Science.gov

Sample records for a7r5 smooth muscle

  1. The anti-proliferative effects of oleanolic acid on A7r5 cells-Role of UCP2 and downstream FGF-2/p53/TSP-1.

    PubMed

    Han, Yantao; Jiang, Qixiao; Wang, Yu; Li, Wenqian; Geng, Min; Han, Zhiwu; Chen, Xuehong

    2017-12-01

    Vascular smooth muscle cell (VSMC) proliferation is a major contributor to atherosclerosis. This study investigated the inhibitory effects of oleanolic acid (OA) against oxidized low-density lipoprotein (ox-LDL)-induced VSMC proliferation in A7r5 cells and explored underlying molecular mechanism. The cell proliferation was quantified with cell counting kit-8 (CCK-8), in which ox-LDL significantly increased A7r5 cells proliferation, while OA pretreatment effectively alleviated such changes without inducing overt cytotoxicity, as indicated by lactate dehydrogenase (LDH) assay. Quantitative real-time RT-PCR (qRT-PCR) and Western blotting revealed increased UCP2 and FGF-2 expression levels as well as decreased p53 and TSP-1 expression levels in A7r5 cells following ox-LDL exposure, while OA pretreatment reversed such changes. Furthermore, inhibiting UCP2 with genipin remarkably reversed the changes in the expression levels of FGF-2, p53, and TSP-1 induced by ox-LDL exposure; silencing FGF-2 with siRNA did not significantly change the expression levels of UCP2 but effectively reversed the changes in the expression levels of p53 and TSP-1, and activation of p53 with PRIMA-1 only significantly affected the changes in the expression levels of TSP-1, but not in UCP2 or FGF-2, suggesting a UCP-2/FGF-2/p53/TSP-1 signaling in A7r5 cells response to ox-LDL exposure. Additionally, co-treatment of OA and genipin exhibited similar effects to the expression levels of UCP2, FGF-2, p53, and TSP-1 as OA or genipin solo treatment in ox-LDL-exposed A7r5 cells, suggesting the involvement of UCP-2/FGF-2/p53/TSP-1 in the mechanism of OA. In conclusion, OA inhibits ox-LDL-induced VSMC proliferation in A7r5 cells, the mechanism involves the changes in UCP-2/FGF-2/p53/TSP-1. © 2017 International Federation for Cell Biology.

  2. Cytotoxic actions of palytoxin on aortic smooth muscle cells in culture.

    PubMed

    Sheridan, Robert E; Deshpande, Sharad S; Adler, Michael

    2005-01-01

    Palytoxin (PTX), isolated from a zoanthid of the genus Palythoa, is the most potent marine toxin known. Intoxication by PTX leads to vasoconstriction, hemorrhage, ataxia, muscle weakness, ventricular fibrillation, pulmonary hypertension, ischemia and death. In this study, clonal A7r5 rat aortic smooth muscle cells were used to study the mechanism of PTX-mediated cytotoxicity. A7r5 cells exposed to PTX for > or = 15 min exhibited surface granularities, vacuoles and rounding. These alterations culminated in a loss of viability as indicated by marked increases in the release of lactate dehydrogenase. Electrophysiological recording from A7r5 cells disclosed a profound membrane depolarization and an increase in conductance to Na+ and K+. PTX-mediated cytotoxicity could not be reversed by washout or by the addition of 10 microM verapamil but was antagonized by 100 microM ouabain or by removal of extracellular Na+ or Ca2+. In light of the involvement of vascular smooth muscle in PTX poisoning, A7r5 cells could serve as a useful model to test specific drugs for treatment of PTX intoxication. 2005 John Wiley & Sons, Ltd.

  3. Vasopressin V1A receptor mediates cell proliferation through GRK2-EGFR-ERK1/2 pathway in A7r5 cells.

    PubMed

    Zhang, Lingling; Wang, Xiaojun; Cao, Hong; Chen, Yunxuan; Chen, Xianfan; Zhao, Xi; Xu, Feifei; Wang, Yifan; Woo, Anthony Yiu-Ho; Zhu, Weizhong

    2016-12-05

    Abnormal proliferation and hypertrophy of vascular smooth muscle (VSMC), as the main structural component of the vasculature, is an important pathological mechanism of hypertension. Recently, increased levels of arginine vasopressin (AVP) and copeptin, the C-terminal fragment of provasopressin, have been shown to correlate with the development of preeclampsia. AVP targets on the G q -coupled vasopressin V 1A receptor and the G s -coupled V 2 receptor in VSMC and the kidneys to regulate vascular tone and water homeostasis. However, the role of the vasopressin receptor on VSM cell proliferation during vascular remodeling is unclear. Here, we studied the effects of AVP on the proliferation of the rat VSMC-derived A7r5 cells. AVP, in a time- and concentration-dependent manner, promoted A7r5 cell proliferation as indicated by the induction of proliferating cell nuclear antigen expression, methylthiazolyldiphenyl-tetrazolium reduction and incorporation of 5'-bromodeoxyuridine into cellular DNA. These effects, coupled with the phosphorylation of extracellular signal-regulated kinase 1/2 (ERK 1/2 ), were blocked by a V 1A receptor antagonist SR45059 but not by a V 2 receptor antagonist lixivaptan. Although acute activation of V 1A receptor induced ERK 1/2 phosphorylation via a protein kinase C-dependent pathway, this effect was not involved in cell proliferation. Cell proliferation and ERK 1/2 phosphorylation in response to prolonged stimulation with AVP were abolished by inhibition of G protein-coupled receptor kinase 2 (GRK2) and epidermal growth factor receptor (EGFR) using specific inhibitors or small hairpin RNA knock-down. These results suggest that activation of V 1A , but not V 2 receptor, produces a cell proliferative signal in A7r5 cells via a GRK2/EGFR/ERK 1/2 -dependent mechanism. Copyright © 2016. Published by Elsevier B.V.

  4. Excitation of Mytilus smooth muscle.

    PubMed

    Twarog, B M

    1967-10-01

    1. Membrane potentials and tension were recorded during nerve stimulation and direct stimulation of smooth muscle cells of the anterior byssus retractor muscle of Mytilus edulis L.2. The resting potential averaged 65 mV (range 55-72 mV).3. Junction potentials reached 25 mV and decayed to one half maximum amplitude in 500 msec. Spatial summation and facilitation of junction potentials were observed.4. Action potentials, 50 msec in duration and up to 50 mV in amplitude were fired at a membrane potential of 35-40 mV. No overshoot was observed.5. Contraction in response to neural stimulation was associated with spike discharge. Measurement of tension and depolarization in muscle bundles at high K(+) indicated that tension is only produced at membrane potentials similar to those achieved by spike discharge.6. Blocking of junction potentials, spike discharge and contraction by methantheline, an acetylcholine antagonist, supports the hypothesis that the muscle is excited by cholinergic nerves. However, evidence of a presynaptic action of methantheline complicates this argument.

  5. Excitation of Mytilus smooth muscle

    PubMed Central

    Twarog, Betty M.

    1967-01-01

    1. Membrane potentials and tension were recorded during nerve stimulation and direct stimulation of smooth muscle cells of the anterior byssus retractor muscle of Mytilus edulis L. 2. The resting potential averaged 65 mV (range 55-72 mV). 3. Junction potentials reached 25 mV and decayed to one half maximum amplitude in 500 msec. Spatial summation and facilitation of junction potentials were observed. 4. Action potentials, 50 msec in duration and up to 50 mV in amplitude were fired at a membrane potential of 35-40 mV. No overshoot was observed. 5. Contraction in response to neural stimulation was associated with spike discharge. Measurement of tension and depolarization in muscle bundles at high K+ indicated that tension is only produced at membrane potentials similar to those achieved by spike discharge. 6. Blocking of junction potentials, spike discharge and contraction by methantheline, an acetylcholine antagonist, supports the hypothesis that the muscle is excited by cholinergic nerves. However, evidence of a presynaptic action of methantheline complicates this argument. PMID:4383455

  6. The bone morphogenetic protein antagonist gremlin promotes vascular smooth muscle cell apoptosis.

    PubMed

    Maciel, Thiago Trovati; Melo, Rosilene Santos; Campos, Alexandre Holthausen

    2009-01-01

    Previous studies from our laboratory demonstrated that gremlin significantly increases vascular smooth muscle cell (VSMC) proliferation and migration. The present study investigates gremlin expression in the initial stages of rat carotid balloon injury and its effects on VSMC apoptosis. Gremlin mRNA expression was evaluated in rat carotids and cultured VSMCs by quantitative PCR. Apoptosis was analyzed in A7r5 cells and rabbit primary VSMCs following gremlin gene overexpression or silencing by chromatin morphology and caspase-3 activity. Vascular injury promoted a significant decrease in gremlin mRNA levels. In addition, platelet-derived growth factor, angiotensin II and transforming growth factor (TGF)-beta1 promoted coordinated regulation of gremlin and bone morphogenetic protein (BMP)-4 expression in opposite directions according to the confluence status of VSMC culture. In A7r5 cells, gremlin overexpression was able to increase apoptosis, as demonstrated by chromatin morphology and caspase-3 activity, while BMP administration promoted opposite effects. Finally, in agreement with our results, gremlin gene silencing effectively suppressed apoptosis in A7r5 cells and rabbit VSMCs. Gremlin is regulated by growth factors and vascular injury and is involved in modulation of VSMC apoptosis. Modifications of gremlin expression during vascular injury may contribute to the apoptosis resistance of VSMCs.

  7. Intercellular calcium waves in primary cultured rat mesenteric smooth muscle cells are mediated by Connexin43

    PubMed Central

    Halidi, Nadia; Alonso, Florian; Burt, Janis M.; Bény, Jean-Louis; Haefliger, Jacques-Antoine; Meister, Jean-Jacques

    2013-01-01

    Intercellular Ca2 + wave propagation between vascular smooth muscle cells (SMCs) is associated with the propagation of contraction along the vessel. Here, we characterize the involvement of gap junctions (GJs) in Ca2 + wave propagation between SMCs at the cellular level. Gap junctional communication was assessed by the propagation of intercellular Ca2 + waves and the transfer of Lucifer Yellow in A7r5 cells, primary rat mesenteric SMCs (pSMCs), and 6B5N cells, a clone of A7r5 cells expressing higher connexin43 (Cx43) to Cx40 ratio. Mechanical stimulation induced an intracellular Ca2 + wave in pSMC and 6B5N cells that propagated to neighboring cells, whereas Ca2 + waves in A7r5 cells failed to progress to neighboring cells. We demonstrate that Cx43 forms the functional GJs that are involved in mediating intercellular Ca2 + waves and that co-expression of Cx40 with Cx43, depending on their expression ratio, may interfere with Cx43 GJ formation, thus altering junctional communication. PMID:22642233

  8. 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. Copyright © 2015 John Wiley & Sons, Inc.

  9. Mechanotransduction, asthma, and airway smooth muscle

    PubMed Central

    Fabry, Ben; Fredberg, Jeffrey J.

    2008-01-01

    Excessive force generation by airway smooth muscle is the main culprit in excessive airway narrowing during an asthma attack. The maximum force the airway smooth muscle can generate is exquisitely sensitive to muscle length fluctuations during breathing, and is governed by complex mechanotransduction events that can best be studied by a hybrid approach in which the airway wall is modeled in silico so as to set a dynamic muscle load comparable to that experienced in vivo. PMID:18836522

  10. Leiomodin and tropomodulin in smooth muscle

    NASA Technical Reports Server (NTRS)

    Conley, C. A.

    2001-01-01

    Evidence is accumulating to suggest that actin filament remodeling is critical for smooth muscle contraction, which implicates actin filament ends as important sites for regulation of contraction. Tropomodulin (Tmod) and smooth muscle leiomodin (SM-Lmod) have been found in many tissues containing smooth muscle by protein immunoblot and immunofluorescence microscopy. Both proteins cofractionate with tropomyosin in the Triton-insoluble cytoskeleton of rabbit stomach smooth muscle and are solubilized by high salt. SM-Lmod binds muscle tropomyosin, a biochemical activity characteristic of Tmod proteins. SM-Lmod staining is present along the length of actin filaments in rat intestinal smooth muscle, while Tmod stains in a punctate pattern distinct from that of actin filaments or the dense body marker alpha-actinin. After smooth muscle is hypercontracted by treatment with 10 mM Ca(2+), both SM-Lmod and Tmod are found near alpha-actinin at the periphery of actin-rich contraction bands. These data suggest that SM-Lmod is a novel component of the smooth muscle actin cytoskeleton and, furthermore, that the pointed ends of actin filaments in smooth muscle may be capped by Tmod in localized clusters.

  11. Modeling myometrial smooth muscle contraction.

    PubMed

    Bursztyn, Limor; Eytan, Osnat; Jaffa, Ariel J; Elad, David

    2007-04-01

    Existing models of uterine contractions assumed a top-down approach in which the function at the organ or tissue level was explained by the behavior of smaller basic units. A new model of the excitation-contraction process in a single myometrial myocyte was recently developed. This model may be used in a bottom-up approach for the description of the contribution of cellular phenomena to the overall performance of the tissue or organ. In this review, we briefly survey current knowledge of uterine electrophysiology and contractility as well as current modeling techniques, which were successfully used to study the function of various types of muscle cells. In the physiological part of the review, we relate to mechanisms of intracellular Ca(2+) control, Ca(2+) oscillations, and Ca(2+) waves and to the various membranal transport mechanisms regulating ion exchange between the intracellular and extracellular spaces. In addition, we describe the process leading from excitation to contraction. In the modeling part of the review, we present the Hodgkin-Huxley (HH) model of excitation in the squid axon as well as models of Ca(2+) control and the latch-bridge model of Hai and Murphy describing the kinetics of smooth muscle cell (SMC) contraction. We also present integrative models describing more than one of these phenomena. Finally, we suggest how these modeling techniques can be applied to modeling myometrial contraction and thus may significantly contribute to current efforts of research of uterine function.

  12. Mechanisms of smooth muscle responses to inflammation.

    PubMed

    Shea-Donohue, T; Notari, L; Sun, R; Zhao, A

    2012-09-01

    Inflammation-induced changes in smooth muscle may be the consequence of changes in the properties of smooth muscle itself, in the control by nerves and hormones, in the microenvironment, or in the balance of constitutive or induced mediators. A general concept is that the specific characteristics and effects of inflammation can be linked to the nature of the infiltrate and the associated mediators, which are dictated predominantly by the immune environment. Inflammatory mediators may regulate smooth muscle function by directly acting on smooth muscle cells or, indirectly, through stimulation of the release of mediators from other cells. In addition, smooth muscle is not a passive bystander during inflammation and our knowledge of molecular signaling pathways that control smooth muscle function, and the contribution of the immune mechanisms to smooth muscle homeostasis, has expanded greatly in the last decade. Recent studies also demonstrated the relevance of extracellular proteases, of endogenous or exogenous origin, redox imbalance, or epigenetic mechanisms, to gastrointestinal dismotility and inflammation in the context of functional and organic disorders. In this review we discuss the various types of inflammation and the established and emerging mechansims of inflammation-induced changes in smooth muscle morphology and function. © 2012 Blackwell Publishing Ltd.

  13. Mediators on human airway smooth muscle.

    PubMed

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

    1997-01-01

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

  14. Autonomic Modification of Intestinal Smooth Muscle Contractility

    ERIC Educational Resources Information Center

    Montgomery, Laura E. A.; Tansey, Etain A.; Johnson, Chris D.; Roe, Sean M.; Quinn, Joe G.

    2016-01-01

    Intestinal smooth muscle contracts rhythmically in the absence of nerve and hormonal stimulation because of the activity of pacemaker cells between and within the muscle layers. This means that the autonomic nervous system modifies rather than initiates intestinal contractions. The practical described here gives students an opportunity to observe…

  15. Interstitial Cells: Regulators of Smooth Muscle Function

    PubMed Central

    Sanders, Kenton M.; Ward, Sean M.; Koh, Sang Don

    2014-01-01

    Smooth muscles are complex tissues containing a variety of cells in addition to muscle cells. Interstitial cells of mesenchymal origin interact with and form electrical connectivity with smooth muscle cells in many organs, and these cells provide important regulatory functions. For example, in the gastrointestinal tract, interstitial cells of Cajal (ICC) and PDGFRα+ cells have been described, in detail, and represent distinct classes of cells with unique ultrastructure, molecular phenotypes, and functions. Smooth muscle cells are electrically coupled to ICC and PDGFRα+ cells, forming an integrated unit called the SIP syncytium. SIP cells express a variety of receptors and ion channels, and conductance changes in any type of SIP cell affect the excitability and responses of the syncytium. SIP cells are known to provide pacemaker activity, propagation pathways for slow waves, transduction of inputs from motor neurons, and mechanosensitivity. Loss of interstitial cells has been associated with motor disorders of the gut. Interstitial cells are also found in a variety of other smooth muscles; however, in most cases, the physiological and pathophysiological roles for these cells have not been clearly defined. This review describes structural, functional, and molecular features of interstitial cells and discusses their contributions in determining the behaviors of smooth muscle tissues. PMID:24987007

  16. Smooth muscle actin and myosin expression in cultured airway smooth muscle cells.

    PubMed

    Wong, J Z; Woodcock-Mitchell, J; Mitchell, J; Rippetoe, P; White, S; Absher, M; Baldor, L; Evans, J; McHugh, K M; Low, R B

    1998-05-01

    In this study, the expression of smooth muscle actin and myosin was examined in cultures of rat tracheal smooth muscle cells. Protein and mRNA analyses demonstrated that these cells express alpha- and gamma-smooth muscle actin and smooth muscle myosin and nonmuscle myosin-B heavy chains. The expression of the smooth muscle specific actin and myosin isoforms was regulated in the same direction when growth conditions were changed. Thus, at confluency in 1 or 10% serum-containing medium as well as for low-density cells (50-60% confluent) deprived of serum, the expression of the smooth muscle forms of actin and myosin was relatively high. Conversely, in rapidly proliferating cultures at low density in 10% serum, smooth muscle contractile protein expression was low. The expression of nonmuscle myosin-B mRNA and protein was more stable and was upregulated only to a small degree in growing cells. Our results provide new insight into the molecular basis of differentiation and contractile function in airway smooth muscle cells.

  17. Origin of spontaneous rhythmicity in smooth muscle

    PubMed Central

    McHale, Noel; Hollywood, Mark; Sergeant, Gerard; Thornbury, Keith

    2006-01-01

    Rhythmic electrical activity is a feature of most smooth muscles but the mechanical consequences can vary from regular rapid phasic contractions to sustained contracture. For many years it was thought that spontaneous electrical activity originated in smooth muscle cells but recently it has become apparent that there are specialized pacemaker cells in many organs that are morphologically and functionally distinct from smooth muscle and that the former cells are the source of spontaneous electrical activity. Such a pacemaker function is well documented for the ICC of the gastrointestinal tract but evidence is accumulating that ICC-like cells play a similar role in other types of smooth muscle. We have recently shown that there are specialized pacemaking cells in the rabbit urethra which are spontaneously active when freshly isolated, readily distinguishable from smooth muscle cells under bright field illumination and relatively easy to study using patch-clamp and confocal imaging techniques. Recent results suggest that calcium oscillations in isolated rabbit urethral interstitial cells are initiated by calcium release from ryanodine sensitive intracellular stores, that oscillation frequency is very sensitive to the external calcium concentration and that conversion of the primary oscillation to a propagated calcium wave depends upon IP3-induced calcium release. PMID:16239271

  18. In situ analysis of smoothelin-like 1 and calmodulin interactions in smooth muscle cells by proximity ligation.

    PubMed

    Ulke-Lemée, Annegret; Turner, Sara R; MacDonald, Justin A

    2015-11-01

    The smoothelin-like 1 (SMTNL1) protein is the newest member of the smoothelin family of muscle proteins. Two calmodulin (CaM)-binding domains (CBD1 for Ca-CaM; CBD2 for apo-CaM) have been described for the SMTNL1 protein using in vitro assays. We now demonstrate in situ associations of SMTNL1 and CaM in A7r5 smooth muscle cells using the proximity ligation assay (PLA). We quantified CaM-SMTNL1 proximity events accurately after taking into account variations in protein expression levels. The refined method allows quantification of in situ proximity after transient transfection with an associated error of <10%. The proximity of SMTNL1 and CaM in A7r5 cells could be reduced by scrambling the amino acid sequence and mutation of large hydrophobic amino acids of CBD1. The truncation of CBD2 did not influence SMTNL1 proximity to CaM. Ultimately, we conclude that SMTNL1 forms complex interactions with CaM in smooth muscle cells, with a role for CBD1 and possibly the intrinsically disordered region. © 2015 Wiley Periodicals, Inc.

  19. Nicotinic Acetylcholine Receptor Mediates Nicotine-Induced Actin Cytoskeletal Remodeling and Extracellular Matrix Degradation by Vascular Smooth Muscle Cells

    PubMed Central

    Gu, Zhizhan; Fonseca, Vera; Hai, Chi-Ming

    2012-01-01

    Cigarette smoking is a significant risk factor for atherosclerosis, which involves the invasion of vascular smooth muscle cells (VSMCs) from the media to intima. A hallmark of many invasive cells is actin cytoskeletal remodeling in the form of podosomes, accompanied by extracellular matrix (ECM) degradation. A7r5 VSMCs form podosomes in response to PKC activation. In this study, we found that cigarette smoke extract, nicotine, and the cholinergic agonist, carbachol, were similarly effective in inducing the formation of podosome rosettes in A7r5 VSMCs. α-Bungarotoxin and atropine experiments confirmed the involvement of nicotinic acetylcholine receptors (nAChRs). Western blotting and immunofluorescence experiments revealed the aggregation of nAChRs at podosome rosettes. Cycloheximide experiments and media exchange experiments suggested that autocrine factor(s) and intracellular phenotypic modulation are putative mechanisms. In situ zymography experiments indicated that, in response to PKC activation, nicotine-treated cells degraded ECM near podosome rosettes, and possibly endocytose ECM fragments to intracellular compartments. Invasion assay of human aortic smooth muscle cells indicated that nicotine and PKC activation individually and synergistically enhanced cell invasion through ECM. Results from this study suggest that nicotine enhances the ability of VSMCs to degrade and invade ECM. nAChR activation, actin cytoskeletal remodeling and phenotypic modulation are possible mechanisms. PMID:22940282

  20. The Smooth Muscle of the Artery

    DTIC Science & Technology

    1975-01-01

    SMOOTH MUSCLE 91 correlate the activity of this enzyme as determined in vitro with the sphingomyelin which accumulates in the artery with age (Table...and sit-ins. Heidelherg has also been a particularly good example of this type of activity . A more physiological tranquilizer for this area seems to be...401), accompanied by the breakdown of ATP by the actin activated myosin ATPase (73). One of the questions facing us, until a few years ago was whether

  1. Endothelium-independent relaxant effect of Rubus coreanus extracts in corpus cavernosum smooth muscle.

    PubMed

    Lee, Jun Ho; Chae, Mee Ree; Sung, Hyun Hwan; Ko, Mikyeong; Kang, Su Jeong; Lee, Sung Won

    2013-07-01

    Rubus coreanus is a perennial shrub native to the southern part of the Korean peninsula. Although it is known that R. coreanus has a dose-dependent relaxation effect on rabbit corpus cavernosum (CC), the exact mechanism of action by which R. coreanus work is not fully known. To elucidate the direct effects of unripe R. coreanus extract (RCE) on CC smooth muscle cells. Dried unripe R. coreanus fruits were pulverized and extracted with 95% ethanol. Isolated rabbit CC strips were mounted in an organ-bath system, and the effects of RCE were evaluated. To estimate [Ca(2+)]i , we used a Fura-2 fluorescent technique. The effects of unripe RCE on ion channels and the intracellular Ca(2+) concentration ([Ca(2+)]i ) of CC. RCE effectively relaxed phenylephrine (PE)-induced tone in rabbit CC, and removal of the endothelium did not completely abolish the relaxation effect of RCE. Tetraethylammonium (1 mM) did not inhibit RCE-induced relaxation in strips precontracted by PE in the organ bath. However, CaCl2 -induced constriction of CC strips, bathed in Ca(2+)-free buffer and primed with PE, was abolished by RCE. In addition, RCE decreased basal [Ca(2+)]i in corporal smooth muscle cells. The increases of [Ca(2+)]i evoked by 60 mM K(+)-containing solution in A7r5 cells were suppressed by RCE, and RCE relaxed KCl-induced tone in endothelium-free CC, which indicated that RCE blocked the voltage-dependent Ca(2+) channels (VDCCs). RCE decreased basal [Ca(2+)]i and the [Arg8]-vasopressin-induced [Ca(2+)]i increases in A7r5 cells, and RCE inhibited the contraction of endothelium-free CC induced by PE in Ca(2+)-free solution, which suggested that RCE might act as a modulator of corporal smooth muscle cell tone by inhibiting Ca(2+) release from sarcoplasmic reticulum. RCE acts through endothelium-independent and endothelium-dependent pathways to relax CC. RCE may inhibit VDCCs and Ca(2+) release from sarcoplasmic reticulum. © 2013 International Society for Sexual Medicine.

  2. 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

  3. Calcium Channels in Vascular Smooth Muscle

    PubMed Central

    Ghosh, D.; Syed, A.U.; Prada, M.P.; Nystoriak, M.A.; Santana, L.F.; Nieves-Cintrón, M.; Navedo, M.F.

    2017-01-01

    Calcium (Ca2+) plays a central role in excitation, contraction, transcription, and proliferation of vascular smooth muscle cells (VSMs). Precise regulation of intracellular Ca2+concentration ([Ca2+]i) is crucial for proper physiological VSM function. Studies over the last several decades have revealed that VSMs express a variety of Ca2+-permeable channels that orchestrate a dynamic, yet finely tuned regulation of [Ca2+]i. In this review, we discuss the major Ca2+-permeable channels expressed in VSM and their contribution to vascular physiology and pathology. PMID:28212803

  4. 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

  5. Aspects of smooth muscle function in molluscan catch muscle.

    PubMed

    Twarog, B M

    1976-10-01

    1) Catch in Mytilus ABRM may be a specialization of a mechanism common to all muscles that gives rise to stretch resistance in the resting state. Catch appears to be due to actin myosin interaction. Since this interaction is regulated by nerves, it provides a convenient model for studying resting stretch resistance. 2) Studies of the structure of Mytilus ABRM revela two types of intercellular connections: a) direct connections between muscle fibers [these nexal (gap) junctions interconnect the muscle cells electrically]; b) muscle fiber-collagen-muscle fiber connections [these provide mechanical connections between muscle cells via collagen fibers]. The structure of Mytilus ABRM supports speculation that smooth muscle filaments are organized into contractile units. 3) A rise in cAMP levels occurs in response to the relaxing transmitter, serotonin. It is not certain whether the cAMP system directly controls the ability of the contractile proteins to interact or whether it regulates intracellular levels of Ca2+. 4) Calcium ions in activation are derived from two sources: an internal source, probably the sarcoplasmic reticulum, and an external source, across the muscle membrane. 5) The nature of catch remains in question, although most evidence favors the linkage hypothesis.

