.4~4.8 J/cm2 were selected in the studies. The morphological character 24h post-PDT was investigated by HE Staining. Annexin V and propidium iodide (PI) binding assays were performed to analyze the characteristics of cell death after HMME-PDT. Furthermore, The intracellular distributions of the HMME were measured by the confocal laser scanning microscope. Result It was showed the photocytotoxity to VSMC cells was dose related by Trypan Blue assay. Histology observing suggests HMME-PDT could induce cell death through apoptosis or necrosis, and the apoptosic rate was up to 50.5% by AnnexinV /PI assay. Moreover, the fluorescence images of HMME intracellular localization demonstrated that the HMME diffused into the mitochondria. Conclusion HMME-PDT could significantly inhibite VSMC proliferation and induce apoptosis.

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

    SciTech Connect

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

    2013-11-22

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

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

    SciTech Connect

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

    1986-07-01

    The distribution and effects of the two neuropeptides, vasoactive intestinal polypeptide (VIP) and peptide histidine isoleucine amide (PHI), on vascular and nonvascular smooth muscle in the urogenital tract of nonpregnant rabbit female, were investigated. Immunoreactive VIP and PHI were present in all regions except the ovary with the highest concentration in the uterin cervix. By using in vitro tension recordings of myometrial specimens, it was demonstrated that both peptides displayed a dose-dependent inhibition of the mechanical activity. The dose-response curves of VIP and PHI were superimposable with and ID50 of 3 x 10 Y mol/l, and their combined effect was additive. In addition, the influence of the two peptides on myometrial blood flow (MBF) was investigated by the xenon-133 washout technique. Both peptides were found to increase MBF with the same potency and efficacy. Their combined effect was additive. In conclusion VIP and PHI are present in the rabbit urogenital tract, and the two peptides are equipotent inhibitors of mechanical nonvascular and vascular smooth muscle activity in the uterus.

  4. Hypoxia stimulates the autocrine regulation of migration of vascular smooth muscle cells via HIF-1alpha-dependent expression of thrombospondin-1.

    PubMed

    Osada-Oka, Mayuko; Ikeda, Takako; Akiba, Satoshi; Sato, Takashi

    2008-08-01

    The migration of vascular smooth muscle cells from the media to intima and their subsequent proliferation are critical causes of arterial wall thickening. In atherosclerotic lesions increases in the thickness of the vascular wall and the impairment of oxygen diffusion capacity result in the development of hypoxic lesions. We investigated the effect of hypoxia on the migration of human coronary artery smooth muscle cells (CASMCs) via HIF-1alpha-dependent expression of thrombospondin-1 (TSP-1). When the cells were cultured under hypoxic conditions, mRNA and protein levels of TSP-1, and mRNA levels of integrin beta(3) were increased with the increase in HIF-1alpha protein. DNA synthesis and migration of the cells were stimulated under the conditions, and a neutralizing anti-TSP-1 antibody apparently suppressed the migration, but not DNA synthesis. The migration was also inhibited by RGD peptide that binds to integrin beta(3). Furthermore, the migration was completely suppressed in HIF-1alpha-knockdown cells exposed to hypoxia, while it was significantly enhanced in HIF-1alpha-overexpressing cells. These results suggest that the hypoxia induces the migration of CASMCs, and that the migration is elicited by TSP-1 of which induction is fully dependent on the stabilization of HIF-1alpha, in autocrine regulation. Thus we suggest that HIF-1alpha plays an important role in the pathogenesis of atherosclerosis.

  5. Consequences of postnatal vascular smooth muscle EGFR deletion on acute angiotensin II action.

    PubMed

    Schreier, Barbara; Hünerberg, Mirja; Rabe, Sindy; Mildenberger, Sigrid; Bethmann, Daniel; Heise, Christian; Sibilia, Maria; Offermanns, Stefan; Gekle, Michael

    2016-01-01

    Epi dermal growth factor (EGF) receptor (EGFR) is activated by its canonical ligands and transactivated by various vasoactive substances, e.g. angiotensin II (Ang II). Vascular EGFR has been proposed to be involved in vascular tissue homoeostasis and remodelling. Thus, most studies have focused on its role during long-term vascular changes whereas the relevance for acute regulation of vascular function in vivo and ex vivo is insufficiently understood. To investigate the postnatal role of VSMCs (vascular smooth muscle cells) EGFR in vivo and ex vivo, we generated a mouse model with cell-specific and inducible deletion of VSMC EGFR and studied the effect on basal blood pressure, acute pressure response to, among others, Ang II in vivo as well as ex vivo, cardiovascular tissue homoeostasis and vessel morphometry in male mice. In knockout (KO) animals, systolic, diastolic and mean blood pressures were reduced compared with wild-type (WT). Furthermore, Ang II-induced pressure load was lower in KO animals, as was Ang II-induced force development and extracellular-signal-regulated kinase 1 and 2 (ERK1/2) phosphorylation in aortic rings from KO animals. By contrast, we observed no difference in force development during application of serotonin, KCl, endothelin-1 or endothelin-1-induced pressure load in KO animals. In addition, nitric oxide (NO)-mediated vasodilation was not affected. Heart weight (HW) increase and up-regulation of aortic and cardiac expression of Ccl2 (chemoattractant protein-2) and serpinE1 (plasminogen activator inhibitor 1) during the transition from 4- to 10-months of age were prevented by VSMC EGFR KO. We conclude that VSMC EGFR is involved in basal blood pressure homoeostasis and acute pressure response to Ang II, and thereby contributes to maturation-related remodelling.

  6. Equol increases cerebral blood flow in rats via activation of large-conductance Ca(2+)-activated K(+) channels in vascular smooth muscle cells.

    PubMed

    Yu, Wei; Wang, Yan; Song, Zheng; Zhao, Li-Mei; Li, Gui-Rong; Deng, Xiu-Ling

    2016-05-01

    The present study was designed to investigate the effect of equol on cerebral blood flow and the underlying molecular mechanisms. The regional cerebral blood flow in parietal lobe of rats was measured by using a laser Doppler flowmetry. Isolated cerebral basilar artery and mesenteric artery rings from rats were used for vascular reactivity measurement with a multi wire myography system. Outward K(+) current in smooth muscle cells of cerebral basilar artery, large-conductance Ca(2+)-activated K(+) (BK) channel current in BK-HEK 293 cells stably expressing both human α (hSlo)- and β1-subunits, and hSlo channel current in hSlo-HEK 293 cells expressing only the α-subunit of BK channels were recorded with whole cell patch-clamp technique. The results showed that equol significantly increased regional cerebral blood flow in rats, and produced a concentration-dependent but endothelium-independent relaxation in rat cerebral basilar arteries. Both paxilline and iberiotoxin, two selective BK channel blockers, significantly inhibited equol-induced vasodilation in cerebral arteries. Outward K(+) currents in smooth muscle cells of cerebral basilar artery were increased by equol and fully reversed by washout or blockade of BK channels with iberiotoxin. Equol remarkably enhanced human BK current in BK-HEK 293 cells, but not hSlo current in hSlo-HEK 293 cells, and the increase was completely abolished by co-application of paxilline. Our findings provide the first information that equol selectively stimulates BK channel current by acting on its β1 subunit, which may in turn contribute to the equol-mediated vasodilation and cerebral blood flow increase.

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

    PubMed

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

    1988-01-01

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

  8. Resveratrol prevents hypoxia-induced arginase II expression and proliferation of human pulmonary artery smooth muscle cells via Akt-dependent signaling.

    PubMed

    Chen, Bernadette; Xue, Jianjing; Meng, Xiaomei; Slutzky, Jessica L; Calvert, Andrea E; Chicoine, Louis G

    2014-08-15

    Pulmonary artery smooth muscle cell (PASMC) proliferation plays a fundamental role in the vascular remodeling seen in pulmonary hypertensive diseases associated with hypoxia. Arginase II, an enzyme regulating the first step in polyamine and proline synthesis, has been shown to play a critical role in hypoxia-induced proliferation of human PASMC (hPASMC). In addition, there is evidence that patients with pulmonary hypertension have elevated levels of arginase in the vascular wall. Resveratrol, a natural polyphenol found in red wine and grape skins, has diverse biochemical and physiological actions including antiproliferative properties. Furthermore, resveratrol has been shown to attenuate right ventricular and pulmonary artery remodeling, both pathological components of pulmonary hypertension. The present studies tested the hypothesis that resveratrol would prevent hypoxia-induced pulmonary artery smooth muscle cell proliferation by inhibiting hypoxia-induced arginase II expression. Our data indicate that hypoxia-induced hPASMC proliferation is abrogated following treatment with resveratrol. In addition, the hypoxic induction of arginase II was directly attenuated by resveratrol treatment. Furthermore, we found that the inhibitory effect of resveratrol on arginase II in hPASMC was mediated through the PI3K-Akt signaling pathway. Supporting these in vitro findings, resveratrol normalized right ventricular hypertrophy in an in vivo neonatal rat model of chronic hypoxia-induced pulmonary hypertension. These novel data support the notion that resveratrol may be a potential therapeutic agent in pulmonary hypertension by preventing PASMC arginase II induction and proliferation.

  9. Sildenafil inhibits chronically hypoxic upregulation of canonical transient receptor potential expression in rat pulmonary arterial smooth muscle

    PubMed Central

    Lu, Wenju; Zhang, Dandan; Peng, Gongyong; Li, Bing; Zhong, Nanshan

    2010-01-01

    In pulmonary arterial smooth muscle cells (PASMCs), Ca2+ influx through store-operated Ca2+ channels thought to be composed of canonical transient receptor potential (TRPC) proteins is an important determinant of intracellular free calcium concentration ([Ca2+]i) and pulmonary vascular tone. Sildenafil, a type V phosphodiesterase inhibitor that increases cellular cGMP, is recently identified as a promising agent for treatment of pulmonary hypertension. We previously demonstrated that chronic hypoxia elevated basal [Ca2+]i in PASMCs due in large part to enhanced store-operated Ca2+ entry (SOCE); moreover, ex vivo exposure to prolonged hypoxia (4% O2 for 60 h) upregulated TRPC1 and TRPC6 expression in PASMCs. We examined the effect of sildenafil on basal [Ca2+]i, SOCE, and the expression of TRPC in PASMCs under prolonged hypoxia exposure. We also examined the effect of sildenafil on TRPC1 and TRPC6 expression in pulmonary arterial smooth muscle (PA) from rats that developed chronically hypoxic pulmonary hypertension (CHPH). Compared with vehicle control, treatment with sildenafil (300 nM) inhibited prolonged hypoxia induced increases of 1) basal [Ca2+]i, 2) SOCE, and 3) mRNA and protein expression of TRPC in PASMCs. Moreover, sildenafil (50 mg · kg−1 · day−1) inhibited mRNA and protein expression of TRPC1 and TRPC6 in PA from chronically hypoxic (10% O2 for 21 days) rats, which was associated with decreased right ventricular pressure and right ventricular hypertrophy. Furthermore, we found, in PASMCs exposed to prolonged hypoxia, that knockdown of TRPC1 or TRPC6 by their specific small interference RNA attenuated the hypoxic increases of SOCE and basal [Ca2+]i, suggesting a cause and effect link between increases of TRPC1 and TRPC6 expression and the hypoxic increases of SOCE and basal [Ca2+]i. These results suggest that sildenafil may alter basal [Ca2+]i in PASMCs by decreasing SOCE through downregulation of TRPC1 and TRPC6 expression, thereby contributing to

  10. TRPM8 downregulation by angiotensin II in vascular smooth muscle cells is involved in hypertension.

    PubMed

    Huang, Fang; Ni, Min; Zhang, Jing-Ming; Li, Dong-Jie; Shen, Fu-Ming

    2017-04-01

    Angiotensin II (Ang II)-induced injury of vascular smooth muscle cells (VSMCs) serves an important role in hypertension and other cardiovascular disorders. Transient receptor potential melastatin 8 (TRPM8) is a thermally‑regulated Ca2+‑permeable channel that is activated by reduced body temperature. Although several recent studies have revealed the regulatory effect of TRPM8 in vascular tone and hypertension, the precise role of TRPM8 in dysfunction of vascular smooth muscle cells (VSMCs) induced by Ang II remains elusive. In the present study, the possible function of TRPM8 in Ang II‑induced VSMCs malfunction in vivo and in vitro was investigated. In the aortae from rats that had undergone a two‑kidney one‑clip operation, which is a widely‑used renovascular hypertension model, the mRNA and protein levels of TRPM8 were reduced. In addition, exogenous Ang II treatment decreased TRPM8 mRNA and protein expression levels in primary cultures of rat VSMCs. TRPM8 activation by menthol, a pharmacological agonist, in VSMCs, significantly attenuated the Ang II‑induced increase in reactive oxygen species and H2O2 production. In addition, TRPM8 activation reduced the Ang II‑induced upregulation of NADPH oxidase (NOX) 1 and NOX4 in VSMCs. Furthermore, TRPM8 activation relieved the Ang II‑induced activation of ras homolog gene family, member A‑rho associated protein kinase 2 and janus kinase 2 signaling pathways in VSMCs. In conclusion, the results presented in the current study indicated that TRPM8 downregulation by Ang II in VSMCs may be involved in hypertension.

  11. Vascular smooth muscle contraction evoked by cell volume modulation: role of the cytoskeleton network.

    PubMed

    Koltsova, Svetlana V; Gusakova, Svetlana V; Anfinogenova, Yana J; Baskakov, Mikhail B; Orlov, Sergei N

    2008-01-01

    Previously, we reported that hyposmotic swelling evoked transient vascular smooth muscle cell (SMC) contraction that was completely abolished by L-type Ca(2+) channel blockers. In contrast, sustained contraction revealed in hyper- and isoosmotically-shrunken SMCs was insensitive to L-type channel blockers and was diminished in Ca(2+)-free medium by only 30-50%. Several research groups reported cell volume-dependent cytoskeleton network rearrangements. This study examines the role of cytoskeleton proteins in cell volume-dependent contraction of endothelium-denuded vascular smooth muscle rings (VSMR) from the rat thoracic aorta. Hyperosmotic shrinkage and hyposmotic swelling were triggered by modulation of medium osmolality; isosmotic shrinkage was induced by VSMR transfer from hypo- to isosmotic medium. The relative content of globular (G) and fibrillar (F) actin was estimated by fluorescence microscopy. Hyperosmotic shrinkage and hyposmotic swelling led to elevation of the F-actin/G-actin ratio by 2.5- and 1.8-fold respectively. Contraction of shrunken and swollen VSMR was insensitive to modulators of microtubules such as vinblastine, colchicine and docetaxel. Microfilament disassembly by cytochalasin B resulted in dramatic attenuation of the maximal amplitude of contraction of hyperosmotically-shrunken and hyposmotically-swollen VSMR, and almost completely abolished the contraction triggered by isosmotic shrinkage. These data suggest that both L-type Ca(2+) channel-mediated contraction of swollen vascular SMC and Ca(2+)(o)-insensitive contractions of shrunken cells are triggered by reorganization of the microfilament network caused by elevation of the F-actin/G-actin ratio.

  12. Benidipine, a calcium channel blocker, regulates proliferation and phenotype of vascular smooth muscle cells.

    PubMed

    Arakawa, Emi; Hasegawa, Kazuhide

    2006-02-01

    Hyperproliferation of phenotypically modified vascular smooth muscle cells (VSMCs) is one of the major factors in the development of atherosclerosis and restenosis. Previously it was demonstrated that benidipine, a dihydropyridine-calcium channel antagonist, reduced neointimal formation in a rat balloon-injury model. In the present study, we examined the effect of benidipine on the phenotypic modulation and proliferation of VSMCs, using primary cultures of rat VSMCs. In the absence of drug treatment, protein levels of the smooth muscle specific markers, such as smooth muscle myosin heavy chain-1 (SM1), calponin 1, and alpha-actin, decreased during culture. However, treatment of VSMCs with benidipine (3 - 10 micromol/L) for 1 week reversed the effect in a concentration-related manner so that high levels of marker proteins were maintained. The expression of calponin mRNAs was reduced markedly during 1-week culture, and treatment with benidipine (3 micromol/L) significantly inhibited the reduction. Treatment with benidipine for 2 days increased the level of p21 protein and partially reduced p70 S6 kinase 1 (p70S6K1) activity. These data suggest that benidipine may arrest the growth of VSMCs, thereby preventing cell dedifferentiation. These additional properties of benidipine suggest that the drug should provide useful therapy for atherosclerosis and restenosis.

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

    PubMed

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

    2004-02-20

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

  14. Functional up-regulation of KCNA gene family expression in murine mesenteric resistance artery smooth muscle

    PubMed Central

    Fountain, S J; Cheong, A; Flemming, R; Mair, L; Sivaprasadarao, A; Beech, D J

    2004-01-01

    This study focused on the hypothesis that KCNA genes (which encode KVα1 voltage-gated K+ channels) have enhanced functional expression in smooth muscle cells of a primary determinant of peripheral resistance – the small mesenteric artery. Real-time PCR methodology was developed to measure cell type-specific in situ gene expression. Profiles were determined for arterial myocyte expression of RNA species encoding KVα1 subunits as well as KVβ1, KVα2.1, KVγ9.3, BKCaα1 and BKCaβ1. The seven major KCNA genes were expressed and more readily detected in endothelium-denuded mesenteric resistance artery compared with thoracic aorta; quantification revealed dramatic differential expression of one to two orders of magnitude. There was also four times more RNA encoding KVα2.1 but less or similar amounts encoding KVβ1, KVγ9.3, BKCaα1 and BKCaβ1. Patch-clamp recordings from freshly isolated smooth muscle cells revealed dominant KVα1 K+ current and current density twice as large in mesenteric cells. Therefore, we suggest the increased RNA production of the resistance artery impacts on physiological function, although there is quantitatively less K+ current than might be expected. The mechanism conferring up-regulated expression of KCNA genes may be common to all the gene family and play a functional role in the physiological control of blood pressure. PMID:14742730

  15. A collagen/smooth muscle cell-incorporated elastic scaffold for tissue-engineered vascular grafts.

    PubMed

    Park, In Su; Kim, Sang-Heon; Kim, Young Ha; Kim, Ik Hwan; Kim, Soo Hyun

    2009-01-01

    Biodegradable tubular scaffolds have been developed for vascular graft application. This study was focused to improve the adhesion and proliferation of vascular smooth muscle cells (SMCs) in a tubular scaffold. Tubular scaffolds (ID 4 mm, OD 6 mm) were fabricated from a biodegradable elastic polymer, poly(L-lactide-co-epsilon-caprolactone) (PLCL) (50:50, M(n) 1.58 x 10(5)), by an extrusion/particulate leaching method. SMCs suspended in a collagen solution were infiltrated in tubular PLCL scaffolds under vacuum and incubated for 1 h at 37 degrees C to form a collagenous gel. Results from SEM image analysis showed that collagen was infiltrated into the inside of the scaffolds. Cell adhesion and proliferation rate increased in collagen/SMC-incorporated tubular PLCL scaffolds as compared with the scaffolds in which only SMCs were seeded. From SEM image and histological analysis, we further found that SMCs grew on the inside as well as on the surface of collagen/SMCs-incorporated scaffolds and the cells continued to grow as a monolayer on collagen fibers. In particular, cell proliferation and elastin contents were the highest in a PLCL scaffold with 50-100 microm pore size than any other scaffolds used in this experiment. A collagen/SMC-incorporated PLCL scaffold may support SMC growth and functions and can be used as a scaffold for tissue engineering to facilitate small-diameter vascular-tissue formation.

  16. Curcumin attenuates osteogenic differentiation and calcification of rat vascular smooth muscle cells.

    PubMed

    Hou, Menglin; Song, Yan; Li, Zhenlin; Luo, Chufan; Ou, Jing-Song; Yu, Huimin; Yan, Jianyun; Lu, Lihe

    2016-09-01

    Vascular calcification has been considered as a biological process resembling bone formation involving osteogenic differentiation. It is a major risk factor for cardiovascular morbidity and mortality. Previous studies have shown the protective effects of curcumin on cardiovascular diseases. However, whether curcumin has effects on osteogenic differentiation and calcification of vascular smooth muscle cells (VSMCs) has not been reported. In the present study, we used an in vitro model of VSMC calcification to investigate the role of curcumin in the progression of rat VSMC calcification. Curcumin treatment significantly reduced calcification of VSMCs in a dose-dependent manner, detected by alizarin red staining and calcium content assay. Similarly, ALP activity and expression of bone-related molecules including Runx2, BMP2, and Osterix were also decreased in VSMCs treated with curcumin. In addition, flow cytometry analysis and caspase-3 activity assay revealed that curcumin treatment significantly suppressed apoptosis of VSMCs, which plays an important role during vascular calcification. Furthermore, we found that pro-apoptotic molecules including p-JNK and Bax were up-regulated in VSMCs treated with calcifying medium, but they were reduced in VSMCs after curcumin treatment. However, curcumin treatment has no effect on expression of NF-κB p65. Taken together, these findings suggest that curcumin attenuates apoptosis and calcification of VSMCs, presumably via inhibition of JNK/Bax signaling pathway.

  17. Vascular Smooth Muscle Cell Stiffness as a Mechanism for Increased Aortic Stiffness with Aging

    PubMed Central

    Qiu, Hongyu; Zhu, Yi; Sun, Zhe; Trzeciakowski, Jerome P.; Gansner, Meredith; Depre, Christophe; Resuello, Ranillo R.G.; Natividad, Filipinas F.; Hunter, William C.; Genin, Guy M.; Elson, Elliot L.; Vatner, Dorothy E.; Meininger, Gerald A.; Vatner, Stephen F.