  6. Pathophysiology of bronchial smooth muscle remodelling in asthma.

    PubMed

    Bara, I; Ozier, A; Tunon de Lara, J-M; Marthan, R; Berger, P

    2010-11-01

    Whereas the role of bronchial smooth muscle remains controversial in healthy subjects its role is well established in asthmatics. Bronchial smooth muscle contraction induces airway narrowing. The smooth muscle also contributes to bronchial inflammation by secreting a range of inflammatory mediators, recruiting and activating inflammatory cells, such as mast cells or T-lymphocytes. In addition, bronchial smooth muscle mass is significantly increased in asthma. Such an increase has been related to a deposition of extracellular matrix proteins, and an increase in both cell size and number. However, the mechanisms of this smooth muscle remodelling are complex and not completely understood. The article will review recent data regarding the pathophysiology of bronchial smooth muscle remodelling in asthma.

  7. Regeneration and Maintenance of Intestinal Smooth Muscle Phenotypes

    NASA Astrophysics Data System (ADS)

    Walthers, Christopher M.

    Tissue engineering is an emerging field of biomedical engineering that involves growing artificial organs to replace those lost to disease or injury. Within tissue engineering, there is a demand for artificial smooth muscle to repair tissues of the digestive tract, bladder, and vascular systems. Attempts to develop engineered smooth muscle tissues capable of contracting with sufficient strength to be clinically relevant have so far proven unsatisfactory. The goal of this research was to develop and sustain mature, contractile smooth muscle. Survival of implanted SMCs is critical to sustain the benefits of engineered smooth muscle. Survival of implanted smooth muscle cells was studied with layered, electrospun polycaprolactone implants with lasercut holes ranging from 0--25% porosity. It was found that greater angiogenesis was associated with increased survival of implanted cells, with a large increase at a threshold between 20% and 25% porosity. Heparan sulfate coatings improved the speed of blood vessel infiltration after 14 days of implantation. With these considerations, thicker engineered tissues may be possible. An improved smooth muscle tissue culture technique was utilized. Contracting smooth muscle was produced in culture by maintaining the native smooth muscle tissue organization, specifically by sustaining intact smooth muscle strips rather than dissociating tissue in to isolated smooth muscle cells. Isolated cells showed a decrease in maturity and contained fewer enteric neural and glial cells. Muscle strips also exhibited periodic contraction and regular fluctuation of intracellular calclium. The muscle strip maturity persisted after implantation in omentum for 14 days on polycaprolactone scaffolds. A low-cost, disposable bioreactor was developed to further improve maturity of cultured smooth muscle cells in an environment of controlled cyclical stress.The bioreactor consistently applied repeated mechanical strain with controllable inputs for strain

  8. Inflammation of bronchial smooth muscle in allergic asthma.

    PubMed

    Begueret, H; Berger, P; Vernejoux, J M; Dubuisson, L; Marthan, R; Tunon-de-Lara, J M

    2007-01-01

    Recent observations in asthma suggest that bronchial smooth muscle is infiltrated by inflammatory cells including mast cells. Such an infiltration may contribute to airway remodelling that is partly due to an increase in smooth muscle mass. Whether muscle increase is the result of smooth muscle cell hypertrophy remains controversial and has not been studied by ultrastructural analysis. A morphometric analysis of airway smooth muscle (ASM) was undertaken in asthmatic patients using electron microscopy to examine the interactions between ASM cells and inflammatory cells. ASM specimens were obtained from 14 asthmatic subjects and nine non-asthmatic controls undergoing fibreoptic endoscopy. Inflammatory cell counts were assessed by immunohistochemistry, and ultrastructural parameters were measured using electron microscopy in a blinded fashion on smooth muscle cells and inflammatory cells. ASM from asthmatic patients was infiltrated by an increased number of mast cells and lymphocytes. Smooth muscle cells and their basal lamina were thicker in asthmatic patients (9.5 (0.8) and 1.4 (0.2) microm) than in controls (6.7 (0.4) and 0.7 (0.1) microm). In asthmatics the extracellular matrix was frequently organised in large amounts between ASM cells. Myofibroblasts within smooth muscle bundles were only observed in asthmatics, some of them displaying a close contact with ASM cells. In asthma, airway myositis is characterised by a direct interaction between ASM cells and mast cells and lymphocytes. Smooth muscle remodelling was present, including cell hypertrophy and abnormal extracellular matrix deposition moulding ASM cells.

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

    PubMed

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

    2012-03-01

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

  10. Vascular Smooth Muscle Cells in Atherosclerosis.

    PubMed

    Bennett, Martin R; Sinha, Sanjay; Owens, Gary K

    2016-02-19

    The historical view of vascular smooth muscle cells (VSMCs) in atherosclerosis is that aberrant proliferation of VSMCs promotes plaque formation, but that VSMCs in advanced plaques are entirely beneficial, for example preventing rupture of the fibrous cap. However, this view has been based on ideas that there is a homogenous population of VSMCs within the plaque, that can be identified separate from other plaque cells (particularly macrophages) using standard VSMC and macrophage immunohistochemical markers. More recent genetic lineage tracing studies have shown that VSMC phenotypic switching results in less-differentiated forms that lack VSMC markers including macrophage-like cells, and this switching directly promotes atherosclerosis. In addition, VSMC proliferation may be beneficial throughout atherogenesis, and not just in advanced lesions, whereas VSMC apoptosis, cell senescence, and VSMC-derived macrophage-like cells may promote inflammation. We review the effect of embryological origin on VSMC behavior in atherosclerosis, the role, regulation and consequences of phenotypic switching, the evidence for different origins of VSMCs, and the role of individual processes that VSMCs undergo in atherosclerosis in regard to plaque formation and the structure of advanced lesions. We think there is now compelling evidence that a full understanding of VSMC behavior in atherosclerosis is critical to identify therapeutic targets to both prevent and treat atherosclerosis. © 2016 American Heart Association, Inc.

  11. ASIC PROTEINS REGULATE SMOOTH MUSCLE CELL MIGRATION

    PubMed Central

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

    2008-01-01

    The purpose of the present study was to investigate Acid Sensing Ion Channel (ASIC) protein expression and importance in cellular migration. We recently demonstrated Epithelial Na+ Channel (ENaC) proteins are required for vascular smooth muscle cell (VSMC) migration, however the role of the closely related ASIC proteins has not been addressed. We used RT-PCR and immunolabeling to determine expression of ASIC1, ASIC2, ASIC3 and ASIC4 in A10 cells. We used small interference RNA to silence individual ASIC expression and determine the importance of ASIC proteins in wound healing and chemotaxis (PDGF-bb) initiated migration. We found ASIC1, ASIC2, and ASIC3, but not ASIC4, expression in A10 cells. ASIC1, ASIC2, and ASIC3 siRNA molecules significantly suppressed expression of their respective proteins compared to non-targeting siRNA (RISC) transfected controls by 63%, 44%, and 55%, respectively. Wound healing was inhibited by 10, 20 and 26% compared to RISC controls following suppression of ASIC1, ASIC2, and ASIC3, respectively. Chemotactic migration was inhibited by 30% and 45%, respectively following suppression of ASIC1 and ASIC3. ASIC2 suppression produced a small, but significant, increase in chemotactic migration (4%). Our data indicate ASIC expression is required for normal migration and may suggest a novel role for ASIC proteins in cellular migration. PMID:17936312

  12. ASIC proteins regulate smooth muscle cell migration.

    PubMed

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

    2008-03-01

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

  13. 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. Copyright © 2016 Elsevier Inc. All rights reserved.

  14. Regulation of gastrointestinal motility—insights from smooth muscle biology

    PubMed Central

    Sanders, Kenton M.; Koh, Sang Don; Ro, Seungil; Ward, Sean M.

    2014-01-01

    Gastrointestinal motility results from coordinated contractions of the tunica muscularis, the muscular layers of the alimentary canal. Throughout most of the gastrointestinal tract, smooth muscles are organized into two layers of circularly or longitudinally oriented muscle bundles. Smooth muscle cells form electrical and mechanical junctions between cells that facilitate coordination of contractions. Excitation–contraction coupling occurs by Ca2+ entry via ion channels in the plasma membrane, leading to a rise in intracellular Ca2+. Ca2+ binding to calmodulin activates myosin light chain kinase; subsequent phosphorylation of myosin initiates cross-bridge cycling. Myosin phosphatase dephosphorylates myosin to relax muscles, and a process known as Ca2+ sensitization regulates the activity of the phosphatase. Gastrointestinal smooth muscles are ‘autonomous’ and generate spontaneous electrical activity (slow waves) that does not depend upon input from nerves. Intrinsic pacemaker activity comes from interstitial cells of Cajal, which are electrically coupled to smooth muscle cells. Patterns of contractile activity in gastrointestinal muscles are determined by inputs from enteric motor neurons that innervate smooth muscle cells and interstitial cells. Here we provide an overview of the cells and mechanisms that generate smooth muscle contractile behaviour and gastrointestinal motility. PMID:22965426

  15. Modeling the dispersion effects of contractile fibers in smooth muscles

    NASA Astrophysics Data System (ADS)

    Murtada, Sae-Il; Kroon, Martin; Holzapfel, Gerhard A.

    2010-12-01

    Micro-structurally based models for smooth muscle contraction are crucial for a better understanding of pathological conditions such as atherosclerosis, incontinence and asthma. It is meaningful that models consider the underlying mechanical structure and the biochemical activation. Hence, a simple mechanochemical model is proposed that includes the dispersion of the orientation of smooth muscle myofilaments and that is capable to capture available experimental data on smooth muscle contraction. This allows a refined study of the effects of myofilament dispersion on the smooth muscle contraction. A classical biochemical model is used to describe the cross-bridge interactions with the thin filament in smooth muscles in which calcium-dependent myosin phosphorylation is the only regulatory mechanism. A novel mechanical model considers the dispersion of the contractile fiber orientations in smooth muscle cells by means of a strain-energy function in terms of one dispersion parameter. All model parameters have a biophysical meaning and may be estimated through comparisons with experimental data. The contraction of the middle layer of a carotid artery is studied numerically. Using a tube the relationships between the internal pressure and the stretches are investigated as functions of the dispersion parameter, which implies a strong influence of the orientation of smooth muscle myofilaments on the contraction response. It is straightforward to implement this model in a finite element code to better analyze more complex boundary-value problems.

  16. Nodular smooth muscle metaplasia in multiple peritoneal endometriosis.

    PubMed

    Kim, Hyun-Soo; Yoon, Gun; Ha, Sang Yun; Song, Sang Yong

    2015-01-01

    We report here an unusual presentation of peritoneal endometriosis with smooth muscle metaplasia as multiple protruding masses on the lateral pelvic wall. Smooth muscle metaplasia is a common finding in rectovaginal endometriosis, whereas in peritoneal endometriosis, smooth muscle metaplasia is uncommon and its nodular presentation on the pelvic wall is even rarer. To the best of our knowledge, this is the first case of nodular smooth muscle metaplasia occurring in peritoneal endometriosis. As observed in this case, when performing laparoscopic surgery in order to excise malignant tumors of intra-abdominal or pelvic organs, it can be difficult for surgeons to distinguish the metastatic tumors from benign nodular pelvic wall lesions, including endometriosis, based on the gross findings only. Therefore, an intraoperative frozen section biopsy of the pelvic wall nodules should be performed to evaluate the peritoneal involvement by malignant tumors. Moreover, this report implies that peritoneal endometriosis, as well as rectovaginal endometriosis, can clinically present as nodular lesions if obvious smooth muscle metaplasia is present. The pathological investigation of smooth muscle cells in peritoneal lesions can contribute not only to the precise diagnosis but also to the structure and function of smooth muscle cells and related cells involved in the histogenesis of peritoneal endometriosis.

  17. Smooth Muscle Differentiation and Patterning in the Urinary Bladder

    PubMed Central

    Tasian, Gregory; Cunha, Gerald; Baskin, Laurence

    2013-01-01

    Smooth muscle differentiation and patterning is a fundamental process in urinary bladder development that involves a complex array of local environmental factors, epithelial-mesenchymal interaction, and signaling pathways. An epithelial signal is necessary to induce smooth muscle differentiation in the adjacent bladder mesenchyme. The bladder epithelium (urothelium) also influences the spatial organization of the bladder wall. Sonic hedgehog (Shh), which is expressed by the urothelium, promotes mesenchymal proliferation and induces differentiation of smooth muscle from embryonic bladder mesenchyme. Shh, whose signal is mediated through various transcription factors including Gli2 and BMP4, is likely also important in the patterning of bladder smooth muscle. However, it is not known to what extent early mediators of mesenchymal migration, other Shh-associated transcription factors, and crosstalk between the Shh signaling cascade and other pathways, are involved in the patterning of bladder smooth muscle. Here we review the role of epithelial-mesenchymal interaction and Shh signaling in smooth muscle differentiation and patterning in the bladder. We also discuss emerging signaling molecules, transcription factors, and mesenchyme properties that might be fruitful areas of future research in the process of smooth muscle formation in the bladder. PMID:20541860

  18. Sphingosylphosphorylcholine inhibits macrophage adhesion to vascular smooth muscle cells.

    PubMed

    Wirrig, Christiane; McKean, Jenny S; Wilson, Heather M; Nixon, Graeme F

    2016-09-01

    Inflammation in de-endothelialised arteries contributes to the development of cardiovascular diseases. The process that initiates this inflammatory response is the adhesion of monocytes/macrophages to exposed vascular smooth muscle cells, typically stimulated by cytokines such as tumour necrosis factor-α (TNF). The aim of this study was to determine the effect of the sphingolipid sphingosylphosphorylcholine (SPC) on the interaction of monocytes/macrophages with vascular smooth muscle cells. Rat aortic smooth muscle cells and rat bone marrow-derived macrophages were co-cultured using an in vitro assay following incubation with sphingolipids to assess inter-cellular adhesion. We reveal that SPC inhibits the TNF-induced adhesion of macrophages to smooth muscle cells. This anti-adhesive effect was the result of SPC-induced changes to the smooth muscle cells (but not the macrophages) and was mediated, at least partly, via the sphingosine 1-phosphate receptor subtype 2. Lipid raft domains were also required. Although SPC did not alter expression or membrane distribution of the adhesion proteins intercellular adhesion molecule-1 and vascular cellular adhesion protein-1 in smooth muscle cells, SPC preincubation inhibited the TNF-induced increase in inducible nitric oxide synthase (NOS2) resulting in a subsequent decrease in nitric oxide production. Inhibiting NOS2 activation in smooth muscle cells led to a decrease in the adhesion of macrophages to smooth muscle cells. This study has therefore delineated a novel pathway which can inhibit the interaction between macrophages and vascular smooth muscle cells via SPC-induced repression of NOS2 expression. This mechanism could represent a potential drug target in vascular disease. Copyright © 2016. Published by Elsevier Inc.

  19. Vascular smooth muscle progenitor cells: building and repairing blood vessels.

    PubMed

    Majesky, Mark W; Dong, Xiu Rong; Regan, Jenna N; Hoglund, Virginia J

    2011-02-04

    Molecular pathways that control the specification, migration, and number of available smooth muscle progenitor cells play key roles in determining blood vessel size and structure, capacity for tissue repair, and progression of age-related disorders. Defects in these pathways produce malformations of developing blood vessels, depletion of smooth muscle progenitor cell pools for vessel wall maintenance and repair, and aberrant activation of alternative differentiation pathways in vascular disease. A better understanding of the molecular mechanisms that uniquely specify and maintain vascular smooth muscle cell precursors is essential if we are to use advances in stem and progenitor cell biology and somatic cell reprogramming for applications directed to the vessel wall.

  20. 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.

  1. Bronchial Smooth Muscle Remodeling in Nonsevere Asthma.

    PubMed

    Girodet, Pierre-Olivier; Allard, Benoit; Thumerel, Matthieu; Begueret, Hugues; Dupin, Isabelle; Ousova, Olga; Lassalle, Régis; Maurat, Elise; Ozier, Annaig; Trian, Thomas; Marthan, Roger; Berger, Patrick

    2016-03-15

    Increased bronchial smooth muscle (BSM) mass is a key feature of airway remodeling that classically distinguishes severe from nonsevere asthma. Proliferation of BSM cells involves a specific mitochondria-dependent pathway in individuals with severe asthma. However, BSM remodeling and mitochondrial biogenesis have not been examined in nonsevere asthma. We aimed to assess whether an increase in BSM mass was also implicated in nonsevere asthma and its relationship with mitochondria and clinical outcomes. We enrolled 34 never-smoker subjects with nonsevere asthma. In addition, we recruited 56 subjects with nonsevere asthma and 19 subjects with severe asthma as comparative groups (COBRA cohort [Cohorte Obstruction Bronchique et Asthme; Bronchial Obstruction and Asthma Cohort; sponsored by the French National Institute of Health and Medical Research, INSERM]). A phenotypic characterization was performed using questionnaires, atopy and pulmonary function testing, exhaled nitric oxide measurement, and blood collection. Bronchial biopsy specimens were processed for immunohistochemistry and electron microscopy analysis. After BSM remodeling assessment, subjects were monitored over a 12-month period. We identified characteristic features of remodeling (BSM area >26.6%) and increased mitochondrial number within BSM in a subgroup of subjects with nonsevere asthma. The number of BSM mitochondria was positively correlated with BSM area (r = 0.78; P < 0.001). Follow-up analysis showed that subjects with asthma with high BSM had worse asthma control and a higher rate of exacerbations per year compared with subjects with low BSM. This study reveals that BSM remodeling and mitochondrial biogenesis may play a critical role in the natural history of nonsevere asthma (Mitasthme study). Clinical trial registered with www.clinicaltrials.gov (NCT00808730).

  2. Inflammation of bronchial smooth muscle in allergic asthma

    PubMed Central

    Begueret, H; Berger, P; Vernejoux, J‐M; Dubuisson, L; Marthan, R; Tunon‐de‐Lara, J M

    2007-01-01

    Background Recent observations in asthma suggest that bronchial smooth muscle is infiltrated by inflammatory cells including mast cells. Such an infiltration may contribute to airway remodelling that is partly due to an increase in smooth muscle mass. Whether muscle increase is the result of smooth muscle cell hypertrophy remains controversial and has not been studied by ultrastructural analysis. A morphometric analysis of airway smooth muscle (ASM) was undertaken in asthmatic patients using electron microscopy to examine the interactions between ASM cells and inflammatory cells. Methods ASM specimens were obtained from 14 asthmatic subjects and nine non‐asthmatic controls undergoing fibreoptic endoscopy. Inflammatory cell counts were assessed by immunohistochemistry, and ultrastructural parameters were measured using electron microscopy in a blinded fashion on smooth muscle cells and inflammatory cells. Results ASM from asthmatic patients was infiltrated by an increased number of mast cells and lymphocytes. Smooth muscle cells and their basal lamina were thicker in asthmatic patients (9.5 (0.8) and 1.4 (0.2) μm) than in controls (6.7 (0.4) and 0.7 (0.1) μm). In asthmatics the extracellular matrix was frequently organised in large amounts between ASM cells. Myofibroblasts within smooth muscle bundles were only observed in asthmatics, some of them displaying a close contact with ASM cells. Conclusion In asthma, airway myositis is characterised by a direct interaction between ASM cells and mast cells and lymphocytes. Smooth muscle remodelling was present, including cell hypertrophy and abnormal extracellular matrix deposition moulding ASM cells. PMID:17189531

  3. Nonparametric Model of Smooth Muscle Force Production During Electrical Stimulation.

    PubMed

    Cole, Marc; Eikenberry, Steffen; Kato, Takahide; Sandler, Roman A; Yamashiro, Stanley M; Marmarelis, Vasilis Z

    2017-03-01

    A nonparametric model of smooth muscle tension response to electrical stimulation was estimated using the Laguerre expansion technique of nonlinear system kernel estimation. The experimental data consisted of force responses of smooth muscle to energy-matched alternating single pulse and burst current stimuli. The burst stimuli led to at least a 10-fold increase in peak force in smooth muscle from Mytilus edulis, despite the constant energy constraint. A linear model did not fit the data. However, a second-order model fit the data accurately, so the higher-order models were not required to fit the data. Results showed that smooth muscle force response is not linearly related to the stimulation power.

  4. Myosin light chain kinase phosphorylation in tracheal smooth muscle

    SciTech Connect

    Stull, J.T.; Hsu, L.C.; Tansey, M.G.

    1990-09-25

    Purified myosin light chain kinase from smooth muscle is phosphorylated by cyclic AMP-dependent protein kinase, protein kinase C, and the multifunctional calmodulin-dependent protein kinase II. Because phosphorylation in a specific site (site A) by any one of these kinases desensitizes myosin light chain kinase to activation by Ca2+/calmodulin, kinase phosphorylation could play an important role in regulating smooth muscle contractility. This possibility was investigated in {sup 32}P-labeled bovine tracheal smooth muscle. Treatment of tissues with carbachol, KCl, isoproterenol, or phorbol 12,13-dibutyrate increased the extent of kinase phosphorylation. Six primary phosphopeptides (A-F) of myosin light chain kinase were identified. Site Amore » was phosphorylated to an appreciable extent only with carbachol or KCl, agents which contract tracheal smooth muscle. The extent of site A phosphorylation correlated to increases in the concentration of Ca2+/calmodulin required for activation. These results show that cyclic AMP-dependent protein kinase and protein kinase C do not affect smooth muscle contractility by phosphorylating site A in myosin light chain kinase. It is proposed that phosphorylation of myosin light chain kinase in site A in contracting tracheal smooth muscle may play a role in the reported desensitization of contractile elements to activation by Ca2+.« less

  5. Rock Tea extract (Jasonia glutinosa) relaxes rat aortic smooth muscle by inhibition of L-type Ca(2+) channels.

    PubMed

    Valero, Marta Sofía; Oliván-Viguera, Aida; Garrido, Irene; Langa, Elisa; Berzosa, César; López, Víctor; Gómez-Rincón, Carlota; Murillo, María Divina; Köhler, Ralf

    2015-12-01

    In traditional herbal medicine, Rock Tea (Jasonia glutinosa) is known for its prophylactic and therapeutic value in various disorders including arterial hypertension. However, the mechanism by which Rock Tea exerts blood pressure-lowering actions has not been elucidated yet. Our aim was to demonstrate vasorelaxing effects of Rock Tea extract and to reveal its possible action mechanism. Isometric myography was conducted on high-K+-precontracted rings from rat thoracic aorta and tested extracts at concentrations of 0.5-5 mg/ml. Whole-cell patch-clamp experiments were performed in rat aortic vascular smooth muscle cells (line A7r5) to determine blocking effects on L-type Ca(2+) channels. Rock Tea extract relaxed the aorta contracted by high [K+] concentration dependently with an EC50 of ≈2.4 mg/ml and produced ≈75 % relaxation at the highest concentration tested. The L-type Ca(2+) channel blocker, verapamil (10(-6) M), had similar effects. Rock Tea extract had no effect in nominally Ca(2+)-free high-K(+) buffer but significantly inhibited contractions to re-addition of Ca(2+). Rock Tea extract inhibited the contractions induced by the L-type Ca(2+) channel activator Bay K 8644 (10(-5) M) and by phenylephrine (10(-6) M). Rock Tea extract and Y-27632 (10(-6) M), Rho-kinase inhibitor, had similar effects and the respective effects were not additive. Patch-clamp experiments demonstrated that Rock Tea extract (2.5 mg/ml) virtually abolished L-type Ca(2+) currents in A7r5. We conclude that Rock Tea extract produced vasorelaxation of rat aorta and that this relaxant effect is mediated by inhibition of L-type Ca(2+) channels. Rock Tea extracts may be of phytomedicinal value for prevention and adjuvant treatment of hypertension and other cardiovascular diseases.

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

    PubMed

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

    2017-04-01

    Vascular smooth muscle cell (VSMC) accumulation and hypertrophy are common in vascular disorders, and inflammation has a crucial role in the development of these diseases. To investigate the effect of inflammation on the neurotransmission of VSMC, bioinformatic analysis was performed, following next generation sequencing. Genes of lipopolysaccharide (LPS)-treated A7r5 cells and phosphate-buffered saline (PBS)-treated A7r5 cells were sequenced via next generation sequencing, and each assay was repeated three times. Differentially expressed genes (DEGs) were obtained using the NOISeq package in R. Subsequently, their potential functions were predicted by functional and pathway enrichment analyses using the Database for Annotation, Visualization and Integrated Discovery online tool. Interaction relationships of the proteins enriched in pathways associated with neurological diseases, the proteins which had interaction relationships with adrenoceptor α 1D ( ADRA1D ) or calcium voltage-gated channel subunit α1 S ( CACNA1S ), separately, were obtained from STRING, and protein-protein interaction (PPI) networks were constructed using Cytoscape software. A total of 2,038 DEGs, including 1,094 upregulated and 944 downregulated genes in the LPS treatment group were identified when compared with the control group. Enrichment analyses showed that NADH:Ubiquinone Oxidoreductase Core Subunit V2 ( NDUFV2 ) was involved in several neurological diseases, including oxidative phosphorylation, Alzheimer's disease, Parkinson's disease and Huntington's disease. Furthermore, NDUFV2 (degree, 20) had a higher degree in the PPI network for DEGs enriched in pathways associated with neurological diseases. In the PPI network for ADRA1D , CACNA1S and the DEGs interacting with them, prohibitin ( PHB ), oxytocin receptor ( OXTR ), collapsin response mediator protein 1 ( CRMP1 ) and dihydropyrimidinase like 2 ( DPYSL2 ) had interaction relationships with both ADRA1D and CACNA1S . To conclude, the

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

    PubMed Central

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

    2017-01-01

    Vascular smooth muscle cell (VSMC) accumulation and hypertrophy are common in vascular disorders, and inflammation has a crucial role in the development of these diseases. To investigate the effect of inflammation on the neurotransmission of VSMC, bioinformatic analysis was performed, following next generation sequencing. Genes of lipopolysaccharide (LPS)-treated A7r5 cells and phosphate-buffered saline (PBS)-treated A7r5 cells were sequenced via next generation sequencing, and each assay was repeated three times. Differentially expressed genes (DEGs) were obtained using the NOISeq package in R. Subsequently, their potential functions were predicted by functional and pathway enrichment analyses using the Database for Annotation, Visualization and Integrated Discovery online tool. Interaction relationships of the proteins enriched in pathways associated with neurological diseases, the proteins which had interaction relationships with adrenoceptor α 1D (ADRA1D) or calcium voltage-gated channel subunit α1 S (CACNA1S), separately, were obtained from STRING, and protein-protein interaction (PPI) networks were constructed using Cytoscape software. A total of 2,038 DEGs, including 1,094 upregulated and 944 downregulated genes in the LPS treatment group were identified when compared with the control group. Enrichment analyses showed that NADH:Ubiquinone Oxidoreductase Core Subunit V2 (NDUFV2) was involved in several neurological diseases, including oxidative phosphorylation, Alzheimer's disease, Parkinson's disease and Huntington's disease. Furthermore, NDUFV2 (degree, 20) had a higher degree in the PPI network for DEGs enriched in pathways associated with neurological diseases. In the PPI network for ADRA1D, CACNA1S and the DEGs interacting with them, prohibitin (PHB), oxytocin receptor (OXTR), collapsin response mediator protein 1 (CRMP1) and dihydropyrimidinase like 2 (DPYSL2) had interaction relationships with both ADRA1D and CACNA1S. To conclude, the present study

  8. Airway smooth muscle dysfunction in Pompe (Gaa-/-) mice.

    PubMed

    Keeler, Allison M; Liu, Donghai; Zieger, Marina; Xiong, Lang; Salemi, Jeffrey; Bellvé, Karl; Byrne, Barry J; Fuller, David D; ZhuGe, Ronghua; ElMallah, Mai K

    2017-06-01

    Pompe disease is an autosomal recessive disorder caused by a deficiency of acid α-glucosidase (GAA), an enzyme responsible for hydrolyzing lysosomal glycogen. Deficiency of GAA leads to systemic glycogen accumulation in the lysosomes of skeletal muscle, motor neurons, and smooth muscle. Skeletal muscle and motor neuron pathology are known to contribute to respiratory insufficiency in Pompe disease, but the role of airway pathology has not been evaluated. Here we propose that GAA enzyme deficiency disrupts the function of the trachea and bronchi and this lower airway pathology contributes to respiratory insufficiency in Pompe disease. Using an established mouse model of Pompe disease, the Gaa -/- mouse, we compared histology, pulmonary mechanics, airway smooth muscle (ASM) function, and calcium signaling between Gaa -/- and age-matched wild-type (WT) mice. Lysosomal glycogen accumulation was observed in the smooth muscle of both the bronchi and the trachea in Gaa -/- but not WT mice. Furthermore, Gaa -/- mice had hyporesponsive airway resistance and bronchial ring contraction to the bronchoconstrictive agents methacholine (MCh) and potassium chloride (KCl) and to a bronchodilator (albuterol). Finally, calcium signaling during bronchiolar smooth muscle contraction was impaired in Gaa -/- mice indicating impaired extracellular calcium influx. We conclude that GAA enzyme deficiency leads to glycogen accumulation in the trachea and bronchi and impairs the ability of lower ASM to regulate calcium and respond appropriately to bronchodilator or constrictors. Accordingly, ASM dysfunction may contribute to respiratory impairments in Pompe disease. Copyright © 2017 the American Physiological Society.