    2010-01-01

    Rationale Increased aortic stiffness, an important feature of many vascular diseases, e.g., aging, hypertension, atherosclerosis and aortic aneurysms, is assumed due to changes in extracellular matrix (ECM). Objective We tested the hypothesis that the mechanisms also involve intrinsic stiffening of vascular smooth muscle cells (VSMCs). Methods and Results Stiffness was measured in vitro both by atomic force microscopy (AFM) and in a reconstituted tissue model, using VSMCs from aorta of young versus old male monkeys (Macaca fascicularis, n=7/group), where aortic stiffness increases by 200 % in vivo. The apparent elastic modulus was increased (P<0.05) in old VSMCs (41.7±0.5 kPa) versus young (12.8±0.3 kPa), but not after disassembly of the actin cytoskeleton with cytochalasin D. Stiffness of the VSMCs in the reconstituted tissue model was also higher (P<0.05) in old (23.3±3.0 kPa) than in young (13.7±2.4 kPa). Conclusions These data support the novel concept, not appreciated previously, that increased vascular stiffness with aging is due not only to changes in ECM, but also to intrinsic changes in VSMCs. PMID:20634486

  18. Kaurane and pimarane-type diterpenes from the Viguiera species inhibit vascular smooth muscle contractility.

    PubMed

    Ambrosio, Sergio R; Tirapelli, Carlos R; da Costa, Fernando B; de Oliveira, Ana M

    2006-08-01

    The research, development and use of natural products as therapeutic agents, especially those derived from plants, have been increasing in recent years. Despite the fact that plants provide a rich source of novel biologically active compounds, only a small percentage have been phytochemically investigated and studied for their medical potential. Viguiera is a genus that belongs to the family Asteraceae and to the sunflower tribe Heliantheae, which is widespread mostly in Mexico and in other areas of the Andes and upland areas of Brazil. A review on the secondary metabolites pointed out that sesquiterpene lactones and diterpenes, of the kaurane and pimarane-type, are the main compounds produced by these plants. Some reports have shown that kaurane- and pimarane-type diterpenes exert several biological activities such as anti-inflammatory action, antimicrobial and antispasmodic activities. Kaurenoic and pimaradienoic acids, which are the main secondary metabolites isolated by our research group from the roots of Viguiera robusta and V. arenaria, respectively, have been evaluated on vascular smooth muscle contractility. We showed that these diterpenoids are able to inhibit the vascular contractility mainly by blocking extracellular Ca(2+) influx. Additionally, in this review we discuss the structure-activity relationship of the diterpenes regarding their inhibitory activity on vascular contractility.

  19. Biphasic responses of human vascular smooth muscle cells to magnesium ion

    PubMed Central

    Ma, Jun; Zhao, Nan; Zhu, Donghui

    2016-01-01

    Magnesium-based alloys are promising in biodegradable cardiovascular stent applications. The degradation products of magnesium stents may have significant impacts on the surrounding vascular cells. However, knowledge on the interactions between magnesium ion and vascular cells at the molecular and cellular levels is still largely missing. Vascular smooth muscle cell (SMC) plays an important role in the pathogenesis of restenosis and wound healing after stent implantation. This study evaluated the short-term effects of extracellular magnesium ion (Mg2+) on the cellular behaviors of SMCs. Cellular responses to Mg2+ were biphasic and in a concentration-dependent manner. Low concentrations (10 mM) of Mg2+ increased cell viability, cell proliferation rate, cell adhesion, cell spreading, cell migration rate, and actin expression. In contrast, higher concentrations (40–60 mM) of Mg2+ had deleterious effects on cells. Gene expression analysis revealed that Mg2+ altered the expressions of genes mostly related to cell adhesion, cell injury, angiogenesis, inflammation, coagulation, and cell growth. Finding from this study provides some valuable information on SMC responses toward magnesium ions at the cellular and molecular levels, and guidance for future controlled release of magnesium from the stent material. PMID:26402437

  20. Biphasic responses of human vascular smooth muscle cells to magnesium ion.

    PubMed

    Ma, Jun; Zhao, Nan; Zhu, Donghui

    2016-02-01

    Magnesium-based alloys are promising in biodegradable cardiovascular stent applications. The degradation products of magnesium stents may have significant impacts on the surrounding vascular cells. However, knowledge on the interactions between magnesium ion and vascular cells at the molecular and cellular levels is still largely missing. Vascular smooth muscle cell (SMC) plays an important role in the pathogenesis of restenosis and wound healing after stent implantation. This study evaluated the short-term effects of extracellular magnesium ion (Mg(2+)) on the cellular behaviors of SMCs. Cellular responses to Mg(2+) were biphasic and in a concentration-dependent manner. Low concentrations (10 mM) of Mg(2+) increased cell viability, cell proliferation rate, cell adhesion, cell spreading, cell migration rate, and actin expression. In contrast, higher concentrations (40-60 mM) of Mg(2+) had deleterious effects on cells. Gene expression analysis revealed that Mg(2+) altered the expressions of genes mostly related to cell adhesion, cell injury, angiogenesis, inflammation, coagulation, and cell growth. Finding from this study provides some valuable information on SMC responses toward magnesium ions at the cellular and molecular levels, and guidance for future controlled release of magnesium from the stent material.

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

    PubMed

    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.

  2. Hydrogen sulfide inhibits the calcification and osteoblastic differentiation of vascular smooth muscle cells

    PubMed Central

    Zavaczki, Erzsébet; Jeney, Viktória; Agarwal, Anupam; Zarjou, Abolfazl; Oros, Melinda; Katkó, Mónika; Varga, Zsuzsa; Balla, György; Balla, József

    2011-01-01

    Osteoblastic differentiation of vascular smooth muscle cells (VSMCs) is involved in the pathogenesis of vascular calcification. Hydrogen sulfide (H2S) is a gas endogenously produced by cystathionine γ-lyase in VSMC. Here we determined whether H2S plays a role in phosphate-induced osteoblastic transformation and mineralization of VSMC. Hydrogen sulfide was found to inhibit calcium deposition in the extracellular matrix and to suppress the induction of the genes involved in osteoblastic transformation of VSMC: alkaline phosphatase, osteocalcin, and Cbfa1. Moreover, phosphate uptake and phosphate-triggered upregulation of the sodium-dependent phosphate cotransporter (Pit-1) were also prevented by H2S. Reduction of endogenous production of H2S by inhibition of cystathionine γ-lyase activity resulted in increased osteoblastic transformation and mineralization. Low plasma levels of H2S, associated with decreased cystathionine γ-lyase enzyme activity, were found in patients with chronic kidney disease receiving hemodialysis. Thus, H2S is a potent inhibitor of phosphate-induced calcification and osteoblastic differentiation of VSMC. This mechanism might contribute to accelerated vascular calcification in chronic kidney disease. PMID:21716261

  3. The role of GRIP1 and ephrin B3 in blood pressure control and vascular smooth muscle cell contractility

    PubMed Central

    Wang, Yujia; Wu, Zenghui; Luo, Hongyu; Peng, Junzheng; Raelson, John; Ehret, Georg B.; Munroe, Patricia B.; Stoyanova, Ekatherina; Qin, Zhao; Cloutier, Guy; Bradley, W. Edward; Wu, Tao; Shen, Jian-Zhong; Hu, Shenjiang; Wu, Jiangping

    2016-01-01

    Several erythropoietin-producing hepatocellular receptor B family (EPHB) and their ligands, ephrinBs (EFNBs), are involved in blood pressure regulation in animal models. We selected 528 single nucleotide polymorphisms (SNPs) within the genes of EPHB6, EFNB2, EFNB3 and GRIP1 in the EPH/EFN signalling system to query the International Blood Pressure Consortium dataset. A SNP within the glutamate receptor interacting protein 1 (GRIP1) gene presented a p-value of 0.000389, approaching the critical p-value of 0.000302, for association with diastolic blood pressure of 60,396 individuals. According to echocardiography, we found that Efnb3 gene knockout mice showed enhanced constriction in the carotid arteries. In vitro studies revealed that in mouse vascular smooth muscle cells, siRNA knockdown of GRIP1, which is in the EFNB3 reverse signalling pathway, resulted in increased contractility of these cells. These data suggest that molecules in the EPHB/EFNB signalling pathways, specifically EFNB3 and GRIP1, are involved blood pressure regulation. PMID:27941904

  4. In vitro photodynamic therapy with chlorin e6 leads to apoptosis of human vascular smooth muscle cells.

    PubMed

    Wawrzyńska, Magdalena; Kałas, Wojciech; Biały, Dariusz; Zioło, Ewa; Arkowski, Jacek; Mazurek, Walentyna; Strzadała, Leon

    2010-02-01

    Percutaneous coronary intervention has become the most common and widely implemented method of heart revascularization. However, the development of restenosis remains the major limitation of this method. Photodynamic therapy (PDT) recently emerged as a new and promising method for the prevention of arterial restenosis. Here the efficacy of chlorin e6 in PDT was investigated in vitro using human vascular smooth muscle cells (TG/HA-VSMCs) as one of the cell types crucial in the development of restenosis. PDT-induced cell death was studied on many levels,including annexin V staining, measurement of the generation reactive oxygen species (ROS) and caspase-3 activity,and assessment of changes in mitochondrial membrane potential and fragmentation of DNA. Photosensitization of TG/HA-VSMCs with a 170 lM of chlorin e6 and subsequent illumination with the light of a 672-nm diode laser(2 J/cm2) resulted in the generation of ROS, a decrease in cell membrane polarization, caspase-3 activation, as well as DNA fragmentation. Interestingly, the latter two apoptotic events could not be observed in photosensitized and illuminated NIH3T3 fibroblasts, suggesting different outcomes of the model of PDT in various types of cells. The results obtained with human VSMCs show that chlorin e6 may be useful in the PDT of aerial restenosis, but its efficacy still needs to be established in an animal model.

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

    PubMed

    Hou, Jianghong; Xue, Xiaolin; Li, Junnong

    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.

  6. Ginsenoside Rb₁ inhibits the carotid neointimal hyperplasia induced by balloon injury in rats via suppressing the phenotype modulation of vascular smooth muscle cells.

    PubMed

    Zhang, Shu; Deng, Jiang; Gao, Yang; Yang, Dan-li; Gong, Qi-hai; Huang, Xie-nan

    2012-06-15

    This study aims to investigate the effects of ginsenoside Rb(1) on vascular intimal hyperplasia in rats and explore the mechanisms. The rat vascular neointimal hyperplasia model was made by rubbing the endothelia of carotid artery with a balloon and Rb(1) (10 and 30 mg/kg/day) was given the day after surgery for 14 consecutive days. The neointimal hyperplasia level and the degree of vascular smooth muscle cells (VSMCs) proliferation were evaluated by histopathology and by calculating the proliferating cell nuclear antigen (PCNA) positive expression percentage; protein expressions of PCNA, phosphorylation extracellular signal-regulated kinase 1/2 (pERK1/2), smooth muscle α-actin (SM α-actin), and the mRNA expressions of proto-oncogene c-myc, SM α-actin, SM-emb (embryonic smooth muscle myosin heavy chain) and p38 MAPK were detected by immunohistochemistry and Real Time RT-PCR, respectively. Compared with the endothelia rubbing model group, Rb(1) 10 and 30 mg/kg/day medication significantly ameliorated the neointimal hyperplasia (P<0.05), and decreased the positive expression percentage of PCNA(P<0.05). Rb(1) medication also significantly decreased the elevated protein expression of pERK1/2 and the mRNA expression of c-myc(P<0.05), and tended to reduce the expression of p38 MAPK mRNA. Endothelial rubbing increased the SM-emb mRNA expression, but decreased the expression of SM α-actin mRNA which was reversed by Rb(1) (P<0.05). The results indicate that Rb(1) inhibits the vascular neointimal hyperplasia induced by balloon-injury in rats via suppressing the VSMC proliferation, which may be involved in part the inhibition of pERK1/2 protein and related to its inhibition on VSMC phenotype modulation.

  7. Andrographolide, a Novel NF-κB Inhibitor, Inhibits Vascular Smooth Muscle Cell Proliferation and Cerebral Endothelial Cell Inflammation

    PubMed Central

    Chang, Chao-Chien; Duann, Yeh-Fang; Yen, Ting-Lin; Chen, Yu-Ying; Jayakumar, Thanasekaran; Ong, Eng-Thiam; Sheu, Joen-Rong

    2014-01-01

    Background Aberrant vascular smooth muscle cell (VSMC) proliferation and cerebral endothelial cell (CEC) dysfunction contribute significantly in the pathogenesis of cardiovascular diseases. Therefore, inhibition of these cellular events would be by candidate agents for treating these diseases. In the present study, the mechanism of anti-proliferative and anti-inflammatory effects of andrographolides, a novel nuclear factor-κB inhibitor, was investigated in VSMC and CEC cells. Methods VSMCs and CECs were isolated from rat artery and mouse brain, respectively, and cultured before experimentation. The effect of andro on platelet-derived growth factor-BB (PDGF-BB) induced VSMC cell proliferation was evaluated by cell number, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. The expression of extracellular signal regulated kinase 1/2 (ERK1/2), proliferating cell nuclear antigen (PCNA), and the effects on lipopolysaccharide (LPS)-induced inducible nitric oxide synthase (iNOS) and, cyclooxygenase-2 (COX2) were detected by Western blotting. Results Andro significantly inhibited PDGF-BB (10 ng/ml) induced cell proliferation in a concentration (20-100 μM) dependent manner, which may be due to reducing the expression of ERK1/2, and by inhibiting the expression of PCNA. Andro also remarkably diminished LPS-induced iNOS and COX2 expression. Conclusions The results of this study suggested that the effects of andro against VSMCs proliferation and CECs dysfunction may represent a promising approach for treatment of vascular diseases. PMID:27122804

  8. Phosphorylation of GATA-6 is required for vascular smooth muscle cell differentiation after mTORC1 inhibition

    PubMed Central

    Xie, Yi; Jin, Yu; Merenick, Bethany L.; Ding, Min; Fetalvero, Kristina M.; Wagner, Robert J.; Mai, Alice; Gleim, Scott; Tucker, David; Birnbaum, Morris J.; Ballif, Bryan A.; Luciano, Amelia K.; Sessa, William C.; Rzucidlo, Eva M.; Powell, Richard J.; Hou, Lin; Zhao, Hongyu; Hwa, John; Yu, Jun; Martin, Kathleen A.

    2015-01-01

    Vascular smooth muscle cells (VSMCs) undergo transcriptionally regulated reversible differentiation in growing and injured blood vessels. This de-differentiation also contributes to VSMC hyperplasia following vascular injury, including that caused by angioplasty and stenting. Stents provide mechanical support and can contain and release rapamycin, an inhibitor of the mammalian target of rapamycin complex 1 (mTORC1). Rapamycin suppresses VSMC hyperplasia and promotes VSMC differentiation. We report that rapamycin-induced differentiation of VSMCs required the transcription factor GATA-6. Inhibition of mTORC1 stabilized GATA-6 and promoted the nuclear accumulation of GATA-6, its binding to DNA, and its transactivation of promoters encoding contractile proteins and inhibitors of proliferation. These effects were mediated by phosphorylation of GATA-6 at Ser290, potentially by Akt2, a kinase that is activated in VSMCs when mTORC1 is inhibited. Rapamycin induced phosphorylation of GATA-6 in wild-type mice, but not in Akt2−/− mice. Intimal hyperplasia after arterial injury was greater in Akt2−/− mice than in wild-type mice, and the exacerbated response in Akt2−/− mice was rescued to a greater extent by local overexpression of the wild-type or phosphomimetic (S290D) mutant GATA-6 than by that of the phosphorylation-deficient (S290A) mutant. Our data indicated that GATA-6 and Akt2 are involved in the mTORC1-mediated regulation of VSMC proliferation and differentiation. Identifying the downstream transcriptional targets of mTORC1 may provide cell type-specific drug targets to combat cardiovascular diseases associated with excessive proliferation of VSMCs. PMID:25969542

  9. Myocardin is required for maintenance of vascular and visceral smooth muscle homeostasis during postnatal development.

    PubMed

    Huang, Jianhe; Wang, Tao; Wright, Alexander C; Yang, Jifu; Zhou, Su; Li, Li; Yang, Jisheng; Small, Aeron; Parmacek, Michael S

    2015-04-07

    Myocardin is a muscle-restricted transcriptional coactivator that activates a serum response factor (SRF)-dependent gene program required for cardiogenesis and embryonic survival. To identify myocardin-dependent functions in smooth muscle cells (SMCs) during postnatal development, mice harboring a SMC-restricted conditional, inducible Myocd null mutation were generated and characterized. Tamoxifen-treated SMMHC-Cre(ERT2)/Myocd(F/F) conditional mutant mice die within 6 mo of Myocd gene deletion, exhibiting profound derangements in the structure of great arteries as well as the gastrointestinal and genitourinary tracts. Conditional mutant mice develop arterial aneurysms, dissection, and rupture, recapitulating pathology observed in heritable forms of thoracic aortic aneurysm and dissection (TAAD). SMCs populating arteries of Myocd conditional mutant mice modulate their phenotype by down-regulation of SMC contractile genes and up-regulation of extracellular matrix proteins. Surprisingly, this is accompanied by SMC autonomous activation of endoplasmic reticulum (ER) stress and autophagy, which over time progress to programmed cell death. Consistent with these observations, Myocd conditional mutant mice develop remarkable dilation of the stomach, small intestine, bladder, and ureters attributable to the loss of visceral SMCs disrupting the muscularis mucosa. Taken together, these data demonstrate that during postnatal development, myocardin plays a unique, and important, role required for maintenance and homeostasis of the vasculature, gastrointestinal, and genitourinary tracts. The loss of myocardin in SMCs triggers ER stress and autophagy, which transitions to apoptosis, revealing evolutionary conservation of myocardin function in SMCs and cardiomyocytes.

  10. MicroRNA 181b promotes vascular smooth muscle cells proliferation through activation of PI3K and MAPK pathways.

    PubMed

    Li, Tie-Jun; Chen, Yan-Li; Gua, Chao-Jun; Xue, Sheng-Jiang; Ma, Shu-Mei; Li, Xiao-Dong

    2015-01-01

    Vascular smooth muscle cells (VSMCs) hyperplasia is a common feature of pathologic cardiovascular event such as restenosis and atherosclerosis. The role and mechanisms of microRNAs (miRs) in VSMCs proliferation are poorly understood. Here, we report that miR-181b promotes VSMCs proliferation and migration. In an animal model, miR-181b was significantly increased in the rat carotid artery after balloon catheter injury. Delivery of miR-181b inhibitor to injured artery exhibited a marked inhibition of neointimal hyperplasia. Transfection of miR-181b with "mimics" to A10 cells accelerated cell proliferation, which was accompanied by an increase of cell migration. The induction of A10 cells proliferation by miR-181b appeared to be involved in activation of S and G2/M checkpoint, concomitant with decreases in cell-cycle inhibitors p21 and p27, and increases in cell-cycle activators CDK4 and cyclinD1. In contract, miR-181b inhibition attenuated A10 cells proliferation, inhibited cell migration and arrested cell cycle transition. Moreover, forced miR-181b expression elevated the phosphorylation levels of Akt and Erk1/2, whereas inhibition of miR-181b produced the opposite effects. Additionally, inhibition of PI3K and MAPK signaling pathways with specific inhibitors, but not inhibition of JNK pathway, significantly abolished the effects of miR-181b in promoting cell proliferation. These findings demonstrate that miR-181b enhances the proliferation and migration of VSMCs through activation of PI3K and MAPK pathways.

  11. VEGF, VEGFR-1 and VEGFR-2 immunoreactivity in the porcine arteries of vascular subovarian plexus (VSP) during the estrous cycle.

    PubMed

    Postek, A; Andronowska, A; Doboszyńska, T; Niewegłowski, H; Jankowska, K

    2006-01-01

    Abstract: Vascular endothelial growth factor (VEGF) is an important angiogenic factor in the female reproductive tract. It binds to cell surface through ligand-stimulatable tyrosine kinase receptors, the most important being VEGFR-1 (flt-1) and VEGFR-2 (flk-1). The broad ligament of the uterus is a dynamic organ consisting of specialized complexes of blood vessels connected functionally to the uterus, oviduct and ovary. Endothelial cells form an inner coating of the vessel walls and thus they stay under the influence of various modulators circulating in blood including ovarian steriods involved in developmental changes in the female reproductive system. The aim of the present study was to immunolocalize VEGF and its two receptors: VEGFR-1 and VEGFR-2 in the broad ligament of the uterus in the area of vascular subovarian plexus during different phases of the estrous cycle in pig and to determine the correlation between immunoreactivity of the investigated factors and phases of the estrous cycle. The study was performed on cryostat sections of vascular subovarian plexus stained immunohistochemically by ABC method. Specific polyclonal antibodies: anti-VEGF, anti-VEGFR-1 and anti-VEGFR-2 were used. Data were subjected to one-way analysis of variance. Our study revealed the presence of VEGF and its receptors in endothelial and smooth muscle cells of VSP arteries. All agents displayed phase-related differences in immunoreactivity suggesting the modulatory effect of VEGF, VEGFR-1 and VEGFR-2 on the arteries of the VSP in the porcine broad ligament of the uterus.