  9. Emblic Leafflower (Phyllanthus emblica L.) Fruits Ameliorate Vascular Smooth Muscle Cell Dysfunction in Hyperglycemia: An Underlying Mechanism Involved in Ellagitannin Metabolite Urolithin A

    PubMed Central

    Zhou, Junxuan; Zhang, Cong

    2018-01-01

    Ellagitannins in Phyllanthus emblica L. (emblic leafflower fruits) have been thought of as the beneficial constituents for ameliorating endocrinal and metabolic diseases including diabetes. However, the effect of emblic leafflower fruits on diabetic vascular complications involved in ellagitannin-derived urolithin metabolites is still rare. In this study, acetylcholine-induced endothelium-independent relaxation in aortas was facilitated upon emblic leafflower fruit consumption in the single dose streptozotocin-induced hyperglycemic rats. Emblic leafflower fruit consumption also suppressed the phosphorylation of Akt (Thr308) in the hyperglycemic aortas. More importantly, urolithin A (UroA) and its derived phase II metabolites were identified as the metabolites upon emblic leafflower fruit consumption by HPLC-ESI-Q-TOF-MS. Moreover, UroA reduced the protein expressions of phosphor-Akt (Thr308) and β-catenin in a high glucose-induced A7r5 vascular smooth muscle cell proliferation model. Furthermore, accumulation of β-catenin protein and activation of Wnt signaling in LiCl-triggered A7r5 cells were also ameliorated by UroA treatment. In conclusion, our data demonstrate that emblic leafflower fruit consumption facilitates the vascular function in hyperglycemic rats by regulating Akt/β-catenin signaling, and the effects are potentially mediated by the ellagitannin metabolite urolithin A.

  10. Smooth muscle adaptation after intestinal transection and resection.

    PubMed

    Thompson, J S; Quigley, E M; Adrian, T E

    1996-09-01

    Changes in motor function occur in the intestinal remnant after intestinal resection. Smooth muscle adaptation also occurs, particularly after extensive resection. The time course of these changes and their interrelationship are unclear. Our aim was to evaluate changes in canine smooth muscle structure and function during intestinal adaptation after transection and resection. Twenty-five dogs underwent either transection (N = 10), 50% distal resection (N = 10), or 50% proximal resection (N = 5). Thickness and length of the circular (CM) and longitudinal (LM) muscle layers were measured four and 12 weeks after resection. In vitro length-tension properties and response to a cholinergic agonist were studied in mid-jejunum and mid-ileum. Transection alone caused increased CM length in the jejunum proximal to the transection but did not affect LM length or muscle thickness. A 50% resection resulted in increased length of CM throughout the intestine and thickening of CM and LM near the anastomosis. Active tension of jejunal CM increased transiently four weeks after resection. Active tension in jejunal LM was decreased 12 weeks after transection and resection. Sensitivity of CM to carbachol was similar after transection and resection. It is concluded that: (1) Structural adaptation of both circular and longitudinal muscle occurs after intestinal resection. (2) This process is influenced by the site of the intestinal remnant. (3) Only minor and transient changes occur in smooth muscle function after resection. (4) Factors other than muscle adaptation are likely involved in the changes in motor function seen following massive bowel resection.

  11. We have contact: endothelial cell-smooth muscle cell interactions.

    PubMed

    Lilly, Brenda

    2014-07-01

    Blood vessels are composed of two primary cell types, endothelial cells and smooth muscle cells, each providing a unique contribution to vessel function. Signaling between these two cell types is essential for maintaining tone in mature vessels, and their communication is critical during development, and for repair and remodeling associated with blood vessel growth. This review will highlight the pathways that endothelial cells and smooth muscle cells utilize to communicate during vessel formation and discuss how disruptions in these pathways contribute to disease. ©2014 Int. Union Physiol. Sci./Am. Physiol. Soc.

  12. Membrane properties of smooth muscle cells in pulmonary hypertensive rats.

    PubMed

    Suzuki, H; Twarog, B M

    1982-05-01

    The membrane properties of smooth muscle cells in rat main pulmonary artery (MPA) and small pulmonary artery (SPA) were investigated during chronic normobaric hypoxia and after monocrotaline injection. As chronic pulmonary hypertension developed, pronounced differences between MPA and SPA were observed. These findings may shed light on mechanisms of smooth muscle hypertrophy. 1) The resting membrane potential of smooth muscle in MPA became less negative than the normal (depolarized), whereas the resting membrane potential of smooth muscle in SPA became more negative (hyperpolarized). 2) In MPA, both the length and time constants diminished. 3) In MPA, the maximum membrane depolarization produced by a 10-fold increase in extracellular [K+] decreased. 4) In SPA, the depolarization observed in K+-free solution was more rapid and greater in amplitude, and the transient hyperpolarization following restoration of K+-containing solution increased. 5) In SPA, initial and sustained depolarization evoked by Na+-deficient solutions were increased. 6) Depolarization in MPA was due to increased membrane permeability, perhaps to Cl-, whereas hyperpolarization in SPA could be attributed to increased activity of an electrogenic Na+-K+ pump.

  13. Smooth Muscle-Mediated Connective Tissue Remodeling in Pulmonary Hypertension

    NASA Astrophysics Data System (ADS)

    Mecham, Robert P.; Whitehouse, Loren A.; Wrenn, David S.; Parks, William C.; Griffin, Gail L.; Senior, Robert M.; Crouch, Edmond C.; Stenmark, Kurt R.; Voelkel, Norbert F.

    1987-07-01

    Abnormal accumulation of connective tissue in blood vessels contributes to alterations in vascular physiology associated with disease states such as hypertension and atherosclerosis. Elastin synthesis was studied in blood vessels from newborn calves with severe pulmonary hypertension induced by alveolar hypoxia in order to investigate the cellular stimuli that elicit changes in pulmonary arterial connective tissue production. A two- to fourfold increase in elastin production was observed in pulmonary artery tissue and medial smooth muscle cells from hypertensive calves. This stimulation of elastin production was accompanied by a corresponding increase in elastin messenger RNA consistent with regulation at the transcriptional level. Conditioned serum harvested from cultures of pulmonary artery smooth muscle cells isolated from hypertensive animals contained one or more low molecular weight elastogenic factors that stimulated the production of elastin in both fibroblasts and smooth muscle cells and altered the chemotactic responsiveness of fibroblasts to elastin peptides. These results suggest that connective tissue changes in the pulmonary vasculature in response to pulmonary hypertension are orchestrated by the medial smooth muscle cell through the generation of specific differentiation factors that alter both the secretory phenotype and responsive properties of surrounding cells.

  14. The pharmacology of a molluscan smooth muscle

    PubMed Central

    Twarog, Betty M.

    1959-01-01

    The effects of a number of pharmacologically active substances on contraction and on membrane polarization of the anterior byssal retractor muscle of Mytilus edulis, L., have been studied. Tetramethylammonium bromide, trimethyl(4-oxopentyl)ammonium chloride and nicotine, like acetylcholine, produced depolarization and sustained contraction. Nicotine, on repeated application, lost acetylcholine-like activity and effectively blocked acetylcholine. In order of decreasing potency, methanthelinium, tubocurarine, benzoquinonium, tetraethylammonium, atropine, pentamethonium, and decamethonium blocked acetylcholine action. These agents did not show initial acetylcholine-like action and did not relax sustained contractions. Adrenaline, noradrenaline, tyramine, dibenamine, phentolamine, and lysergic acid diethylamide relaxed sustained contractions without reducing initial depolarization and tension development in response to acetylcholine or electrical stimuli. Adrenaline and noradrenaline often caused depolarization and contraction when first applied, and displayed relaxing action on subsequent application. PMID:13840060

  15. The pharmacology of a molluscan smooth muscle.

    PubMed

    TWAROG, B M

    1959-09-01

    The effects of a number of pharmacologically active substances on contraction and on membrane polarization of the anterior byssal retractor muscle of Mytilus edulis, L., have been studied. Tetramethylammonium bromide, trimethyl(4-oxopentyl)ammonium chloride and nicotine, like acetylcholine, produced depolarization and sustained contraction. Nicotine, on repeated application, lost acetylcholine-like activity and effectively blocked acetylcholine. In order of decreasing potency, methanthelinium, tubocurarine, benzoquinonium, tetraethylammonium, atropine, pentamethonium, and decamethonium blocked acetylcholine action. These agents did not show initial acetylcholine-like action and did not relax sustained contractions. Adrenaline, noradrenaline, tyramine, dibenamine, phentolamine, and lysergic acid diethylamide relaxed sustained contractions without reducing initial depolarization and tension development in response to acetylcholine or electrical stimuli. Adrenaline and noradrenaline often caused depolarization and contraction when first applied, and displayed relaxing action on subsequent application.

  16. Ablation of Smooth Muscle Caldesmon Affects the Relaxation Kinetics of Arterial Muscle

    PubMed Central

    Guo, Hongqiu; Huang, Renjian; Semba, Shingo; Kordowska, Jolata; Huh, Yang Hoon; Khalina-Stackpole, Yana; Mabuchi, Katsuhide; Kitazawa, Toshio; Wang, C.-L. Albert

    2012-01-01

    Smooth muscle caldesmon (h-CaD) is an actin- and myosin-binding protein that reversibly inhibits the actomyosin ATPase activity in vitro. To test the function of h-CaD in vivo, we eliminated its expression in mice. The h-CaD-null animals appeared normal and fertile, although the litter size was smaller. Tissues from the homozygotes lacked h-CaD and exhibited up-regulation of the non-muscle isoform, l-CaD, in visceral, but not vascular tonic smooth muscles. While the Ca2+-sensitivity of force generation of h-CaD-deficient smooth muscle remained largely unchanged, the kinetic behavior during relaxation in arteries was different. Both intact and permeabilized arterial smooth muscle tissues from the knockout animals relaxed more slowly than those of the wild-type. Since this difference occurred after myosin dephosphorylation was complete, the kinetic effect most likely resulted from slower detachment of unphosphorylated cross-bridges. Detailed analyses revealed that the apparently slower relaxation of h-CaD-null smooth muscle was due to an increase in the amplitude of a slower component of the biphasic tension decay. While the identity of this slower process has not been unequivocally determined, we propose it reflects a thin-filament state that elicits fewer re-attached cross-bridges. Our finding that h-CaD modulates the rate of smooth muscle relaxation clearly supports a role in the control of vascular tone. PMID:23149489

  17. Superoxide production: a procalcifying cell signalling event in osteoblastic differentiation of vascular smooth muscle cells exposed to calcification media.

    PubMed

    Sutra, Thibault; Morena, Marion; Bargnoux, Anne-Sophie; Caporiccio, Bertrand; Canaud, Bernard; Cristol, Jean-Paul

    2008-09-01

    Recent studies showed that hydrogen peroxide (H(2)O(2)) enhanced bone markers expression in vascular smooth muscle cells (VSMCs) implicated in osteoblastic differentiation. This study aimed at investigating the role of NAD(P)H oxidase in vascular calcification processes. A7r5 rat VSMCs were incubated with beta-glycerophosphate (10 mm) or uremic serum to induce a diffuse mineralization. H(2)O(2) production by VSMCs was determinated by chemiluminescence. NAD(P)H oxidase sub-unit (p22(phox)), Cbfa-1, ERK phosphorylation and bone alkaline phosphatase (ALP) expressions were measured by Western blotting. VSMCs exhibited higher production of H(2)O(2) and early expression of p22(phox) with beta-glycerophosphate or uremic serum within 24 h of treatment. beta-glycerophosphate-induced oxidative stress was associated with Cbfa-1 expression followed by ALP expression and activity, meanwhile the VSMCs expressing ALP diffusely calcified their extracellular matrix. Interestingly, diphenyleneiodonium partly prevented the osteoblastic differentiation. Results from this model strongly suggest a major implication of vascular NAD(P)H oxidase in vascular calcification supported by VSMCs osteoblastic differentiation.

  18. Effects of cyclopiazonic acid and dexamethasone on serotonin-induced calcium responses in vascular smooth muscle cells.

    PubMed

    Selli, Cigdem; Tosun, Metiner

    2016-06-01

    We previously observed that sarcoendoplasmic reticulum Ca(2+) ATPase (SERCA) blockade by cyclopiazonic acid (CPA) significantly potentiates serotonin (5-hydroxytryptamine (5-HT))-induced vascular contractions. Furthermore, 5-HT receptor antagonist methysergide partially inhibited CPA-potentiated 5-HT contractions. In the present study, we further investigated whether SERCA inhibition potentiates 5-HT-induced Ca(2+) responses along with attenuating the receptor antagonism by store-operated Ca(2+) (SOC) entry and protein kinase C (PKC)-mediated mechanisms. The effects of dexamethasone that was previously shown to induce SOC entry and enhance 5-HT responses were also tested. For this purpose, intracellular Ca(2+) levels were monitored in A7r5 embryonic rat vascular smooth muscle cells by spectrofluorometry using the fluorescent indicator fura-2. The results showed that CPA, although not dexamethasone, significantly potentiated 5-HT-induced Ca(2+) elevations. Ketanserin partially decreased 5-HT-induced and CPA-potentiated Ca(2+) elevations whereas both PKC inhibitor D-sphingosine and SOC entry blocker 2-aminoethoxydiphenyl borate (2-APB) abolished the remaining responses. The data suggests that diminished antagonistic effect on 5-HT-induced Ca(2+) elevations in the presence of SERCA inhibition is induced by SOC entry and PKC activation.

  19. GLP-1 promotes mitochondrial metabolism in vascular smooth muscle cells by enhancing endoplasmic reticulum-mitochondria coupling.

    PubMed

    Morales, Pablo E; Torres, Gloria; Sotomayor-Flores, Cristian; Peña-Oyarzún, Daniel; Rivera-Mejías, Pablo; Paredes, Felipe; Chiong, Mario

    2014-03-28

    Incretin GLP-1 has important metabolic effects on several tissues, mainly through the regulation of glucose uptake and usage. One mechanism for increasing cell metabolism is modulating endoplasmic reticulum (ER)-mitochondria communication, as it allows for a more efficient transfer of Ca(2+) into the mitochondria, thereby increasing activity. Control of glucose metabolism is essential for proper vascular smooth muscle cell (VSMC) function. GLP-1 has been shown to produce varied metabolic actions, but whether it regulates glucose metabolism in VSMC remains unknown. In this report, we show that GLP-1 increases mitochondrial activity in the aortic cell line A7r5 by increasing ER-mitochondria coupling. GLP-1 increases intracellular glucose and diminishes glucose uptake without altering glycogen content. ATP, mitochondrial potential and oxygen consumption increase at 3h of GLP-1 treatment, paralleled by increased Ca(2+) transfer from the ER to the mitochondria. Furthermore, GLP-1 increases levels of Mitofusin-2 (Mfn2), an ER-mitochondria tethering protein, via a PKA-dependent mechanism. Accordingly, PKA inhibition and Mfn2 down-regulation prevented mitochondrial Ca(2+) increases in GLP-1 treated cells. Inhibiting both Ca(2+) release from the ER and Ca(2+) entry into mitochondria as well as diminishing Mfn2 levels blunted the increase in mitochondrial activity in response to GLP-1. Altogether, these results strongly suggest that GLP-1 increases ER-mitochondria communication in VSMC, resulting in higher mitochondrial activity. Copyright © 2014 Elsevier Inc. All rights reserved.

  20. Effects of oxymetazoline on isolated rat's tracheal smooth muscle.

    PubMed

    Wang, Hsing-Won; Wu, Chi-Chung

    2008-06-01

    Oxymetazoline is often used as a decongestant in rhinitis patients who are suffering from nasal obstruction. It is used as a nasal drop or spray solution. The effect on nasal mucosa in vitro or in vivo is well known. However, the effect of the drug on tracheal smooth muscle has rarely been explored. During administration of the drug to the nose, it might affect the trachea via inhalation. We used our preparation to test the effectiveness of oxymetazoline on isolated rat's tracheal smooth muscle. A 5 mm long portion of rat trachea was submersed in 30 ml Kreb's solution in a muscle bath at 37 degrees C. Changes in tracheal contractility in response to the application of parasympathetic mimetic agents were measured using a transducer connected to a Pentium III computer equipped with polygraphy software. The following assessments were performed: (1) effect on tracheal smooth muscle resting tension; (2) effect on contraction caused by 10(-6)M methacholine as a parasympathetic mimetic; (3) effect of oxymetazoline on electrically induced tracheal smooth muscle contractions. Addition of parasympathetic mimetics to the incubation medium caused the trachea to contract in a dose-dependent manner. Addition of oxymetazoline induced a significant relaxation response when the preparation was up to 10(-4) M. At the same concentration, the drug also could inhibit EFS induced spike contraction. Oxymetazoline had negligible effect on the basal tension of trachea as the concentration increased. The degree of drug-induced tracheal contraction or relaxation was dose-dependent. The study indicated that high concentrations of oxymetazoline might actually antagonize cholinergic receptors of the trachea.

  1. Structural Limits on Force Production and Shortening of Smooth Muscle

    PubMed Central

    Siegman, Marion J.; Davidheiser, Sandra; Mooers, Susan U.; Butler, Thomas M.

    2012-01-01

    This study determined the factors that limit force production and shortening in two smooth muscles having very different relationships between active and passive force as a function of muscle length. The rat anococcygeus muscle develops active force over the range of lengths 0.2 – 2.0× the optimum length for force production (Lo). Passive tension due to extension of the resting muscle occurs only at lengths exceeding Lo. In contrast, the rabbit taenia coli develops force in the range of lengths 0.4– 1.1 Lo, and passive force which is detectable at 0.56 Lo, increases to ~0.45 maximum active force at Lo, and increases sharply with further extension. The anococcygeus muscle can shorten to 0.2 Lo and the taenia coli to 0.4 Lo. Dynamic stiffness and energy usage at short muscle lengths suggest that the limit of shortening in the taenia coli, in contrast to the anococcygeus muscle, is not due to a failure of cross bridge interaction. Phosphorylation of the regulatory myosin light chains in intact muscles decreased to a small extent at short lengths compared to the decrease in force production. The differences in force production and the extent of shortening in the two muscles was maintained even when, following permeabilization, the myosin light chains were irreversibly phosphorylated with ATPγS, indicating that differences in activation played little, if any role. Ultrastructural studies on resting and activated muscles show that the taenia coli, which is rich in connective tissue (unlike the anococcygeus muscle) undergoes marked cellular twisting and contractile filament misalignment at short lengths with compression of the extracellular matrix. As a result, force is not transmitted in the longitudinal axis of the muscle, but is dissipated against an internal load provided by the compressed extracellular matrix. These observations on two very different normal smooth muscles reveal how differences in the relative contribution of active and passive structural

  2. Bladder smooth muscle organ culture preparation maintains the contractile phenotype

    PubMed Central

    Wang, Tanchun; Kendig, Derek M.; Chang, Shaohua; Trappanese, Danielle M.; Chacko, Samuel

    2012-01-01

    Smooth muscle cells, when subjected to culture, modulate from a contractile to a secretory phenotype. This has hampered the use of cell culture for molecular techniques to study the regulation of smooth muscle biology. The goal of this study was to develop a new organ culture model of bladder smooth muscle (BSM) that would maintain the contractile phenotype and aid in the study of BSM biology. Our results showed that strips of BSM subjected to up to 9 days of organ culture maintained their contractile phenotype, including the ability to achieve near-control levels of force with a temporal profile similar to that of noncultured tissues. The technical aspects of our organ culture preparation that were responsible, in part, for the maintenance of the contractile phenotype were a slight longitudinal stretch during culture and subjection of the strips to daily contraction-relaxation. The tissues contained viable cells throughout the cross section of the strips. There was an increase in extracellular collagenous matrix, resulting in a leftward shift in the passive length-tension relationship. There were no significant changes in the content of smooth muscle-specific α-actin, calponin, h-caldesmon, total myosin heavy chain, protein kinase G, Rho kinase-I, or the ratio of SM1 to SM2 myosin isoforms. Moreover the organ cultured tissues maintained functional voltage-gated calcium channels and large-conductance calcium-activated potassium channels. Therefore, we propose that this novel BSM organ culture model maintains the contractile phenotype and will be a valuable tool for the use in cellular/molecular biology studies of bladder myocytes. PMID:22896042

  3. [Smooth muscle hamartoma: anatomoclinical characteristics and nosological limits].

    PubMed

    de la Espriella, J; Grossin, M; Marinho, E; Belaïch, S

    1993-01-01

    Smooth muscle hamartoma is an uncommon cutaneous dysembryoplasia usually diagnosed in infancy. Among the 61 cases published since 1923, 56 were congenital and 3 appeared in young adults. We report a case in which the lesions started at the age of 15 years as a papular plaque in the right mammary region of a young woman. A review of the literature showed that the usual clinical presentation is a frequently pigmented plaque made of often follicular papules and measuring 1 to 10 centimeters on average. Excessive hairiness is the most frequent sign, being observed in more than two-thirds of the cases, and Darier's pseudo-sign is present in about 53 p. 100 of the patients. The disease is electively located on the lumbar region, the back and the root of the limbs. In 3 cases the lesions were generalized and the patients looked like fatty "Michelin-Tire Babies". The course of the disease is always favourable, and associated pathologies remain exceptional: urticaria pigmentosa and psychomotor retardation have been reported in two cases of the generalized form. Histology is characterized by the presence of numerous smooth muscle fibres disseminated in the dermis and diversely oriented, sometimes in contact with hair follicles which retain their normal morphology. The differential clinical diagnosis is with naevocytic naevus, café-au-lait spots, mastocytosis and connective tissue hamartoma. Belatedly revealed forms of the disease must be distinguished from Becker's hamartoma, but it must be known that in certain cases the classification is so difficult that some authors have suggested that smooth muscle hamartoma and Becker's hamartoma are only two poles of a single spectrum of dysembryoplastic lesions involving to varying degrees the epidermic and hair structures. Finally, the distinction between the localized forms of late onset smooth muscle hamartoma and multiple leiomyomas "en plaques" remains difficult both anatomico-clinically and nosologically.

  4. Voltage-independent calcium influx in smooth muscle.

    PubMed

    Guibert, Christelle; Ducret, Thomas; Savineau, Jean-Pierre

    2008-09-01

    In smooth muscle cells, agonists such as neurotransmitters or hormones can induce an increase in [Ca(2+)](i) via a release of intracellular stored calcium or/and an influx of extracellular calcium. The calcium entry pathway operates through a variety of plasmalemmal calcium channels which involve voltage-dependent and voltage-independent calcium channels. Voltage-independent calcium channels include (1) receptor-operated channels (ROCs) activated by agonist-receptor interaction and, in the majority of cases, the downstream signal transduction proteins, (2) store-operated channels (SOCs) activated by the emptying of intracellular Ca(2+) store (mainly the sarcoplasmic reticulum), (3) mechanosensitive or stretch-activated channels (SACs) activated by membrane stretch. Generally, voltage-independent calcium channels are calcium permeable non-selective cation channels with electrophysiological differences, complex regulatory mechanisms and pharmacology. Although the molecular identity of voltage-independent calcium channels is not yet fully elucidated, there are growing evidences that these channels correspond to a new family of membrane proteins encoded by mammalian homologues of specific transient receptor potential (TRP) genes. Several types of TRP proteins are ubiquitously expressed in smooth muscle cells and variations in the expression depend on tissue and species. More recently, other proteins such as Orai1 and STIM1 proteins have been also proposed as participating in the molecular identity of voltage-independent calcium channels. These channels control phenomena such as smooth muscle cells proliferation and/or contraction.

  5. MicroRNA Expression in Human Airway Smooth Muscle Cells

    PubMed Central

    Kuhn, Andrew R.; Schlauch, Karen; Lao, Ronna; Halayko, Andrew J.; Gerthoffer, William T.; Singer, Cherie A.

    2010-01-01

    Defining mechanisms by which differentiated, contractile smooth muscle cells become proliferative and secretory in response to mechanical and environmental stress is crucial for determining the contribution of airway smooth muscle (ASM) to inflammatory responses that result in airway disease. Regulation by microRNAs (miRNAs) has emerged as an important post-transcriptional mechanism regulating gene expression that may modulate ASM phenotype, but little is known about the expression and functions of miRNA in smooth muscle. In the present study we used microarrays to determine whether miRNAs in human ASM cells are altered by a proinflammatory stimulus. In ASM cells exposed to IL-1β, TNF-α, and IFN-γ, we found 11 miRNAs to be significantly down-regulated. We verified decreased expression of miR-25, miR-140*, mir-188, and miR-320 by quantitative PCR. Analysis of miR-25 expression indicates that it has a broad role in regulating ASM phenotype by modulating expression of inflammatory mediators such as RANTES, eotaxin, and TNF-α; genes involved in extracellular matrix turnover; and contractile proteins, most notably myosin heavy chain. miRNA binding algorithms predict that miR-25 targets Krüppel-like factor 4 (KLF4), a potent inhibitor of smooth muscle–specific gene expression and mediator of inflammation. Our study demonstrates that inhibition of miR-25 in cytokine-stimulated ASM cells up-regulates KLF4 expression via a post-transcriptional mechanism. This provides novel evidence that miR-25 targets KLF4 in ASM cells and proposes that miR-25 may be an important mediator of ASM phenotype. PMID:19541842

  6. Angiogenesis is induced by airway smooth muscle strain.

    PubMed

    Hasaneen, Nadia A; Zucker, Stanley; Lin, Richard Z; Vaday, Gayle G; Panettieri, Reynold A; Foda, Hussein D

    2007-10-01

    Angiogenesis is an important feature of airway remodeling in both chronic asthma and chronic obstructive pulmonary disease (COPD). Airways in those conditions are exposed to excessive mechanical strain during periods of acute exacerbations. We recently reported that mechanical strain of human airway smooth muscle (HASM) led to an increase in their proliferation and migration. Sustained growth in airway smooth muscle in vivo requires an increase in the nutritional supply to these muscles, hence angiogenesis. In this study, we examined the hypothesis that cyclic mechanical strain of HASM produces factors promoting angiogenic events in the surrounding vascular endothelial cells. Our results show: 1) a significant increase in human lung microvascular endothelial cell (HMVEC-L) proliferation, migration, and tube formation following incubation in conditioned media (CM) from HASM cells exposed to mechanical strain; 2) mechanical strain of HASM cells induced VEGF expression and release; 3) VEGF neutralizing antibodies inhibited the proliferation, migration, and tube formations of HMVEC-L induced by the strained airway smooth muscle CM; 4) mechanical strain of HASM induced a significant increase in hypoxia-inducible factor-1alpha (HIF-1alpha) mRNA and protein, a transcription factor required for VEGF gene transcription; and 5) mechanical strain of HASM induced HIF-1alpha/VEGF through dual phosphatidylinositol 3-kinase (PI3K)/Akt/mammalian target of rapamycin (mTOR) and ERK pathways. In conclusion, exposing HASM cells to mechanical strain induces signal transduction pathway through PI3K/Akt/mTOR and ERK pathways that lead to an increase in HIF-1alpha, a transcription factor required for VEGF expression. VEGF release by mechanical strain of HASM may contribute to the angiogenesis seen with repeated exacerbation of asthma and COPD.

  7. The magnetic field of gastrointestinal smooth muscle activity

    NASA Astrophysics Data System (ADS)

    Bradshaw, Alan; Ladipo, Jk; Richards, William; Wikswo, John

    1997-11-01

    The gastrointestinal (GI) tract controls the absorption and transport of ingested materials. Its function is determined largely by the electrical activity of the smooth muscle that lines the GI tract. GI electrical activity consists of an omnipresent slowly oscillating wave known as the basic electrical rhythm (BER) that modulates a higher-frequency spiking activity associated with muscle contraction. The BER has been shown to be a reliable indicator of intestinal viability, and thus, recording of smooth muscle activity may have clinical value. The BER is difficult to measure with cutaneous electrodes because layers of low-conductivity fat between the GI tract and the abdominal surface attenuate the potential. On the other hand, the magnetic field associated with GI electrical activity is mostly unaffected by intervening fat layers. We recorded the magnetic fields from GI activity in 12 volunteers using a multichannel Superconducting QUantum Interference Device (SQUID) magnetometer. Characteristics typical of gastric and intestinal BER were apparent in the data. Channels near the epigastrium recorded gastric BER, and channels in intestinal areas recorded small bowel BER. These results suggest that a single multichannel SQUID magnetometer is able to measure gastrointestinal electrical activity from multiple locations around the abdomen simultaneously.

  8. 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

  9. Distinct Cellular Mechanisms Underlie Smooth Muscle Turnover in Vascular Development and Repair.

    PubMed

    Roostalu, Urmas; Aldeiri, Bashar; Albertini, Alessandra; Humphreys, Neil; Simonsen-Jackson, Maj; Wong, Jason K F; Cossu, Giulio

    2018-01-19

    Vascular smooth muscle turnover has important implications for blood vessel repair and for the development of cardiovascular diseases, yet lack of specific transgenic animal models has prevented it's in vivo analysis. The objective of this study was to characterize the dynamics and mechanisms of vascular smooth muscle turnover from the earliest stages of embryonic development to arterial repair in the adult. We show that CD146 is transiently expressed in vascular smooth muscle development. By using CRISPR-Cas9 genome editing and in vitro smooth muscle differentiation assay, we demonstrate that CD146 regulates the balance between proliferation and differentiation. We developed a triple-transgenic mouse model to map the fate of NG2 + CD146 + immature smooth muscle cells. A series of pulse-chase experiments revealed that the origin of aortic vascular smooth muscle cells can be traced back to progenitor cells that reside in the wall of the dorsal aorta of the embryo at E10.5. A distinct population of CD146 + smooth muscle progenitor cells emerges during embryonic development and is maintained postnatally at arterial branch sites. To characterize the contribution of different cell types to arterial repair, we used 2 injury models. In limited wire-induced injury response, existing smooth muscle cells are the primary contributors to neointima formation. In contrast, microanastomosis leads to early smooth muscle death and subsequent colonization of the vascular wall by proliferative adventitial cells that contribute to the repair. Extensive proliferation of immature smooth muscle cells in the primitive embryonic dorsal aorta establishes the long-lived lineages of smooth muscle cells that make up the wall of the adult aorta. A discrete population of smooth muscle cells forms in the embryo and is postnatally sustained at arterial branch sites. In response to arterial injuries, existing smooth muscle cells give rise to neointima, but on extensive damage, they are replaced by

  10. The inhibitory actions of prostaglandins on respiratory smooth muscle

    PubMed Central

    Main, I. H. M.

    1964-01-01

    Prostaglandin E1, in concentrations as low as 1 ng/ml., relaxed isolated tracheal muscle from cat, monkey, rabbit, guinea-pig and ferret. Tracheal muscle from the cat, monkey and rabbit did not exhibit inherent tone and the effect of prostaglandin E1 on these preparations was seen only after a sustained contraction had been produced by a previous dose of acetylcholine or of another agonist. Prostaglandins E2, E3 and F1α also relaxed isolated cat tracheal muscle which had been stimulated by acetylcholine: their activities relative to that of prostaglandin E1 were, respectively, 1.0, 0.2 and 0.002. In the anaesthetized cat prostaglandin E1 increased lung “resistance to inflation” (presumably comparable to bronchial resistance) and the heart rate. In the anaesthetized rabbit and guinea-pig, prostaglandin E1 antagonized the rise in resistance to inflation of the lungs obtained after vagal stimulation or after the intravenous injection of histamine; it sometimes lowered the resistance to inflation in these species. The possibility that prostaglandin may have a local physiological role in the control of bronchial smooth muscle tone is discussed. ImagesFig. 5Fig. 7 PMID:14211681

  11. Defining an olfactory receptor function in airway smooth muscle cells

    PubMed Central

    Aisenberg, William H.; Huang, Jessie; Zhu, Wanqu; Rajkumar, Premraj; Cruz, Randy; Santhanam, Lakshmi; Natarajan, Niranjana; Yong, Hwan Mee; De Santiago, Breann; Oh, Jung Jin; Yoon, A-Rum; Panettieri, Reynold A.; Homann, Oliver; Sullivan, John K.; Liggett, Stephen B.; Pluznick, Jennifer L.; An, Steven S.

    2016-01-01

    Pathways that control, or can be exploited to alter, the increase in airway smooth muscle (ASM) mass and cellular remodeling that occur in asthma are not well defined. Here we report the expression of odorant receptors (ORs) belonging to the superfamily of G-protein coupled receptors (GPCRs), as well as the canonical olfaction machinery (Golf and AC3) in the smooth muscle of human bronchi. In primary cultures of isolated human ASM, we identified mRNA expression for multiple ORs. Strikingly, OR51E2 was the most highly enriched OR transcript mapped to the human olfactome in lung-resident cells. In a heterologous expression system, OR51E2 trafficked readily to the cell surface and showed ligand selectivity and sensitivity to the short chain fatty acids (SCFAs) acetate and propionate. These endogenous metabolic byproducts of the gut microbiota slowed the rate of cytoskeletal remodeling, as well as the proliferation of human ASM cells. These cellular responses in vitro were found in ASM from non-asthmatics and asthmatics, and were absent in OR51E2-deleted primary human ASM. These results demonstrate a novel chemo-mechanical signaling network in the ASM and serve as a proof-of-concept that a specific receptor of the gut-lung axis can be targeted to treat airflow obstruction in asthma. PMID:27905542

  12. Effects of palytoxin on isolated intestinal and vascular smooth muscles.

    PubMed

    Ito, K; Karaki, H; Ishida, Y; Urakawa, N; Deguchi, T

    1976-12-01

    Palytoxin (PTX), the most potent marine toxin isolated from the Zoanthid, Palythoa tuberculosa, was studied to determine the effect on isolated smooth muscles. In guinea pig taenia coli PTX at above 3 X 10(-10) g/ml caused a contraction which slowly subsided under isotonic recording. Under isometric recording PTX at above 1 X 10(-10) g/ml caused a contraction which depended on the spontaneous activity. The PTX-induced contraction was not affected by atropine, tripelenmamine or tetrodotoxin but was inhibited by 5 mM Mg, norephinrphrine, isoprenaline or papaverine. PTX at above 1 X 10(-9) g/ml induced an increase in spike frequency and a slight depolarization accompanied with a contraction when measured using a sucrose gap method. In some cases the spike generation was almost abolished after a long exposure to higher dose of PTX and the developed tension gradually decreased. Under isometric recording PTX caused a sustained contraction in rabbit aorta, dog mesenteric and coronary arteries at above 1 X 10(-10) and 1 X 10(-11) g/ml, respectively, in a dose-dependent manner. The coronary artery was most sensitive among the preparation used. PTX-induced contraction in aorta was irreversible, was not influenced by phentolamine but diminished with 5 mM Mg and disappeared in a D-600 or Ca-free medium. PTX is thus an extremely potent and direct stimulant which acts on smooth muscles.

  13. Myocardin Regulates Vascular Smooth Muscle Cell Inflammatory Activation and Disease

    PubMed Central

    Ackers-Johnson, Matthew; Talasila, Amarnath; Sage, Andrew P; Long, Xiaochun; Bot, Ilze; Morrell, Nicholas W; Bennett, Martin R; Miano, Joseph M.; Sinha, Sanjay

    2015-01-01

    Objective Atherosclerosis, the cause of 50% of deaths in westernised societies, is widely regarded as a chronic vascular inflammatory disease. Vascular smooth muscle cell (VSMC) inflammatory activation in response to local pro-inflammatory stimuli contributes to disease progression and is a pervasive feature in developing atherosclerotic plaques. Therefore, it is of considerable therapeutic importance to identify mechanisms that regulate the VSMC inflammatory response. Approach and Results We report that myocardin, a powerful myogenic transcriptional coactivator, negatively regulates VSMC inflammatory activation and vascular disease. Myocardin levels are reduced during atherosclerosis, in association with phenotypic switching of smooth muscle cells. Myocardin deficiency accelerates atherogenesis in hypercholesterolemic ApoE−/− mice. Conversely, increased myocardin expression potently abrogates the induction of an array of inflammatory cytokines, chemokines and adhesion molecules in VSMCs. Expression of myocardin in VSMCs reduces lipid uptake, macrophage interaction, chemotaxis and macrophage-endothelial tethering in vitro, and attenuates monocyte accumulation within developing lesions in vivo. These results demonstrate that endogenous levels of myocardin are a critical regulator of vessel inflammation. Conclusions We propose myocardin as a guardian of the contractile, non-inflammatory VSMC phenotype, with loss of myocardin representing a critical permissive step in the process of phenotypic transition and inflammatory activation, at the onset of vascular disease. PMID:25614278

  14. Endothelin ETA receptor expression in human cerebrovascular smooth muscle cells.

    PubMed Central

    Yu, J. C.; Pickard, J. D.; Davenport, A. P.

    1995-01-01

    1. Endothelin (ET) has been implicated in cerebrovasospasm for example, following subarachnoid haemorrhage, and blocking the interaction of ET with its receptors on cerebral vessels, may be of therapeutic benefit. The aim of our study was to characterize endothelin receptor sub-types on medial smooth muscle cells of human cerebral vessels. Cultures of vascular smooth muscle cells were explanted from human cerebral resistance vessels and characterized as human brain smooth muscle cells (HBSMCs). 2. Over a 48 h incubation period, HBSMC cultures secreted comparable levels of immunoreactive (IR) big endothelin-1 (big ET-1) and IR endothelin (ET): 12.7 +/- 10.3 and 8.3 +/- 5.6 pmol/10(6) cells, respectively (mean +/- s.e. mean from three different individuals), into the culture medium. 3. Total RNA was extracted from cultures of human brain smooth muscle cells. Reverse-transcriptase polymerase chain reaction (RI-PCR) assays and subsequent product separation by agarose gel electrophoresis revealed single bands corresponding to the expected product sizes encoding cDNA for ETA (299 base pairs) and ETB (428 base pairs) (n = 3 different cultures). 4. Autoradiography demonstrated the presence of specific binding sites for [125I]-ET-1 which labels all ET receptors, and [125I]-PD151242, an ETA subtype-selective antagonist which exclusively labels ETA receptors, but no specific-binding was detected using ETB subtype-selective [125I]-BQ3020 (n = 3 different cultures, in duplicate). 5. In saturation binding assays, [123I]-ET-1 bound with high affinity: KD = 0.8 +/- 0.1 nM and Bmax = 690 +/- 108 fmol mg-1. A one-site fit was preferred and Hill slopes were close to unity over the concentration range (10(-12) to 10(-8) M). [125I]-PD151242 also bound with similar affinity: KD = 0.4 +/- 0.1 nM and Bmax = 388 +/- 68 fmol mg-1 (mean +/- s.e. mean, n = 3 different cultures). Again, a one-site fit was preferred and Hill slopes were close to unity over the concentration range. Unlabelled PD

  15. Phosphate and ADP Differently Inhibit Coordinated Smooth Muscle Myosin Groups

    PubMed Central

    Hilbert, Lennart; Balassy, Zsombor; Zitouni, Nedjma B.; Mackey, Michael C.; Lauzon, Anne-Marie

    2015-01-01

    Actin filaments propelled in vitro by groups of skeletal muscle myosin motors exhibit distinct phases of active sliding or arrest, whose occurrence depends on actin length (L) within a range of up to 1.0 μm. Smooth muscle myosin filaments are exponentially distributed with ≈150 nm average length in vivo—suggesting relevance of the L-dependence of myosin group kinetics. Here, we found L-dependent actin arrest and sliding in in vitro motility assays of smooth muscle myosin. We perturbed individual myosin kinetics with varying, physiological concentrations of phosphate (Pi, release associated with main power stroke) and adenosine diphosphate (ADP, release associated with minor mechanical step). Adenosine triphosphate was kept constant at physiological concentration. Increasing [Pi] lowered the fraction of time for which actin was actively sliding, reflected in reduced average sliding velocity (ν) and motile fraction (fmot, fraction of time that filaments are moving); increasing [ADP] increased the fraction of time actively sliding and reduced the velocity while sliding, reflected in reduced ν and increased fmot. We introduced specific Pi and ADP effects on individual myosin kinetics into our recently developed mathematical model of actin propulsion by myosin groups. Simulations matched our experimental observations and described the inhibition of myosin group kinetics. At low [Pi] and [ADP], actin arrest and sliding were reflected by two distinct chemical states of the myosin group. Upon [Pi] increase, the probability of the active state decreased; upon [ADP] increase, the probability of the active state increased, but the active state became increasingly similar to the arrested state. PMID:25650929

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

    PubMed

    Elofsson, Rolf; Kröger, Ronald H H

    2017-01-01

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

  17. Superparamagnetic iron oxide nanoparticles regulate smooth muscle cell phenotype

    PubMed Central

    Angelopoulos, Ioannis; Southern, Paul; Pankhurst, Quentin A.

    2016-01-01

    Abstract Superparamagnetic iron oxide nanoparticles (SPION) are used for an increasing range of biomedical applications, from imaging to mechanical actuation of cells and tissue. The aim of this study was to investigate the loading of smooth muscle cells (SMC) with SPION and to explore what effect this has on the phenotype of the cells. Adherent human SMC were loaded with ∼17 pg of unconjugated, negatively charged, 50 nm SPION. Clusters of the internalized SPION particles were held in discrete cytoplasmic vesicles. Internalized SPION did not cause any change in cell morphology, proliferation, metabolic activity, or staining pattern of actin and calponin, two of the muscle contractile proteins involved in force generation. However, internalized SPION inhibited the increased gene expression of actin and calponin normally observed when cells are incubated under differentiation conditions. The observed change in the control of gene expression of muscle contractile apparatus by SPION has not previously been described. This finding could offer novel approaches for regulating the phenotype of SMC and warrants further investigation. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 104A: 2412–2419, 2016. PMID:27176658

  18. Regulation of human penile smooth muscle tone byprostanoid receptors

    PubMed Central

    Angulo, Javier; Cuevas, Pedro; La Fuente, Jose M; Pomerol, Jose M; Ruiz-Castañé, Eduardo; Puigvert, Ana; Gabancho, Sonia; Fernández, Argentina; Ney, Peter; Sáenz de Tejada, Iñigo

    2002-01-01

    We have characterized the prostanoid receptors involved in the regulation of human penile arterial and trabecular smooth muscle tone.Arachidonic acid induced relaxation of human corpus cavernosum strips (HCCS) that was blocked by the cyclo-oxygenase inhibitor, indomethacin, and augmented by the thromboxane receptor (TP) antagonist, SQ29548, suggesting that endogenous production of prostanoids regulates penile smooth muscle tone.TP-receptors mediate contraction of HCCS and penile resistance arteries (HPRA), since the agonist of these receptors, U46619, potently contracted HCCS (EC50 8.3±2.8 nM) and HPRA (EC50 6.2±2.2 nM), and the contractions produced by prostaglandin F2α at high concentrations (EC50 6460±3220 nM in HCCS and 8900±6700 nM in HPRA) were inhibited by the selective TP-receptor antagonist, SQ29548 (0.02 μM).EP-receptors are responsible for prostanoid-induced relaxant effects in HCCS because only prostaglandin E1 (PGE1), prostaglandin E2 and the EP2/EP4-receptor agonist, butaprost, produced consistent relaxation of this tissue (EC50 93.8±31.5, 16.3±3.8 and 1820±1284 nM, respectively). In HPRA, both prostacyclin and PGE1 (EC50 60.1±18.4 and 109.0±30.9 nM, respectively) as well as the selective IP receptor agonist, cicaprost, and butaprost (EC50 25.2±15.2 and 7050±6020 nM, respectively) caused relaxation, suggesting co-existence of IP- and EP-receptors (EP2 and/or EP4).In summary, endogenous production of prostanoids may regulate penile smooth muscle contractility by way of specific receptors. TP-receptors mediate contraction in HCCS and HPRA, while the relaxant effects of prostanoids are mediated by EP2- and/or EP4-receptors in HCCS and by EP- and IP-receptors in HPRA. PMID:11976264

  19. Leukotriene B4 mediates vascular smooth muscle cell migration through αvβ3 integrin transactivation.

    PubMed

    Moraes, João; Assreuy, Jamil; Canetti, Cláudio; Barja-Fidalgo, Christina

    2010-10-01

    Vascular injury leads to a local inflammatory response, characterized by endothelial damage, extracellular matrix exposition and aggregation/adhesion of platelets and circulating leukocytes. The release of inflammatory mediators amplifies the process, and can induce vascular smooth muscle cells (SMC) migration and proliferation. Released by leukocytes, leukotriene B4 (LTB4) induces reactive oxygen species production and SMC chemotaxis. This study was conducted to elucidate the molecular mechanisms involved in the effect of LTB4 on SMC migration, and a rat linage of vascular SMC (A7r5) were used throughout. The chemotactic effect of LTB4 was dependent on the concentration used, being comparable to AngII at 100 nM. Migration induced by LTB4 was inhibited in the presence of pertussis toxin, CP-105696, a BLT1 receptor antagonist, and by LY294002 or PD98059, two inhibitors of PI3K and MEK1/2, respectively. Stimulation of SMC with LTB4 triggered integrin-associated signaling pathways, inducing focal adhesion kinase (FAK) phosphorylation, mobilization of actin cytoskeleton, association of FAK to PI3K, ERK-2 phosphorylation and nuclear translocation, and also NFκB pathway activation. Pretreatment of SMC with a selective ligand of αvβ3 integrin, kistrin, inhibited LTB4-induced chemotaxis, FAK phosphorylation, FAK-PI3K association, and also inhibited ERK-2 and NFκB pathways activation. Taken together, the data demonstrated, for the first time, that the effect of LTB4 on SMC migration is modulated by integrin signaling activation, suggesting that these adhesion molecules might be important target for therapeutic intervention in cardiovascular diseases. Copyright © 2010 Elsevier Ireland Ltd. All rights reserved.

  20. Notch2 and Notch3 Function Together to Regulate Vascular Smooth Muscle Development

    PubMed Central

    Wang, Qingqing; Zhao, Ning; Kennard, Simone; Lilly, Brenda

    2012-01-01

    Notch signaling has been implicated in the regulation of smooth muscle differentiation, but the precise role of Notch receptors is ill defined. Although Notch3 receptor expression is high in smooth muscle, Notch3 mutant mice are viable and display only mild defects in vascular patterning and smooth muscle differentiation. Notch2 is also expressed in smooth muscle and Notch2 mutant mice show cardiovascular abnormalities indicative of smooth muscle defects. Together, these findings infer that Notch2 and Notch3 act together to govern vascular development and smooth muscle differentiation. To address this hypothesis, we characterized the phenotype of mice with a combined deficiency in Notch2 and Notch3. Our results show that when Notch2 and Notch3 genes are simultaneously disrupted, mice die in utero at mid-gestation due to severe vascular abnormalities. Assembly of the vascular network occurs normally as assessed by Pecam1 expression, however smooth muscle cells surrounding the vessels are grossly deficient leading to vascular collapse. In vitro analysis show that both Notch2 and Notch3 robustly activate smooth muscle differentiation genes, and Notch3, but not Notch2 is a target of Notch signaling. These data highlight the combined actions of the Notch receptors in the regulation of vascular development, and suggest that while these receptors exhibit compensatory roles in smooth muscle, their functions are not entirely overlapping. PMID:22615991

  1. 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.

  2. Hypoxic contraction of cultured pulmonary vascular smooth muscle cells

    SciTech Connect

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

    1990-11-01

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

  3. Aging impairs Ca2+ sensitization pathways in gallbladder smooth muscle.

    PubMed

    Macias, Beatriz; Gomez-Pinilla, Pedro J; Camello-Almaraz, Cristina; Pascua, Patricia; Tresguerres, Jesus Af; Camello, Pedro J; Pozo, Maria J

    2012-08-01

    Calcium sensitization is an important physiological process in agonist-induced contraction of smooth muscle. In brief, calcium sensitization is a pathway that leads to smooth muscle contraction independently of changes in [Ca(2+)](i) by mean of inhibition of myosin light chain phosphatase. Aging has negative impacts on gallbladder contractile response due to partial impairment in calcium signaling and alterations in the contractile machinery. However, information regarding aging-induced alterations in calcium sensitization is scanty. We hypothesized that the calcium sensitization system is negatively affected by age. To investigate this, gallbladders were collected from adult (4 months old) and aged (22-24 months old) guinea pigs. To evaluate the contribution of calcium sensitization pathways we assayed the effect of the specific inhibitors Y-27632 and GF109203X on the "in vitro" isometric gallbladder contractions induced by agonist challenges. In addition, expression and phosphorylation (as activation index) of proteins participating in the calcium sensitization pathways were quantified by Western blotting. Aging reduced bethanechol- and cholecystokinin-evoked contractions, an effect associated with a reduction in MLC20 phosphorylation and in the effects of both Y-27632 and GF109203X. In addition, there was a drop in ROCK I, ROCK II, MYPT-1 and PKC expression and in the activation/phosphorylation of MYPT-1, PKC and CPI-17 in response to agonists. Interestingly, melatonin treatment for 4 weeks restored gallbladder contractile responses due to re-establishment of calcium sensitization pathways. These results demonstrate that age-related gallbladder hypocontractility is associated to alterations of calcium sensitization pathways and that melatonin treatment exerts beneficial effects in the recovery of gallbladder contractility.

  4. The structure of Mytilus smooth muscle and the electrical constants of the resting muscle.

    PubMed

    Twarog, B M; Dewey, M M; Hidaka, T

    1973-02-01

    The individual muscle fibers of the anterior byssus retractor muscle (ABRM) of Mytilus edulis L. are uninucleate, 1.2-1.8 mm in length, 5 microm in diameter, and organized into bundles 100-200 microm in diameter, surrounded by connective tissue. Some bundles run the length of the whole muscle. Adjacent muscle cell membranes are interconnected by nexuses at frequent intervals. Specialized attachments exist between muscle fibers and connective tissue. Electrical constants of the resting muscle membrane were measured with intracellular recording electrodes and both extracellular and intracellular current-passing electrodes. With an intracellular current-passing electrode, the time constant tau, was 4.3 +/- 1.5 ms. With current delivered via an extracellular electrode tau was 68.3 +/- 15 ms. The space constant, lambda, was 1.8 mm +/- 0.4. The membrane input resistance, R(eff), ranged from 23 to 51 MOmega. The observations that values of tau depend on the method of passing current, and that the value of lambda is large relative to fiber length and diameter are considered evidence that the individual muscle fibers are electrically interconnected within bundles in a three-dimensional network. Estimations are made of the membrane resistance, R(m), to compare the values to fast and slow striated muscle fibers and mammalian smooth muscles. The implications of this study in reinterpreting previous mechanical and electrical studies are discussed.