  12. Platelet-localized FXI promotes a vascular coagulation-inflammatory circuit in arterial hypertension.

    PubMed

    Kossmann, Sabine; Lagrange, Jeremy; Jäckel, Sven; Jurk, Kerstin; Ehlken, Moritz; Schönfelder, Tanja; Weihert, Yvonne; Knorr, Maike; Brandt, Moritz; Xia, Ning; Li, Huige; Daiber, Andreas; Oelze, Matthias; Reinhardt, Christoph; Lackner, Karl; Gruber, Andras; Monia, Brett; Karbach, Susanne H; Walter, Ulrich; Ruggeri, Zaverio M; Renné, Thomas; Ruf, Wolfram; Münzel, Thomas; Wenzel, Philip

    2017-02-01

    Multicellular interactions of platelets, leukocytes, and the blood vessel wall support coagulation and precipitate arterial and venous thrombosis. High levels of angiotensin II cause arterial hypertension by a complex vascular inflammatory pathway that requires leukocyte recruitment and reactive oxygen species production and is followed by vascular dysfunction. We delineate a previously undescribed, proinflammatory coagulation-vascular circuit that is a major regulator of vascular tone, blood pressure, and endothelial function. In mice with angiotensin II-induced hypertension, tissue factor was up-regulated, as was thrombin-dependent endothelial cell vascular cellular adhesion molecule 1 expression and integrin αMβ2- and platelet-dependent leukocyte adhesion to arterial vessels. The resulting vascular inflammation and dysfunction was mediated by activation of thrombin-driven factor XI (FXI) feedback, independent of factor XII. The FXI receptor glycoprotein Ibα on platelets was required for this thrombin feedback activation in angiotensin II-infused mice. Inhibition of FXI synthesis with an antisense oligonucleotide was sufficient to prevent thrombin propagation on platelets, vascular leukocyte infiltration, angiotensin II-induced endothelial dysfunction, and arterial hypertension in mice and rats. Antisense oligonucleotide against FXI also reduced the increased blood pressure and attenuated vascular and kidney dysfunction in rats with established arterial hypertension. Further, platelet-localized thrombin generation was amplified in an FXI-dependent manner in patients with uncontrolled arterial hypertension, suggesting that platelet-localized thrombin generation may serve as an inflammatory marker of high blood pressure. Our results outline a coagulation-inflammation circuit that promotes vascular dysfunction, and highlight the possible utility of FXI-targeted anticoagulants in treating hypertension, beyond their application as antithrombotic agents in

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

    SciTech Connect

    Sung, Jin Young; Woo, Chang-Hoon; Kang, Young Jin; Lee, Kwang Youn; Choi, Hyoung Chul

    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.

  14. Free Fatty Acids Induce Autophagy and LOX-1 Upregulation in Cultured Aortic Vascular Smooth Muscle Cells.

    PubMed

    Cheng, Cheng-I; Lee, Yueh-Hong; Chen, Po-Han; Lin, Yu-Chun; Chou, Ming-Huei; Kao, Ying-Hsien

    2016-11-05

    Elevation of free fatty acids (FFAs) is known to affect microvascular function and contribute to obesity-associated insulin resistance, hypertension, and microangiopathy. Proliferative and synthetic vascular smooth muscle cells (VSMCs) increase intimal thickness and destabilize atheromatous plaques. This study aimed to investigate whether saturated palmitic acid (PA) and monounsaturated oleic acid (OA) modulate autophagy activity, cell proliferation, and vascular tissue remodeling in an aortic VSMC cell line. Exposure to PA and OA suppressed growth of VSMCs without apoptotic induction, but enhanced autophagy flux with elevation of Beclin-1, Atg5, and LC3I/II. Cotreatment with autophagy inhibitors potentiated the FFA-suppressed VSMC growth and showed differential actions of PA and OA in autophagy flux retardation. Both FFAs upregulated lectin-like oxidized low-density lipoprotein receptor 1 (LOX-1) but only OA increased LDL uptake by VSMCs. Mechanistically, FFAs induced hyperphosphorylation of Akt, ERK1/2, JNK1/2, and p38 MAPK. All pathways, except OA-activated PI3K/Akt cascade, were involved in the LOX-1 upregulation, whereas blockade of PI3K/Akt and MEK/ERK cascades ameliorated the FFA-induced growth suppression on VSMCs. Moreover, both FFAs exhibited tissue remodeling effect through increasing MMP-2 and MMP-9 expression and their gelatinolytic activities, whereas high-dose OA significantly suppressed collagen type I expression. Conversely, siRNA-mediated LOX-1 knockdown significantly attenuated the OA-induced tissue remodeling effects in VSMCs. In conclusion, OA and PA enhance autophagy flux, suppress aortic VSMC proliferation, and exhibit vascular remodeling effect, thereby leading to the loss of VSMCs and interstitial ECM in vascular walls and eventually the instability of atheromatous plaques. J. Cell. Biochem. 9999: 1-13, 2016. © 2016 Wiley Periodicals, Inc.

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

    SciTech Connect

    Ashino, T.; Varadarajan, S.; Urao, N.; Oshikawa, J.; Chen, G. -F.; Wang, H.; Huo, Y.; Finney, L.; Vogt, S.; McKinney, R. D.; Maryon, E. B.; Kaplan, J. H.; Ushio-Fukai, M.; Fukai, T.

    2010-09-09

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

  16. Clinical Outcomes of Cryopreserved Arterial Allograft Used as a Vascular Conduit for Hemodialysis

    PubMed Central

    Ha, Tae-Yong; Kim, Young Hoon; Chang, Jai Won; Park, Yangsoon; Han, Youngjin; Kwon, Hyunwook; Kwon, Tae-Won; Han, Duck Jong; Lee, Sung-Gyu

    2016-01-01

    This single center cohort study aimed to test the hypothesis that use of a cryopreserved arterial allograft could avoid the maturation or healing process of a new vascular access and to evaluate the patency of this technique compared with that of vascular access using a prosthetic graft. Between April 2012 and March 2013, 20 patients underwent an upper arm vascular access using a cryopreserved arterial allograft for failed or failing vascular accesses and 53 using a prosthetic graft were included in this study. The mean duration of catheter dependence, calculated as the time interval from upper arm access placement to removal of the tunneled central catheter after successful cannulation of the access, was significantly longer for accesses using a prosthetic graft than a cryopreserved arterial allograft (34.4 ± 11.39 days vs. 4.9 ± 8.5 days, P < 0.001). In the allograft group, use of vascular access started within 7 days in 16 patients (80%), as soon as from the day of surgery in 10 patients. Primary (unassisted; P = 0.314) and cumulative (assisted; P = 0.673) access survivals were similar in the two groups. There were no postoperative complications related to the use of a cryopreserved iliac arterial allograft except for one patient who experienced wound hematoma. In conclusion, upper arm vascular access using a cryopreserved arterial allograft may permit immediate hemodialysis without the maturation or healing process, resulting in access survival comparable to that of an access using a prosthetic graft. PMID:27478338

  17. Medical Textiles as Vascular Implants and Their Success to Mimic Natural Arteries.

    PubMed

    Singh, Charanpreet; Wong, Cynthia S; Wang, Xungai

    2015-06-30

    Vascular implants belong to a specialised class of medical textiles. The basic purpose of a vascular implant (graft and stent) is to act as an artificial conduit or substitute for a diseased artery. However, the long-term healing function depends on its ability to mimic the mechanical and biological behaviour of the artery. This requires a thorough understanding of the structure and function of an artery, which can then be translated into a synthetic structure based on the capabilities of the manufacturing method utilised. Common textile manufacturing techniques, such as weaving, knitting, braiding, and electrospinning, are frequently used to design vascular implants for research and commercial purposes for the past decades. However, the ability to match attributes of a vascular substitute to those of a native artery still remains a challenge. The synthetic implants have been found to cause disturbance in biological, biomechanical, and hemodynamic parameters at the implant site, which has been widely attributed to their structural design. In this work, we reviewed the design aspect of textile vascular implants and compared them to the structure of a natural artery as a basis for assessing the level of success as an implant. The outcome of this work is expected to encourage future design strategies for developing improved long lasting vascular implants.

  18. Medical Textiles as Vascular Implants and Their Success to Mimic Natural Arteries

    PubMed Central

    Singh, Charanpreet; Wong, Cynthia S.; Wang, Xungai

    2015-01-01

    Vascular implants belong to a specialised class of medical textiles. The basic purpose of a vascular implant (graft and stent) is to act as an artificial conduit or substitute for a diseased artery. However, the long-term healing function depends on its ability to mimic the mechanical and biological behaviour of the artery. This requires a thorough understanding of the structure and function of an artery, which can then be translated into a synthetic structure based on the capabilities of the manufacturing method utilised. Common textile manufacturing techniques, such as weaving, knitting, braiding, and electrospinning, are frequently used to design vascular implants for research and commercial purposes for the past decades. However, the ability to match attributes of a vascular substitute to those of a native artery still remains a challenge. The synthetic implants have been found to cause disturbance in biological, biomechanical, and hemodynamic parameters at the implant site, which has been widely attributed to their structural design. In this work, we reviewed the design aspect of textile vascular implants and compared them to the structure of a natural artery as a basis for assessing the level of success as an implant. The outcome of this work is expected to encourage future design strategies for developing improved long lasting vascular implants. PMID:26133386

  19. Eugenol dilates rat cerebral arteries by inhibiting smooth muscle cell voltage-dependent calcium channels.

    PubMed

    Peixoto-Neves, Dieniffer; Leal-Cardoso, Jose Henrique; Jaggar, Jonathan H

    2014-11-01

    Plants high in eugenol, a phenylpropanoid compound, are used as folk medicines to alleviate diseases including hypertension. Eugenol has been demonstrated to relax conduit and ear arteries and reduce systemic blood pressure, but mechanisms involved are unclear. Here, we studied eugenol regulation of resistance-size cerebral arteries that control regional brain blood pressure and flow and investigated mechanisms involved. We demonstrate that eugenol dilates arteries constricted by either pressure or membrane depolarization (60 mM K) in a concentration-dependent manner. Experiments performed using patch-clamp electrophysiology demonstrated that eugenol inhibited voltage-dependent calcium (Ca) currents, when using Ba as a charge carrier, in isolated cerebral artery smooth muscle cells. Eugenol inhibition of voltage-dependent Ca currents involved pore block, a hyperpolarizing shift (∼-10 mV) in voltage-dependent inactivation, an increase in the proportion of steady-state inactivating current, and acceleration of inactivation rate. In summary, our data indicate that eugenol dilates cerebral arteries by means of multimodal inhibition of voltage-dependent Ca channels.

  20. The effect of cadmium and other metals on vascular smooth muscle of the dogfish shark, Squalus acanthias.

    PubMed

    Evans, D H; Weingarten, K

    1990-04-30

    The effect of Cd2+ and related metals (Ni2+, Hg2+, Pb2+, Co2+, Sn2+ Cu2+ and Zn2+) on vascular tension was studied using isolated rings of endothelium-free, smooth muscle from the ventral aorta of the shark, Squalus acanthias. Both Cd2+ and Ni2+ produced significant vasoconstriction at concentrations at or above 10(-6) M (112 and 59 ppb, respectively); the other metals were either marginally constrictive (Hg2+ and Sn2+) or were without effect (Pb2+, Co2+, Cu2+, and Zn2+). We suggest that previously published vascular effects of Hg2+ and Pb2+ may have been secondary to responses of the vascular endothelium, and that the role of Ni2+ in hypertension should be investigated further. Our data indicate that the effects of metals on this vascular smooth muscle are specific and not generic. Moreover, this system could be utilized to investigate the mechanisms of metal-induced vasoconstriction.

  1. Peach (Prunus persica) extract inhibits angiotensin II-induced signal transduction in vascular smooth muscle cells.

    PubMed

    Kono, Ryohei; Okuno, Yoshiharu; Nakamura, Misa; Inada, Ken-ichi; Tokuda, Akihiko; Yamashita, Miki; Hidaka, Ryu; Utsunomiya, Hirotoshi

    2013-08-15

    Angiotensin II (Ang II) is a vasoactive hormone that has been implicated in cardiovascular diseases. Here, the effect of peach, Prunus persica L. Batsch, pulp extract on Ang II-induced intracellular Ca(2+) mobilization, reactive oxygen species (ROS) production and signal transduction events in cultured vascular smooth muscle cells (VSMCs) was investigated. Pretreatment of peach ethyl acetate extract inhibited Ang II-induced intracellular Ca(2+) elevation in VSMCs. Furthermore, Ang II-induced ROS generation, essential for signal transduction events, was diminished by the peach ethyl acetate extract. The peach ethyl acetate extract also attenuated the Ang II-induced phosphorylation of epidermal growth factor receptor and myosin phosphatase target subunit 1, both of which are associated with atherosclerosis and hypertension. These results suggest that peach ethyl acetate extract may have clinical potential for preventing cardiovascular diseases by interfering with Ang II-induced intracellular Ca(2+) elevation, the generation of ROS, and then blocking signal transduction events.

  2. [3H]ouabain binding to cultured rat vascular smooth muscle cells.

    PubMed

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

    1984-05-01

    The number of Na+ pump units (Bmax) and the equilibrium dissociation constant (Kd) for ouabain as well as parameters of K+ binding to the Na+ pump were examined in in vitro-grown vascular smooth muscle cells ( VSMC ) derived from Sprague-Dawley rats. The technique to measure these variables utilizes analyses of [3H]ouabain displacement from its VSMC receptors by nonlabeled ouabain and K+. The mean values for Bmax and Kd in the cultured VSMCs were 1.95 X 10(5) receptor sites per single VSMC and 2.68 X 10(-6) M, respectively. The equilibrium dissociation constant for K+ (Ki) was 0.92 mM. K+ binding to the cultured VSMCs demonstrated positive cooperativity with a Hill coefficient (n) of 1.78.

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

    SciTech Connect

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

    2007-10-05

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

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

    PubMed

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

    2000-08-11

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

  5. Injury of myocardial conduction tissue and coronary artery smooth muscle following brain death in the baboon.

    PubMed

    Novitzky, D; Rose, A G; Cooper, D K

    1988-05-01

    Experimental brain death was induced in 36 chacma baboons. In group A (n = 17), brain death was induced with no pharmacologic or surgical manipulation. Group B (n = 7) underwent bilateral vagotomy, unilateral left cardiac sympathectomy, or bilateral adrenalectomy before induction of brain death. Group C (n = 7) underwent total cardiac sympathectomy. Group D (n = 5) was pretreated with verapamil hydrochloride. Following induction of brain death, group A animals were maintained on a ventilator for a mean of 12 hr and 6 hr for the remaining groups. At the end of the experiment, the heart was excised, and tissue blocks were examined with light microscopy at (A) the atriaventricular node-bundle of His; (B) the major coronary arteries; and (C) myocardial tissue from the ventricular septum or left ventricular wall. In group A, 41% of the hearts showed histologic features of injury to the conduction tissue, 70% presented contraction band necrosis of the smooth muscle of the coronary arteries, and an incidence of 100% of the groups showed myocyte injury, more evident in the subendocardial area. In group B animals, conduction tissue injury was seen in 6 animals; the coronary arteries were not examined in this group; the incidence of myocyte injury was seen in 80% of the animals. Animals in groups C and D show no histopathologic injury in the conduction tissue (group A vs. C P less than 0.04), nor in the coronary arteries (group A vs. C P less than 0.002; group A vs. D P less than 0.01), preserving the myocytes (P less than 0.001). The catecholamine storm associated to acute increment of the endocranial pressure at the time of induction of brain death induces major histopathologic changes in the myocardium, as a result of endogenous catecholamines released inducing calcium overflow injury, affecting the conduction tissue, the smooth muscle of the coronary arteries, and the contractile myocardium. This can be prevented by calcium blockers or cardiac denervation.

  6. COX-2 is involved in vascular oxidative stress and endothelial dysfunction of renal interlobar arteries from obese Zucker rats.

    PubMed

    Muñoz, Mercedes; Sánchez, Ana; Pilar Martínez, María; Benedito, Sara; López-Oliva, Maria-Elvira; García-Sacristán, Albino; Hernández, Medardo; Prieto, Dolores

    2015-07-01

    Obesity is related to vascular dysfunction through inflammation and oxidative stress and it has been identified as a risk factor for chronic renal disease. In the present study, we assessed the specific relationships among reactive oxygen species (ROS), cyclooxygenase 2 (COX-2), and endothelial dysfunction in renal interlobar arteries from a genetic model of obesity/insulin resistance, the obese Zucker rats (OZR). Relaxations to acetylcholine (ACh) were significantly reduced in renal arteries from OZR compared to their counterpart, the lean Zucker rat (LZR), suggesting endothelial dysfunction. Blockade of COX with indomethacin and with the selective blocker of COX-2 restored the relaxations to ACh in obese rats. Selective blockade of the TXA2/PGH2 (TP) receptor enhanced ACh relaxations only in OZR, while inhibition of the prostacyclin (PGI2) receptor (IP) enhanced basal tone and inhibited ACh vasodilator responses only in LZR. Basal production of superoxide was increased in arteries of OZR and involved NADPH and xanthine oxidase activation and NOS uncoupling. Under conditions of NOS blockade, ACh induced vasoconstriction and increased ROS generation that were augmented in arteries from OZR and blunted by COX-2 inhibition and by the ROS scavenger tempol. Hydrogen peroxide (H2O2) evoked both endothelium- and vascular smooth muscle (VSM)-dependent contractions, as well as ROS generation that was reduced by COX-2 inhibition. In addition, COX-2 expression was enhanced in both VSM and endothelium of renal arteries from OZR. These results suggest that increased COX-2-dependent vasoconstriction contributes to renal endothelial dysfunction through enhanced (ROS) generation in obesity. COX-2 activity is in turn upregulated by ROS.

  7. NO Hyperpolarizes Pulmonary Artery Smooth Muscle Cells and Decreases the Intracellular Ca2+ Concentration by Activating Voltage-Gated K+ Channels

    NASA Astrophysics Data System (ADS)

    Yuan, Xiao-Jian; Tod, Mary L.; Rubin, Lewis J.; Blaustein, Mordecai P.

    1996-09-01

    NO causes pulmonary vasodilation in patients with pulmonary hypertension. In pulmonary arterial smooth muscle cells, the activity of voltage-gated K+ (KV) channels controls resting membrane potential. In turn, membrane potential is an important regulator of the intracellular free calcium concentration ([Ca2+]i) and pulmonary vascular tone. We used patch clamp methods to determine whether the NO-induced pulmonary vasodilation is mediated by activation of KV channels. Quantitative fluorescence microscopy was employed to test the effect of NO on the depolarization-induced rise in [Ca2+]i. Blockade of KV channels by 4-aminopyridine (5 mM) depolarized pulmonary artery myocytes to threshold for initiation of Ca2+ action potentials, and thereby increased [Ca2+]i. NO (≈ 3 μ M) and the NO-generating compound sodium nitroprusside (5-10 μ M) opened KV channels in rat pulmonary artery smooth muscle cells. The enhanced K+ currents then hyperpolarized the cells, and blocked Ca2+-dependent action potentials, thereby preventing the evoked increases in [Ca2+]i. Nitroprusside also increased the probability of KV channel opening in excised, outside-out membrane patches. This raises the possibility that NO may act either directly on the channel protein or on a closely associated molecule rather than via soluble guanylate cyclase. In isolated pulmonary arteries, 4-aminopyridine significantly inhibited NO-induced relaxation. We conclude that NO promotes the opening of KV channels in pulmonary arterial smooth muscle cells. The resulting membrane hyperpolarization, which lowers [Ca2+]i, is apparently one of the mechanisms by which NO induces pulmonary vasodilation.

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

    PubMed

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

    2013-01-01

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

  9. T3 inhibits the calcification of vascular smooth muscle cells and the potential mechanism

    PubMed Central

    Chang, Xiaodan; Zhang, Baohong; Lihua, Li; Feng, Zhichun

    2016-01-01

    Objective: This study aimed to investigate the potential molecular mechanism underlying the T3 induced vascular calcification and phenotype transformation of vascular smooth muscle cells (VSMCs). Methods: Rat thoracic aortic smooth muscle cells (A7r5) were cultured in vitro and randomly assigned into normal control group, calcification group, T3 group and inhibitor group. Results: When compared with normal control group, the osteocalcin content, ALP activity, Osterix and Runx2 mRNA expression and OPN protein expression increased significantly (P<0.01), and the protein expression of SMα and SM22α reduced dramatically in A7r5 cells of calcification group (P<0.01). After T3 treatment, the osteocalcin content and ALP activity reduced markedly, mRNA expression of Osterix and Runx2 and OPN protein expression reduced significantly. However, MMI (inhibitor of T3) was able to block the above effects of T3. When compared with calcification group, Osterix and Runx2 mRNA expression and OPN protein expression increased markedly (P<0.01). In addition, the protein expression of ERK1/2, p-ERK, Akt and p-Akt increased significantly in calcification group. In the presence of integrin αvβ3/ERK blocker (PD98059) and/or PI3K/Akt antagonist (LY294002), T3 was still able to inhibit the calcification, and this effect was similar to that after treatment with inhibitors alone. Moreover, LY294002 had a better inhibitory effect as compared to PD98059. Conclusion: T3 may act on PI3K/Akt signaling pathway to inhibit the phenotype transformation of VSMC, which then suppresses the calcium/phosphate induced calcification of rat VSMCs. Thus, T3 is an endogenous molecule that can protect the blood vessels against calcification. PMID:27904672

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

    SciTech Connect

    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

    2012-10-19

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

  11. Cell biology of Smad2/3 linker region phosphorylation in vascular smooth muscle.

    PubMed

    Rezaei, Hossein B; Kamato, Danielle; Ansari, Ghazaleh; Osman, Narin; Little, Peter J

    2012-08-01

    The transforming growth factor (TGF)-β superfamily of ligands regulates a diverse set of cellular functions. Transforming growth factor-β induces its biological effects through Type I and Type II transmembrane receptors that have serine/threonine kinase activities and weak tyrosine kinase activity. In vascular smooth muscle, TGF-β binds to the TGF-β Type II receptor (TβRII) at the cell surface, recruiting the Type I receptor (TβRI) to form a heterocomplex. Consequently, after phosphorylation and activation of TβRI, the transcription factors receptor activated (R-) Smad2 and Smad3 are recruited and activated through phosphorylation of C terminal residues. Overall, Smad2/3 and co-Smad4 have similar structures consisting of three regions an N-terminal MH1 domain, a C-terminal MH2 domain and a central linker region. Phosphorylation of the Smad linker region appears to have an important role in the regulation of Smad activity and function. The mitogen-activated protein kinase (MAPK) family, CDK2, CDK4 and calcium-calmodulin dependent kinase are the main kinases that phosphorylate sites in the linker region. The role of the linker region includes enabling the formation of Smad homo-oligomers and provision of phosphorylation sites for MAPK and other kinases. In some instances, linker region phosphorylation regulates the inhibition of the nuclear translocation of Smads. In the present review, we describe TGF-β signalling through Smad2/3 and the importance of the linker region in the regulation and expression of genes induced by TGF-β superfamily ligands in the context of vascular smooth muscle.