  5. The Structure of Mytilus Smooth Muscle and the Electrical Constants of the Resting Muscle

    PubMed Central

    Twarog, Betty M.; Dewey, Maynard M.; Hidaka, Tohoru

    1973-01-01

    The individual muscle fibers of the anterior byssus retractor muscle (ABRM) of Mytilus edulis L. are uninucleate, 1.2–1.8 mm in length, 5 µm in diameter, and organized into bundles 100–200 µm in diameter, surrounded by connective tissue. Some bundles run the length of the whole muscle. Adjacent muscle cell membranes are interconnected by nexuses at frequent intervals. Specialized attachments exist between muscle fibers and connective tissue. Electrical constants of the resting muscle membrane were measured with intracellular recording electrodes and both extracellular and intracellular current-passing electrodes. With an intracellular current-passing electrode, the time constant τ, was 4.3 ± 1.5 ms. With current delivered via an extracellular electrode τ was 68.3 ± 15 ms. The space constant, λ, was 1.8 mm ± 0.4. The membrane input resistance, R eff, ranged from 23 to 51 MΩ. The observations that values of τ depend on the method of passing current, and that the value of λ is large relative to fiber length and diameter are considered evidence that the individual muscle fibers are electrically interconnected within bundles in a three-dimensional network. Estimations are made of the membrane resistance, Rm, to compare the values to fast and slow striated muscle fibers and mammalian smooth muscles. The implications of this study in reinterpreting previous mechanical and electrical studies are discussed. PMID:4688321

  6. Circular smooth muscle contributes to esophageal shortening during peristalsis.

    PubMed

    Vegesna, Anil K; Chuang, Keng-Yu; Besetty, Ramashesai; Phillips, Steven J; Braverman, Alan S; Barbe, Mary F; Ruggieri, Michael R; Miller, Larry S

    2012-08-28

    To study the angle between the circular smooth muscle (CSM) and longitudinal smooth muscle (LSM) fibers in the distal esophagus. In order to identify possible mechanisms for greater shortening in the distal compared to proximal esophagus during peristalsis, the angles between the LSM and CSM layers were measured in 9 cadavers. The outer longitudinal layer of the muscularis propria was exposed after stripping the outer serosa. The inner circular layer of the muscularis propria was then revealed after dissection of the esophageal mucosa and the underlying muscularis mucosa. Photographs of each specimen were taken with half of the open esophagus folded back showing both the outer longitudinal and inner circular muscle layers. Angles were measured every one cm for 10 cm proximal to the squamocolumnar junction (SCJ) by two independent investigators. Two human esophagi were obtained from organ transplant donors and the angles between the circular and longitudinal smooth muscle layers were measured using micro-computed tomography (micro CT) and Image J software. All data are presented as mean ± SE. The CSM to LSM angle at the SCJ and 1 cm proximal to SCJ on the autopsy specimens was 69.3 ± 4.62 degrees vs 74.9 ± 3.09 degrees, P = 0.32. The CSM to LSM angle at SCJ were statistically significantly lower than at 2, 3, 4 and 5 cm proximal to the SCJ, 69.3 ± 4.62 degrees vs 82.58 ± 1.34 degrees, 84.04 ± 1.64 degrees, 84.87 ± 1.04 degrees and 83.72 ± 1.42 degrees, P = 0.013, P = 0.008, P = 0.004, P = 0.009 respectively. The CSM to LSM angle at SCJ was also statistically significantly lower than the angles at 6, 7 and 8 cm proximal to the SCJ, 69.3 ± 4.62 degrees vs 80.18 ± 2.09 degrees, 81.81 ± 1.75 degrees and 80.96 ± 2.04 degrees, P = 0.05, P = 0.02, P = 0.03 respectively. The CSM to LSM angle at 1 cm proximal to SCJ was statistically significantly lower than at 3, 4 and 5 cm proximal to the SCJ, 74.94 ± 3.09 degrees vs 84.04 ± 1.64 degrees, 84.87 ± 1

  7. Circular smooth muscle contributes to esophageal shortening during peristalsis

    PubMed Central

    Vegesna, Anil K; Chuang, Keng-Yu; Besetty, Ramashesai; Phillips, Steven J; Braverman, Alan S; Barbe, Mary F; Ruggieri, Michael R; Miller, Larry S

    2012-01-01

    AIM: To study the angle between the circular smooth muscle (CSM) and longitudinal smooth muscle (LSM) fibers in the distal esophagus. METHODS: In order to identify possible mechanisms for greater shortening in the distal compared to proximal esophagus during peristalsis, the angles between the LSM and CSM layers were measured in 9 cadavers. The outer longitudinal layer of the muscularis propria was exposed after stripping the outer serosa. The inner circular layer of the muscularis propria was then revealed after dissection of the esophageal mucosa and the underlying muscularis mucosa. Photographs of each specimen were taken with half of the open esophagus folded back showing both the outer longitudinal and inner circular muscle layers. Angles were measured every one cm for 10 cm proximal to the squamocolumnar junction (SCJ) by two independent investigators. Two human esophagi were obtained from organ transplant donors and the angles between the circular and longitudinal smooth muscle layers were measured using micro-computed tomography (micro CT) and Image J software. RESULTS: All data are presented as mean ± SE. The CSM to LSM angle at the SCJ and 1 cm proximal to SCJ on the autopsy specimens was 69.3 ± 4.62 degrees vs 74.9 ± 3.09 degrees, P = 0.32. The CSM to LSM angle at SCJ were statistically significantly lower than at 2, 3, 4 and 5 cm proximal to the SCJ, 69.3 ± 4.62 degrees vs 82.58 ± 1.34 degrees, 84.04 ± 1.64 degrees, 84.87 ± 1.04 degrees and 83.72 ± 1.42 degrees, P = 0.013, P = 0.008, P = 0.004, P = 0.009 respectively. The CSM to LSM angle at SCJ was also statistically significantly lower than the angles at 6, 7 and 8 cm proximal to the SCJ, 69.3 ± 4.62 degrees vs 80.18 ± 2.09 degrees, 81.81 ± 1.75 degrees and 80.96 ± 2.04 degrees, P = 0.05, P = 0.02, P = 0.03 respectively. The CSM to LSM angle at 1 cm proximal to SCJ was statistically significantly lower than at 3, 4 and 5 cm proximal to the SCJ, 74.94 ± 3.09 degrees vs 84.04 ± 1

  8. Modeling the impairment of airway smooth muscle force by stretch

    PubMed Central

    2015-01-01

    Imposed length changes of only a small percent produce transient reductions in active force in strips of airway smooth muscle (ASM) due to the temporary detachment of bound cross-bridges caused by the relative motion of the actin and myosin fibers. More dramatic and sustained reductions in active force occur following large changes in length. The Huxley two-state model of skeletal muscle originally proposed in 1957 and later adapted to include a four-state description of cross-bridge kinetics has been widely used to model the former phenomenon, but is unable to account for the latter unless modified to include mechanisms by which the contractile machinery in the ASM cell becomes appropriately rearranged. Even so, the Huxley model itself is based on the assumption that the contractile proteins are all aligned precisely in the direction of bulk force generation, which is not true for ASM. The present study derives a coarse-grained version of the Huxley model that is free of inherent assumptions about cross-bridge orientation. This simplified model recapitulates the key features observed in the force-length behavior of activated strips of ASM and, in addition, provides a mechanistically based way of accounting for the sustained force reductions that occur following large stretch. PMID:25571992

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

    PubMed

    Wang, Fang; Zachar, Vladimir; Pennisi, Cristian Pablo; Fink, Trine; Maeda, Yasuko; Emmersen, Jeppe

    2018-02-08

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

  10. The persistence of active smooth muscle in the female rat cervix through pregnancy.

    PubMed

    Ferland, David J; Darios, Emma S; Watts, Stephanie W

    2015-02-01

    A controversy exists as to whether functional smooth muscle exists in the cervix before and during pregnancy, potentially continuous with the uterus. We hypothesized that cervical smooth muscle persists through pregnancy and is functional. Uteri and cervices were taken from female virgin, 11 day, and 20 day (near labor) pregnant rats. All experiments used the uterus as a positive control. Three different smooth muscle proteins (smooth muscle α-actin, SM-22α, and calponin-1) allowed immunohistochemical detection of the continuous nature of the smooth muscle from the vagina, cervix, and uterus. Tissues were also hung in isolated tissue baths for the measurement of isometric smooth muscle contraction. Uterine and cervical homogenates were also used in Western analyses to measure protein expression. Immunohistochemistry revealed there to be smooth muscle as validated by an expression of all 3 markers in the cervix. This smooth muscle was continuous with that of the vagina and uterus. Smooth muscle α-actin was detected in virgin tissue (291.3 ± 32.2 arbitrary densitometry units/β-actin), day 11 (416.8 ± 19.5), and day 20 pregnant tissue (293.0 ± 34.4). The virgin, day 11, and day 20 cervices contracted 2.18 ± 0.24 g, 1.46 ± 0.08 g, and 3.88 ± 0.49 g (respectively) to depolarizing KCl. Cervices contracted at day 20 to the cholinergic muscarinic agonist carbamylcholine (maximum, 133% ± 18.2% KCl contraction, n = 4). These findings strongly support that smooth muscle is present in the cervix through pregnancy and continuous with the uterus. Copyright © 2015 Elsevier Inc. All rights reserved.

  11. Functional effects of KCNQ K+ channels in airway smooth muscle

    PubMed Central

    Evseev, Alexey I.; Semenov, Iurii; Archer, Crystal R.; Medina, Jorge L.; Dube, Peter H.; Shapiro, Mark S.; Brenner, Robert

    2013-01-01

    KCNQ (Kv7) channels underlie a voltage-gated K+ current best known for control of neuronal excitability, and its inhibition by Gq/11-coupled, muscarinic signaling. Studies have indicated expression of KCNQ channels in airway smooth muscle (ASM), a tissue that is predominantly regulated by muscarinic receptor signaling. Therefore, we investigated the function of KCNQ channels in rodent ASM and their interplay with Gq/11-coupled M3 muscarinic receptors. Perforated-patch clamp of dissociated ASM cells detected a K+ current inhibited by the KCNQ antagonist, XE991, and augmented by the specific agonist, flupirtine. KCNQ channels begin to activate at voltages near resting potentials for ASM cells, and indeed XE991 depolarized resting membrane potentials. Muscarinic receptor activation inhibited KCNQ current weakly (~20%) at concentrations half-maximal for contractions. Thus, we were surprised to see that KCNQ had no affect on membrane voltage or muscle contractility following muscarinic activation. Further, M3 receptor-specific antagonist J104129 fumarate alone did not reveal KCNQ effects on muscarinic evoked depolarization or contractility. However, a role for KCNQ channels was revealed when BK-K+ channel activities are reduced. While KCNQ channels do control resting potentials, they appear to play a redundant role with BK calcium-activated K+ channels during ASM muscarinic signaling. In contrast to effect of antagonist, we observe that KCNQ agonist flupirtine caused a significant hyperpolarization and reduced contraction in vitro irrespective of muscarinic activation. Using non-invasive whole animal plethysmography, the clinically approved KCNQ agonist retigabine caused a transient reduction in indexes of airway resistance in both wild type and BK β1 knockout (KO) mice treated with the muscarinic agonist. These findings indicate that KCNQ channels can be recruited via agonists to oppose muscarinic evoked contractions and may be of therapeutic value as bronchodilators

  12. Cardiac, skeletal, and smooth muscle mitochondrial respiration: are all mitochondria created equal?

    PubMed Central

    Park, Song-Young; Gifford, Jayson R.; Andtbacka, Robert H. I.; Trinity, Joel D.; Hyngstrom, John R.; Garten, Ryan S.; Diakos, Nikolaos A.; Ives, Stephen J.; Dela, Flemming; Larsen, Steen; Drakos, Stavros

    2014-01-01

    Unlike cardiac and skeletal muscle, little is known about vascular smooth muscle mitochondrial respiration. Therefore, the present study examined mitochondrial respiratory rates in smooth muscle of healthy human feed arteries and compared with that of healthy cardiac and skeletal muscles. Cardiac, skeletal, and smooth muscles were harvested from a total of 22 subjects (53 ± 6 yr), and mitochondrial respiration was assessed in permeabilized fibers. Complex I + II, state 3 respiration, an index of oxidative phosphorylation capacity, fell progressively from cardiac to skeletal to smooth muscles (54 ± 1, 39 ± 4, and 15 ± 1 pmol·s−1·mg−1, P < 0.05, respectively). Citrate synthase (CS) activity, an index of mitochondrial density, also fell progressively from cardiac to skeletal to smooth muscles (222 ± 13, 115 ± 2, and 48 ± 2 μmol·g−1·min−1, P < 0.05, respectively). Thus, when respiration rates were normalized by CS (respiration per mitochondrial content), oxidative phosphorylation capacity was no longer different between the three muscle types. Interestingly, complex I state 2 normalized for CS activity, an index of nonphosphorylating respiration per mitochondrial content, increased progressively from cardiac to skeletal to smooth muscles, such that the respiratory control ratio, state 3/state 2 respiration, fell progressively from cardiac to skeletal to smooth muscles (5.3 ± 0.7, 3.2 ± 0.4, and 1.6 ± 0.3 pmol·s−1·mg−1, P < 0.05, respectively). Thus, although oxidative phosphorylation capacity per mitochondrial content in cardiac, skeletal, and smooth muscles suggest all mitochondria are created equal, the contrasting respiratory control ratio and nonphosphorylating respiration highlight the existence of intrinsic functional differences between these muscle mitochondria. This likely influences the efficiency of oxidative phosphorylation and could potentially alter ROS production. PMID:24906913

  13. Cardiac, skeletal, and smooth muscle mitochondrial respiration: are all mitochondria created equal?

    PubMed

    Park, Song-Young; Gifford, Jayson R; Andtbacka, Robert H I; Trinity, Joel D; Hyngstrom, John R; Garten, Ryan S; Diakos, Nikolaos A; Ives, Stephen J; Dela, Flemming; Larsen, Steen; Drakos, Stavros; Richardson, Russell S

    2014-08-01

    Unlike cardiac and skeletal muscle, little is known about vascular smooth muscle mitochondrial respiration. Therefore, the present study examined mitochondrial respiratory rates in smooth muscle of healthy human feed arteries and compared with that of healthy cardiac and skeletal muscles. Cardiac, skeletal, and smooth muscles were harvested from a total of 22 subjects (53 ± 6 yr), and mitochondrial respiration was assessed in permeabilized fibers. Complex I + II, state 3 respiration, an index of oxidative phosphorylation capacity, fell progressively from cardiac to skeletal to smooth muscles (54 ± 1, 39 ± 4, and 15 ± 1 pmol·s(-1)·mg(-1), P < 0.05, respectively). Citrate synthase (CS) activity, an index of mitochondrial density, also fell progressively from cardiac to skeletal to smooth muscles (222 ± 13, 115 ± 2, and 48 ± 2 μmol·g(-1)·min(-1), P < 0.05, respectively). Thus, when respiration rates were normalized by CS (respiration per mitochondrial content), oxidative phosphorylation capacity was no longer different between the three muscle types. Interestingly, complex I state 2 normalized for CS activity, an index of nonphosphorylating respiration per mitochondrial content, increased progressively from cardiac to skeletal to smooth muscles, such that the respiratory control ratio, state 3/state 2 respiration, fell progressively from cardiac to skeletal to smooth muscles (5.3 ± 0.7, 3.2 ± 0.4, and 1.6 ± 0.3 pmol·s(-1)·mg(-1), P < 0.05, respectively). Thus, although oxidative phosphorylation capacity per mitochondrial content in cardiac, skeletal, and smooth muscles suggest all mitochondria are created equal, the contrasting respiratory control ratio and nonphosphorylating respiration highlight the existence of intrinsic functional differences between these muscle mitochondria. This likely influences the efficiency of oxidative phosphorylation and could potentially alter ROS production.

  14. Gene expression in asthmatic airway smooth muscle: a mixed bag.

    PubMed

    Pascoe, Christopher D; Swyngedouw, Nicholas E; Seow, Chun Y; Paré, Peter D

    2015-02-01

    It has long been known that airway smooth muscle (ASM) contraction contributes significantly to the reversible airflow obstruction that defines asthma. It has also been postulated that phenotypic changes in ASM contribute to the airway hyper-responsiveness (AHR) that is a characteristic feature of asthma. Although there is agreement that the mass of ASM surrounding the airways is significantly increased in asthmatic compared with non-asthmatic airways, it is still uncertain whether there are quantitative or qualitative changes in the level of expression of the genes and proteins involved in the canonical contractile pathway in ASM that could account for AHR. This review will summarize past attempts at quantifying gene expression changes in the ASM of asthmatic lungs as well as non-asthmatic ASM cells stimulated with various inflammatory cytokines. The lack of consistent findings in asthmatic samples coupled with the relative concordance of results from stimulated ASM cells suggests that changes to the contractility of ASM tissues in asthma may be dependent on the presence of an inflammatory environment surrounding the ASM layer. Removal of the ASM from this environment could explain why hypercontractility is rarely seen ex vivo.

  15. SREBP inhibits VEGF expression in human smooth muscle cells

    SciTech Connect

    Motoyama, Koka; Fukumoto, Shinya; Koyama, Hidenori

    2006-03-31

    Sterol regulatory element-binding proteins (SREBPs) are transcription factors that regulate expression of genes encoding enzymes for lipid biosynthesis. SREBPs are activated by HMG-CoA reductase inhibitors (statins). Statins have been also reported to suppress vascular endothelial growth factor (VEGF) expression in vascular smooth muscle cells (VSMCs). Therefore, we hypothesized that SREBPs are involved in statin-mediated regulation of VEGF production in VSMCs. SREBP1 was robustly expressed, and was activated by atorvastatin in VSMCs, as demonstrated by increased levels of the mature nuclear form of SREBP1, and increased promoter activities of a reporter containing sterol regulatory elements by atorvastatin. Moreover, overexpression of SREBP1amore » dose-dependently suppressed VEGF promoter activity. Site-specific mutation or deletion of the proximal Sp1 sites reduced the inhibitory effects of SREBP1a on VEGF promoter activity. These data demonstrated that SREBP1, activated by atorvastatin, suppressed VEGF expression through the indirect interaction with the proximal tandem Sp1 sites in VSMCs.« less

  16. Impaired arterial smooth muscle cell vasodilatory function in methamphetamine users.

    PubMed

    Nabaei, Ghaemeh; Oveisgharan, Shahram; Ghorbani, Askar; Fatehi, Farzad

    2016-11-15

    Methamphetamine use is a strong risk factor for stroke. This study was designed to evaluate arterial function and structure in methamphetamine users ultrasonographically. In a cross-sectional study, 20 methamphetamine users and 21 controls, aged between 20 and 40years, were enrolled. Common carotid artery intima-media thickness (CCA-IMT) marker of early atherogenesis, flow-mediated dilatation (FMD) determinants of endothelium-dependent vasodilation, and nitroglycerine-mediated dilatation (NMD) independent marker of vasodilation were measured in two groups. There were no significant differences between the two groups regarding demographic and metabolic characteristics. The mean (±SD) CCA-IMT in methamphetamine users was 0.58±0.09mm, versus 0.59±0.07mm in the controls (p=0.84). Likewise, FMD% was not significantly different between the two groups [7.6±6.1% in methamphetamine users vs. 8.2±5.1% in the controls; p=0.72], nor were peak flow and shear rate after hyperemia. However, NMD% was considerably decreased in the methamphetamine users [8.5±7.8% in methamphetamine users vs. 13.4±6.2% in controls; p=0.03]. According to our results, NMD is reduced among otherwise healthy methamphetamine users, which represents smooth muscle dysfunction in this group. This may contribute to the high risk of stroke among methamphetamine users. Copyright © 2016 Elsevier B.V. All rights reserved.

  17. Does atorvastatin induce aortic smooth muscle cell apoptosis in vivo?

    PubMed

    Doyon, Marielle; Hale, Taben Mary; Huot-Marchand, Julie-Emilie; Wu, Rong; de Champlain, Jacques; DeBlois, Denis

    2011-01-01

    It has been reported that HMG-CoA reductase inhibitors such as atorvastatin induce vascular smooth muscle cell (SMC) apoptosis in vitro. However, this effect remains to be demonstrated in vivo. The present studies were designed to test the ability of atorvastatin to induce SMC apoptosis in vivo, using the spontaneously hypertensive rat (SHR) as a well-known reference model of SMC apoptosis induction in vivo by cardiovascular drugs including the calcium channel blocker amlodipine. Atorvastatin was administered to SHR for 3 or 6 weeks either alone or together with amlodipine, a drug combination clinically available to patients. Primary endpoints included aortic medial hypertrophy and aortic SMC hyperplasia, internucleosomal DNA fragmentation and expression of the apoptosis regulatory proteins Bax and Bcl-2. The SHR aorta showed no evidence of SMC apoptosis induction by atorvastatin, even at the high dose of 50 mg kg(-1) day(-1), although the statin significantly reduced oxidative stress after 3 weeks and blood pressure after 6 weeks of administration. Amlodipine-induced regression of aortic hypertophy and aortic SMC hyperplasia were dose- and time-dependent, but there was no interaction between atorvastatin and amlodipine in modulating the primary endpoints. These results do not support the notion that atorvastatin induces SMC apoptosis in the aortic media in vivo. Copyright © 2010. Published by Elsevier Inc.

  18. [Targeting of myosin light chain kinase in smooth muscle cell].

    PubMed

    Kohama, K

    1999-10-01

    We constructed a plasmid vector to have a 1.4 kb insert of myosin light chain kinase (MLCK) cDNA in an antisense direction to express antisense mRNA. The construct was then transfected to SM3, a cell line from vascular smooth muscle cells (VSMCs), producing a few stable transfectants. The down-regulation of MLCK expression in the transfectants was confirmed by both Northern and Western blots. The control SM3 showed chemotaf1p4++ motility to the platelet derived growth factor (PDGF), which was supported by the membrane ruffling. However, the transfectants showed neither chemotaxic motility nor developed membrane ruffling, indicating the essential role of MLCK in the motility. The specificity for the targeting was assessed by demonstrating that Rho-kinase activity, which also phosphorylates the myosin light chain (MLC), was well preserved in both SM3 and the transfectants. In spite of this importance of MLCK, PDGF failed to induce MLC phosphorylation in not only the transfectants but also in SM3. The mode in which MLCK was involved in the development of membrane ruffling is discussed with special reference to myosin-binding property of MLCK (Ye et al. Proc Natl. Acad. Sci. USA 96 6666-6671, 1999).

  19. Ageing induced vascular smooth muscle cell senescence in atherosclerosis

    PubMed Central

    Uryga, Anna K.

    2015-01-01

    Abstract 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. PMID:26174609

  20. The importance of the smooth muscle cytoskeleton to preterm labour.

    PubMed

    Morgan, Kathleen G

    2014-03-01

    Multiple mechanisms have been shown to regulate the onset of labour in a co-operative and complex manner. One factor, myometrial stretch and associated increases in wall tension, has been implicated clinically in the initiation of labour and especially the aetiology of preterm labour. Recent work on the mechanisms involved has led to the finding that the intracellular Ca(2+) requirement for activation of the myometrial contractile filaments increases during gestation. The decreased Ca(2+) sensitivity correlates with an increase in the expression of caldesmon, an actin-binding protein and inhibitor of myosin activation, during pregnancy. In late pregnancy, an increase in extracellular signal-regulated kinase-mediated caldesmon phosphorylation occurs, which appears to reverse the inhibitory action of caldesmon during labour. Force generated by the myometrial contractile filaments is communicated across the plasmalemma to the uterine wall through focal adhesions. Phospho-tyrosine screening and mass spectrometry of stretched myometrial samples identified several stretch-activated focal adhesion proteins. This Src-mediated focal adhesion signalling appears to provide a tunable, i.e. regulated, tension sensor and force transmitter in the myometrial cell. In other parallel studies, biophysical measurements of smooth muscle compliance at both the cellular and tissue levels suggest that decreases in cellular compliance due to changing interactions of the actin cytoskeleton with the focal adhesions may also promote increases in uterine wall tension. These results, taken together, suggest that focal adhesion proteins and their interaction with the cytoskeleton may present a new mode of regulation of uterine contractility.

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

    SciTech Connect

    Smajilovic, Sanela; Danish National Research Foundation Centre for Cardiac Arrhythmia; Hansen, Jakob Lerche

    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. Wemore » 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.« less

  2. Traction in smooth muscle cells varies with cell spreading

    NASA Technical Reports Server (NTRS)

    Tolic-Norrelykke, Iva Marija; Wang, Ning

    2005-01-01

    Changes in cell shape regulate cell growth, differentiation, and apoptosis. It has been suggested that the regulation of cell function by the cell shape is a result of the tension in the cytoskeleton and the distortion of the cell. Here we explore the association between cell-generated mechanical forces and the cell morphology. We hypothesized that the cell contractile force is associated with the degree of cell spreading, in particular with the cell length. We measured traction fields of single human airway smooth muscle cells plated on a polyacrylamide gel, in which fluorescent microbeads were embedded to serve as markers of gel deformation. The traction exerted by the cells at the cell-substrate interface was determined from the measured deformation of the gel. The traction was measured before and after treatment with the contractile agonist histamine, or the relaxing agonist isoproterenol. The relative increase in traction induced by histamine was negatively correlated with the baseline traction. On the contrary, the relative decrease in traction due to isoproterenol was independent of the baseline traction, but it was associated with cell shape: traction decreased more in elongated than in round cells. Maximum cell width, mean cell width, and projected area of the cell were the parameters most tightly coupled to both baseline and histamine-induced traction in this study. Wide and well-spread cells exerted larger traction than slim cells. These results suggest that cell contractility is controlled by cell spreading.

  3. Ultrasound induces contraction of the bladder smooth muscle.

    PubMed

    Ren, Yan; Zhu, Yi; Liu, Li; Yu, Tinghe; Dong, Xiaojing

    2016-08-01

    To investigate whether the treatment of overt postpartum urinary retention (PUR) with low-intensity pulsed ultrasound (LIPUS) was clinically effective and whether LIPUS could accelerate bladder smooth muscle (BSM) contraction by opening the L-type calcium channels and activating the Ca(2+) signaling pathway. Records of 136 patients undergoing PUR were retrospectively reviewed in two different groups for LIPUS and neostigmine between from 2014 to July 2015. The rats BSM strips in vitro were irradiated by LIPUS. The contraction frequency and amplitude were recorded with BL-410F biological experimental system. The BSM cells were constructed and identified by α-actin-specific antibody staining, and the intracellular Ca(2+) concentration was analyzed by flow cytometry. The clinical trial indicated that LIPUS had potential therapeutic effect on PUR (80.6 vs. 64.1 %, p < 0.05), and the BSM strip contractility was increased by LIPUS (p < 0.001), and the concentration of Ca(2+) was markedly enhanced by about twofold than that without LIPUS exposure (p < 0.01). Besides, nimodipine could suppress the contraction of BSM and the concentration of intracellular Ca(2+) which was caused by ultrasound. The results suggested LIPUS had potential therapeutic effect on PUR and the Ca(2+) signaling pathway was involved in the mechanism. The ultrasound irradiation may provide a new method for PUR therapy.

  4. Vascular smooth muscle cell phenotypic plasticity: focus on chromatin remodelling

    PubMed Central

    Spin, Joshua M.; Maegdefessel, Lars; Tsao, Philip S.

    2012-01-01

    Differentiated vascular smooth muscle cells (SMCs) retain the capacity to modify their phenotype in response to inflammation or injury. This phenotypic switching is a crucial component of vascular disease, and is partly dependent on epigenetic regulation. An appreciation has been building in the literature for the essential role chromatin remodelling plays both in SMC lineage determination and in influencing changes in SMC behaviour and state. This process includes numerous chromatin regulatory elements and pathways such as histone acetyltransferases, deacetylases, and methyltransferases and other factors that act at SMC-specific marker sites to silence or permit access to the cellular transcriptional machinery and on other key regulatory elements such as myocardin and Kruppel-like factor 4 (KLF4). Various stimuli known to alter the SMC phenotype, such as transforming growth factor beta (TGF-β), platelet-derived growth factor (PDGF), oxidized phospholipids, and retinoic acid, appear to act in part through effects upon SMC chromatin structure. In recent years, specific covalent histone modifications that appear to establish SMC determinacy have been identified, while others alter the differentiation state. In this article, we review the mechanisms of chromatin remodelling as it applies to the SMC phenotype. PMID:22362814

  5. Fragmented esophageal smooth muscle contraction segments on high resolution manometry: a marker of esophageal hypomotility.

    PubMed

    Porter, R F; Kumar, N; Drapekin, J E; Gyawali, C P

    2012-08-01

    Esophageal peristalsis consists of a chain of contracting striated and smooth muscle segments on high resolution manometry (HRM). We compared smooth muscle contraction segments in symptomatic subjects with reflux disease to healthy controls. High resolution manometry Clouse plots were analyzed in 110 subjects with reflux disease (50 ± 1.4 years, 51.5% women) and 15 controls (27 ± 2.1 years, 60.0% women). Using the 30 mmHg isobaric contour tool, sequences were designated fragmented if either smooth muscle contraction segment was absent or if the two smooth muscle segments were separated by a pressure trough, and failed if both smooth muscle contraction segments were absent. The discriminative value of contraction segment analysis was assessed. A total of 1115 swallows were analyzed (reflux group: 965, controls: 150). Reflux subjects had lower peak and averaged contraction amplitudes compared with controls (P < 0.0001 for all comparisons). Fragmented sequences followed 18.4% wet swallows in the reflux group, compared with 7.5% in controls (P < 0.0001), and were seen more frequently than failed sequences (7.9% and 2.5%, respectively). Using a threshold of 30% in individual subjects, a composite of failed and/or fragmented sequences was effective in segregating reflux subjects from control subjects (P = 0.04). Evaluation of smooth muscle contraction segments adds value to HRM analysis. Specifically, fragmented smooth muscle contraction segments may be a marker of esophageal hypomotility. © 2012 Blackwell Publishing Ltd.

  6. Characterization of smooth muscle-like cells in circulating human peripheral blood.

    PubMed

    Sugiyama, Seigo; Kugiyama, Kiyotaka; Nakamura, Shinichi; Kataoka, Keiichiro; Aikawa, Masanori; Shimizu, Koichi; Koide, Shunichi; Mitchell, Richard N; Ogawa, Hisao; Libby, Peter

    2006-08-01

    Smooth muscle cells play an important role in human vascular diseases. Several lines of evidence demonstrate that circulating smooth muscle precursor cells contribute to intimal hyperplasia in animal models. We obtained large spindle cells expressing alpha-smooth muscle actin (alpha-SMA), denoted here as "smooth muscle-like cells" (SMLC), from human peripheral blood mononuclear cells (PBMC). SMLC derived from human PBMC proliferated readily and expressed pro-inflammatory genes during early culture. After long-term culture, SMLC could contract and express characteristic smooth muscle cell markers. We found peripheral blood mononuclear cell expressing alpha-smooth muscle actin in the circulating blood that bore CD14 and CD105. Sorted CD14/CD105 double-positive PBMC could differentiate into SMLC. The number of CD14-CD105-bearing PBMC increased significantly in patients with coronary artery disease compared to patients without coronary artery disease. These results support the novel concept that smooth muscle precursor cells exist in circulating human blood and may contribute to the pathogenesis of vascular diseases.

  7. Matrix Metalloproteinase-1 Activation Contributes to Airway Smooth Muscle Growth and Asthma Severity

    PubMed Central

    Naveed, Shams-un-nisa; Clements, Debbie; Jackson, David J.; Philp, Christopher; Billington, Charlotte K.; Soomro, Irshad; Reynolds, Catherine; Harrison, Timothy W.; Johnston, Sebastian L.; Shaw, Dominick E.

    2017-01-01

    Rationale: Matrix metalloproteinase-1 (MMP-1) and mast cells are present in the airways of people with asthma. Objectives: To investigate whether MMP-1 could be activated by mast cells and increase asthma severity. Methods: Patients with stable asthma and healthy control subjects underwent spirometry, methacholine challenge, and bronchoscopy, and their airway smooth muscle cells were grown in culture. A second asthma group and control subjects had symptom scores, spirometry, and bronchoalveolar lavage before and after rhinovirus-induced asthma exacerbations. Extracellular matrix was prepared from decellularized airway smooth muscle cultures. MMP-1 protein and activity were assessed. Measurements and Main Results: Airway smooth muscle cells generated pro–MMP-1, which was proteolytically activated by mast cell tryptase. Airway smooth muscle treated with activated mast cell supernatants produced extracellular matrix, which enhanced subsequent airway smooth muscle growth by 1.5-fold (P < 0.05), which was dependent on MMP-1 activation. In asthma, airway pro–MMP-1 was 5.4-fold higher than control subjects (P = 0.002). Mast cell numbers were associated with airway smooth muscle proliferation and MMP-1 protein associated with bronchial hyperresponsiveness. During exacerbations, MMP-1 activity increased and was associated with fall in FEV1 and worsening asthma symptoms. Conclusions: MMP-1 is activated by mast cell tryptase resulting in a proproliferative extracellular matrix. In asthma, mast cells are associated with airway smooth muscle growth, MMP-1 levels are associated with bronchial hyperresponsiveness, and MMP-1 activation are associated with exacerbation severity. Our findings suggest that airway smooth muscle/mast cell interactions contribute to asthma severity by transiently increasing MMP activation, airway smooth muscle growth, and airway responsiveness. PMID:27967204

  8. Decreased trabecular smooth muscle and caveolin-1 expression in the penile tissue of aged rats.

    PubMed

    Bakircioglu, M E; Sievert, K D; Nunes, L; Lau, A; Lin, C S; Lue, T F

    2001-08-01

    Because decreased trabecular smooth muscle content is reportedly associated with vasculogenic impotence in men, we performed a rodent study to investigate the effect of aging on trabecular smooth muscle content and caveolin-1 protein expression in penile smooth muscle cells. In 6 young (age 3 months) and 6 old (age 24 months) rats erectile function was evaluated by cavernous nerve stimulation. At sacrifice penile tissue samples were collected for Western blot analysis, Masson's trichrome staining, caveolin-1 immunostaining and electron microscopy. The percent of smooth muscle in the trabecular tissue was assessed by computer assisted image analysis. In the aged rats mean intracavernous pressure plus or minus standard deviation was decreased (70 +/- 8.8 versus 107 +/- 12.3 cm. water) and the latency period was increased (7.8 +/- 1.2 versus 4.5 +/- 0.5 seconds) significantly compared to values in the young rats (p <0.001). The mean ratio of trabecular smooth muscle-to-connective tissue was also significantly altered in old versus young rats (27% +/- 2.9% versus 42.1% +/- 5.1%, p <0.001). Immunostaining for caveolin-1 was noted in each group in the sarcolemma of smooth muscle cells and endothelium of trabecular sinusoids but the staining pattern was less intense and the percent of smooth muscle positive for caveolin-1 was decreased in aged versus young rats (17.9% +/- 2.5% versus 27.5% +/- 3.6%, p <0.001). Moreover, young trabecular smooth muscle cells had more caveolae in the sarcolemma on electron microscopy and a higher expression of caveolin-1 protein on Western blot analysis. In contrast, higher endothelial nitric oxide synthase protein expression was noted in the penile tissue of old rats. In these aged rats the decreased ratio of trabecular smooth muscle-to-collagen and the reduced expression of caveolin-1 may contribute to erectile dysfunction.

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

    PubMed

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

    2010-11-01

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

  10. Epigenetics and miRNA emerge as key regulators of smooth muscle cell phenotype and function

    PubMed Central

    Clifford, Rachel L.; Singer, Cherie A.; John, Alison E.

    2014-01-01

    Regulation of phenotypic plasticity in smooth muscle requires an understanding of the mechanisms regulating phenotype-specific genes and the processes dysregulated during pathogenesis. Decades of study in airway smooth muscle has provided extensive knowledge of the gene expression patterns and signaling pathways necessary to maintain and alter smooth muscle cell phenotype. With this solid foundation, the importance and complexity of inheritable epigenetic modifications and mechanisms silencing gene expression have now emerged as fundamental components regulating aspects of inflammation, proliferation and remodeling. PMID:22800879

  11. Smooth muscle physiology and effect of bladder and urethra muscle length/tension on response to stimulation. Part I. Review.

    PubMed

    Bissada, N K; Finkbeiner, A E

    1980-09-01

    With particular reference to the lower urinary tract, a review of basic anatomy and physiology of smooth muscle is presented. The relationship as altered by electrica and pharmacologic stimulation is discussed.

  12. Smooth muscle antibodies and type 1 autoimmune hepatitis.

    PubMed

    Muratori, Paolo; Muratori, Luigi; Agostinelli, Daniela; Pappas, Georgios; Veronesi, Lorenza; Granito, Alessandro; Cassani, Fabio; Terlizzi, Paolo; Lenzi, Marco; Bianchi, Francesco B

    2002-12-01

    Smooth muscle antibodies (SMA) characterize type 1 autoimmune hepatitis. Our aim was to evaluate sensitivity and specificity of different immunofluorescence substrates for the detection of SMA. Sera from 55 patients with type 1 AIH 20 with primary biliary cirrhosis, 20 with HCV-related chronic hepatitis and 25 blood donors were studied for SMA and anti-microfilaments reactivity by immunofluorescence on rat tissue sections, cultured fibroblasts and commercially available HEp-2 cells (collectively revealing the so called anti-actin pattern), and for the XR1 system by counterimmunoelectrophoresis. SMA was classified on the basis of its immunofluorescence pattern (V--vessels, G--glomerular, T--tubular). As further control group, we studied 26 patients with a diagnosis other than AIH, selected on the basis of a SMA-non-T/XR1 positivity. In patients with AIH the SMA-T pattern on rodent tissue, and anti-MF on fibroblasts and on HEp-2 cells were present in 80, 82 and 80%, respectively. Five out of 11 SMA-non T positive AIH patients were anti-MF positive. None of the pathological and healthy controls was positive for SMA-T or anti-MF reactivity. XR1 system was present in 84% of AIH patients and in 5% of pathological controls (p = 0.01). Two out of 26 SMA-non-T/XR1 positive sera were positive for anti-MF by fibroblasts and HEp-2 cells. A significant correlation was found between SMA-T pattern and anti-MF reactivity; no correlation was found between XR1 system and SMA-T pattern or anti-MF reactivity. SMA-T pattern is highly sensitive and specific first diagnostic test for type 1 AIH; anti-MF can be used as additional tool for the diagnosis, particularly when, despite the absence of the SMA-T pattern, AIH is strongly suspected.

  13. Airways and vascular smooth muscles relaxant activities of Gaultheria trichophylla.

    PubMed

    Alam, Fiaz; Saqib, Qazi Najumus; Shah, Abdul Jabbar

    2017-01-01

    The aim of this experimental work was to explore the potential pharmacological activities of Gaultheria trichophylla Royle in hyperactive respiratory and vascular conditions. Gaultheria trichophylla was extracted with solvents, phytochemical detection tests were performed, and rabbit trachea and aorta strips were used to evaluate its effects on airways and vascular smooth muscles. Qualitative phytochemical tests showed the presence of flavonoids, alkaloids, anthraquinones, saponins, terpenoids, and condensed tannins. The methanol extract caused inhibition (EC 50 values of 3.12 mg/mL) of carbachol (1 μM) and partial relaxation of K + (80 mM) caused contractions in tracheal strips. The chloroform extract was comparatively more potent against carbachol than K+ induced contraction with EC 50 values of 0.64 and 2.26 mg/mL, respectively. However, the n-hexane extract showed more potency against K + than cabachol induced contractions, as in case with verapamil, with EC 50 values of 0.61 and 6.58 mg/mL, respectively. In isolated prepared trachea, the extracts displaced the carbachol concentration response curves and maximum response was suppressed. In rabbit aorta preparations, methanol and n-hexane extracts partially relaxed phenylephrine (1 μM) and K + induced vasoconstrictions. However, the chloroform extract inhibited phenylephrine induced contractions and exhibited a vasoconstrictor effect at lower concentrations and a relaxant effect at higher concentrations against K + precontractions. The data indicates that, in addition to others, the extracts of G .trichophylla possess verapamil like Ca ++ channel blocking components which explain the possible role of this plant in respiratory and vascular conditions.

  14. Morphological variability of smooth muscle cells in human nasal swell bodies.

    PubMed

    Grevers, G; Kamargakis, W; Welsch, U

    1996-01-01

    The complex functional behavior of nasal swell bodies is still not completely understood. In the present study the histology of the vessels involved in the swelling mechanism is examined and the ultrastructural appearances described of the different types of smooth muscle cells located in the vascular wall of swell bodies in the human inferior turbinate. Even though the majority of smooth muscle cells of the nasal swell bodies showed a normal, elongated appearance comparable to other smooth muscle cells elsewhere in the body, a variety of cells with atypical shapes could be detected that have not been described previously in vessels of the nasal mucosa. The diameters of the smooth muscle cells in general were strikingly variable. The individual smooth muscle cells were surrounded by a basal lamina that was occasionally disrupted or doubled. Myoblasts were separated by a connective tissue space containing collagen fibrils, mature elastin fibers and bundles of microfibrils. The latter two types of fibers and fibrils occurred mainly in the outer parts of the muscular coat. The endowment of cytoplasmic components was similar in all smooth muscle cells of the vascular wall in the swell bodies. These findings indicate that the specific feature of smooth musculature presumably resides in the unusual morphological variability of the single cells present, as well as in the striking heterogeneity of the arrangement of bundles of these cells in the vascular wall.

  15. The effect of caffeine on excitation-contraction coupling in skeletal and smooth muscle.

    PubMed

    Syson, A J; Huddart, H

    1976-06-01

    1. For cockroach skeletal muscle, 2 mM caffeine considerably lowered the mechanical threshold without affecting the membrane potential. Constractures were induced by 8-10 mM caffeine. 2. In rat ileal smooth muscle, 1-10 mM caffeine inhibited spontaneous contractile behaviour, abolished spike activity and reduced KCl-induced contracture tension. 3. Enhanced spike activity associated with the KCl-induced phasic contraction was abolished by caffeine, the degree of caffeine-induced relaxation being proportional to the concentration employed. These relaxations were not accompanied by membrane hyperpolarization. 4. The present results accord with previous work which has shown that caffeine increases myoplasmic free calcium in the skeletal muscle and lowers it in the smooth muscle. It is suggested that caffiene releases bound calcium in the former muscle and promotes binding in the latter. 5. It is further suggested that in the smooth muscle caffeine may reduce the membrane permeability to calcium.

  16. Not all vascular smooth muscle cell exosomes calcify equally in chronic kidney disease.

    PubMed

    Dusso, Adriana; Colombo, Maria Isabel; Shanahan, Catherine M

    2018-02-01

    Prevention of medial calcification in patients with chronic kidney disease requires the maintenance of vascular smooth muscle cell fitness. To preserve viability under chronic kidney disease-induced stress, vascular smooth muscle cells increase exosome formation and release, but the result is aggravated pathological calcification. Now Chen et al. report that microvesicles from calcifying vascular smooth muscle cells may propagate procalcifying signals to normal vascular smooth muscle cells. To help design effective strategies to impair procalcifying cell-to-cell communication, this commentary updates the current understanding of the main regulators of microvesicle/exosome biogenesis and secretion. Copyright © 2017 International Society of Nephrology. Published by Elsevier Inc. All rights reserved.

  17. The function of intimal longitudinal smooth muscles of the human coronary artery.

    PubMed

    Kawasaki, K; Iino, T; Hasegawa, H; Miyazawa, I; Hosoda, S

    1986-12-01

    Bundles of smooth muscles in the intimal layer of the human coronary artery contracted in a longitudinal direction on the application of vasoactive substances. The data indicate that the human coronary artery contracts not only transversely but also longitudinally.

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

    PubMed

    Tolg, Cornelia; Ahsan, Alya; Dworski, Shaalee; Kirwan, Tyler; Yu, Jeffery; Aitken, Karen; Bägli, Darius Jehan

    2013-01-01

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

  19. Genetic approaches to identify pathological limitations in aortic smooth muscle contraction.

    PubMed

    Huang, Jian; Gao, Ning; Wang, Shanzhi; Milewicz, Dianna M; Kamm, Kristine E; Stull, James T

    2018-01-01

    Aortic smooth muscle contains limiting amounts of myosin light chain kinase (MLCK) for myosin regulatory light chain (RLC) phosphorylation and contraction that predisposes to thoracic aortic disease in humans containing heterozygous loss-of-function mutations in MYLK. We tested the hypothesis that thoracic aortic smooth muscle contraction may also be susceptible to variations in the smooth muscle-specific isoform of the motor protein myosin where inactivation of one Myh11 allele or the presence of one Myh11 missense variant associated with an increased risk of human aortic disease may result in a reduced force development response. Additionally, other kinds of smooth muscles may be less sensitive to the effects of mutations in one smooth muscle myosin allele, similar to results obtained with Mylk. Force development responses were reduced in aortic tissue from a conditional knockout of smooth muscle myosin heavy chain in adult mice (Myh11+/- or Myh11-/-) with a greater reduction with homozygous vs heterozygous tissues. Similar reductions in force responses were obtained with tissues containing either a heterozygous or homozygous knockin mutation in smooth muscle myosin heavy chain (Myh11+/R247C or Myh11R247C/R247C mutations that cause human aortic disease) with no significant changes in RLC phosphorylation. Agonist-dependent force responses were not reduced significantly in urinary bladder, ileal, or tracheal tissues from Myh11+/- mice while only ileal tissue showed a reduced force response in Myh11R247C/R247C mice. Thus, heterozygous mutations in Myh11 associated with reduced myosin function result in compromised contractile function primarily in aortic smooth muscle.

  20. Voltage dependent potassium channel remodeling in murine intestinal smooth muscle hypertrophy induced by partial obstruction.

    PubMed

    Liu, Dong-Hai; Huang, Xu; Guo, Xin; Meng, Xiang-Min; Wu, Yi-Song; Lu, Hong-Li; Zhang, Chun-Mei; Kim, Young-chul; Xu, Wen-Xie

    2014-01-01

    Partial obstruction of the small intestine causes obvious hypertrophy of smooth muscle cells and motility disorder in the bowel proximate to the obstruction. To identify electric remodeling of hypertrophic smooth muscles in partially obstructed murine small intestine, the patch-clamp and intracellular microelectrode recording methods were used to identify the possible electric remodeling and Western blot, immunofluorescence and immunoprecipitation were utilized to examine the channel protein expression and phosphorylation level changes in this research. After 14 days of obstruction, partial obstruction caused obvious smooth muscle hypertrophy in the proximally located intestine. The slow waves of intestinal smooth muscles in the dilated region were significantly suppressed, their amplitude and frequency were reduced, whilst the resting membrane potentials were depolarized compared with normal and sham animals. The current density of voltage dependent potassium channel (KV) was significantly decreased in the hypertrophic smooth muscle cells and the voltage sensitivity of KV activation was altered. The sensitivity of KV currents (IKV) to TEA, a nonselective potassium channel blocker, increased significantly, but the sensitivity of IKv to 4-AP, a KV blocker, stays the same. The protein levels of KV4.3 and KV2.2 were up-regulated in the hypertrophic smooth muscle cell membrane. The serine and threonine phosphorylation levels of KV4.3 and KV2.2 were significantly increased in the hypertrophic smooth muscle cells. Thus this study represents the first identification of KV channel remodeling in murine small intestinal smooth muscle hypertrophy induced by partial obstruction. The enhanced phosphorylations of KV4.3 and KV2.2 may be involved in this process.

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

    PubMed

    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

    2015-09-16

    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. 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. 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. © 2015 The Authors. Published on behalf of the American Heart Association, Inc., by Wiley Blackwell.

  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. CD98 regulates vascular smooth muscle cell proliferation in atherosclerosis.

    PubMed

    Baumer, Yvonne; McCurdy, Sara; Alcala, Martin; Mehta, Nehal; Lee, Bog-Hieu; Ginsberg, Mark H; Boisvert, William A

    2017-01-01

    Vascular smooth muscle cells (VSMC) migrate and proliferate to form a stabilizing fibrous cap that encapsulates atherosclerotic plaques. CD98 is a transmembrane protein made of two subunits, CD98 heavy chain (CD98hc) and one of six light chains, and is known to be involved in cell proliferation and survival. Because the influence of CD98hc on atherosclerosis development is unknown, our aim was to determine if CD98hc expressed on VSMC plays a role in shaping the morphology of atherosclerotic plaques by regulating VSMC function. In addition to determining the role of CD98hc in VSMC proliferation and apoptosis, we utilized mice with SMC-specific deletion of CD98hc (CD98hc fl/fl SM22αCre + ) to determine the effects of CD98hc deficiency on VSMC function in atherosclerotic plaque. After culturing for 5 days in vitro, CD98hc -/- VSMC displayed dramatically reduced cell counts, reduced proliferation, as well as reduced migration compared to control VSMC. Analysis of aortic VSCM after 8 weeks of HFD showed a reduction in CD98hc -/- VSMC proliferation as well as increased apoptosis compared to controls. A long-term atherosclerosis study using SMC-CD98hc -/- /ldlr -/- mice was performed. Although total plaque area was unchanged, CD98hc -/- mice showed reduced presence of VSMC within the plaque (2.1 ± 0.4% vs. 4.3 ± 0.4% SM22α-positive area per plaque area, p < 0.05), decreased collagen content, as well as increased necrotic core area (25.8 ± 1.9% vs. 10.9 ± 1.6%, p < 0.05) compared to control ldlr -/- mice. We conclude that CD98hc is required for VSMC proliferation, and that its deficiency leads to significantly reduced presence of VSMC in the neointima. Thus, CD98hc expression in VSMC contributes to the formation of plaques that are morphologically more stable, and thereby protects against atherothrombosis. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

  4. TRPC3 overexpression and intervention in airway smooth muscle of ovalbumin-induced hyperresponsiveness and remodeling.

    PubMed

    Xu, Bo-Ming; Zhang, Jia-Hua; Wang, Jia-Ling; Xiao, Jun-Hua

    2018-04-06

    Transient receptor potential canonical channel 3 (TRPC3) proteins function as non-voltage-gated Ca 2+ -permeable channels and play divergent roles in many processes of pathophysiology. The purpose of this study was to determine the relationship between TRPC3 expression and airway hyperresponsiveness and remodeling in ovalbumin-induced asthmatic Kunming mice. Mice were sensitized and challenged by ovalbumin to establish asthmatic model. Hematoxylin-eosin staining, hydroxyproline assay, and isometric tracheal ring force measurement were used to evaluate airway remodeling and hyperresponsiveness in asthmatic mice. Western blot was performed to detect the expression of TRPC3 proteins. MTT assay was used to measure the proliferation of airway smooth muscle cells. TRPC3 protein expression increased in airway smooth muscle of asthmatic mice. GdCl 3 , a nonspecific TRPC blocker, attenuated the contractile force of airway smooth muscle. Fetal bovine serum stimulated airway smooth muscle cells proliferation and augmented TRPC3 protein expression. Both TRPC3 blockade by GdCl 3 or specific TRPC3 antibodies and gene silencing by siRNA inhibited the proliferation of airway smooth muscle cells. In contrast, the current drugs treatment for asthma such as Dexamethasone and Aminophylline had no effects on TRPC3 protein overexpression. Therefore, TRPC3 protein overexpression may be involved in airway smooth muscle hyperresponsiveness and remodeling in asthmatic mice, providing evidence for a new direction of asthma pathogenesis research and a new target for drug intervention. © 2018 International Federation for Cell Biology.

  5. Bimodal distribution of the prostaglandin I2 synthase antigen in smooth muscle cells.

    PubMed

    Smith, W L; DeWitt, D L; Allen, M L

    1983-05-10

    Vascular and nonvascular smooth muscles in 12 different organs from the gastrointestinal, respiratory, and urogenital tracts of the rabbit, cow, dog, sheep, pig, and rat were examined for prostaglandin I2 (PGI2) synthase immunoreactivity by indirect immunocytofluorescence using monoclonal antibodies against the enzyme. Each of 35 different smooth muscle layers tested stained for the PGI2 synthase antigen except the circular smooth muscle of the rabbit large intestine. Interestingly, PGI2 synthase-positive fluorescent staining of smooth muscle was always observed to be associated with both the nuclear and plasma membranes. Our results indicate that most smooth muscle, both vascular and nonvascular, has the capacity to synthesize PGI2. Moreover, the fact that the PGI2 synthase antigen is associated with at least two different organelles in each cell suggests that there are two independent PGI2-synthesizing systems in smooth muscle; one on the nuclear membrane, and one on the plasma membrane. PGI2 formed at these different sites may subserve different functions within the same cell.

  6. KIR channels function as electrical amplifiers in rat vascular smooth muscle

    PubMed Central

    Smith, Pamela D; Brett, Suzanne E; Luykenaar, Kevin D; Sandow, Shaun L; Marrelli, Sean P; Vigmond, Edward J; Welsh, Donald G

    2008-01-01

    Strong inward rectifying K+ (KIR) channels have been observed in vascular smooth muscle and can display negative slope conductance. In principle, this biophysical characteristic could enable KIR channels to ‘amplify’ responses initiated by other K+ conductances. To test this, we have characterized the diversity of smooth muscle KIR properties in resistance arteries, confirmed the presence of negative slope conductance and then determined whether KIR inhibition alters the responsiveness of middle cerebral, coronary septal and third-order mesenteric arteries to K+ channel activators. Our initial characterization revealed that smooth muscle KIR channels were highly expressed in cerebral and coronary, but not mesenteric arteries. These channels comprised KIR2.1 and 2.2 subunits and electrophysiological recordings demonstrated that they display negative slope conductance. Computational modelling predicted that a KIR-like current could amplify the hyperpolarization and dilatation initiated by a vascular K+ conductance. This prediction was consistent with experimental observations which showed that 30 μm Ba2+ attenuated the ability of K+ channel activators to dilate cerebral and coronary arteries. This attenuation was absent in mesenteric arteries where smooth muscle KIR channels were poorly expressed. In summary, smooth muscle KIR expression varies among resistance arteries and when channel are expressed, their negative slope conductance amplifies responses initiated by smooth muscle and endothelial K+ conductances. These findings highlight the fact that the subtle biophysical properties of KIR have a substantive, albeit indirect, role in enabling agonists to alter the electrical state of a multilayered artery. PMID:18063660

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

    PubMed

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

    2015-01-01

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

  8. Emodin augments calcium activated chloride channel in colonic smooth muscle cells by Gi/Go protein.

    PubMed

    Xu, Long; Ting-Lou; Lv, Nonghua; Zhu, Xuan; Chen, Youxiang; Yang, Jing

    2009-08-01

    Emodin is a natural anthraquinone in rhubarb. It has been identified as a prokinetic drug for gastrointestinal motility in Chinese traditional medicine. Emodin contracts smooth muscle by increasing the concentration of intracellular Ca(2+). In many smooth muscles, increasing intracellular Ca(2+) activates Ca(2+)-activated Cl(-) channels (ClCA). The study was aimed to investigate the effects of emodin on ClCA channels in colonic smooth muscle. 4 channel physiology signal acquire system was used to measure isometric contraction of smooth muscle strips. ClCA currents were recorded by EPC10 with perforated whole cell model. Emodin contracted strips and cells in colonic smooth muscle and augmented ClCA currents. Niflumic acid (NFA) and 4', 4'-diisothiostilbene-2, 2-disulfonic acid (DIDS) blocked the effects. Gi/Go protein inhibits protein kinase A (PKA) and protein kinase C (PKC), and PKA and PKC reduced ClCA currents. Pertussis toxin (PTX, a special inhibitor of Gi/Go protein), 8-bromoadenosine 38, 58-cyclic monophosphate (8-BrcAMP, a membrane-permeant protein kinase A activator) and Phorbol-12-myristate-13-acetate (PMA, a membrane-permeant protein kinase C activator) inhibited the effects on ClCA currents significantly. Our findings suggest that emodin augments ClCA channels to contract smooth muscle in colon, and the effect is induced mostly by enhancement of membrane Gi/Go protein signal transducer pathway.