  12. Cyclooxygenase-2 in Endothelial and Vascular Smooth Muscle Cells Restrains Atherogenesis in Hyperlipidemic Mice

    PubMed Central

    Tang, Soon Yew; Monslow, James; Todd, Leslie; Lawson, John; Puré, Ellen; FitzGerald, Garret A.

    2014-01-01

    Background Placebo controlled trials of nonsteroidal antinflammatory drugs (NSAIDs) selective for inhibition of COX-2 reveal an emergent cardiovascular hazard in patients selected for low risk of heart disease. Postnatal global deletion of COX-2 accelerates atherogenesis in hyperlipidemic mice, a process delayed by selective enzyme deletion in macrophages. Methods and Results Here, selective depletion of COX-2 in vascular smooth muscle cells (VSMCs) and endothelial cells (ECs) depressed biosynthesis of prostaglandin (PG)I2 and PGE2, elevated blood pressure and accelerated atherogenesis in Ldlr knockout (KO) mice. Deletion of COX-2 in VSMCs and ECs coincided with an increase in COX-2 expression in lesional macrophages and increased biosynthesis of thromboxane. Increased accumulation of less organized intimal collagen, laminin, α-smooth muscle actin and matrix-rich fibrosis was also apparent in lesions of the mutants. Conclusions Although atherogenesis is accelerated in global COX-2 KOs, consistent with evidence of risk transformation during chronic NSAID administration, this masks the contrasting effects of enzyme depletion in macrophages versus VSMCs and ECs. Targeting delivery of COX-2 inhibitors to macrophages may conserve their efficacy while limiting cardiovascular risk. PMID:24519928

  13. Phytoncide, Nanochemicals from Chamaecyparis obtusa, Inhibits Proliferation and Migration of Vascular Smooth Muscle Cells.

    PubMed

    Lim, Leejin; Jang, Young-Su; Yun, Je-Jung; Song, Heesang

    2015-01-01

    Phytoncide, nanochemicals extracted from Chamaecyparis obtusa (C. obtusa), is reported to possess many pharmacological activities including immunological stimulating, anti-cancer, antioxidant, and antiinflammatory activities. However, the effect of phytoncide in vascuar diseases, especially on the behavior of vascular smooth muscle cells, has not yet been clearly elucidated. Therefore, in the present study, we investigated the effects of 15 kinds of phytoncide by various extraction conditions from C. obtusa on the proliferation and migration in rat aortic smooth muscle cells (RAoSMCs). First of all, we determined the concentration of each extracts not having cytotoxicity by MTT assay. We observed that the proliferation rate measured using BrdU assay was significantly reduced by supercritical fluid, steam distillation, Me-OH, and hexane extraction fraction in order with higher extent, respectively. Moreover, the treatment of above nanofractions inhibit the migration of RAoSMCs by 40%, 60%, and 30%, respectively, both in 2-D wound healing assay and 3-D boyden chamber assay. Immunoblot revealed that the phosphorylated levels of Akt and ERK were significantly reduced in nanofractions treated RAoSMCs. Taken together, these data suggest that phytoncide extracted from C. obtusa inhibits proliferation and migration in RAoSMCs via the modulation of phosphorylated levels of Akt and ERK. Therefore, phytoncide nanomolecules might be a potential therapeutic approach to prevent or treat atheroscrelosis and restenosis.

  14. Effects of dexamethasone on the synthesis and secretion of galaptin by vascular smooth muscle cells

    SciTech Connect

    Sanford, G.L.; Harris-Hooker, S.A.

    1986-05-01

    The effects of dexamethasone (Dex) on the synthesis and secretion of beta-galactoside specific lectin (galaptin) was examined in cultured primate aortic smooth muscle cells (SMC). SMC cells were treated with 0.15 ..mu..M Dex during their proliferative phase to confluency, and after reaching confluency. Both cultures were labeled with (/sup 3/H)-phenylalanine (phe) for 24 h following exposure to Dex. Incorporation of phe into galaptin increased twofold in the medium from Dex treated confluent cultures, when serum was present. No change was found in incorporation when serum was removed prior to Dex treatment. Phe incorporation into total protein was also increased twofold by Dex treatment of SMC in the presence of serum, but there was a 1.4-fold increase when serum was absent. Dex did not affect the incorporation of phe into either total protein or galaptin in the cell layer of confluent cultures in the presence of serum, but caused a twofold increase in its absence. There was no effect of Dex on the incorporation of phe into galaptin or total protein in either the medium or cell layer of cultures given Dex during their proliferative phase. Dex retarded the growth of SMC, and lowered the total protein content of the cell layer. The results show that vascular smooth muscle cells synthesize and secrete galaptin and that Dex acts directly on confluent SMC to increase galaptin synthesis and secretion. Serum seems to modulate the effect of Dex.

  15. Glucagon-like peptide-1 inhibits vascular smooth muscle cell dedifferentiation through mitochondrial dynamics regulation.

    PubMed

    Torres, Gloria; Morales, Pablo E; García-Miguel, Marina; Norambuena-Soto, Ignacio; Cartes-Saavedra, Benjamín; Vidal-Peña, Gonzalo; Moncada-Ruff, David; Sanhueza-Olivares, Fernanda; San Martín, Alejandra; Chiong, Mario

    2016-03-15

    Glucagon-like peptide-1 (GLP-1) is a neuroendocrine hormone produced by gastrointestinal tract in response to food ingestion. GLP-1 plays a very important role in the glucose homeostasis by stimulating glucose-dependent insulin secretion, inhibiting glucagon secretion, inhibiting gastric emptying, reducing appetite and food intake. Because of these actions, the GLP-1 peptide-mimetic exenatide is one of the most promising new medicines for the treatment of type 2 diabetes. In vivo treatments with GLP-1 or exenatide prevent neo-intima layer formation in response to endothelial damage and atherosclerotic lesion formation in aortic tissue. Whether GLP-1 modulates vascular smooth muscle cell (VSMC) migration and proliferation by controlling mitochondrial dynamics is unknown. In this report, we showed that GLP-1 increased mitochondrial fusion and activity in a PKA-dependent manner in the VSMC cell line A7r5. GLP-1 induced a Ser-637 phosphorylation in the mitochondrial fission protein Drp1, and decreased Drp1 mitochondrial localization. GLP-1 inhibited PDGF-BB-induced VSMC migration and proliferation, actions inhibited by overexpressing wild type Drp1 and mimicked by the Drp1 inhibitor Mdivi-1 and by overexpressing dominant negative Drp1. These results show that GLP-1 stimulates mitochondrial fusion, increases mitochondrial activity and decreases PDGF-BB-induced VSMC dedifferentiation by a PKA/Drp1 signaling pathway. Our data suggest that GLP-1 inhibits vascular remodeling through a mitochondrial dynamics-dependent mechanism.

  16. Peroxynitrite induces apoptosis in rat aortic smooth muscle cells: possible relation to vascular diseases.

    PubMed

    Li, Jianfeng; Li, Wenyan; Su, Jialin; Liu, Weimin; Altura, Bella T; Altura, Burton M

    2004-03-01

    An emerging body of evidence is accumulating to suggest that in vivo formation of free radicals in the vasculature, such as peroxynitrite (ONOO-), and programmed cell death (i.e., apoptosis) play important roles in vascular diseases such as atherosclerosis, hypertension, and restenosis. The present study was designed to determine whether primary rat aortic smooth muscle cells (SMCs) undergo apoptosis following treatment with ONOO-. Direct exposure of primary rat aortic SMCs to ONOO--induced apoptosis in a concentration-dependent manner, as confirmed by means of quantitative fluorescence staining and TUNEL assays. ONOO--induced apoptosis in rat aortic SMCs appears to involve activation of Ca2+-dependent endonucleases. Although the precise mechanisms by which peroxynitrite induces apoptosis in rat aortic SMCs need to be further investigated, the present, preliminary findings could be used to suggest that ONOO- formation in the vasculature may play roles in the processes of vascular diseases, such as atherosclerosis, hypertension, and restenosis, via adverse actions on blood vessels.

  17. Growth arrest of vascular smooth muscle cells in suspension culture using low-acyl gellan gum.

    PubMed

    Natori, Tomomi; Fujiyoshi, Masachika; Uchida, Masashi; Abe, Natsuki; Kanaki, Tatsuro; Fukumoto, Yasunori; Ishii, Itsuko

    2017-03-01

    The proliferation of vascular smooth muscle cells (SMCs) causes restenosis in biomaterial vascular grafts. The purposes of this study were to establish a suspension culture system for SMCs by using a novel substrate, low-acyl gellan gum (GG) and to maintain SMCs in a state of growth inhibition. When SMCs were cultured in suspension with GG, their proliferation was inhibited. Their viability was 70% at day 2, which was maintained at more than 50% until day 5. In contrast, the viability of cells cultured in suspension without GG was 5.6% at day 2. By cell cycle analysis, the ratio of SMCs in the S phase when cultured in suspension with GG was lower than when cultured on plastic plates. In SMCs cultured in suspension with GG, the ratio of phosphorylated retinoblastoma (Rb) protein to Rb protein was decreased and p27(Kip1) expression was unchanged in comparison with SMCs cultured on plastic plates. In addition, SMCs could be induced to proliferate again by changing the culture condition from suspension with GG to plastic plates. These results suggest that our established culturing method for SMCs is useful to maintain SMCs in a state of growth inhibition with high viability.

  18. Aldosterone-induced osteopontin gene transcription in vascular smooth muscle cells involves glucocorticoid response element.

    PubMed

    Kiyosue, Arihiro; Nagata, Daisuke; Myojo, Masahiro; Sato, Tomohiko; Takahashi, Masao; Satonaka, Hiroshi; Nagai, Ryozo; Hirata, Yasunobu

    2011-12-01

    Osteopontin (OPN) is known to be one of the cytokines that is involved in the vascular inflammation caused by aldosterone (Aldo). Previous reports have shown that Aldo increases OPN transcripts, and the mechanisms for this remain to be clarified. In this study, we investigated how Aldo increases OPN transcripts in the vascular smooth muscle cells of rats. Aldosterone increased OPN transcripts time-dependently as well as dose-dependently. This increase was diminished by eplerenone, a mineralocorticoid receptor (MR) antagonist. Luciferase promoter assays showed that the OPN promoter deleted to the -1599 site retained the same promoting ability as the full-length OPN promoter when stimulated by 10(-7) M Aldo, but the promoter deleted to the -1300 site lost the promoting ability. A glucocorticoid response element (GRE) is located in that deleted region. Luciferase assays of a mutated promoter without the GRE lost the luciferase upregulation, although mutated promoters with the deletion of other consensus sites maintained the promoter activity. The binding of the Aldo-MR complex to the GRE fragment was confirmed by an electrophoretic-mobility shift assay. This is the first report showing that Aldo regulates the transcriptional levels of OPN and inflammatory responses in the vasculature through a specific GRE site in the OPN promoter region.

  19. Expression of apolipoprotein E by cultured vascular smooth muscle cells is controlled by growth state

    PubMed Central

    1988-01-01

    Rat vascular smooth muscle cells (SMC) in culture synthesize and secrete a approximately 38,000-Mr protein doublet or triplet that, as previously described (Majack and Bornstein. 1984. J. Cell Biol. 99:1688- 1695), rapidly and reversibly accumulates in the SMC culture medium upon addition of heparin. In the present study, we show that this approximately 38,000-Mr heparin-regulated protein is electrophoretically and immunologically identical to apolipoprotein E (apo-E), a major plasma apolipoprotein involved in cholesterol transport. In addition, we show that expression of apo-E by cultured SMC varies according to growth state: while proliferating SMC produced little apo-E and expressed low levels of apo-E mRNA, quiescent SMC produced significantly more apo-E (relative to other proteins) and expressed markedly increased levels of apo-E mRNA. Northern analysis of RNA extracted from aortic tissue revealed that fully differentiated, quiescent SMC contain significant quantities of apo-E mRNA. These data establish aortic SMC as a vascular source for apo-E and suggest new functional roles for this apolipoprotein, possibly unrelated to traditional concepts of lipid metabolism. PMID:2458361

  20. Lanthanum chloride bidirectionally influences calcification in bovine vascular smooth muscle cells.

    PubMed

    Zhao, Wen-Hua; Gou, Bao-Di; Zhang, Tian-Lan; Wang, Kui

    2012-05-01

    Vascular calcification (VC) is frequent prevalence in patients with chronic kidney disease (CKD) and atherosclerosis. Lanthanum carbonate is used as an orally administered phosphate-binding agent to reduce the gastrointestinal absorption of phosphate and ameliorate VC in advanced CKD. In this study, we used bovine vascular smooth muscle cells as a model VC in vitro and studied the effects of lanthanum chloride on calcium deposition. Exposure of cells to LaCl(3) at the concentration of 0.1 µM suppressed the β-glycerophosphate-induced alkaline phosphatase activity and calcium deposition. Furthermore, LaCl(3) upregulated the β-glycerophosphate-suppressed expression of calcium-sensing receptor. In contrast to the inhibitory effect of LaCl(3) on calcium deposition, higher level lanthanum (50 µM) was found to promote immediately precipitation of calcium phosphate in cell culture medium. At this concentration, LaCl(3) was found to induce cell apoptosis which involves caspases-9 and -3. These data indicate that the promotory effect of LaCl(3) on calcium deposition is likely mediated by induction of apoptosis. Our in vitro findings do suggest that, in the context of raised lanthanum, greater attention should be paid to potential toxic effects associated to the use of lanthanide-based drugs.

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

    PubMed Central

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

    2014-01-01

    Phenotypic plasticity in vascular smooth muscle cells (VSMC) is necessary for vessel maintenance, repair and adaptation to vascular changes associated with aging. De-differentiated VSMC contribute to pathologies including atherosclerosis and intimal hyperplasia. As resveratrol has been reported to have cardio- protective effects, we investigated its role in VSMC phenotypic modulation. We demonstrated the novel finding that resveratrol promoted VSMC differentiation as measured by contractile 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. PMID:24416418

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

    SciTech Connect

    Patecki, Margret; Schaewen, Markus von; Tkachuk, Sergey; Jerke, Uwe; Dietz, Rainer; Dumler, Inna; Kusch, Angelika . E-mail: angelika.kusch@charite.de

    2007-08-03

    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.

  3. Bioabsorbable zinc ion induced biphasic cellular responses in vascular smooth muscle cells

    PubMed Central

    Ma, Jun; Zhao, Nan; Zhu, Donghui

    2016-01-01

    Bioabsorbable metal zinc (Zn) is a promising new generation of implantable scaffold for cardiovascular and orthopedic applications. In cardiovascular stent applications, zinc ion (Zn2+) will be gradually released into the surrounding vascular tissues from such Zn-containing scaffolds after implantation. However, the interactions between vascular cells and Zn2+ are still largely unknown. We explored the short-term effects of extracellular Zn2+ on human smooth muscle cells (SMCs) up to 24 h, and an interesting biphasic effect of Zn2+ was observed. Lower concentrations (<80 μM) of Zn2+ had no adverse effects on cell viability but promoted cell adhesion, cell spreading, cell proliferation, cell migration, and enhanced the expression of F-actin and vinculin. Cells treated with such lower concentrations of Zn2+ displayed an elongated shape compared to controls without any treatment. In contrast, cells treated with higher Zn2+ concentrations (80–120 μM) had opposite cellular responses and behaviors. Gene expression profiles revealed that the most affected functional genes were related to angiogenesis, inflammation, cell adhesion, vessel tone, and platelet aggregation. Results indicated that Zn has interesting concentration-dependent biphasic effects on SMCs with low concentrations being beneficial to cellular functions. PMID:27248371

  4. Rat vascular smooth muscle cells in culture contract upon Ca2+ repletion after depletion.

    PubMed Central

    Kobayashi, S.; Kanaide, H.; Hasegawa, M.; Yamamoto, H.; Nakamura, M.

    1985-01-01

    We investigated the effects of Ca2+-repletion following depletion on cultured vascular smooth muscle cells (SMCs) from the rat aorta. With Ca2+-repletion, the cells in primary cultures contracted, as indicated by a decrease in cell area. The process was slow (30 min to maximum effect) and reversible (relaxation completed by 120 min). Contraction during Ca2+-repletion was never observed in subcultured cells. The SMCs in primary culture after treatment maintained the ability to grow and to exclude dye, with a normal plating efficiency. There was no treatment-related additional leakage of intracellular enzymes, LDH and CPK, into the medium. Ca2+-repletion at first accelerated the 45Ca uptake by SMCs (1-5 min after repletion) and then increased Ca2+ efflux after about 10 min of Ca2+-repletion. We conclude that Ca2+-repletion after depletion induces a transient and reversible contraction of vascular SMCs in primary culture, without cell injury and in association with a transient increase in Ca2+ influx and then efflux. This phenomenon may relate to the decrease in perfusion flow in hearts and kidneys during Ca2+-repletion after depletion (Ca2+-paradox). Images Fig. 1 Fig. 3 PMID:4084451

  5. Magnesium prevents β-glycerophosphate-induced calcification in rat aortic vascular smooth muscle cells.

    PubMed

    Bai, Yaling; Zhang, Junxia; Xu, Jinsheng; Cui, Liwen; Zhang, Huiran; Zhang, Shenglei; Feng, Xunwei

    2015-07-01

    Vascular calcification (VC), in which high serum phosphate plays a critical role, is one major problem in patients with chronic kidney disease. Clinical studies report that magnesium has a protective effect on VC. However, the studies regarding the impact of high serum magnesium on VC at a cellular level are few and require further investigation. Therefore, the present study explored the effect of magnesium on calcification induced by β-glycerophosphate (BGP) in rat aortic vascular smooth muscle cells (RAVSMCs). In the present study, the addition of magnesium decreased calcium deposition, which was increased by BGP. Higher magnesium levels inhibited BGP-induced alkaline phosphatase (ALP) activity and decreased the expression of core-binding factor α-1 (Cbfα1). In conclusion, higher magnesium levels prevented BGP-induced calcification in RAVSMCs and inhibited the expression of Cbfα1 and ALP. Thus, magnesium is influencing the expression of Cbfα1 and ALP associated with VC and may have the potential to serve as a role for VC in clinical situations.

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

    PubMed

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

    2007-08-03

    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.

  7. Tropomyosin variants describe distinct functional subcellular domains in differentiated vascular smooth muscle cells.

    PubMed

    Gallant, Cynthia; Appel, Sarah; Graceffa, Philip; Leavis, Paul; Lin, Jim Jung-Ching; Gunning, Peter W; Schevzov, Galina; Chaponnier, Christine; DeGnore, Jon; Lehman, William; Morgan, Kathleen G

    2011-06-01

    Tropomyosin (Tm) is known to be an important gatekeeper of actin function. Tm isoforms are encoded by four genes, and each gene produces several variants by alternative splicing, which have been proposed to play roles in motility, proliferation, and apoptosis. Smooth muscle studies have focused on gizzard smooth muscle, where a heterodimer of Tm from the α-gene (Tmsm-α) and from the β-gene (Tmsm-β) is associated with contractile filaments. In this study we examined Tm in differentiated mammalian vascular smooth muscle (dVSM). Liquid chromatography-tandem mass spectrometry (LC MS/MS) analysis and Western blot screening with variant-specific antibodies revealed that at least five different Tm proteins are expressed in this tissue: Tm6 (Tmsm-α) and Tm2 from the α-gene, Tm1 (Tmsm-β) from the β-gene, Tm5NM1 from the γ-gene, and Tm4 from the δ-gene. Tm6 is by far most abundant in dVSM followed by Tm1, Tm2, Tm5NM1, and Tm4. Coimmunoprecipitation and coimmunofluorescence studies demonstrate that Tm1 and Tm6 coassociate with different actin isoforms and display different intracellular localizations. Using an antibody specific for cytoplasmic γ-actin, we report here the presence of a γ-actin cortical cytoskeleton in dVSM cells. Tm1 colocalizes with cortical cytoplasmic γ-actin and coprecipitates with γ-actin. Tm6, on the other hand, is located on contractile bundles. These data indicate that Tm1 and Tm6 do not form a classical heterodimer in dVSM but rather describe different functional cellular compartments.

  8. Up-regulation of the mammalian target of rapamycin complex 1 subunit Raptor by aldosterone induces abnormal pulmonary artery smooth muscle cell survival patterns to promote pulmonary arterial hypertension.