  9. Ca2+ sparks act as potent regulators of excitation-contraction coupling in airway smooth muscle.

    PubMed

    Zhuge, Ronghua; Bao, Rongfeng; Fogarty, Kevin E; Lifshitz, Lawrence M

    2010-01-15

    Ca2+ sparks are short lived and localized Ca2+ transients resulting from the opening of ryanodine receptors in sarcoplasmic reticulum. These events relax certain types of smooth muscle by activating big conductance Ca2+-activated K+ channels to produce spontaneous transient outward currents (STOCs) and the resultant closure of voltage-dependent Ca2+ channels. But in many smooth muscles from a variety of organs, Ca2+ sparks can additionally activate Ca2+-activated Cl(-) channels to generate spontaneous transient inward current (STICs). To date, the physiological roles of Ca2+ sparks in this latter group of smooth muscle remain elusive. Here, we show that in airway smooth muscle, Ca2+ sparks under physiological conditions, activating STOCs and STICs, induce biphasic membrane potential transients (BiMPTs), leading to membrane potential oscillations. Paradoxically, BiMPTs stabilize the membrane potential by clamping it within a negative range and prevent the generation of action potentials. Moreover, blocking either Ca2+ sparks or hyperpolarization components of BiMPTs activates voltage-dependent Ca2+ channels, resulting in an increase in global [Ca2+](i) and cell contraction. Therefore, Ca2+ sparks in smooth muscle presenting both STICs and STOCs act as a stabilizer of membrane potential, and altering the balance can profoundly alter the status of excitability and contractility. These results reveal a novel mechanism underlying the control of excitability and contractility in smooth muscle.

  10. Homeostatic and therapeutic roles of VIP in smooth muscle function: myo-neuroimmune interactions

    PubMed Central

    Shi, Xuan-Zheng

    2009-01-01

    We tested the hypothesis that spontaneous release of vasoactive intestinal peptide (VIP) from enteric neurons maintains homeostasis in smooth muscle function in mild inflammatory insults and that infusion of exogenous VIP has therapeutic effects on colonic smooth muscle dysfunction in inflammation. In vitro experiments were performed on human colonic circular smooth muscle tissues and in vivo on rats. The incubation of human colonic circular smooth muscle strips with TNF-α suppressed their contractile response to ACh and the expression of the pore-forming α1C subunit of Cav1.2 channels. VIP reversed both effects by blocking the translocation of NF-κB to the nucleus and its binding to the κB recognition sites on hα1C1b promoter. The translocation of NF-κB was inhibited by blocking the degradation of IκBβ. Induction of inflammation by a subthreshold dose of 17 mg/kg trinitrobenzene sulfonic acid (TNBS) in rats moderately decreased muscularis externa concentration of VIP, and it had little effect on the contractile response of circular smooth muscle strips to ACh. The blockade of VIP and pituitary adenylate cyclase-activating peptide receptors 1/2 during mild inflammatory insult significantly worsened the suppression of contractility and the inflammatory response. The induction of more severe inflammation by 68 mg/kg TNBS induced marked suppression of colonic circular muscle contractility and decrease in serum VIP. Exogenous infusion of VIP by an osmotic pump reversed these effects. We conclude that the spontaneous release of VIP from the enteric motor neurons maintains homeostasis in smooth muscle function in mild inflammation by blocking the activation of NF-κB. The infusion of exogenous VIP mitigates colonic inflammatory response and smooth muscle dysfunction. PMID:19661154

  11. A Simple, Inexpensive Model to Demonstrate How Contraction of GI Longitudinal Smooth Muscle Promotes Propulsion

    ERIC Educational Resources Information Center

    Lujan, Heidi L.; DiCarlo, Stephen E.

    2015-01-01

    Peristalis is a propulsive activity that involves both circular and longitudinal muscle layers of the esophagus, distal stomach, and small and large intestines. During peristalsis, the circular smooth muscle contracts behind (on the orad side) the bolus and relaxes in front (on the aborad side) of the bolus. At the same time, the longitudinal…

  12. Autologous Smooth Muscle Progenitor Cells Enhance Regeneration of Tissue-Engineered Bladder.

    PubMed

    Zhou, Liuhua; Xia, Jiadong; Wang, Pengji; Jia, Ruipeng; Zheng, Junhua; Yao, Xudong; Chen, Yun; Dai, Yutian; Yang, Bin

    2018-03-01

    Tissue engineering techniques provide a great potential to de novo construct a histological bladder. Smooth muscle regeneration is extremely important for the functional recovery of engineered neobladder. However, many challenges remain for the use of bladder smooth muscle cells (SMCs) as the cell sources. Recent evidences showed that smooth muscle progenitor cells (SPCs) in the peripheral blood have the capacity of differentiating into SMCs, while their use for bladder regeneration has not yet been reported. The aim of our study was to investigate the effect and mechanism of autologous SPCs on bladder regeneration in a rabbit model. In this study, autologous SPCs were isolated and cultured from the peripheral blood, labeled with CM-DiI, and then seeded into a porcine bladder acellular matrix (BAM) to construct a SPC-BAM complex, which was finally implanted to substitute the partial bladder with an equivalent size. In the results, SPCs demonstrated the phenotype of stem/progenitor cells, expressed SMs markers (alpha-smooth muscle actin [α-SMA], desmin, calponin, SM22α, and smooth muscle myosin heavy chain [SMMHC]), and displayed carbachol-induced contraction. Compared with cell-free BAM, the SPC-BAM was able to improve histological regeneration (smooth muscle regeneration, vascularization, and nerve formation) and functional recovery (urodynamic function and smooth muscle contraction) of the engineered neobladder. Cell tracing indicated that seeded SPCs could survive and directly integrated into the regenerated neobladder. In addition, SPCs could also promote proliferation and migration of rabbit bladder SMCs through the paracrine platelet-derived growth factor-BB (PDGF-BB). In conclusion, our study first demonstrated that SPCs from the peripheral blood could enhance histological regeneration and functional recovery of the tissue-engineered neobladder through both the direct integration and indirect paracrine effect, supporting the use of SPCs as the cell sources

  13. A new paradigm for the role of smooth muscle cells in the human cervix.

    PubMed

    Vink, Joy Y; Qin, Sisi; Brock, Clifton O; Zork, Noelia M; Feltovich, Helen M; Chen, Xiaowei; Urie, Paul; Myers, Kristin M; Hall, Timothy J; Wapner, Ronald; Kitajewski, Jan K; Shawber, Carrie J; Gallos, George

    2016-10-01

    Premature cervical remodeling resulting in spontaneous preterm birth may begin with premature failure or relaxation at the internal os (termed "funneling"). To date, we do not understand why the internal os fails or why funneling occurs in some cases of premature cervical remodeling. Although the human cervix is thought to be mostly collagen with minimal cellular content, cervical smooth muscle cells are present in the cervix and can cause cervical tissue contractility. To understand why the internal os relaxes or why funneling occurs in some cases of premature cervical remodeling, we sought to evaluate cervical smooth muscle cell content and distribution throughout human cervix and correlate if cervical smooth muscle organization influences regional cervical tissue contractility. Using institutional review board-approved protocols, nonpregnant women <50 years old undergoing hysterectomy for benign indications were consented. Cervical tissue from the internal and external os were immunostained for smooth muscle cell markers (α-smooth muscle actin, smooth muscle protein 22 calponin) and contraction-associated proteins (connexin 43, cyclooxygenase-2, oxytocin receptor). To evaluate cervical smooth muscle cell morphology throughout the entire cervix, whole cervical slices were obtained from the internal os, midcervix, and external os and immunostained with smooth muscle actin. To correlate tissue structure with function, whole slices from the internal and external os were stimulated to contract with 1 μmol/L of oxytocin in organ baths. In separate samples, we tested if the cervix responds to a common tocolytic, nifedipine. Cervical slices from the internal os were treated with oxytocin alone or oxytocin + increasing doses of nifedipine to generate a dose response and half maximal inhibitory concentration. Student t test was used where appropriate. Cervical tissue was collected from 41 women. Immunohistochemistry showed cervical smooth muscle cells at the internal

  14. Evidence Supports Tradition: The in Vitro Effects of Roman Chamomile on Smooth Muscles

    PubMed Central

    Sándor, Zsolt; Mottaghipisheh, Javad; Veres, Katalin; Hohmann, Judit; Bencsik, Tímea; Horváth, Attila; Kelemen, Dezső; Papp, Róbert; Barthó, Loránd; Csupor, Dezső

    2018-01-01

    The dried flowers of Chamaemelum nobile (L.) All. have been used in traditional medicine for different conditions related to the spasm of the gastrointestinal system. However, there have been no experimental studies to support the smooth muscle relaxant effect of this plant. The aim of our research was to assess the effects of the hydroethanolic extract of Roman chamomile, its fractions, four of its flavonoids (apigenin, luteolin, hispidulin, and eupafolin), and its essential oil on smooth muscles. The phytochemical compositions of the extract and its fractions were characterized and quantified by HPLC-DAD, the essential oil was characterized by GC and GC-MS. Neuronally mediated and smooth muscle effects were tested in isolated organ bath experiments on guinea pig, rat, and human smooth muscle preparations. The crude herbal extract induced an immediate, moderate, and transient contraction of guinea pig ileum via the activation of cholinergic neurons of the gut wall. Purinoceptor and serotonin receptor antagonists did not influence this effect. The more sustained relaxant effect of the extract, measured after pre-contraction of the preparations, was remarkable and was not affected by an adrenergic beta receptor antagonist. The smooth muscle-relaxant activity was found to be associated with the flavonoid content of the fractions. The essential oil showed only the relaxant effect, but no contracting activity. The smooth muscle-relaxant effect was also detected on rat gastrointestinal tissues, as well as on strip preparations of human small intestine. These results suggest that Roman chamomile extract has a direct and prolonged smooth muscle-relaxant effect on guinea pig ileum which is related to its flavonoid content. In some preparations, a transient stimulation of enteric cholinergic motoneurons was also detected. The essential oil also had a remarkable smooth muscle relaxant effect in this setting. Similar relaxant effects were also detected on other visceral

  15. Evidence Supports Tradition: The in Vitro Effects of Roman Chamomile on Smooth Muscles.

    PubMed

    Sándor, Zsolt; Mottaghipisheh, Javad; Veres, Katalin; Hohmann, Judit; Bencsik, Tímea; Horváth, Attila; Kelemen, Dezső; Papp, Róbert; Barthó, Loránd; Csupor, Dezső

    2018-01-01

    The dried flowers of Chamaemelum nobile (L.) All. have been used in traditional medicine for different conditions related to the spasm of the gastrointestinal system. However, there have been no experimental studies to support the smooth muscle relaxant effect of this plant. The aim of our research was to assess the effects of the hydroethanolic extract of Roman chamomile, its fractions, four of its flavonoids (apigenin, luteolin, hispidulin, and eupafolin), and its essential oil on smooth muscles. The phytochemical compositions of the extract and its fractions were characterized and quantified by HPLC-DAD, the essential oil was characterized by GC and GC-MS. Neuronally mediated and smooth muscle effects were tested in isolated organ bath experiments on guinea pig, rat, and human smooth muscle preparations. The crude herbal extract induced an immediate, moderate, and transient contraction of guinea pig ileum via the activation of cholinergic neurons of the gut wall. Purinoceptor and serotonin receptor antagonists did not influence this effect. The more sustained relaxant effect of the extract, measured after pre-contraction of the preparations, was remarkable and was not affected by an adrenergic beta receptor antagonist. The smooth muscle-relaxant activity was found to be associated with the flavonoid content of the fractions. The essential oil showed only the relaxant effect, but no contracting activity. The smooth muscle-relaxant effect was also detected on rat gastrointestinal tissues, as well as on strip preparations of human small intestine. These results suggest that Roman chamomile extract has a direct and prolonged smooth muscle-relaxant effect on guinea pig ileum which is related to its flavonoid content. In some preparations, a transient stimulation of enteric cholinergic motoneurons was also detected. The essential oil also had a remarkable smooth muscle relaxant effect in this setting. Similar relaxant effects were also detected on other visceral

  16. The action of 5-hydroxytryptamine on Mytilus smooth muscle

    PubMed Central

    Hidaka, T.; Osa, T.; Twarog, Betty M.

    1967-01-01

    1. In the nerve—muscle preparation, where catch was characteristically minimal, 5-hydroxytryptamine (5-HT) had no effect on resting membrane potential, junction potentials, spikes or contraction. 2. In muscle bundles, where catch was prominent, 5-HT did not change membrane potentials, but prolonged junction potentials and lowered the threshold for spike discharge and contraction. 3. In muscle bundles, exposed to high concentrations of 5-HT, depolarization evoked repetitive spikes, while in low 5-HT, spikes were seldom fired even with much greater depolarization. 4. In muscle bundles, the effective membrane resistance, Reff., decreased from 45-60 to 23-35 MΩ as 5-HT concentration was increased. 5. It is suggested that 5-HT may facilitate spike discharge by lowering the internal free Ca2+ concentration. PMID:6059006

  17. Factors influencing contraction and catch in Mytilus smooth muscle

    PubMed Central

    Twarog, Betty M.

    1967-01-01

    1. Conditions are defined which determine the level of catch after acetylcholine stimulation of Mytilus muscle. 2. Catch tension in dissected muscle is absent when connexions with ganglia are intact. 3. Catch tension is absent at temperatures above 30° C. 4. Catch tension decreases when intervals between stimuli are increased. 5. Increasing concentrations of 5-hydroxytryptamine (5-HT) from 10-8M to 10-6M quantitatively decreases catch tension. 6. The length—tension curve of ganglion-free Mytilus muscle bundles suggests that catch tension varies in proportion to the tension developed in contraction. 7. External Ca2+ concentration has no selective influence on catch. 8. All factors which reduce catch also increase muscle excitability, suggesting that catch may depend on a mechanism controlling the intracellular concentration of an activator such as Ca2+. PMID:6059005

  18. The action of 5-hydroxytryptamine on Mytilus smooth muscle.

    PubMed

    Hidaka, T; Osa, T; Twarog, B M

    1967-10-01

    1. In the nerve-muscle preparation, where catch was characteristically minimal, 5-hydroxytryptamine (5-HT) had no effect on resting membrane potential, junction potentials, spikes or contraction.2. In muscle bundles, where catch was prominent, 5-HT did not change membrane potentials, but prolonged junction potentials and lowered the threshold for spike discharge and contraction.3. In muscle bundles, exposed to high concentrations of 5-HT, depolarization evoked repetitive spikes, while in low 5-HT, spikes were seldom fired even with much greater depolarization.4. In muscle bundles, the effective membrane resistance, R(eff.), decreased from 45-60 to 23-35 MOmega as 5-HT concentration was increased.5. It is suggested that 5-HT may facilitate spike discharge by lowering the internal free Ca(2+) concentration.

  19. Factors influencing contraction and catch in Mytilus smooth muscle.

    PubMed

    Twarog, B M

    1967-10-01

    1. Conditions are defined which determine the level of catch after acetylcholine stimulation of Mytilus muscle.2. Catch tension in dissected muscle is absent when connexions with ganglia are intact.3. Catch tension is absent at temperatures above 30 degrees C.4. Catch tension decreases when intervals between stimuli are increased.5. Increasing concentrations of 5-hydroxytryptamine (5-HT) from 10(-8)M to 10(-6)M quantitatively decreases catch tension.6. The length-tension curve of ganglion-free Mytilus muscle bundles suggests that catch tension varies in proportion to the tension developed in contraction.7. External Ca(2+) concentration has no selective influence on catch.8. All factors which reduce catch also increase muscle excitability, suggesting that catch may depend on a mechanism controlling the intracellular concentration of an activator such as Ca(2+).

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

    PubMed

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

    2011-09-01

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

  1. Captopril augments acetylcholine-induced bronchial smooth muscle contractions in vitro via kinin-dependent mechanisms.

    PubMed

    Agrawal, Naman; Akella, Aparna; Deshpande, Shripad B

    2016-06-01

    Angiotensin converting enzyme (ACE) inhibitors therapy is aassociated with bothersome dry cough as an adverse effect. The mechanisms underlying this adverse effect are not clear. Therefore, influence of captopril (an ACE inhibitor) on acetylcholine (ACh)-induced bronchial smooth muscle contractions was investigated. Further, the mechanisms underlying the captopril-induced changes were also explored. In vitro contractions of rat bronchial smooth muscle to cumulative concentrations of ACh were recorded before and after exposure to captopril. Further, the involvement of kinin and inositol triphosphate (IP₃) pathways for captopril-induced alterations were explored. ACh produced concentration-dependent (5-500 µM) increase in bronchial smooth muscle contractions. Pre-treatment with captopril augmented the ACh-induced contractions at each concentration significantly. Pre-treatment with aprotinin (kinin synthesis inhibitor) or heparin (inositol triphosphate, IP₃-inhibitor), blocked the captopril-induced augmentation of bronchial smooth muscle contractions evoked by ACh. Further, captopril-induced augmentation was absent in calcium-free medium. These results suggest that captopril sensitizes bronchial smooth muscles to ACh-induced contractions. This sensitization may be responsible for dry cough associated with captopril therapy.

  2. Inhibitory effect of potassium alum on smooth muscle contraction of rabbit and its mechanism.

    PubMed

    Tang, Zhong-Yuan; Lin, Yuan; Yang, Xiao-Li; Wei, Wei; Tang, Ze-Yao

    2014-07-10

    To investigate the effects of potassium alum (Alunite) on smooth muscle contraction and phosphorylation of myosin light chain by myosin light chain kinase (MLCK) and to try to find out the clue of its mechanism. An isolated rabbit duodenum smooth muscle strip was selected to study the effects of potassium alum on its contractile activity under the condition of Krebs' solution using HW-400S constant temperature smooth muscle trough. The myosin and MLCK were purified from chicken gizzard smooth muscle. Myosin light chain phosphorylation was determined by glycerol-polyacrylamide gel electrophoresis; myosin Mg 2+ -ATPase activity was measured by inorganic phosphate liberation method. Potassium alum (2.5-20 mmol/L) inhibited the contraction on duodenum in a dose-related and a time-dependent manner; potassium alum could also inhibit the extent of phosphorylation of myosin light chain in a dose-related and a time-dependent manner; and potassium alum inhibited the extent of Mg 2+ -ATPase activity in a dose-related manner. Potassium alum inhibited smooth muscle contraction in a way of inhibiting phosphorylation of myosin light chain and Mg 2+ -ATPase activity. This has revealed the molecular mechanism of treatment of gastrointestinal spastic disorders by potassium alum.

  3. Role of SM22 in the differential regulation of phasic vs. tonic smooth muscle

    PubMed Central

    Ali, Mehboob

    2015-01-01

    Preliminary proteomics studies between tonic vs. phasic smooth muscles identified three distinct protein spots identified to be those of transgelin (SM22). The latter was found to be distinctly downregulated in the internal anal sphincter (IAS) vs. rectal smooth muscle (RSM) SMC. The major focus of the present studies was to examine the differential molecular control mechanisms by SM22 in the functionality of truly tonic smooth muscle of the IAS vs. the adjoining phasic smooth muscle of the RSM. We monitored SMC lengths before and after incubation with pFLAG-SM22 (for SM22 overexpression), and SM22 small-interfering RNA. pFLAG-SM22 caused concentration-dependent and significantly greater relaxation in the IAS vs. the RSM SMCs. Conversely, temporary silencing of SM22 caused contraction in both types of the SMCs. Further studies revealed a significant reverse relationship between the levels of SM22 phosphorylation and the amount of SM22-actin binding in the IAS and RSM SMC. Data showed higher phospho-SM22 levels and decreased SM22-actin binding in the IAS, and reverse to be the case in the RSM SMCs. Experiments determining the mechanism for SM22 phosphorylation in these smooth muscles revealed that Y-27632 (Rho kinase inhibitor) but not Gö-6850 (protein kinase C inhibitor) caused concentration-dependent decreased phosphorylation of SM22. We speculate that SM22 plays an important role in the regulation of basal tone via Rho kinase-induced phosphorylation of SM22. PMID:25617350

  4. Differential effects of thin and thick filament disruption on zebrafish smooth muscle regulatory proteins

    PubMed Central

    Davuluri, G.; Seiler, C.; Abrams, J.; Soriano, A. J.; Pack, M.

    2013-01-01

    Background The smooth muscle actin binding proteins Caldesmon and Tropomyosin (Tm) promote thin filament assembly by stabilizing actin polymerization, however, whether filament assembly affects either the stability or activation of these and other smooth muscle regulatory proteins is not known. Methods Measurement of smooth muscle regulatory protein levels in wild type zebrafish larvae following antisense knockdown of smooth muscle actin (Acta2) and myosin heavy chain (Myh11) proteins, and in colourless mutants that lack enteric nerves. Comparison of intestinal peristalsis in wild type and colourless larvae. Key Results Knockdown of Acta2 led to reduced levels of phospho-Caldesmon and Tm. Total Caldesmon and phospho-myosin light chain (p-Mlc) levels were unaffected. Knockdown of Myh11 had no effect on the levels of either of these proteins. Phospho-Caldesmon and p-Mlc levels were markedly reduced in colourless mutants that have intestinal motility comparable with wild type larvae. Conclusions & Inferences These in vivo findings provide new information regarding the activation and stability of smooth muscle regulatory proteins in zebrafish larvae and their role in intestinal peristalsis in this model organism. PMID:20591105

  5. Discs-large homolog 1 regulates smooth muscle orientation in the mouse ureter

    PubMed Central

    Mahoney, Zhen X.; Sammut, Bénédicte; Xavier, Ramnik J.; Cunningham, Jeanette; Go, Gloriosa; Brim, Karry L.; Stappenbeck, Thaddeus S.; Miner, Jeffrey H.; Swat, Wojciech

    2006-01-01

    Discs-large homolog 1 (DLGH1) is a mouse ortholog of the Drosophila discs-large (DLG) tumor suppressor protein, a founding member of the PDZ and MAGUK protein families. DLG proteins play important roles in regulating cell proliferation, epithelial cell polarity, and synapse formation and function. Here, we generated a null allele of Dlgh1 and studied its role in urogenital development. Dlgh1−/− mice developed severe urinary tract abnormalities, including congenital hydronephrosis, which is the leading cause of renal failure in infants and children. DLGH1 is expressed in the developing ureter; in its absence, the stromal cells that normally lie between the urothelial and smooth muscle layers were missing. Moreover, in ureteric smooth muscle, the circular smooth muscle cells were misaligned in a longitudinal orientation. These abnormalities in the ureter led to severely impaired ureteric peristalsis. Similar smooth muscle defects are observed frequently in patients with ureteropelvic junction obstruction, a common form of hydronephrosis. Our results suggest that (i) besides its well documented role in regulating epithelial polarity, Dlgh1 also regulates smooth muscle orientation, and (ii) human DLG1 mutations may contribute to hereditary forms of hydronephrosis. PMID:17172448

  6. The relationship between bronchial hyperresponsiveness to methacholine and airway smooth muscle structure and reactivity.

    PubMed

    Armour, C L; Black, J L; Berend, N; Woolcock, A J

    1984-11-01

    The airway responsiveness of a group of 25 patients scheduled for lung resection was studied. 10 of 25 patients had a greater than or equal to 20% fall in FEV1 in response to inhaled methacholine (responders), with PD20 FEV1 values ranging from 0.6 to 7.3 mumol. Methacholine did not induce a 20% fall in FEV1 in 15 patients (non-responders). The sensitivity to carbachol and histamine of the bronchial smooth muscle resected from these patients was similar in tissue from responders and non-responders. There was no correlation between in vivo responsiveness to methacholine and in vitro sensitivity to carbachol or histamine. The volume of smooth muscle in some of these airway preparations was quantitated. There was a significant correlation between the maximum tension change in response to histamine and the volume of smooth muscle in each airway. There was no similar correlation for carbachol. The in vivo responsiveness to methacholine and in vitro sensitivity to histamine or carbachol was not related to the degree of inflammation in the airways studied. It is concluded that in vivo responsiveness cannot be explained in terms of smooth muscle sensitivity and that there may be differences between histamine and carbachol in the mechanism of contraction of airway smooth muscle.

  7. Urinary Bladder Smooth Muscle Engineered from Adipose Stem Cells and a Three Dimensional Synthetic Composite

    PubMed Central

    Jack, Gregory S.; Zhang, Rong; Lee, Min; Xu, Yuhan; Wu, Ben; Rodríguez, Larissa V.

    2009-01-01

    Human adipose stem cells were cultured in smooth muscle inductive media and seeded into synthetic bladder composites to tissue engineer bladder smooth muscle. 85:15 poly-lactic-glycolic acid bladder dome composites were cast using an electropulled microfiber luminal surface combined with an outer porous sponge. Cell seeded bladders expressed smooth muscle actin, myosin heavy chain, calponinin, and caldesmon via RT-PCR and immunoflourescence. Nude rats (n=45) underwent removal of half their bladder and repair using: (i) augmentation with the adipose stem cell seeded composites, (ii) augmentation with a matched acellular composite, or (iii) suture closure. Animals were followed for 12 weeks post-implantation and bladders were explanted serially. Results showed that bladder capacity and compliance were maintained in the cell seeded group throughout the 12 weeks, but deteriorated in the acellular scaffold group sequentially with time. Control animals repaired with sutures regained their baseline bladder capacities by week 12, demonstrating a long term limitation of this model. Histological analysis of explanted materials demonstrated viable adipose stem cells and increasing smooth muscle mass in the cell seeded scaffolds with time. Tissue bath stimulation demonstrated smooth muscle contraction of the seeded implants but not the acellular implants after 12 weeks in vivo. Our study demonstrates the feasibility and short term physical properties of bladder tissue engineered from adipose stem cells. PMID:19345408

  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. EMMPRIN (CD147) Expression in Smooth Muscle Tumors of the Uterus.