    PubMed

    Aghamohammadzadeh, Reza; Zhang, Ying-Yi; Stephens, Thomas E; Arons, Elena; Zaman, Paula; Polach, Kevin J; Matar, Majed; Yung, Lai-Ming; Yu, Paul B; Bowman, Frederick P; Opotowsky, Alexander R; Waxman, Aaron B; Loscalzo, Joseph; Leopold, Jane A; Maron, Bradley A

    2016-07-01

    Activation of the mammalian target of rapamycin complex 1 (mTORC1) subunit Raptor induces cell growth and is a downstream target of Akt. Elevated levels of aldosterone activate Akt, and, in pulmonary arterial hypertension (PAH), correlate with pulmonary arteriole thickening, which suggests that mTORC1 regulation by aldosterone may mediate adverse pulmonary vascular remodeling. We hypothesized that aldosterone-Raptor signaling induces abnormal pulmonary artery smooth muscle cell (PASMC) survival patterns to promote PAH. Remodeled pulmonary arterioles from SU-5416/hypoxia-PAH rats and monocrotaline-PAH rats with hyperaldosteronism expressed increased levels of the Raptor target, p70S6K, which provided a basis for investigating aldosterone-Raptor signaling in human PASMCs. Aldosterone (10(-9) to 10(-7) M) increased Akt/mTOR/Raptor to activate p70S6K and increase proliferation, viability, and apoptosis resistance in PASMCs. In PASMCs transfected with Raptor-small interfering RNA or treated with spironolactone/eplerenone, aldosterone or pulmonary arterial plasma from patients with PAH failed to increase p70S6K activation or to induce cell survival in vitro Optimal inhibition of pulmonary arteriole Raptor was achieved by treatment with Staramine-monomethoxy polyethylene glycol that was formulated with Raptor-small interfering RNA plus spironolactone in vivo, which decreased arteriole muscularization and pulmonary hypertension in 2 experimental animal models of PAH in vivo Up-regulation of mTORC1 by aldosterone is a critical pathobiologic mechanism that controls PASMC survival to promote hypertrophic vascular remodeling and PAH.-Aghamohammadzadeh, R., Zhang, Y.-Y., Stephens, T. E., Arons, E., Zaman, P., Polach, K. J., Matar, M., Yung, L.-M., Yu, P. B., Bowman, F. P., Opotowsky, A. R., Waxman, A. B., Loscalzo, J., Leopold, J. A., Maron, B. A. Up-regulation of the mammalian target of rapamycin complex 1 subunit Raptor by aldosterone induces abnormal pulmonary artery smooth

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

    PubMed

    Toniolo, Alice; Warden, Erica Alessia; Nassi, Alberto; Cignarella, Andrea; Bolego, Chiara

    2013-01-01

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

  10. The vascular smooth muscle cell: a therapeutic target in Type 2 diabetes?

    PubMed

    Porter, Karen E; Riches, Kirsten

    2013-08-01

    The rising epidemic of T2DM (Type 2 diabetes mellitus) worldwide is of significant concern. The inherently silent nature of the disease in its early stages precludes early detection; hence cardiovascular disease is often established by the time diabetes is diagnosed. This increased cardiovascular risk leads to significant morbidity and mortality in these individuals. Progressive development of complications as a result of previous exposure to metabolic disturbances appears to leave a long-lasting impression on cells of the vasculature that is not easily reversed and is termed 'metabolic memory'. SMCs (smooth muscle cells) of blood vessel walls, through their inherent ability to switch between a contractile quiescent phenotype and an active secretory state, maintain vascular homoeostasis in health and development. This plasticity also confers SMCs with the essential capacity to adapt and remodel in pathological states. Emerging clinical and experimental studies propose that SMCs in diabetes may be functionally impaired and thus contribute to the increased incidence of macrovascular complications. Although this idea has general support, the underlying molecular mechanisms are currently unknown and hence are the subject of intense research. The aim of the present review is to explore and evaluate the current literature relating to the problem of vascular disease in T2DM and to discuss the critical role of SMCs in vascular remodelling. Possibilities for therapeutic strategies specifically at the level of T2DM SMCs, including recent novel advances in the areas of microRNAs and epigenetics, will be evaluated. Since restoring glucose control in diabetic patients has limited effect in ameliorating their cardiovascular risk, discovering alternative strategies that restrict or reverse disease progression is vital. Current research in this area will be discussed.

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

    SciTech Connect

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

    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.

  12. Human Embryonic Stem Cell Derived Vascular Progenitor Cells Capable of Endothelial and Smooth Muscle Cell Function

    PubMed Central

    Hill, Katherine L; Obrtlikova, Petra; Alvarez, Diego F; King, Judy A; Keirstead, Susan A; Allred, Jeremy R; Kaufman, Dan S

    2010-01-01

    OBJECTIVE Previous studies have demonstrated development of endothelial cells (ECs) and smooth muscle cells (SMCs) as separate cell lineages derived from human embryonic stem cells (hESCs). We demonstrate CD34+ cells isolated from differentiated hESCs function as vascular progenitor cells capable of producing both ECs and SMCs. These studies better define the developmental origin and reveal the relationship between these two cell types, as well as provide a more complete biological characterization. MATERIALS AND METHODS hESCs are co-cultured on M2-10B4 stromal cells or Wnt1 expressing M2-10B4 for 13–15 days to generate a CD34+ cell population. These cells are isolated using a magnetic antibody separation kit and cultured on fibronectin coated dishes in EC medium. To induce SMC differentiation, culture medium is changed and a morphological and phenotypic change occurs within 24–48 hours. RESULTS CD34+ vascular progenitor cells give rise to ECs and SMCs. The two populations express respective cell specific transcripts and proteins, exhibit intracellular calcium in response to various agonists, and form robust tube-like structures when co-cultured in Matrigel. Human umbilical vein endothelial cells (HUVEC) cultured under SMC conditions do not exhibit a change in phenotype or genotype. Wnt1 overexpressing stromal cells produced an increased number of progenitor cells. CONCLUSIONS The ability to generate large numbers of ECs and SMCs from a single vascular progenitor cell population is promising for therapeutic use to treat a variety of diseased and ischemic conditions. The step-wise differentiation outlined here is an efficient, reproducible method with potential for large scale cultures suitable for clinical applications. PMID:20067819

  13. Simultaneous pressure and /sup 19/F NMR pH measurements of smooth muscle cells of intact hog carotid arteries at rest and during contractions with norepinephrine

    SciTech Connect

    Grieder, T.A.; Evans, C.A.; Greenberg, S.S.; Diecke, F.P.J.

    1988-01-01

    Using /sup 19/F NMR we have measured the intracellular pH of the vascular smooth muscle cells of hog carotid arteries at rest and during contractions induced with norepinephrine. Experiments were performed on single, intact arteries closed at both ends, superfused from the lumen and loaded with the /sup 19/F NMR pH indicator ..cap alpha..-difluoromethylalanine. At rest, luminal pressure was maintained at 100 +/-2 mm Hg and intracellular pH was 7.12 +/- 0.04. Contractions elicited with 10/sup -5/ M norepinephrine were associated with a pressure increase of 18 +/-6 mm Hg and a decrease in pH of 0.04 +/- 0.02 units.

  14. Macrophage secretory products selectively stimulate dermatan sulfate proteoglycan production in cultured arterial smooth muscle cells.

    PubMed Central

    Edwards, I. J.; Wagner, W. D.; Owens, R. T.

    1990-01-01

    Arterial dermatan sulfate proteoglycan has been shown to increase with atherosclerosis progression, but factors responsible for this increase are unknown. To test the hypothesis that smooth muscle cell proteoglycan synthesis may be modified by macrophage products, pigeon arterial smooth muscle cells were exposed to the media of either cholesteryl ester-loaded pigeon peritoneal macrophages or a macrophage cell line P388D1. Proteoglycans radiolabeled with [35S]sulfate and [3H]serine were isolated from culture media and smooth muscle cells and purified following precipitation with 1-hexadecylpyridinium chloride and chromatography. Increasing concentrations of macrophage-conditioned media were associated with a dose-response increase in [35S]sulfate incorporation into secreted proteoglycans, but there was no change in cell-associated proteoglycans. Incorporation of [3H]serine into total proteoglycan core proteins was not significantly different (5.2 X 10(5) dpm and 5.5 X 10(5) disintegrations per minute (dpm) in control and conditioned media-treated cultures, respectively), but selective effects were observed on individual proteoglycan types. Twofold increases in dermatan sulfate proteoglycan and limited degradation of chondroitin sulfate proteoglycan were apparent based on core proteins separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Immunoinhibition studies indicated that interleukin-1 was involved in the modulation of proteoglycan synthesis by macrophage-conditioned media. These data provide support for the role of macrophages in alteration of the matrix proteoglycans synthesized by smooth muscle cells and provide a mechanism to account for the reported increased dermatan sulfate/chondroitin sulfate ratios in the developing atherosclerotic lesion. Images Figure 6 PMID:2316626

  15. Macrophage secretory products selectively stimulate dermatan sulfate proteoglycan production in cultured arterial smooth muscle cells

    SciTech Connect

    Edwards, I.J.; Wagner, W.D.; Owens, R.T. )

    1990-03-01

    Arterial dermatan sulfate proteoglycan has been shown to increase with atherosclerosis progression, but factors responsible for this increase are unknown. To test the hypothesis that smooth muscle cell proteoglycan synthesis may be modified by macrophage products, pigeon arterial smooth muscle cells were exposed to the media of either cholesteryl ester-loaded pigeon peritoneal macrophages or a macrophage cell line P388D1. Proteoglycans radiolabeled with (35S)sulfate and (3H)serine were isolated from culture media and smooth muscle cells and purified following precipitation with 1-hexadecylpyridinium chloride and chromatography. Increasing concentrations of macrophage-conditioned media were associated with a dose-response increase in (35S)sulfate incorporation into secreted proteoglycans, but there was no change in cell-associated proteoglycans. Incorporation of (3H)serine into total proteoglycan core proteins was not significantly different (5.2 X 10(5) dpm and 5.5 X 10(5) disintegrations per minute (dpm) in control and conditioned media-treated cultures, respectively), but selective effects were observed on individual proteoglycan types. Twofold increases in dermatan sulfate proteoglycan and limited degradation of chondroitin sulfate proteoglycan were apparent based on core proteins separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Immunoinhibition studies indicated that interleukin-1 was involved in the modulation of proteoglycan synthesis by macrophage-conditioned media. These data provide support for the role of macrophages in alteration of the matrix proteoglycans synthesized by smooth muscle cells and provide a mechanism to account for the reported increased dermatan sulfate/chondroitin sulfate ratios in the developing atherosclerotic lesion.

  16. Impaired function of alpha-2 adrenoceptors in smooth muscle of mesenteric arteries from spontaneously hypertensive rats.

    PubMed

    Feres, T; Borges, A C; Silva, E G; Paiva, A C; Paiva, T B

    1998-11-01

    The alpha2-adrenoceptor function in mesenteric arteries of spontaneously hypertensive rats (SHR) was investigated by comparing membrane potential changes in response to adrenergic agonists in preparations from female SHR, Wistar-Kyoto (WKY) and normotensive Wistar rats (NWR). Resting membrane potential was found to be less negative in mesenteric arteries from SHR than in those from NWR and WKY. Apamin induced a decrease in the membrane potential of mesenteric artery rings without endothelium from NWR and WKY, but had no effects in those from SHR. Both UK 14,304 and adrenaline, in the presence of prazosin, induced a hyperpolarization that was significantly lower in de-endothelialized mesenteric rings from SHR than in those from NWR and WKY. In mesenteric rings with endothelium, however, similar hyperpolarization was observed in the three strains. In NWR mesenteric rings with endothelium the hyperpolarization induced by activation of alpha2-adrenoceptors was abolished by apamin, whereas in intact SHR mesenteric rings this hyperpolarization was slightly reduced by apamin and more efficiently reduced by Nomega-nitro-L-arginine. It is concluded that the activity of potassium channels coupled to alpha2-adrenoceptors is altered in the smooth muscle cells of SHR mesenteric arteries, contributing to their less negative membrane potential. On the other hand, the endothelial alpha2-receptors are functioning in mesenteric vessels from SHR and their stimulation induces a hyperpolarization mainly through the release of nitric oxide.

  17. Minimally Modified LDL Upregulates Endothelin Type A Receptors in Rat Coronary Arterial Smooth Muscle Cells

    PubMed Central

    Li, Jie; Cao, Lei; Xu, Cang-Bao; Wang, Jun-Jie

    2013-01-01

    Minimally modified low-density lipoprotein (mmLDL) is a risk factor for cardiovascular disease. The present study investigated the effects of mmLDL on the expression of endothelin type A (ETA) receptors in coronary arteries. Rat coronary arteries were organ-cultured for 24 h. The contractile responses were recorded using a myographic system. ETA receptor mRNA and protein expressions were determined using real-time PCR and western blotting, respectively. The results showed that organ-culturing in the presence of mmLDL enhanced the arterial contractility mediated by the ETA receptor in a concentration-dependent and time-dependent manner. Culturing with mmLDL (10 μg/mL) for 24 h shifted the concentration-contractile curves toward the left significantly with increased Emax of 228% ± 20% from control of 100% ± 10% and significantly increased ETA receptor mRNA and protein levels. Inhibition of the protein kinase C, extracellular signal-related kinases 1 and 2 (ERK1/2), or NF-κB activities significantly attenuated the effects of mmLDL. The c-Jun N-terminal kinase inhibitor or the p38 pathway inhibitor, however, had no such effects. The results indicate that mmLDL upregulates the ETA receptors in rat coronary arterial smooth muscle cells mainly via activating protein kinase C, ERK1/2, and the downstream transcriptional factor, NF-κB. PMID:23861561

  18. Periostin mediates cigarette smoke extract-induced proliferation and migration in pulmonary arterial smooth muscle cells.

    PubMed

    Wang, Xiao-Dong; Li, Fang; Ma, Dong-Bo; Deng, Xiang; Zhang, Hui; Gao, Jia; Hao, Li; Liu, Dan-Dan; Wang, Jing

    2016-10-01

    Cigarette smoking is an important risk factor for pulmonary arterial hypertension (PAH). Pulmonary arterial smooth muscle cells (PASMCs) play a critical role in the pathogenesis of PAH-associated arterial remodeling. This study was done to explore the expression and biological roles of periostin in PASMCs following exposure to cigarette smoke extract (CSE). PASMCs were exposed to different concentrations of CSE and tested for gene expression and reactive oxygen species (ROS) production. PASMCs were incubated with recombinant periostin protein or transfected with small interfering RNA targeting periostin before CSE exposure and then examined for cell proliferation and migration. Compared to control cells, exposure to CSE led to a significant upregulation of periostin. Pretreatment with 5mM N-acetyl-l-cysteine (an inhibitor of ROS formation) or 10μM U0126 (an inhibitor of ERK1/2) significantly prevented the induction of periostin in CSE-treated PASMCs. The addition of recombinant periostin protein significantly enhanced the proliferation and migration of PASMCs. In contrast, knockdown of endogenous periostin counteracted the proliferation and migration of PASMCs induced by CSE treatment. In conclusion, CSE induces the expression of periostin in PASMCs via promotion of ROS and activation of ERK1/2. Periostin mediates the effects of CSE on PASMC proliferation and migration. These findings warrant further exploration of the roles of periostin in cigarette smoking-associated pulmonary arterial remodeling.

  19. Adenosine Attenuates Human Coronary Artery Smooth Muscle Cell Proliferation by Inhibiting Multiple Signaling Pathways That Converge on Cyclin D.

    PubMed

    Dubey, Raghvendra K; Fingerle, Jürgen; Gillespie, Delbert G; Mi, Zaichuan; Rosselli, Marinella; Imthurn, Bruno; Jackson, Edwin K

    2015-12-01

    The goal of this study was to determine whether and how adenosine affects the proliferation of human coronary artery smooth muscle cells (HCASMCs). In HCASMCs, 2-chloroadenosine (stable adenosine analogue), but not N(6)-cyclopentyladenosine, CGS21680, or N(6)-(3-iodobenzyl)-adenosine-5'-N-methyluronamide, inhibited HCASMC proliferation (A2B receptor profile). 2-Chloroadenosine increased cAMP, reduced phosphorylation (activation) of ERK and Akt (protein kinases known to increase cyclin D expression and activity, respectively), and reduced levels of cyclin D1 (cyclin that promotes cell-cycle progression in G1). Moreover, 2-chloroadenosine inhibited expression of S-phase kinase-associated protein-2 (Skp2; promotes proteolysis of p27(Kip1)) and upregulated levels of p27(Kip1) (cell-cycle regulator that impairs cyclin D function). 2-Chloroadenosine also inhibited signaling downstream of cyclin D, including hyperphosphorylation of retinoblastoma protein and expression of cyclin A (S phase cyclin). Knockdown of A2B receptors prevented the effects of 2-chloroadenosine on ERK1/2, Akt, Skp2, p27(Kip1), cyclin D1, cyclin A, and proliferation. Likewise, inhibition of adenylyl cyclase and protein kinase A abrogated 2-chloroadenosine's inhibitory effects on Skp2 and stimulatory effects on p27(Kip1) and rescued HCASMCs from 2-chloroadenosine-mediated inhibition. Knockdown of p27(Kip1) also reversed the inhibitory effects of 2-chloroadenosine on HCASMC proliferation. In vivo, peri-arterial (rat carotid artery) 2-chloroadenosine (20 μmol/L for 7 days) downregulated vascular expression of Skp2, upregulated vascular expression of p27(Kip1), and reduced neointima hyperplasia by 71% (P<0.05; neointimal thickness: control, 37 424±18 371 pixels; treated, 10 352±2824 pixels). In conclusion, the adenosine/A2B receptor/cAMP/protein kinase A axis inhibits HCASMC proliferation by blocking multiple signaling pathways (ERK1/2, Akt, and Skp2) that converge at cyclin D, a key G1 cyclin

  20. Leukotriene B4 signaling through NF-κB-dependent BLT1 receptors on vascular smooth muscle cells in atherosclerosis and intimal hyperplasia

    PubMed Central

    Bäck, Magnus; Bu, De-xiu; Bränström, Robert; Sheikine, Yuri; Yan, Zhong-Qun; Hansson, Göran K.

    2005-01-01

    Leukotriene B4 (LTB4), a potent leukocyte chemoattractant derived from the 5-lipoxygenase metabolism of arachidonic acid, exerts its action by means of specific cell surface receptors, denoted BLT1 and BLT2. In this study, BLT1 receptor proteins were detected in human carotid artery atherosclerotic plaques, colocalizing with markers for macrophages, endothelial cells, and vascular smooth muscle cells (SMC). Challenge of human coronary artery SMC with either LTB4 or U75302, a partial agonist that is selective for the BLT1 receptor, induced an ≈4-fold increase of whole-cell currents by using the patch-clamp technique, indicating that these cells express functional BLT1 receptors. LTB4 induced migration and proliferation of SMC in vitro, and treatment with the BLT receptor antagonist BIIL 284 (10 mg/kg, once daily) for 14 days after carotid artery balloon injury in vivo inhibited intimal hyperplasia in rats. In the latter model, SMC derived from the intima exhibited increased levels of BLT1 receptor mRNA compared with medial SMC. BLT receptor up-regulation in the intima in vivo, as well as that induced by IL-1β in vitro, were prevented by transfection with a dominant-negative form of Iκ kinase β carried by adenovirus, indicating that BLT1 receptor expression depends on NF-κΒ. These results show that LTB4 activates functional BLT1 receptors on vascular SMC, inducing chemotaxis and proliferation, and that BLT1 receptors were up-regulated through an Iκ kinase β/NF-κB-dependent pathway. Inhibition of LTB4/BLT1 signaling during the response to vascular injury reduced intimal hyperplasia, suggesting this pathway as a possible target for therapy. PMID:16293697

  1. [Progress of study on inhibitory effects of traditional Chinese herbs on growth factor induced proliferation of vascular smooth muscle cells].

    PubMed

    Yang, Guang; Zhang, Min-zhou; Jiang, Wei

    2005-10-01

    This paper sums up some studies in the last decade regarding the inhibitory effects of traditional Chinese herbs on growth factor induced proliferation of vascular smooth muscle cell (VSMC) via directly measuring the mRNA expression of its growth factors and the related receptors by electron microscope, immunohistochemistry, blot and hybridization in situ.

  2. UAP56 is a novel interacting partner of Bcr in regulating vascular smooth muscle cell DNA synthesis

    SciTech Connect

    Sahni, Abha; Wang, Nadan; Alexis, Jeffrey D.

    2012-04-13

    Highlights: Black-Right-Pointing-Pointer UAP56 is an important regulator of DNA synthesis in vascular smooth muscle cells. Black-Right-Pointing-Pointer UAP56 binds to Bcr. Black-Right-Pointing-Pointer Interaction between Bcr and UAP56 is critical for Bcr induced DNA synthesis. -- Abstract: Bcr is a serine/threonine kinase that is a critical regulator of vascular smooth muscle cell inflammation and proliferation. We have previously demonstrated that Bcr acts in part via phosphorylation and inhibition of PPAR{gamma}. We have identified the RNA helicase UAP56 as another substrate of Bcr. In this report we demonstrate that knockdown of UAP56 blocks Bcr induced DNA synthesis in vascular smooth muscle cells (VSMC). We also found that over expression of Bcr increased the expression of cyclin E and decreased the expression of p27. Knockdown of UAP56 reversed the effect of Bcr on cyclin E and p27 expression. Furthermore, we found that Bcr binds to UAP56 and demonstrate that binding of UAP56 to Bcr is critical for Bcr induced DNA synthesis in VSMC. Our data identify UAP56 as an important binding partner of Bcr and a novel target for inhibiting vascular smooth muscle cell proliferation.

  3. Palmitic acid induces osteoblastic differentiation in vascular smooth muscle cells through ACSL3 and NF-κB, novel targets of eicosapentaenoic acid.

    PubMed

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

    2013-01-01

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

  4. Function and expression of ryanodine receptors and inositol 1,4,5-trisphosphate receptors in smooth muscle cells of murine feed arteries and arterioles

    PubMed Central

    Westcott, Erika B; Goodwin, Erica L; Segal, Steven S; Jackson, William F

    2012-01-01

    We tested the hypothesis that vasomotor control is differentially regulated between feed arteries and downstream arterioles from the cremaster muscle of C57BL/6 mice. In isolated pressurized arteries, confocal Ca2+ imaging of smooth muscle cells (SMCs) revealed Ca2+ sparks and Ca2+ waves. Ryanodine receptor (RyR) antagonists (ryanodine and tetracaine) inhibited both sparks and waves but increased global Ca2+ and myogenic tone. In arterioles, SMCs exhibited only Ca2+ waves that were insensitive to ryanodine or tetracaine. Pharmacological interventions indicated that RyRs are functionally coupled to large-conductance, Ca2+-activated K+ channels (BKCa) in SMCs of arteries, whereas BKCa appear functionally coupled to voltage-gated Ca2+ channels in SMCs of arterioles. Inositol 1,4,5-trisphosphate receptor (IP3R) antagonists (xestospongin D or 2-aminoethoxydiphenyl borate) or a phospholipase C inhibitor (U73122) attenuated Ca2+ waves, global Ca2+ and myogenic tone in arteries and arterioles but had no effect on arterial sparks. Real-time PCR of isolated SMCs revealed RyR2 as the most abundant isoform transcript; arteries expressed twice the RyR2 but only 65% the RyR3 of arterioles and neither vessel expressed RyR1. Immunofluorescent localisation of RyR protein indicated bright, clustered staining of arterial SMCs in contrast to diffuse staining in arteriolar SMCs. Expression of IP3R transcripts and protein immunofluorescence were similar in SMCs of both vessels with IP3R1>>IP3R2>IP3R3. Despite similar expression of IP3Rs and dependence of Ca2+ waves on IP3Rs, these data illustrate pronounced regional heterogeneity in function and expression of RyRs between SMCs of the same vascular resistance network. We conclude that vasomotor control is differentially regulated in feed arteries vs. downstream arterioles. PMID:22331418

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

    PubMed Central

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

    2016-01-01

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

  6. Kv7.5 Potassium Channel Subunits Are the Primary Targets for PKA-Dependent Enhancement of Vascular Smooth Muscle Kv7 Currents

    PubMed Central

    Mani, Bharath K.; Robakowski, Christina; Brueggemann, Lyubov I.; Cribbs, Leanne L.; Tripathi, Abhishek; Majetschak, Matthias

    2016-01-01

    Kv7 (KCNQ) channels, formed as homo- or heterotetramers of Kv7.4 and Kv7.5 α-subunits, are important regulators of vascular smooth muscle cell (VSMC) membrane voltage. Recent studies demonstrate that direct pharmacological modulation of VSMC Kv7 channel activity can influence blood vessel contractility and diameter. However, the physiologic regulation of Kv7 channel activity is still poorly understood. Here, we study the effect of cAMP/protein kinase A (PKA) activation on whole cell K+ currents through endogenous Kv7.5 channels in A7r5 rat aortic smooth muscle cells or through Kv7.4/Kv7.5 heteromeric channels natively expressed in rat mesenteric artery smooth muscle cells. The contributions of specific α-subunits are further dissected using exogenously expressed human Kv7.4 and Kv7.5 homo- or heterotetrameric channels in A7r5 cells. Stimulation of Gαs-coupled β-adrenergic receptors with isoproterenol induced PKA-dependent activation of endogenous Kv7.5 currents in A7r5 cells. The receptor-mediated enhancement of Kv7.5 currents was mimicked by pharmacological agents that increase [cAMP] (forskolin, rolipram, 3-isobutyl-1-methylxanthine, and papaverine) or mimic cAMP (8-bromo-cAMP); the 2- to 4-fold PKA-dependent enhancement of currents was also observed with exogenously expressed Kv7.5 channels. In contrast, exogenously-expressed heterotetrameric Kv7.4/7.5 channels in A7r5 cells or native mesenteric artery smooth muscle Kv7.4/7.5 channels were only modestly enhanced, and homo-tetrameric Kv7.4 channels were insensitive to this regulatory pathway. Correspondingly, proximity ligation assays indicated that isoproterenol induced PKA-dependent phosphorylation of exogenously expressed Kv7.5 channel subunits, but not of Kv7.4 subunits. These results suggest that signal transduction-mediated responsiveness of vascular smooth muscle Kv7 channel subunits to cAMP/PKA activation follows the order of Kv7.5 >> Kv7.4/Kv7.5 > Kv7.4. PMID:26700561

  7. /sup 22/Na+ and /sup 86/Rb+ transport in vascular smooth muscle of SHR, Wistar Kyoto, and Wistar rats

    SciTech Connect

    Kuriyama, S.; Denny, T.N.; Aviv, A.

    1988-06-01

    To gain further insight into differences in cellular Na+ and K+ regulation between the spontaneously hypertensive rat (SHR), Wistar Kyoto (WKY), and American Wistar (W) rats, 22Na+ and 86Rb+ washouts were performed under steady-state conditions in cultured vascular smooth muscle cells from the three rat strains. SHR vascular smooth muscle cells showed significantly higher bumetanide sensitive 86Rb+ washout rate constant (x 10(-4)/min; mean +/- SEM) than WKY cells (-38.6 +/- 2.84 and -23.8 +/- 3.58, respectively; p less than 0.005). SHR vascular smooth muscle cells also exhibited significantly higher values than WKY cells in the total 22Na+ washout rate constant (x 10(-2)/min) (-61.0 +/- 1.57 vs. -53.8 +/- 1.24; p less than 0.005). The amiloride sensitive component of the 22Na+ washout rate constant accounted for these differences (-18.6 +/- 1.04 for SHR and -12.1 +/- 2.00 for WKY; p less than 0.05). There were no apparent differences in cellular Na+ concentrations between WKY and SHR cells. In general, the 86Rb+ and 22Na+ washout parameters of W rat cells were quite similar to those of cells from SHR. We conclude that the bumetanide-sensitive 86Rb+ washout (the Na+ K+-cotransport), the overall, and the amiloride-sensitive 22Na+ washout (the latter primarily represents the Na+/H+ antiport) are higher in SHR than WKY rat vascular smooth muscle cells. These findings indicate innate differences in cellular Na+ and K+ transport in vascular smooth muscle cells of the SHR and WKY rat. The mechanisms responsible for these differences are yet to be determined.

  8. Activation of AMPK inhibits PDGF-induced pulmonary arterial smooth muscle cells proliferation and its potential mechanisms.

    PubMed

    Song, Yang; Wu, Yuanyuan; Su, Xiaofan; Zhu, Yanting; Liu, Lu; Pan, Yilin; Zhu, Bo; Yang, Lan; Gao, Li; Li, Manxiang

    2016-05-01

    The aims of the present study were to examine signaling mechanisms for PDGF-induced pulmonary arterial smooth muscle cells (PASMC) proliferation and to determine the effect of AMPK activation on PDGF-induced PASMC proliferation and its underlying mechanisms. PDGF activated PI3K/Akt/mTOR signaling pathway, and this in turn up-regulated Skp2 and consequently reduced p27 leading to PASMC proliferation. Prior incubation of PASMC with metformin induced a dramatic AMPK activation and significantly blocked PDGF-induced cell proliferation. PASMC lacking AMPKα2 were resistant to the inhibitory effect of metformin on PDGF-induced cell proliferation. Metformin did not affect Akt activation but blocked mTOR phosphorylation in response to PDGF; these were accompanied by the reversion of Skp2 up-regulation and p27 reduction. Our study suggests that the activation of AMPK negatively regulates mTOR activity to suppress PASMC proliferation and therefore has a potential value in the prevention and treatment of pulmonary hypertension by negatively modulating pulmonary vascular remodeling.

  9. Thrombin immobilized to extracellular matrix is a potent mitogen for vascular smooth muscle cells: nonenzymatic mode of action.

    PubMed Central

    Bar-Shavit, R; Benezra, M; Eldor, A; Hy-Am, E; Fenton, J W; Wilner, G D; Vlodavsky, I

    1990-01-01

    Esterolytically inactive diisopropyl fluorophosphate-conjugated thrombin (DIP-alpha-thrombin) stimulated 3H-thymidine incorporation and proliferation of growth-arrested vascular smooth muscle cells (SMCs), similar to native alpha-thrombin. Half-maximal mitogenic response of SMCs was obtained at 1 nM thrombin and was specifically blocked by the leech-derived, high-affinity thrombin inhibitor, hirudin. Native thrombin and a variety of thrombin species that were chemically modified to alter thrombin procoagulant or esterolytic functions were found to induce 3H-thymidine incorporation to a similar extent. Exposure of SMCs to DIP-alpha-thrombin caused a rapid and transient expression of the c-fos protooncogene, determined by Northern blot analysis. These results indicate that thrombin is a potent mitogen for SMCs through a distinct non-enzymatic domain. Binding of 125I-alpha-thrombin to SMC cultures revealed an apparent dissociation constant of 6 nM and an estimated 5.4 x 10(5) binding sites per cell. This binding was inhibited to the same extent by native thrombin and by its nonenzymatic form, DIP-alpha-thrombin. Moreover, the chemotactic fragment of thrombin (CB67-129), which failed to elicit a mitogenic response, competed for 125I-alpha-thrombin binding to SMCs. Cross-linking analysis of 125I-alpha-thrombin to SMCs revealed a specific cell-surface binding site 55 kDa in size. Finally, thrombin immobilized to a naturally produced extracellular matrix retained potent mitogenic activity toward SMCs. These observations lend support to the possibility that in vivo, subendothelial basement membranes sequester thrombin (as well as other bioactive molecules), which may stimulate localized and persistent growth of arterial SMCs. Thrombin may thus be involved directly in progression of atherosclerotic plaque formation. Images PMID:1963793

  10. MicroRNA-15b/16 attenuates vascular neointima formation by promoting the contractile phenotype of vascular smooth muscle through targeting YAP

    PubMed Central

    Xu, Fei; Ahmed, Abu Shufian Ishtiaq; Kang, Xiuhua; Hu, Guoqing; Liu, Fang; Zhang, Wei; Zhou, Jiliang

    2015-01-01

    Objective To investigate the functional role of the miR-15b/16 in vascular smooth muscle phenotypic modulation. Approach and Results We found that miR-15b/16 is the one of most abundant microRNAs expressed in contractile vascular smooth muscle cells (VSMCs). However, when contractile VSMCs convert to a synthetic phenotype miR-15b/16 expression is significantly reduced. Knocking-down endogenous miR-15b/16 in VSMCs attenuates smooth muscle-specific gene expression but promotes VSMC proliferation and migration. Conversely, over-expression of miR-15b/16 promotes smooth muscle contractile gene expression while attenuating VSMC migration and proliferation. Consistent with this, over-expression of miR-15b/16 in a rat carotid balloon injury model markedly attenuates injury-induced smooth muscle de-differentiation and neointima formation. Mechanistically, we identified the potent oncoprotein yes-associated protein (YAP) as a downstream target of miR-15b/16 in VSMCs. Reporter assays validated that miR-15b/16 targets YAP’s 3′-untranslated region. Moreover, overexpression of miR-15b/16 significantly represses YAP expression, whereas conversely, depletion of endogenous miR-15b/16 results in up-regulation of YAP expression. Conclusions These results indicate that miR-15b/16 plays a critical role in smooth muscle phenotypic modulation at least partly through targeting YAP. Restoring expression of miR-15b/16 would be a potential therapeutic approach for treatment of proliferative vascular diseases. PMID:26293467

  11. Arterial vascularization of the pineal gland in the fetus of Zavot-bred cattle.

    PubMed

    Aslan, K; Ozcan, S; Aksoy, G; Kurtul, I; Dursun, N

    2003-04-01

    This study aimed at revealing arterial vascularization of the pineal gland of the Zavot-bred foetus. Twenty foetuses, regardless of their sex, at the age of 2-7 months were used. Coloured-latex was injected by way of both the right and left common carotid arteries. Then, dissection was performed and vessels nourishing the pineal gland were documented. The pineal gland is vascularized by a number of 2-5 central rami. A small vessel arising from each of the central rami in two foetuses (10%) was shown anastomosing with a branch of the cranial cerebral artery, which advances in cranio-caudal direction in the callosal groove. Hence, anastomoses were observed between several sub-branches of each caudal cerebral and cranial cerebellar arteries.

  12. Rab5a-mediated autophagy regulates the phenotype and behavior of vascular smooth muscle cells

    PubMed Central

    Tan, Jin-Yun; Jia, Luo-Qi; Shi, Wei-Hao; He, Qing; Zhu, Lei; Yu, Bo

    2016-01-01

    Rab5a, a key member of the Rab family of GTPases, was determined to be a regulator of vascular smooth muscle cell (VSMC) proliferation and migration. However, the exact regulatory mechanism remains unclear. As Rab5a has been shown to be associated with autophagy, which is essential for the conversion of VSMCs from a contractile to a synthetic phenotype in order to prevent cell death due to oxidative stress. The present study hypothesized that autophagy may be responsible for the proliferation and migration of VSMCs via the Rab5a protein. The aim of the present study was to evaluate the effect of Rab5a on autophagy in VSMCs. The human aorta vascular smooth muscle cell line, T/G HA-VSMCs, was treated with small interfering (si)RNA against Rab5a and/or platelet-derived growth factor (PDGF). Following treatment, the phenotype transition of the VSMCs was evaluated by detecting the mRNA and protien expression levels of VSMC molecular markers using reverse transcription-quantitative polymerase chain reaction and western blotting, respectively. In addition, autophagy in VSMCs was evaluated by western blotting for autophagy-associated proteins, flow cytometry of acidic vesicular organelles, punctate fluorescence of microtubule associated protein light chain 3 and transmission electron microscopy of typical scattered double-membrane vacuolar structures. Additionally, the proliferation, migration, cell cycle and apoptotic response of VSMCs were detected by sulforhodamine B assay, transwell assay and flow cytometry, respectively. The results revealed that transfection with siRNA against Rab5a led to a significant decrease in Rab5a protein expression, while the reduced expression trend of Rab5a was rescued by intervention with PDGF. Furthermore, cells transfected with siRNA against Rab5a inhibited the autophagy of VSMCs. Downregulated Rab5a inhibited the phenotype transition of VSMCs. Additionally, downregulated Rab5a led to slowed cell growth, decreased numbers of migrated

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

    SciTech Connect

    Salabei, Joshua K.; Balakumaran, Arun; Frey, Justin C.; Boor, Paul J.; Treinen-Moslen, Mary; Conklin, Daniel J.

    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

  14. PDGF induces SphK1 expression via Egr-1 to promote pulmonary artery smooth muscle cell proliferation.

    PubMed

    Sysol, Justin R; Natarajan, Viswanathan; Machado, Roberto F

    2016-06-01

    Pulmonary arterial hypertension (PAH) is a progressive, life-threatening disease for which there is currently no curative treatment available. Pathologic changes in this disease involve remodeling of the pulmonary vasculature, including marked proliferation of pulmonary artery smooth muscle cells (PASMCs). Recently, the bioactive lipid sphingosine-1-phosphate (S1P) and its activating kinase, sphingosine kinase 1 (SphK1), have been shown to be upregulated in PAH and promote PASMC proliferation. The mechanisms regulating the transcriptional upregulation of SphK1 in PASMCs are unknown. In this study, we investigated the role of platelet-derived growth factor (PDGF), a PAH-relevant stimuli associated with enhanced PASMC proliferation, on SphK1 expression regulation. In human PASMCs (hPASMCs), PDGF significantly increased SphK1 mRNA and protein expression and induced cell proliferation. Selective inhibition of SphK1 attenuated PDGF-induced hPASMC proliferation. In silico promoter analysis for SphK1 identified several binding sites for early growth response protein 1 (Egr-1), a PDGF-associated transcription factor. Luciferase assays demonstrated that PDGF activates the SphK1 promoter in hPASMCs, and truncation of the 5'-promoter reduced PDGF-induced SphK1 expression. Stimulation of hPASMCs with PDGF induced Egr-1 protein expression, and direct binding of Egr-1 to the SphK1 promoter was confirmed by chromatin immunoprecipitation analysis. Inhibition of ERK signaling prevented induction of Egr-1 by PDGF. Silencing of Egr-1 attenuated PDGF-induced SphK1 expression and hPASMC proliferation. These studies demonstrate that SphK1 is regulated by PDGF in hPASMCs via the transcription factor Egr-1, promoting cell proliferation. This novel mechanism of SphK1 regulation may be a therapeutic target in pulmonary vascular remodeling in PAH.

  15. Role of platelet-derived growth factor-BB (PDGF-BB) in human pulmonary artery smooth muscle cell proliferation.

    PubMed

    Zhao, Yan; Lv, Wentao; Piao, Hongying; Chu, Xiaojie; Wang, Hao

    2014-08-01

    Pulmonary arterial hypertension (PAH) is a vascular remodeling disease characterized by enhanced proliferation of pulmonary artery smooth muscle cells (PASMCs) and suppressed apoptosis. Platelet-derived growth factor (PDGF) is a potent mitogen involved in cell proliferation and migration. PDGF-BB induces the proliferation and migration of PASMCs and has been proposed to be a key mediator in the progression of PAH. Previous studies have shown that PDGF and its receptor are substantially elevated in lung tissues and PASMCs isolated from patients and animals with PAH, but the underlying mechanisms are still poorly manifested. MAP kinases, including extracellular signal-regulated kinase1/2 (ERK1/2), c-Jun NH2-terminal kinase1/2 (JNK1/2), and p38 are the key intracellular signals for stimuli-induced cell proliferation, survival, and apoptosis. Therefore, the purpose of this study is to determine whether PDGF-BB on cell proliferation process is mediated through the MAP kinases pathway in human PASMCs (HPASMCs). Our results showed PDGF-BB-induced proliferating cell nuclear antigen (PCNA), Cyclin A and Cyclin E expression in a concentration-dependent manner. The expression levels of phosphorylated JNK (p-JNK) was upregulated with 20 ng/ml PDGF-BB treatment, while PDGF-BB could not increase phosphorylated ERK1/2 (p-ERK1/2) and p-38 (p-p38) expression. The effects of PDGF-BB on cell proliferation and survival were weakened after the administration of antagonist of the JNK pathway or si-JNK. In addition, PDGF-BB protected against the loss of mitochondrial membrane potentials evoked by serum deprivation (SD) in a JNK-dependent manner. These results suggest that PDGF-BB promotes HPASMCs proliferation and survival, which is likely to be mediated via the JNK pathway.

  16. Continuous exposure to low amplitude extremely low frequency electrical fields characterizing the vascular streaming potential alters elastin accumulation in vascular smooth muscle cells.

    PubMed

    Bergethon, Peter R; Kindler, Dean D; Hallock, Kevin; Blease, Susan; Toselli, Paul

    2013-07-01

    In normal development and pathology, the vascular system depends on complex interactions between cellular elements, biochemical molecules, and physical forces. The electrokinetic vascular streaming potential (EVSP) is an endogenous extremely low frequency (ELF) electrical field resulting from blood flowing past the vessel wall. While generally unrecognized, it is a ubiquitous electrical biophysical force to which the vascular tree is exposed. Extracellular matrix elastin plays a central role in normal blood vessel function and in the development of atherosclerosis. It was hypothesized that ELF fields of low amplitude would alter elastin accumulation, supporting a link between the EVSP and the biology of vascular smooth muscle cells. Neonatal rat aortic smooth muscle cell cultures were exposed chronically to electrical fields characteristic of the EVSP. Extracellular protein accumulation, DNA content, and electron microscopic (EM) evaluation were performed after 2 weeks of exposure. Stimulated cultures showed no significant change in cellular proliferation as measured by the DNA concentration. The per-DNA normalized protein in the extracellular matrix was unchanged while extracellular elastin accumulation decreased 38% on average. EM analysis showed that the stimulated cells had a 2.85-fold increase in mitochondrial number. These results support the formulation that ELF fields are a potential factor in both normal vessel biology and in the pathogenesis of atherosclerotic diseases including heart disease, stroke, and peripheral vascular disease.

  17. Thrombin potently stimulates cytokine production in human vascular smooth muscle cells but not in mononuclear phagocytes.

    PubMed

    Kranzhöfer, R; Clinton, S K; Ishii, K; Coughlin, S R; Fenton, J W; Libby, P

    1996-08-01

    Thrombosis frequently occurs during atherogenesis and in response to vascular injury. Accumulating evidence supports a role for inflammation in the same situation. The present study therefore sought links between thrombosis and inflammation by determining whether thrombin, which is present in active form at sites of thrombosis, can elicit inflammatory functions of human monocytes and vascular smooth muscle cells (SMCs), two major constituents of advanced atheroma. Human alpha-thrombin (EC50, approximately equal to 500 pmol/L) potently induced interleukin (IL)-6 release from SMCs. The tethered-ligand thrombin receptor appeared to mediate this effect. Furthermore, alpha-thrombin also rapidly increased levels of mRNA encoding IL-6 and monocyte chemotactic protein-1 (MCP-1) in SMCs. In contrast, only alpha-thrombin concentrations of > or = 100 nmol/L could stimulate release of IL-6 or tumor necrosis factor-alpha (TNF alpha) in peripheral blood monocytes or monocyte-derived macrophages. Lipid loading of macrophages did not augment thrombin responsiveness. Likewise, only alpha-thrombin concentrations of > or = 100 nmol/L increased levels of IL-6, IL-1 beta, MCP-1, or TNF alpha mRNA in monocytes. Differential responses of SMCs and monocytes to thrombin extended to early agonist-mediated increases in [Ca2+]i. SMCs and endothelial cells, but not monocytes, contained abundant mRNA encoding the thrombin receptor and displayed cell surface thrombin receptor expression detected with a novel monoclonal antibody. Thus, the level of thrombin receptors appeared to account for the differential thrombin susceptibility of SMCs and monocytes. These data suggest that SMCs may be more sensitive than monocytes/macrophages to thrombin activation in human atheroma. Cytokines produced by thrombin-activated SMCs may contribute to ongoing inflammation in atheroma complicated by thrombosis or subjected to angioplasty.

  18. Cyclin-dependent kinase inhibitor, p21Waf1, regulates vascular smooth muscle cell hypertrophy.

    PubMed

    Okamoto, Kenichi; Kato, Seiya; Arima, Nobuyuki; Fujii, Teruhiko; Morimatsu, Minoru; Imaizumi, Tsutomu

    2004-04-01

    In the process of vascular diseases, smooth muscle cells (SMC) undergo not only hyperplasia but also hypertrophy, resulting in vascular remodeling. A cyclin-dependent kinase inhibitor (CDKI), p21Waf1, has been shown to play an important role in SMC hyperplasia. Here we investigated a potential role of p21Waf1 in SMC hypertrophy. An exposure of cultured rat SMC to serum drove the cell cycle progression with up-regulation of various cell cycle markers and increased activities of cyclin-dependent kinases, but did not cause SMC hypertrophy. In contrast, incubation of SMC for 48 h with angiotensin II (AII, 100 nmol/l) resulted in a significant increase in the cell size measured by flowcytometric forward-angle light scatter assay, in association with an increase in the ratio of [3H]leucine/[3H]thymidine uptake, indicating SMC hypertrophy. At 48 h, p21Waf1 expression was up-regulated in SMC exposed to AII but not in those exposed to serum. These results suggest that p21Waf1 may be involved in hypertrophy. To further investigate this issue, two manipulations of the p21Waf1 gene were performed. Adenovirus-mediated over-expression of p21Waf1 not only reduced S-phasic cells but also caused hypertrophy, despite the exposure to serum. Antisense oligodeoxynucleotide for p21Waf1 inhibited the hypertrophy of SMC exposed to AII. Our data suggest that p21Waf1 may play a role in SMC hypertrophy as well.

  19. Zoledronate upregulates MMP-9 and -13 in rat vascular smooth muscle cells by inducing oxidative stress

    PubMed Central

    Arun, Mehmet Zuhuri; Reel, Buket; Sala-Newby, Graciela B; Bond, Mark; Tsaousi, Aikaterini; Maskell, Perry; Newby, Andrew C

    2016-01-01

    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 zoledronate. Conclusion We conclude that zoledronate increases MMP-9 and MMP-13 expressions in rat VSMCs dependent upon stimulation of the NF-κB pathway by reactive oxygen species. Effects on MMP expression may contribute to the pharmacologic profile of bisphosphonates. PMID:27143852

  20. Extracellular matrix-specific focal adhesions in vascular smooth muscle produce mechanically active adhesion sites

    PubMed Central

    Sun, Zhe; Martinez-Lemus, Luis A.; Hill, Michael A.; Meininger, Gerald A.

    2008-01-01

    Integrin-mediated mechanotransduction in vascular smooth muscle cells (VSMCs) plays an important role in the physiological control of tissue blood flow and vascular resistance. To test whether force applied to specific extracellular matrix (ECM)-integrin interactions could induce myogenic-like mechanical activity at focal adhesion sites, we used atomic force microscopy (AFM) to apply controlled forces to specific ECM adhesion sites on arteriolar VSMCs. The tip of AFM probes were fused with a borosilicate bead (2∼5 μm) coated with fibronectin (FN), collagen type I (CNI), laminin (LN), or vitronectin (VN). ECM-coated beads induced clustering of α5- and β3-integrins and actin filaments at sites of bead-cell contact indicative of focal adhesion formation. Step increases of an upward (z-axis) pulling force (800∼1,600 pN) applied to the bead-cell contact site for FN-specific focal adhesions induced a myogenic-like, force-generating response from the VSMC, resulting in a counteracting downward pull by the cell. This micromechanical event was blocked by cytochalasin D but was enhanced by jasplakinolide. Function-blocking antibodies to α5β1- and αvβ3-integrins also blocked the micromechanical cell event in a concentration-dependent manner. Similar pulling experiments with CNI, VN, or LN failed to induce myogenic-like micromechanical events. Collectively, these results demonstrate that mechanical force applied to integrin-FN adhesion sites induces an actin-dependent, myogenic-like, micromechanical event. Focal adhesions formed by different ECM proteins exhibit different mechanical characteristics, and FN appears of particular relevance in its ability to strongly attach to VSMCs and to induce myogenic-like, force-generating reactions from sites of focal adhesion in response to externally applied forces. PMID:18495809

  1. Thiamylal sodium increased inflammation and the proliferation of vascular smooth muscle cells

    PubMed Central

    Hoka, Sumio

    2016-01-01

    Background Thiamylal sodium is a common anesthetic barbiturate prepared in alkaline solution for clinical use. There is no previously reported study on the effects of barbiturates on the inflammation and proliferation of vascular smooth muscle cells (VSMCs). Here, we examined the effects of clinical-grade thiamylal sodium solution (TSS) on the inflammation and proliferation of rat VSMCs. Methods Expression levels of interleukin (IL)-1α, IL-1β, IL-6, and toll-like receptors in rat VSMCs were detected by quantitative reverse transcription-polymerase chain reaction and microarray analyses. The production of IL-6 by cultured VSMCs or ex vivo-cultured rat aortic segments was detected in supernatants by enzyme-linked immunosorbent assay. VSMC proliferation and viability were determined by the water-soluble tetrazolium-1 assay and trypan blue staining, respectively. Results TSS increased expression of IL-1α, IL-6, and TLR4 in VSMCs in a dose-dependent manner, and reduced IL-1β expression. Ex vivo TSS stimulation of rat aorta also increased IL-6. Low concentrations of TSS enhanced VSMC proliferation, while high concentrations reduced both cell proliferation and viability. Expression of IL-1 receptor antagonist, which regulates cell proliferation, was not increased by TSS stimulation. Exposure of cells to the TSS additive, sodium carbonate, resulted in significant upregulation of IL-1α and IL-6 mRNA levels, to a greater extent than TSS. Conclusions TSS-induced proinflammatory cytokine production by VSMCs is caused by sodium carbonate. However, pure thiamylal sodium has an anti-inflammatory effect in VSMCs. TSS exposure to VSMCs may promote vascular inflammation, leading to the progression of atherosclerosis or in-stent restenosis, resulting in vessel bypass graft failure. PMID:27274372

  2. NOX4 downregulation leads to senescence of human vascular smooth muscle cells

    PubMed Central

    Przybylska, Dorota; Janiszewska, Dorota; Goździk, Aleksandra; Bielak-Zmijewska, Anna; Sunderland, Piotr; Sikora, Ewa; Mosieniak, Grażyna

    2016-01-01

    Senescence is a stress response characterized by an irreversible growth arrest and alterations in certain cell functions. It is believed that both double-strand DNA breaks (DSB) and increased ROS level are the main culprit of senescence. Excessive ROS production is also particularly important in the development of a number of cardiovascular disorders. In this context the involvement of professional ROS-producing enzymes, NADPH oxidases (NOX), was postulated. In contrary to the common knowledge, we have shown that not only increased ROS production but also diminished ROS level could be involved in the induction of senescence. Accordingly, our studies revealed that stress-induced premature senescence (SIPS) of vascular smooth muscle cells (VSMCs) induced by doxorubicin or H2O2, correlates with increased level of DSB and ROS. On the other hand, both SIPS and replicative senescence were accompanied by diminished expression of NOX4. Moreover, inhibition of NOX activity or decrease of NOX4 expression led to permanent growth arrest of VSMCs and secretion of interleukins and VEGF. Interestingly, cells undergoing senescence due to NOX4 depletion neither acquired DSB nor activated DNA damage response. Instead, transient induction of the p27, upregulation of HIF-1alpha, decreased expression of cyclin D1 and hypophosphorylated Rb was observed. Our results showed that lowering the level of ROS-producing enzyme - NOX4 oxidase below physiological level leads to cellular senescence of VSMCs which is correlated with secretion of pro-inflammatory cytokines. Thus the use of specific NOX4 inhibitors for pharmacotherapy of vascular diseases should be carefully considered. PMID:27655718

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

    PubMed Central

    Hortells, Luis; Sosa, Cecilia; Millán, Ángel; Sorribas, Víctor

    2015-01-01

    Background 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. Experimental Procedures and Results 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. Conclusions 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

  4. Oxidized Phospholipid Species Promote in Vivo Differential Cx43 Phosphorylation and Vascular Smooth Muscle Cell Proliferation

    PubMed Central

    Johnstone, Scott R.; Ross, Jeremy; Rizzo, Michael J.; Straub, Adam C.; Lampe, Paul D.; Leitinger, Norbert; Isakson, Brant E.

    2009-01-01

    Regulation of both the expression and function of connexins in the vascular wall is important during atherosclerosis. Progression of the disease state is marked by vascular smooth muscle cell (VSMC) proliferation, which coincides with the reduced expression levels of connexin 43 (Cx43). However, nothing is currently known about the factors that regulate post-translational modifications of Cx43 in atherogenesis, which could be of particular importance, due to the association between site-specific Cx43 phosphorylation and cellular proliferation. We compared the effects of direct carotid applications of two oxidized phospholipid derivatives, 1-palmitoyl-2-oxovaleroyl-sn-glycero-3-phosphorylcholine (POVPC) and 1-palmitoyl-2-glutaroyl-sn-glycero-3-phosphorylcholine (PGPC), on Cx43 expression and phosphorylation, and on cell proliferation. Since both POVPC and PGPC have been shown to act through different intracellular pathways, we hypothesized that each oxidized phospholipid species could induce differential Cx43 phosphorylation events in the cytoplasmically located carboxyl-terminal region of the protein, which could potentially enhance cell proliferation. Application of POVPC caused a reduction in VSMC Cx43 levels, enhanced its phosphorylation at serine (pS) 279/282, and increased VSMC proliferation both in vivo and in vitro. Treatment with PGPC enhanced VSMC pS368 levels with no associated change in proliferation. These oxidized phospholipid-induced Cx43 post-translational changes in VSMCs were consistent with those identified in ApoE−/− mice. Taken together, these results demonstrate that post-translational phosphorylation of Cx43 could be a key factor in the pathogenesis of atherosclerosis. PMID:19608875

  5. Essential Role of TGF-β/Smad Pathway on Statin Dependent Vascular Smooth Muscle Cell Regulation

    PubMed Central

    Rodríguez-Vita, Juan; Sánchez-Galán, Eva; Santamaría, Beatriz; Sánchez-López, Elsa; Rodrigues-Díez, Raquel; Blanco-Colio, Luís Miguel; Egido, Jesús; Ortiz, Alberto; Ruiz-Ortega, Marta

    2008-01-01

    Background The 3-hydroxy-3-methylglutaryl CoA reductase inhibitors (also called statins) exert proven beneficial effects on cardiovascular diseases. Recent data suggest a protective role for Transforming Growth Factor-β (TGF-β) in atherosclerosis by regulating the balance between inflammation and extracellular matrix accumulation. However, there are no studies about the effect of statins on TGF-β/Smad pathway in atherosclerosis and vascular cells. Methodology In cultured vascular smooth muscle cells (VSMCs) statins enhanced Smad pathway activation caused by TGF-β. In addition, statins upregulated TGF-β receptor type II (TRII), and increased TGF-β synthesis and TGF-β/Smad-dependent actions. In this sense, statins, through Smad activation, render VSMCs more susceptible to TGF-β induced apoptosis and increased TGF-β-mediated ECM production. It is well documented that high doses of statins induce apoptosis in cultured VSMC in the presence of serum; however the precise mechanism of this effect remains to be elucidated. We have found that statins-induced apoptosis was mediated by TGF-β/Smad pathway. Finally, we have described that RhoA inhibition is a common intracellular mechanisms involved in statins effects. The in vivo relevance of these findings was assessed in an experimental model of atherosclerosis in apolipoprotein E deficient mice: Treatment with Atorvastatin increased Smad3 phosphorylation and TRII overexpression, associated to elevated ECM deposition in the VSMCs within atheroma plaques, while apoptosis was not detected. Conclusions Statins enhance TGF-β/Smad pathway, regulating ligand levels, receptor, main signaling pathway and cellular responses of VSMC, including apoptosis and ECM accumulation. Our findings show that TGF-β/Smad pathway is essential for statins-dependent actions in VSMCs. PMID:19088845

  6. Functional regulation of ClC-3 in the migration of vascular smooth muscle cells.

    PubMed

    Ganapathi, Sindura B; Wei, Shun-Guang; Zaremba, Angelika; Lamb, Fred S; Shears, Stephen B

    2013-01-01

    Migration of vascular smooth muscle cells (VSMCs) into neointima contributes to atherosclerosis and restenosis. This migration requires coordinated plasmalemmal fluxes of water and ions. Here, we show that aortic VSMC migration depends on the regulation of transmembrane Cl(-) flux by ClC-3, a Cl(-) channel/transporter. The contribution of ClC-3 to plasmalemmal Cl(-) current was studied in VSMCs by electrophysiological recordings. Cl(-) current was negligible in cells perfused with 0 [Ca(2+)]. Raising intracellular [Ca(2+)] to 0.5 μM activated a Cl(-) current (I(Cl.Ca)), approximately half of which was eliminated on inhibition by KN-93 of calmodulin-dependent protein kinase II. I(Cl.Ca) was also halved by inositol-3,4,5,6-tetrakisphosphate, a cellular signal with the biological function of specifically preventing calmodulin-dependent protein kinase II from activating I(Cl.Ca). Gene disruption of ClC-3 reduced I(Cl.Ca) by 50%. Moreover, I(Cl.Ca) in the ClC-3 null VSMCs was not affected by either KN-93 or inositol-3,4,5,6-tetrakisphosphate. We conclude that I(Cl.Ca) is composed of 2 components, one is ClC-3 independent whereas the other is ClC-3 dependent, activated by calmodulin-dependent protein kinase II and inhibited by inositol-3,4,5,6-tetrakisphosphate. We also assayed VSMC migration in transwell assays. Migration was halved in ClC-3 null cells versus wild-type cells. In addition, inhibition of ClC-3 by niflumic acid, KN-93, or inositol-3,4,5,6-tetrakisphosphate each reduced cell migration in wild-type cells but not in ClC-3 null cells. These cell-signaling roles of ClC-3 in VSMC migration suggest new therapeutic approaches to vascular remodeling diseases.

  7. [Spontaneous recanalization after embolization of the renal artery with an Amplatzer vascular plug 4].

    PubMed

    Gómez-Martínez, Pablo; Ciampi Dopazo, Juan José; González Fejás, Ariel; Lanciego, Carlos

    2014-01-01

    The Amplatzer vascular plug (AVP) is an occluding device used in vascular embolizations. Thanks to its excellent maneuverability and effectiveness, it is being used more and more often. The latest version, the AVP 4, enables access to smaller and more tortuous vessels. To date, the only cases of spontaneous recanalization published occurred with earlier versions of the AVP. We present a case of recanalization after renal artery embolization with an AVP 4.

  8. Effect of low dose intra-arterial reserpine on vascular wall norepinephrine content.

    PubMed Central

    Porter, J M; Reiney, C G

    1975-01-01

    A number of reports in recent years have indicated that the administration of low dose intra-arterial reserpine has resulted in significant clinical improvement in patients with symptomatic vasospasm, with the benefits presumably resulting from regional vascular wall norepinephrine depletion with resultant vasodilatation. However, to date, there has been no evidence that such low dose reserpine actually alters vascular wall norepinephrine content. This study was performed to determine both regional and systemic effects of low dose intra-arterial reserpine on vascular-wall norepinephrine content, and the duration of any alterations. Twenty-four mongrel dogs had vascular segments excised and assayed for norepinephrine content, before and for up to 4 weeks following a single injection of reserpine, 0.01 mgm/kg, into one femoral artery. The results indicate a pronounced norepinephrine depletion in the injected femoral arterial system, with the reduction persisting for 2-4 weeks, at which time complete norepinephrine recovery occurred. The visceral vessels sampled also showed considerable norepinephrine depletion, indicating systemic spill-over of the drug from the injected peripheral arterial tree. The visceral vessels, however, showed maximal depletion at 24 hours, with recovery by 7 days. Images Fig. 1a. Fig. 1b. Fig. 1c. PMID:1147709

  9. Characterization of (/sup 3/H)nifedipine binding to intact vascular smooth muscle cells

    SciTech Connect

    Sumimoto, K.; Hirata, M.; Kuriyama, H.

    1988-01-01

    Specific binding of the dihydropyridine Ca2+ antagonist (/sup 3/H)nifedipine to dispersed smooth muscle cells of the porcine coronary artery was investigated and the findings were compared with the binding to microsomes of smooth muscles. Specific binding to intact cells was saturable and reversible. The dissociation constant was 1.93 +/- 0.42 nM and the maximal binding capacity was 59.6 +/- 12.4 fmol/10(6) cells, as assessed by Scatchard analysis of the equilibrium binding at 25 degrees C. The Kd value with intact cells was slightly higher than that observed with microsomes. Specific binding of (/sup 3/H)nifedipine to intact cells was completely displaced by unlabeled dihydropyridine derivatives. Among other Ca2+ antagonists, verapamil and d-cis-diltiazem partially and flunarizine completely inhibited the binding. In the case of microsomes, d-cis-diltiazem stimulated the binding of (/sup 3/H)nifedipine. These results suggest that there may be multiple binding sites for different subclasses of Ca2+ antagonists. Polyvalent cations had no effect on the binding to intact cells. In the case of ethylene glycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid (EGTA)-treated microsomes, the addition of CaCl/sub 2/ and BaCl/sub 2/ increased the Bmax, but the Kd value remained unchanged. MnCl/sub 2/ and CdCl/sub 2/ had stimulatory or inhibitory effects, depending on the concentrations, whereas LaCl3 had no effect. The effect of membrane depolarization on the binding was also examined. When the intact cells were incubated in high (K+)o solution for 60 min, the Kd was lowered to 1.4 nM from the control value of 2.0 nM, thereby indicating that (/sup 3/H)nifedipine binds to Ca2+ channels, with a higher affinity, at depolarized states.

  10. Omega-3 and omega-6 DPA equally inhibit the sphingosylphosphorylcholine-induced Ca2+-sensitization of vascular smooth muscle contraction via inhibiting Rho-kinase activation and translocation

    PubMed Central

    Zhang, Ying; Zhang, Min; Lyu, Bochao; Kishi, Hiroko; Kobayashi, Sei

    2017-01-01

    We previously reported that eicosapentaenoic acid (EPA), an omega-3 polyunsaturated fatty acid (n-3 PUFA), effectively inhibits sphingosylphosphorylcholine (SPC)-induced Ca2+-sensitization of vascular smooth muscle (VSM) contraction which is a major cause of cardiovascular and cerebrovascular vasospasm, and EPA is utilized clinically to prevent cerebrovascular vasospasm. In this study, we clearly demonstrate that docosapentaenoic acid (DPA), which exists in two forms as omega-3 (n-3) and omega-6 (n-6) PUFA, strongly inhibits SPC-induced contraction in VSM tissue and human coronary artery smooth muscle cells (CASMCs), with little effect on Ca2+-dependent contraction. Furthermore, n-3 and n-6 DPA inhibited the activation and translocation of Rho-kinase from cytosol to cell membrane. Additionally, SPC-induced phosphorylation of myosin light chain (MLC) was inhibited in n-3 and n-6 DPA pretreated smooth muscleVSM cells and tissues. In summary, we provide direct evidence that n-3 and n-6 DPA effectively equally inhibits SPC-induced contraction by inhibiting Rho-kinase activation and translocation to the cell membrane. PMID:28169288

  11. Direct demonstration of β1- and evidence against β2- and β3-adrenoceptors, in smooth muscle cells of rat small mesenteric arteries

    PubMed Central

    Briones, Ana M; Daly, Craig J; Jimenez-Altayo, Francesc; Martinez-Revelles, Sonia; Gonzalez, Jose M; McGrath, John C; Vila, Elisabet

    2005-01-01

    Recent evidence supports additional subtypes of vasodilator β-adrenoceptor (β-AR) besides the ‘classical' β2. The aim of this study was to investigate the distribution of β-ARs in the wall of rat mesenteric resistance artery (MRA), to establish the relative roles of β-ARs in smooth muscle and other cell types in mediating vasodilatation and to analyse this in relation to the functional pharmacology. We first examined the vasodilator β-AR subtype using ‘subtype-selective' agonists against the, commonly employed, phenylephrine-induced tone. Concentration-related relaxation was produced by isoprenaline (pEC50: 7.70±0.1) (β1 and β2). Salbutamol (β2), BRL 37344 (β3) and CGP 12177 (atypical β) caused relaxation but were 144, 100 and 263 times less potent than isoprenaline; the ‘β3-adrenoceptor agonist' CL 316243 was ineffective. In arteries precontracted with 5-HT or U 46619, isoprenaline produced concentration-related relaxation but salbutamol, BRL 37344, CGP 12177 and CL 316243 did not. SR 59230A, CGP 12177 and BRL 37344 caused a parallel rightward shift in the concentration–response curve to phenylephrine indicating competitive α1-AR antagonism, explaining the false-positive ‘vasodilator' action against phenylephrine-induced tone. Endothelial denudation but not L-NAME slightly attenuated isoprenaline-mediated vasodilatation in phenylephrine and U 46619 precontracted MRA. The β-AR fluorescent ligand BODIPY TMR-CGP 12177 behaved as an irreversible β1-AR antagonist in MRA and bound to the surface and inside vascular smooth muscle cells in intact vascular wall. β-ARs in smooth muscle cells were observed in a perinuclear location, consistent with the location of Golgi and endoplasmic reticulum. Binding of BODIPY TMR-CGP 12177 was inhibited by BAAM (1 μM) in all three vascular tunics, confirming the presence of β-ARs in adventitia, media and intima. Binding in adventitia was observed in both neuronal and non-neuronal cell types. Lack of co

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

    PubMed

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

    2015-12-29

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

  13. Hemoglobin induced cell trauma indirectly influences endothelial TLR9 activity resulting in pulmonary vascular smooth muscle cell activation

    PubMed Central

    Loomis, Zoe; Eigenberger, Paul; Redinius, Katherine; Lisk, Christina; Karoor, Vijaya; Nozik-Grayck, Eva; Ferguson, Scott K.; Hassell, Kathryn; Nuss, Rachelle; Stenmark, Kurt; Buehler, Paul; Irwin, David C.

    2017-01-01

    It is now well established that both inherited and acquired forms of hemolytic disease can promote pulmonary vascular disease consequent of free hemoglobin (Hb) induced NO scavenging, elevations in reactive oxygen species and lipid peroxidation. It has recently been reported that oxidative stress can activate NFkB through a toll-like receptor 9 (TLR9) mediated pathway; further, TLR9 can be activated by either nuclear or mitochondrial DNA liberated by stress induced cellular trauma. We hypothesis that Hb induced lipid peroxidation and subsequent endothelial cell trauma is linked to TLR9 activation, resulting in IL-6 mediated pulmonary smooth muscle cell proliferation. We examined the effects of Hb on rat pulmonary artery endothelial and smooth muscle cells (rPAEC and rPASMC, respectively), and then utilized TLR9 and IL6 inhibitors, as well as the Hb and heme binding proteins (haptoglobin (Hp) and hemopexin (Hpx), respectively) to further elucidate the aforementioned mediators. Further, we explored the effects of Hb in vivo utilizing endothelial cell (EC) specific myeloid differentiation primary response gene-88 (MyD88) and TLR9 null mice. Our data show that oxidized Hb induces lipid peroxidation, cellular toxicity (5.5 ± 1.7 fold; p≤0.04), increased TLR9 activation (60%; p = 0.01), and up regulated IL6 expression (1.75±0.3 fold; p = 0.04) in rPAEC. Rat PASMC exhibited a more proliferative state (13 ± 1%; p = 0.01) when co-cultured with Hb activated rPAEC. These effects were attenuated with the sequestration of Hb or heme by Hp and Hpx as well as with TLR9 an IL-6 inhibition. Moreover, in both EC-MyD88 and TLR9 null mice Hb-infusion resulted in less lung IL-6 expression compared to WT cohorts. These results demonstrate that Hb-induced lipid peroxidation can initiate a modest TLR9 mediated inflammatory response, subsequently generating an activated SMC phenotype. PMID:28152051

  14. IP-10/CXCR3 Axis Promotes the Proliferation of Vascular Smooth Muscle Cells through ERK1/2/CREB Signaling Pathway.

    PubMed

    Wang, Hui-Jin; Zhou, Yu; Liu, Rui-Ming; Qin, Yuan-Sen; Cen, Ying-Huan; Hu, Ling-Yu; Wang, Shen-Ming; Hu, Zuo-Jun

    2017-03-01

    Excessive proliferation of vascular smooth muscle cells is one of the main pathological processes leading to atherosclerosis and intimal hyperplasia after vascular interventional therapy. Our previous study has shown that interferon-γ inducible protein-10 contributes to the proliferation of vascular smooth muscle cell. However, the underlying mechanisms remain unclear. Extracellular signal-regulated kinase 1/2, serine/threonine kinase Akt, and cAMP response element binding protein are signaling pathways, which are considered to play important roles in the processes of vascular smooth muscle cell proliferation. Moreover, chemokine receptor 3 and Toll-like receptor 4 are potential receptors of inducible protein-10 in this process. In the present study, IP-10 was found to directly induce vascular smooth muscle cell proliferation, and exposure to inducible protein-10 activated extracellular signal-regulated kinase 1/2, serine/threonine kinase, and cAMP response element binding protein signaling. Inhibitor of extracellular signal-regulated kinase 1/2, rather than inhibitor of serine/threonine kinase, inhibited the phosphorylation of cAMP response element binding protein and reduced inducible protein-10-stimulated vascular smooth muscle cell proliferation. Knockdown of cAMP response element binding protein by siRNA inhibited inducible protein-10-induced vascular smooth muscle cell proliferation. Moreover, anti-CXCR3 IgG, instead of anti-Toll-like receptor 4 IgG, reduced inducible protein-10-induced vascular smooth muscle cell proliferation and inducible protein-10-stimulated extracellular signal-regulated kinase 1/2 and cAMP response element binding protein activation. Together, these results indicate that inducible protein-10 promotes vascular smooth muscle cell proliferation via chemokine receptor 3 and activation of extracellular signal-regulated kinase 1/2 inducible protein-10-induced vascular smooth muscle cell proliferation. These data provide important targets

  15. A Novel System for Studying Mechanical Strain Waveform-Dependent Responses in Vascular Smooth Muscle Cells

    PubMed Central

    Lee, Jason; Wong, Mitchell; Smith, Quentin; Baker, Aaron B.

    2013-01-01

    While many studies have examined the effects mechanical forces on vSMCs, there is a limited understanding of how the different arterial strain waveforms that occur in disease and different vascular beds alter vSMC mechanotransduction and phenotype. Here, we present a novel system for applying complex, time-varying strain waveforms to cultured cells and use this system to understand how these waveforms can alter vSMC phenotype and signaling. We have developed a highly adaptable cell culture system that allows the application of mechanical strain to cells in culture and can reproduce the complex dynamic mechanical environment experienced by arterial cells in the body. Using this system, we examined whether the type of applied strain waveform altered phenotypic modulation of vSMCs by mechanical forces. Cells exposed to the brachial waveform had increased phosphorylation of AKT, EGR-1, c-Fos expression and cytoskeletal remodeling in comparison to cells treated with the aortic waveform. In addition, vSMCs exposed to physiological waveforms had adopted a more differentiated phenotype in comparison to those treated with static or sinusoidal cyclic strain, with increased expression of vSMC markers desmin, calponin and SM-22 as well as increased expression of regulatory miRNAs including miR-143, -145 and -221. Taken together, our studies demonstrate the development of a novel system for applying complex, timevarying mechanical forces to cells in culture. In addition, we have shown that physiological strain waveforms have powerful effects on vSMC phenotype. PMID:24096612

  16. Proliferating or interleukin 1-activated human vascular smooth muscle cells secrete copious interleukin 6.

    PubMed Central

    Loppnow, H; Libby, P

    1990-01-01

    The cells that make up blood vessel walls appear to participate actively in local immune and inflammatory responses, as well as in certain vascular diseases. We tested here whether smooth muscle cells (SMC) can produce the important inflammatory mediator IL6. Unstimulated SMC in vitro elaborated 5 X 10(3) pg recIL6/24h (i.e., biological activity equivalent to 5 X 10(3) pg recombinant IL6 (recIL6), as determined in B9-assay with a recIL6 standard). Several pathophysiologically relevant factors augmented IL6 release from SMC including 10 micrograms LPS/ml (10(4) pg recIL6), 10 ng tumor necrosis factor/ml (4 X 10(4) pg recIL6), and most notably 10 ng IL1/ml (greater than or equal to 3.2 X 10(5) pg recIL6). Production of IL6 activity corresponded to IL6 mRNA accumulation and de novo synthesis. SMC released newly synthesized IL6 rapidly, as little metabolically labeled material remained cell-associated. In supernatants of IL1-stimulated SMC, IL6 accounted for as much as 4% of the secreted proteins. In normal vessels SMC seldom divide, but SMC proliferation can occur in hypertension or during atherogenesis. We therefore tested the relationship between IL6 production and SMC proliferation in response to platelet-derived growth factor (PDGF) in vitro. Quiescent SMC released scant IL6 activity, whereas PDGF (1-100 ng/ml) produced concentration-dependent and coordinate enhancement of SMC proliferation and IL6 release (linear regression of growth vs. IL6 release yielded r greater than 0.9). IL6 itself neither stimulated nor inhibited SMC growth or IL6 production. Intact medial strips studied in short-term organoid culture produced large quantities of IL6, similar to the results obtained with cultured SMC. These findings illustrate a new function of vascular SMC by which these cells might participate in local immunoregulation and in the pathogenesis of various important vascular diseases as well as in inflammatory responses generally. Images PMID:2312724

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

    SciTech Connect

    Nagayama, Daiji; Ishihara, Noriko; Bujo, Hideaki; Shirai, Kohji; Tatsuno, Ichiro

    2014-04-18

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

  18. Benefit of SERCA2a gene transfer to vascular endothelial and smooth muscle cells: a new aspect in therapy of cardiovascular diseases.

    PubMed

    Lipskaia, Larissa; Hadri, Lahouaria; Lopez, Jose J; Hajjar, Roger J; Bobe, Regis

    2013-07-01

    Despite the great progress in cardiovascular health and clinical care along with marked decline in morbidity and mortality, cardiovascular diseases remain the leading causes of death and disability in the developed world. New therapeutic approaches, targeting not only systematic but also causal dysfunction, are ultimately needed to provide a valuable alternative for treatment of complex cardiovascular diseases. In heart failure, there are currently a number of trials that have been either completed or are ongoing targeting the sarcoplasmic reticulum calcium ATPase pump (SERCA2a) gene transfer in the context of heart failure. Recently, a phase 2 trial was completed, demonstrating safety and suggested benefit of adeno-associated virus type 1/SERCA2a gene transfer in advanced heart failure, supporting larger confirmatory trials. The experimental and clinical data suggest that, when administrated through perfusion, virus vector carrying SERCA2a can also transduce vascular endothelial and smooth muscle cells (EC and SMC) thereby improving the clinical benefit of gene therapy. Indeed, recent advances in understanding the molecular basis of vascular dysfunction point towards a reduction of sarcoplasmic reticulum Ca2+ uptake and an impairment of Ca2+ cycling in vascular EC and SMC from patients and preclinical models with cardiac diseases or with cardiovascular risk factors such as diabetes, hypercholesterolemia, coronary artery diseases, as well as other conditions such as pulmonary hypertension. In recent years, several studies have established that SERCA2a gene-based therapy could be an efficient option to treat vascular dysfunction. This review focuses on the recent finding showing the beneficial effects of SERCA2a gene transfer in vascular EC and SMC.

  19. Protease-Activated Receptor 2 Promotes Pro-Atherogenic Effects through Transactivation of the VEGF Receptor 2 in Human Vascular Smooth Muscle Cells

    PubMed Central

    Indrakusuma, Ira; Romacho, Tania; Eckel, Jürgen

    2017-01-01

    Background: Obesity is associated with impaired vascular function. In the cardiovascular system, protease-activated receptor 2 (PAR2) exerts multiple functions such as the control of the vascular tone. In pathological conditions, PAR2 is related to vascular inflammation. However, little is known about the impact of obesity on PAR2 in the vasculature. Therefore, we explored the role of PAR2 as a potential link between obesity and cardiovascular diseases. Methods: C57BL/6 mice were fed with either a chow or a 60% high fat diet for 24 weeks prior to isolation of aortas. Furthermore, human coronary artery endothelial cells (HCAEC) and human coronary smooth muscle cells (HCSMC) were treated with conditioned medium obtained from in vitro differentiated primary human adipocytes. To investigate receptor interaction vascular endothelial growth factor receptor 2 (VEGFR2) was blocked by exposure to calcium dobesilate and a VEGFR2 neutralization antibody, before treatment with PAR2 activating peptide. Student's t-test or one-way were used to determine statistical significance. Results: Both, high fat diet and exposure to conditioned medium increased PAR2 expression in aortas and human vascular cells, respectively. In HCSMC, conditioned medium elicited proliferation as well as cyclooxygenase 2 induction, which was suppressed by the PAR2 antagonist GB83. Specific activation of PAR2 by the PAR2 activating peptide induced proliferation and cyclooxygenase 2 expression which were abolished by blocking the VEGFR2. Additionally, treatment of HCSMC with the PAR2 activating peptide triggered VEGFR2 phosphorylation. Conclusion: Under obesogenic conditions, where circulating levels of pro-inflammatory adipokines are elevated, PAR2 arises as an important player linking obesity-related adipose tissue inflammation to atherogenesis. We show for the first time that the underlying mechanisms of these pro-atherogenic effects involve a potential transactivation of the VEGFR2 by PAR2. PMID

  20. Role of cAMP-Phosphodiesterase 1C Signaling in Regulating Growth Factor Receptor Stability, Vascular Smooth Muscle Cell Growth, Migration, and Neointimal Hyperplasia

    PubMed Central

    Cai, Yujun; Nagel, David J.; Zhou, Qian; Cygnar, Katherine D.; Zhao, Haiqing; Li, Faqian; Pi, Xinchun; Knight, Peter A.; Yan, Chen

    2015-01-01

    Objective Neointimal hyperplasia characterized by abnormal accumulation of vascular smooth muscle cells (SMCs) is a hallmark of occlusive disorders such as atherosclerosis, post-angioplasty restenosis, vein graft stenosis, and allograft vasculopathy. Cyclic nucleotides are vital in SMC proliferation and migration, which are regulated by cyclic nucleotide phosphodiesterases (PDEs). Our goal is to understand the regulation and function of PDEs in SMC pathogenesis of vascular diseases. Methods & Results We performed screening for genes differentially expressed in normal contractile versus proliferating synthetic SMCs. We observed that PDE1C expression was low in contractile SMCs but drastically elevated in synthetic SMCs in vitro and in various mouse vascular injury models in vivo. Additionally, PDE1C was highly induced in neointimal SMCs of human coronary arteries. More importantly, injury-induced neointimal formation was significantly attenuated by PDE1C deficiency or PDE1 inhibition in vivo. PDE1 inhibition suppressed vascular remodeling of human saphenous vein explants ex vivo. In cultured SMCs, PDE1C deficiency or PDE1 inhibition attenuated SMC proliferation and migration. Mechanistic studies revealed that PDE1C plays a critical role in regulating the stability of growth factor receptors, such as PDGF-receptor-beta (PDGFRβ) known to be important in pathological vascular remodeling. PDE1C interacts with LDL-receptor-related-protein-1 (LRP1) and PDGFRβ, thus regulating PDGFRβ endocytosis and lysosome-dependent degradation in an LRP1-dependent manner. A transmembrane-adenylyl-cyclase (tmAC)-cAMP-PKA cascade modulated by PDE1C is critical in regulating PDGFRβ degradation. Conclusion These findings demonstrated that PDE1C is an important regulator of SMC proliferation, migration, and neointimal hyperplasia, in part through modulating endosome/lysosome dependent PDGFRβ protein degradation via LRP1. PMID:25608528

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

    SciTech Connect

    Guo, Xiaoxia; Zhou, Chunyan; Sun, Ningling

    2011-04-22

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

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

    PubMed

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

    2017-04-01

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

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

    SciTech Connect

    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.

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

    PubMed

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

    2012-08-01

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

  5. Insulin-independent GLUT4 translocation in proliferative vascular smooth muscle cells involves SM22α.

    PubMed

    Zhao, Li-Li; Zhang, Fan; Chen, Peng; Xie, Xiao-Li; Dou, Yong-Qing; Lin, Yan-Ling; Nie, Lei; Lv, Pin; Zhang, Dan-Dan; Li, Xiao-Kun; Miao, Sui-Bing; Yin, Ya-Juan; Dong, Li-Hua; Song, Yu; Shu, Ya-Nan; Han, Mei

    2017-02-01

    The insulin-sensitive glucose transporter 4 (GLUT4) is a predominant facilitative glucose transporter in vascular smooth muscle cells (VSMCs) and is significantly upregulated in rabbit neointima. This study investigated the role of GLUT4 in VSMC proliferation, the cellular mechanism underlying PDGF-BB-stimulated GLUT4 translocation, and effects of SM22α, an actin-binding protein, on this process. Chronic treatment of VSMCs with PDGF-BB significantly elevated GLUT4 expression and glucose uptake. PDGF-BB-induced VSMC proliferation was dependent on GLUT4-mediated glucose uptake. Meanwhile, the response of GLUT4 to insulin decreased in PDGF-BB-stimulated VSMCs. PDGF-BB-induced GLUT4 translocation partially rescued glucose utilization in insulin-resistant cells. Immunofluorescence and western blot analysis revealed that PDGF-BB induced GLUT4 translocation in an actin dynamics-dependent manner. SM22α disruption facilitated GLUT4 translocation and glucose uptake by promoting actin dynamics and cortical actin polymerization. Similar results were observed in VSMCs of SM22α (-/-) mice. The in vivo experiments showed that the glucose level in the neointima induced by ligation was significantly increased in SM22α (-/-) mice, accompanied by increased neointimal thickness, compared with those in wild-type mice. These findings suggest that GLUT4-mediated glucose uptake is involved in VSMC proliferation, and provide a novel link between SM22α and glucose utilization in PDGF-BB-triggered proliferation.

  6. Autocrine secretion of osteopontin by vascular smooth muscle cells regulates their adhesion to collagen gels.

    PubMed Central

    Weintraub, A. S.; Giachelli, C. M.; Krauss, R. S.; Almeida, M.; Taubman, M. B.

    1996-01-01

    Osteopontin (OPN) is a secreted protein postulated to facilitate vascular smooth muscle cell (VSMC) adhesion and migration. Rat aortic VSMC lines were isolated after infection with recombinant retroviruses harboring OPN sense and antisense constructs. All lines grew normally in monolayer culture. On three-dimensional collagen gels, normal VSMCs and lines containing sense constructs (n=15) or empty vector (n=10) attached to gel and invaded the matrix. Four of five antisense clones did not adhere or invade. Antisense clones had lower OPN levels after stimulation with angiotensin II than sense clones or clones containing the empty vector (antisense, 257+/-102 ng/ml; sense, 473+/-104; vector, 434+/-66). Non-adhering antisense clones had lower mean OPN levels after angiotensin II stimulation (161+/-47 ng/ml) than sense or antisense lines with normal adhesion (486+/-63 ng/ml). The ability to adhere correlated with OPN levels >250 ng/ml. Adhesion and invasion were fully restored with addition of 100 to 200 ng/ml of exogenous OPN and were inhibited in normal VSMCs by incubation with 1 microgram/ml anti-OPN antibody. The autocrine secretion of OPN appears to play an important role in VSMC adhesion, spreading, and invasion. Images Figure 2 Figure 3 Figure 5 Figure 6 Figure 7 PMID:8686750

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

    SciTech Connect

    Eto, Hideyuki; Miyata, Masaaki . E-mail: miyatam@m3.kufm.kagoshima-u.ac.jp; Kume, Noriaki; Minami, Manabu; Itabe, Hiroyuki; Orihara, Koji; Hamasaki, Shuichi; Biro, Sadatoshi; Otsuji, Yutaka; Kita, Toru; Tei, Chuwa

    2006-03-10

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

  8. Poly(ADP-ribose) protects vascular smooth muscle cells from oxidative DNA damage

    PubMed Central

    Zhang, Chao; Luo, Tao; Cui, Shijun; Gu, Yongquan; Bian, Chunjing; Chen, Yibin; Yu, Xiaochun; Wang, Zhonggao

    2015-01-01

    Vascular smooth muscle cells (VSMCs) undergo death during atherosclerosis, a widespread cardiovascular disease. Recent studies suggest that oxidative damage occurs in VSMCs and induces atherosclerosis. Here, we analyzed oxidative damage repair in VSMCs and found that VSMCs are hypersensitive to oxidative damage. Further analysis showed that oxidative damage repair in VSMCs is suppressed by a low level of poly (ADP-ribosyl)ation (PARylation), a key post-translational modification in oxidative damage repair. The low level of PARylation is not caused by the lack of PARP-1, the major poly(ADP-ribose) polymerase activated by oxidative damage. Instead, the expression of poly(ADP-ribose) glycohydrolase, PARG, the enzyme hydrolyzing poly(ADP-ribose), is significantly higher in VSMCs than that in the control cells. Using PARG inhibitor to suppress PARG activity facilitates oxidative damage-induced PARylation as well as DNA damage repair. Thus, our study demonstrates a novel molecular mechanism for oxidative damage-induced VSMCs death. This study also identifies the use of PARG inhibitors as a potential treatment for atherosclerosis. [BMB Reports 2015; 48(6): 354-359] PMID:25748172

  9. Shikonin inhibits TNF-α-induced growth and invasion of rat aortic vascular smooth muscle cells.

    PubMed

    Zhang, Xuemin; Hu, Wenyu; Wu, Fang; Yuan, Xue; Hu, Jian

    2015-08-01

    Shikonin is a naphthoquinone compound extracted from the Chinese herb purple gromwell. Shikonin has broad antibacterial, anti-inflammatory, and antitumor activities. The tumor necrosis factor-α (TNF-α)-induced proliferation and invasion of vascular smooth muscle cells (VSMCs) is an important factor that contributes to atherosclerosis. The effects of shikonin on the proliferation and apoptosis of VSMCs have been reported; however, the function of shikonin on TNF-α-mediated growth and invasion of VSMCs during atherosclerosis remains unclear. In this study, we used Western blot, flow cytometry, real-time quantitative PCR, and enzyme-linked immunosorbent assay to investigate the effect of shikonin on the TNF-α-induced growth and invasion of VSMCs and to determine the underlying mechanism. Our results showed that shikonin inhibits the TNF-α-mediated growth and invasion. Further study revealed that shikonin regulates the activation of nuclear factor kappa B and phosphatidyl inositol 3-kinase signaling pathways; modulates the expression of cyclin D1, cyclin E, B-cell lymphoma 2, and Bax; activates caspase-3 and caspase-9; induces cell cycle arrest; and promotes the apoptosis of VSMCs. Together, our results indicate that shikonin may become a promising agent for the treatment of atherosclerosis and they also establish foundation for the development of anti-atherosclerosis drugs.

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

    PubMed Central

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

    2015-01-01

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