    PubMed

    Kefeli, Mehmet; Yildiz, Levent; Gun, Seda; Ozen, Fatma Z; Karagoz, Filiz

    2016-01-01

    Smooth muscle tumors of the uterus are the most common mesenchymal tumors of the gynecologic tract. The vast majority of these are benign leiomyomas that present no diagnostic difficulty. Because some benign smooth muscle tumors may degenerate and uncommon variants exist, the diagnosis can be challenging in some cases. The goal of this research was to investigate EMMPRIN expression in leiomyomas, leiomyoma variants, and leiomyosarcomas (LMS) to determine whether it has a potential role in differential diagnosis. EMMPRIN expression was investigated with immunohistochemistry in 103 uterine smooth muscle tumors, which included 19 usual leiomyomas, 52 leiomyoma variants, and 32 LMS. They were evaluated on the basis of staining extent, intensity, and also their combined score, and the groups were compared. EMMPRIN expression was present in 3 of 19 (15.7%) usual leiomyomas, 23 of 52 (44.3%) leiomyoma variants, and 28 of 32 (87.5%) LMS. There were statistically significant differences in staining extent and intensity, and also for their combined scores, between the LMS and benign groups. Although uterine smooth muscle tumors are usually diagnosed easily with conventional diagnostic criteria, the differentiation of LMS from some variants of leiomyoma can be challenging based soley on morphology. EMMPRIN may be a valuable immunohistochemical marker for differentiating LMS from benign smooth muscle tumors in problematic cases.

  10. Pathogenesis of Focal Cytoplasmic Necrosis of the Smooth Muscle Cells in Hypertensive Rat Arterial Media

    PubMed Central

    Suzuki, Keiji; Sakata, Noriyuki; Hiraishi, Katsuya; Mori, Ichiro; Takatama, Masamitsu

    2014-01-01

    Hypertensive rat arteries exhibited severe medial smooth muscle cell injury and necrosis. Electron microscopic observations showed the smooth muscle cells of these arteries exhibited characteristics of focal cytoplasmic necrosis forming new cytodemarcating membrane between the healthy cytoplasm and necrotic cytoplasm. When the focal necrotic cytoplasm disappeared from the injured smooth muscle cells, it left it with a moth-eaten leaf-like appearance (moth-eaten necrosis). At an advanced stage of injury, smooth muscle cells changed to islet-like cell bodies with newly formed basement membranes around them, and further islet-like cell bodies and cell debris disappeared leaving lamellar and reticular basement membranes. In hypertensive rats injected with nitroblue tetrazolium (NBT), formazan deposits were observed in the medial cells and nitrotyrosine, a biomarker of peroxynitrite, were immunohistochemically observed in the arterial media. Nick-end positive extranuclear small granular bodies, which might have derived from focal necrotic cytoplasm and nucleus, were detected in the arterial media using DNA nick-end labeling method. Based on electron microscopical and histochemical findings, we conjectured that the focal cytoplasmic necrosis of the smooth muscle cells in the arterial media depended on injury arising from mitochondria-derived oxidants. PMID:25861127

  11. An extensive system of extravascular smooth muscle cells exists in the choroid of the rabbit eye.

    PubMed

    Haddad, A; Laicine, E M; Tripathi, B J; Tripathi, R C

    2001-09-01

    We investigated the structural organization of the choroid especially with regard to the presence of extravascular smooth muscle (EVSM) cells in albino rabbits. The eyes were fixed by intracardiac perfusion and processed for light, confocal, and electron microscopy. An unlabeled monoclonal antibody against alpha-actin of smooth muscle and a horseradish-peroxidase-conjugated secondary antibody were used for immunodetection of smooth muscle actin by light microscopy. For confocal microscopy, whole mount choroids were immunostained with a fluorescein isothiocyanate-conjugated (FITC) antibody. Our investigations revealed that the choroidal vessels are enveloped by bundles of EVSM. In contrast to the circular orientation of the smooth muscle cells of the tunica media of the choroidal vessels, the cells of the EVSM system were oriented longitudinally along the external surface of the vessel wall. The EVSM cells were strongly immunopositive for smooth muscle alpha-actin and exhibited a green fluorescence of the FITC-labeled anti-alpha-actin antibody. Individual cells were elongated and spindle-shaped, had the usual ultrastructural features of smooth muscle cells and, in places, were organized as 20 layers. EVSM cells were present throughout the thickness of the choroid, but not between the fenestrated endothelial lining of the choriocapillaris and Bruch's membrane, and extended from the optic nerve to the ciliary body where they merged with the ciliary muscles. Based on the three dimensional organization, immunoreactivity, and cellular and subcellular features of the EVSM system as well as information in the literature, we hypothesize that, functionally, this system, in conjunction with the choroidal vasculature, contributes to the myogenic control of choroidal blood flow and tissue volume, and also affects the intraocular pressure as well as the refractive and accommodative state of the eye.

  12. Ranolazine inhibits voltage-gated mechanosensitive sodium channels in human colon circular smooth muscle cells

    PubMed Central

    Neshatian, Leila; Strege, Peter R.; Rhee, Poong-Lyul; Kraichely, Robert E.; Mazzone, Amelia; Bernard, Cheryl E.; Cima, Robert R.; Larson, David W.; Dozois, Eric J.; Kline, Crystal F.; Mohler, Peter J.; Beyder, Arthur

    2015-01-01

    Human jejunum smooth muscle cells (SMCs) and interstitial cells of Cajal (ICCs) express the SCN5A-encoded voltage-gated, mechanosensitive sodium channel NaV1.5. NaV1.5 contributes to small bowel excitability, and NaV1.5 inhibitor ranolazine produces constipation by an unknown mechanism. We aimed to determine the presence and molecular identity of Na+ current in the human colon smooth muscle and to examine the effects of ranolazine on Na+ current, mechanosensitivity, and smooth muscle contractility. Inward currents were recorded by whole cell voltage clamp from freshly dissociated human colon SMCs at rest and with shear stress. SCN5A mRNA and NaV1.5 protein were examined by RT-PCR and Western blots, respectively. Ascending human colon strip contractility was examined in a muscle bath preparation. SCN5A mRNA and NaV1.5 protein were identified in human colon circular muscle. Freshly dissociated human colon SMCs had Na+ currents (−1.36 ± 0.36 pA/pF), shear stress increased Na+ peaks by 17.8 ± 1.8% and accelerated the time to peak activation by 0.7 ± 0.3 ms. Ranolazine (50 μM) blocked peak Na+ current by 43.2 ± 9.3% and inhibited shear sensitivity by 25.2 ± 3.2%. In human ascending colon strips, ranolazine decreased resting tension (31%), reduced the frequency of spontaneous events (68%), and decreased the response to smooth muscle electrical field stimulation (61%). In conclusion, SCN5A-encoded NaV1.5 is found in human colonic circular smooth muscle. Ranolazine blocks both peak amplitude and mechanosensitivity of Na+ current in human colon SMCs and decreases contractility of human colon muscle strips. Our data provide a likely mechanistic explanation for constipation induced by ranolazine. PMID:26185330

  13. Smooth muscle architecture within cell-dense vascular tissues influences functional contractility.

    PubMed

    Win, Zaw; Vrla, Geoffrey D; Steucke, Kerianne E; Sevcik, Emily N; Hald, Eric S; Alford, Patrick W

    2014-12-01

    The role of vascular smooth muscle architecture in the function of healthy and dysfunctional vessels is poorly understood. We aimed at determining the relationship between vascular smooth muscle architecture and contractile output using engineered vascular tissues. We utilized microcontact printing and a microfluidic cell seeding technique to provide three different initial seeding conditions, with the aim of influencing the cellular architecture within the tissue. Cells seeded in each condition formed confluent and aligned tissues but within the tissues, the cellular architecture varied. Tissues with a more elongated cellular architecture had significantly elevated basal stress and produced more contractile stress in response to endothelin-1 stimulation. We also found a correlation between the contractile phenotype marker expression and the cellular architecture, contrary to our previous findings in non-confluent tissues. Taken with previous results, these data suggest that within cell-dense vascular tissues, smooth muscle contractility is strongly influenced by cell and tissue architectures.

  14. 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.

  15. Primary Intraosseous Smooth Muscle Tumor of Uncertain Malignant Potential: Original Report and Molecular Characterization

    PubMed Central

    Kropp, Lauren; Siegal, Gene P.; Frampton, Garrett M.; Rodriguez, Michael G.; McKee, Svetlana; Conry, Robert M.

    2016-01-01

    We report the first case of primary intraosseous smooth muscle tumor of uncertain malignant potential (STUMP) which is analogous to borderline malignant uterine smooth muscle tumors so designated. The tumor presented in the femur of an otherwise healthy 30-year-old woman. Over a 3-year period, the patient underwent 11 biopsies or resections and 2 cytologic procedures. Multiple pathologists reviewed the histologic material including musculoskeletal pathologists but could not reach a definitive diagnosis. However, metastases eventually developed and were rapidly progressive and responsive to gemcitabine and docetaxel. Molecular characterization and ultrastructural analysis was consistent with smooth muscle origin, and amplification of unmutated chromosome 12p and 12q segments appears to be the major genomic driver of this tumor. Primary intraosseous STUMP is thought to be genetically related to leiomyosarcoma of bone, but likely representing an earlier stage of carcinogenesis. Wide excision and aggressive follow-up is warranted for this potentially life-threatening neoplasm. PMID:27994831

  16. Primary Intraosseous Smooth Muscle Tumor of Uncertain Malignant Potential: Original Report and Molecular Characterization.

    PubMed

    Kropp, Lauren; Siegal, Gene P; Frampton, Garrett M; Rodriguez, Michael G; McKee, Svetlana; Conry, Robert M

    2016-11-17

    We report the first case of primary intraosseous smooth muscle tumor of uncertain malignant potential (STUMP) which is analogous to borderline malignant uterine smooth muscle tumors so designated. The tumor presented in the femur of an otherwise healthy 30-year-old woman. Over a 3-year period, the patient underwent 11 biopsies or resections and 2 cytologic procedures. Multiple pathologists reviewed the histologic material including musculoskeletal pathologists but could not reach a definitive diagnosis. However, metastases eventually developed and were rapidly progressive and responsive to gemcitabine and docetaxel. Molecular characterization and ultrastructural analysis was consistent with smooth muscle origin, and amplification of unmutated chromosome 12p and 12q segments appears to be the major genomic driver of this tumor. Primary intraosseous STUMP is thought to be genetically related to leiomyosarcoma of bone, but likely representing an earlier stage of carcinogenesis. Wide excision and aggressive follow-up is warranted for this potentially life-threatening neoplasm.

  17. Smooth muscle relaxant activity of Crocus sativus (saffron) and its constituents: possible mechanisms

    PubMed Central

    Mokhtari-Zaer, Amin; Khazdair, Mohammad Reza; Boskabady, Mohammad Hossein

    2015-01-01

    Saffron, Crocus sativus L. (C. sativus) is rich in carotenoids and used in traditional medicine for treatment of various conditions such as coughs, stomach disorders, amenorrhea, asthma and cardiovascular disorders. These therapeutic effects of the plant are suggested to be due to its relaxant effect on smooth muscles. The effect of C. sativus and its constituents on different smooth muscles and the underlying mechanisms have been studied. Several studies have shown the relaxant effects of C. sativus and its constituents including safranal, crocin, crocetin and kaempferol on blood vessels. In addition, it was reported that saffron stigma lowers systolic blood pressure. The present review highlights the relaxant effects of C. sativus and its constituents on various smooth muscles. The possible mechanisms of this relaxing effect including activation of ß2-adrenoceptors, inhibition of histamine H1 and muscarinic receptors and calcium channels and modulation of nitric oxide (NO) are also reviewed. PMID:26468456

  18. Increased IGF-IEc expression and mechano-growth factor production in intestinal muscle of fibrostenotic Crohn's disease and smooth muscle hypertrophy

    PubMed Central

    Li, Chao; Vu, Kent; Hazelgrove, Krystina

    2015-01-01

    The igf1 gene is alternatively spliced as IGF-IEa and IGF-IEc variants in humans. In fibrostenotic Crohn's disease, the fibrogenic cytokine TGF-β1 induces IGF-IEa expression and IGF-I production in intestinal smooth muscle and results in muscle hyperplasia and collagen I production that contribute to stricture formation. Mechano-growth factor (MGF) derived from IGF-IEc induces skeletal and cardiac muscle hypertrophy following stress. We hypothesized that increased IGF-IEc expression and MGF production mediated smooth muscle hypertrophy also characteristic of fibrostenotic Crohn's disease. IGF-IEc transcripts and MGF protein were increased in muscle cells isolated from fibrostenotic intestine under regulation by endogenous TGF-β1. Erk5 and MEF2C were phosphorylated in vivo in fibrostenotic muscle; both were phosphorylated and colocalized to nucleus in response to synthetic MGF in vitro. Smooth muscle-specific protein expression of α-smooth muscle actin, γ-smooth muscle actin, and smoothelin was increased in affected intestine. Erk5 inhibition or MEF2C siRNA blocked smooth muscle-specific gene expression and hypertrophy induced by synthetic MGF. Conditioned media of cultured fibrostenotic muscle induced muscle hypertrophy that was inhibited by immunoneutralization of endogenous MGF or pro-IGF-IEc. The results indicate that TGF-β1-dependent IGF-IEc expression and MGF production in patients with fibrostenotic Crohn's disease regulates smooth muscle cell hypertrophy a critical factor that contributes to intestinal stricture formation. PMID:26428636

  19. Role of Telokin in Regulating Murine Gastric Fundus Smooth Muscle Tension

    PubMed Central

    An, Changlong; Bhetwal, Bhupal P.; Sanders, Kenton M.; Somlyo, Avril V.; Perrino, Brian A.

    2015-01-01

    Telokin phosphorylation by cyclic GMP-dependent protein kinase facilitates smooth muscle relaxation. In this study we examined the relaxation of gastric fundus smooth muscles from basal tone, or pre-contracted with KCl or carbachol (CCh), and the phosphorylation of telokin S13, myosin light chain (MLC) S19, MYPT1 T853, T696, and CPI-17 T38 in response to 8-Bromo-cGMP, the NO donor sodium nitroprusside (SNP), or nitrergic neurotransmission. We compared MLC phosphorylation and the contraction and relaxation responses of gastric fundus smooth muscles from telokin-/- mice and their wild-type littermates to KCl or CCh, and 8-Bromo-cGMP, SNP, or nitrergic neurotransmission, respectively. We compared the relaxation responses and telokin phosphorylation of gastric fundus smooth muscles from wild-type mice and W/WV mice which lack ICC-IM, to 8-Bromo-cGMP, SNP, or nitrergic neurotransmission. We found that telokin S13 is basally phosphorylated and that 8-Bromo-cGMP and SNP increased basal telokin phosphorylation. In muscles pre-contracted with KCl or CCh, 8-Bromo-cGMP and SNP had no effect on CPI-17 or MYPT1 phosphorylation, but increased telokin phosphorylation and reduced MLC phosphorylation. In telokin-/- gastric fundus smooth muscles, basal tone and constitutive MLC S19 phosphorylation were increased. Pre-contracted telokin-/- gastric fundus smooth muscles have increased contractile responses to KCl, CCh, or cholinergic neurotransmission and reduced relaxation to 8-Bromo-cGMP, SNP, and nitrergic neurotransmission. However, basal telokin phosphorylation was not increased when muscles were stimulated with lower concentrations of SNP or when the muscles were stimulated by nitrergic neurotransmission. SNP, but not nitrergic neurotransmission, increased telokin Ser13 phosphorylation in both wild-type and W/WV gastric fundus smooth muscles. Our findings indicate that telokin may play a role in attenuating constitutive MLC phosphorylation and provide an additional mechanism to

  20. Galectin‑3 induces the phenotype transformation of human vascular smooth muscle cells via the canonical Wnt signaling.

    PubMed

    Tian, Lei; Chen, Kan; Cao, Jiatian; Han, Zhihua; Wang, Yue; Gao, Lin; Fan, Yuqi; Wang, Changqian

    2017-06-01

    Galectin‑3, a galactoside‑binding protein, is highly expressed in carotid plaques and plays an important role in the atherosclerotic lesions. The phenotype transformation of vascular smooth muscle cells is the basic pathological change of atherosclerosis. This study investigated the effects of exogenous galectin‑3 on the function and phenotype transformation of human umbilical vascular smooth muscle cells (HUSMC). In this study, we treated vascular smooth muscle cells with recombinant galectin‑3 and tested its effect on cell proliferation, migration, and phenotype transformation. Our results showed that exogenous galectin‑3 promoted human umbilical vascular smooth muscle cells (HUSMC) proliferation and migration. Exogenous galectin‑3 enhanced the expression of the smooth muscle synthetic protein osteopontin, smooth muscle contractile proteins calponin and smooth muscle α‑actin. The galectin‑3‑induced change in cell phenotype was associated with the activation of canonical Wnt signaling, as measured by β‑catenin axin2 and cyclin D1 expression. β‑catenin inhibition by small interfering RNA reduced cell proliferation, decreased cell motility, and blocked galectin‑3‑induced phenotype transformation of human umbilical vascular smooth muscle cells (HUSMC). Our data suggest galectin‑3 promotes the phenotype transformation of human umbilical vascular smooth muscle cells (HUSMC) by activating Wnt/β‑catenin signaling pathway.

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

    PubMed

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

    2004-05-01

    Bronchial hyperresponsiveness in mild to moderate asthma may result from airway smooth muscle cell proliferation or acquisition of a hypercontractile phenotype. Because these cells have not been well characterized in mild to moderate asthma, we examined the morphometric and gene expression characteristics of smooth muscle cells in this subgroup of patients with asthma. Using bronchial biopsies from 14 subjects with mild to moderate asthma and 15 control subjects, we quantified smooth muscle cell morphology by stereology and the expression of a panel of genes related to a hypercontractile phenotype of airway smooth muscle, using laser microdissection and two-step real-time polymerase chain reaction. We found that airway smooth muscle cell size was similar in both groups, but cell number was nearly twofold higher in subjects with asthma (p = 0.03), and the amount of smooth muscle in the submucosa was increased 50-83% (p < 0.005). Gene expression profiling in smooth muscle cells showed no difference in the expression of genes encoding phenotypic markers in cells from healthy subjects and subjects with asthma (all p > 0.1). We conclude that airway smooth muscle proliferation is a pathologic characteristic of subjects with mild to moderate asthma. However, smooth muscle cells in mild to moderate asthma do not show hypertrophy or gene expression changes of a hypercontractile phenotype observed in vitro.

  2. 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

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

  4. 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. © 2016 American Heart Association, Inc.

  5. mir145 Regulates TGFBR2 Expression and Matrix Synthesis in Vascular Smooth Muscle Cells

    PubMed Central

    Zhao, Ning; Koenig, Sara N.; Trask, Aaron J.; Lin, Cho-Hao; Hans, Chetan P.; Garg, Vidu; Lilly, Brenda

    2014-01-01

    Rationale MicroRNA miR145 has been implicated in vascular smooth muscle cell differentiation, but its mechanisms of action and downstream targets have not been fully defined. Objective Here, we sought to explore and define the mechanisms of miR145 function in smooth muscle cells. Methods and Results Using a combination of cell culture assays and in vivo mouse models to modulate miR145, we characterized its downstream actions on smooth muscle phenotypes. Our results show that the miR-143/145 gene cluster is induced in smooth muscle cells by coculture with endothelial cells. Endothelial cell-induced expression of miR-143/145 is augmented by Notch signaling and accordingly expression is reduced in Notch receptor-deficient cells. Screens to identify miR145-regulated genes revealed that the TGFβ pathway has a significantly high number of putative target genes, and we show that TGFβ receptor II (TGFBR2) is a direct target of miR145. Extracellular matrix (ECM) genes that are regulated by TGFBR2 were attenuated by miR145 overexpression, and miR145 mutant mice exhibit an increase in ECM synthesis. Furthermore, activation of TGFβ signaling via angiotensin II infusion revealed a pronounced fibrotic response in the absence of miR145. Conclusions These data demonstrate a specific role for miR145 in the regulation of matrix gene expression in smooth muscle cells, and suggest that miR145 acts to suppress TGFβ-dependent ECM accumulation and fibrosis, while promoting TGFβ-induced smooth muscle cell differentiation. Our findings offer evidence to explain how TGFβ signaling exhibits distinct downstream actions via its regulation by a specific microRNA. PMID:25323858

  6. Mixed stromal and smooth muscle tumours of the uterus: a report of two cases.

    PubMed

    Abid, N; Kallel, R; Mellouli, M; Mnif, H; Ayedi, L; Khabir, A; Boudawara, T

    2014-12-01

    Mixed stromal and smooth muscle uterine tumours, defined as those containing at least 30% of each component as seen by routine light microscopy, are rare. This report describes the morphological features of two such tumours diagnosed in 44-year-old and 50-year-old females complaining from recurrent uterine bleeding that was unresponsive to medical treatment. Morphological and immunohistochemical evaluations were performed, and a final diagnosis of mixed endometrial stromal nodule and smooth muscle tumour of the uterus was rendered in both cases.

  7. Rac1 modulates G-protein-coupled receptor-induced bronchial smooth muscle contraction.

    PubMed

    Sakai, Hiroyasu; Kai, Yuki; Sato, Ken; Ikebe, Mitsuo; Chiba, Yohihiko

    2018-01-05

    Increasing evidence suggests a functional role of RhoA/Rho-kinase signalling as a mechanism for smooth muscle contraction; however, little is known regarding the roles of Rac1 and other members of the Rho protein family. This study aimed to examine whether Rac1 modulates bronchial smooth muscle contraction. Ring preparations of bronchi isolated from rats were suspended in an organ bath, and isometric contraction of circular smooth muscle was measured. Immunoblotting was used to examine myosin light chain phosphorylation in bronchial smooth muscle. Our results demonstrated that muscle contractions induced by carbachol (CCh) and endothelin-1 (ET-1) were inhibited by EHT1864, a selective Rac1 inhibitor, and NSC23766, a selective inhibitor of Rac1-specific guanine nucleotide exchange factors. Similarly, myosin light chain and myosin phosphatase target subunit 1 (MYPT1) at Thr853 phosphorylation induced by contractile agonist were inhibited with Rac1 inhibition. However, contractions induced by high K + , calyculin A (a potent protein phosphatase inhibitor) and K + /PDBu were not inhibited by these Rac1 inhibitors. Interestingly, NaF (a G-protein activator)-induced contractions were inhibited by EHT1864 but not by NSC23766. We next examined the effects of a trans-acting activator of transcription protein transduction domain (PTD) fusion protein with Rac1 (PTD-Rac1) on muscle contraction. The constitutively active form of PTD-Rac1 directly induced force development and contractions were abolished by EHT1864. These results suggest that Rac1, activated by G protein-coupled receptor agonists, such as CCh and ET-1, may induce myosin light chain and MYPT phosphorylation and modulate the contraction of bronchial smooth muscle. Copyright © 2017 Elsevier B.V. All rights reserved.

  8. 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.

  9. Estimation of airway smooth muscle stiffness changes due to length oscillation using artificial neural network.

    PubMed

    Al-Jumaily, Ahmed; Chen, Leizhi

    2012-10-07

    This paper presents a novel approach to estimate stiffness changes in airway smooth muscles due to external oscillation. Artificial neural networks are used to model the stiffness changes due to cyclic stretches of the smooth muscles. The nonlinear relationship between stiffness ratios and oscillation frequencies is modeled by a feed-forward neural network (FNN) model. The structure of the FNN is selected through the training and validation using literature data from 11 experiments with different muscle lengths, muscle masses, oscillation frequencies and amplitudes. Data pre-processing methods are used to improve the robustness of the neural network model to match the non-linearity. The validation results show that the FNN model can predict the stiffness ratio changes with a mean square error of 0.0042. Copyright © 2012 Elsevier Ltd. All rights reserved.

  10. Crystal Structure of a Phosphorylated Light Chain Domain of Scallop Smooth-Muscle Myosin

    SciTech Connect

    Kumar, V.S.; Robinson, H.; O-Neall-Hennessey, E.

    2011-11-02

    We have determined the crystal structure of a phosphorylated smooth-muscle myosin light chain domain (LCD). This reconstituted LCD is of a sea scallop catch muscle myosin with its phosphorylatable regulatory light chain (RLC SmoA). In the crystal structure, Arg{sup 16}, an arginine residue that is present in this isoform but not in vertebrate smooth-muscle RLC, stabilizes the phosphorylation site. This arginine interacts with the carbonyl group of the phosphorylation-site serine in the unphosphorylated LCD (determined previously), and with the phosphate group when the serine is phosphorylated. However, the overall conformation of the LCD is essentially unchanged upon phosphorylation. This resultmore » provides additional evidence that phosphorylation of the RLC is unlikely to act as an on-switch in regulation of scallop catch muscle myosin.« less

  11. Embracing change: striated-for-smooth muscle replacement in esophagus development.

    PubMed

    Krauss, Robert S; Chihara, Daisuke; Romer, Anthony I

    2016-01-01

    The esophagus functions to transport food from the oropharyngeal region to the stomach via waves of peristalsis and transient relaxation of the lower esophageal sphincter. The gastrointestinal tract, including the esophagus, is ensheathed by the muscularis externa (ME). However, while the ME of the gastrointestinal tract distal to the esophagus is exclusively smooth muscle, the esophageal ME of many vertebrate species comprises a variable amount of striated muscle. The esophageal ME is initially composed only of smooth muscle, but its developmental maturation involves proximal-to-distal replacement of smooth muscle with striated muscle. This fascinating phenomenon raises two important questions: what is the developmental origin of the striated muscle precursor cells, and what are the cellular and morphogenetic mechanisms underlying the process? Studies addressing these questions have provided controversial answers. In this review, we discuss the development of ideas in this area and recent work that has shed light on these issues. A working model has emerged that should permit deeper understanding of the role of ME development and maturation in esophageal disorders and in the functional and evolutionary underpinnings of the variable degree of esophageal striated myogenesis in vertebrate species.

  12. Subchronic exposure to diisocyanates increases guinea pig tracheal smooth muscle responses to acetylcholine.

    PubMed

    Marek, W; Potthast, J; Marczynski, B; Mensing, T; Baur, X

    1999-01-01

    In order to study the threshold concentrations of isocyanates (IC) for induction of lung disorders, constrictive responses of tracheal smooth muscles to acetylcholine (ACH) in guinea pigs with and without diisocyanate [toluene diisocyanate (TDI), hexamethylene diisocyanate (HDI) and diphenylmethane diisocyanate (MDI)] exposure were investigated. An IC-induced increase in smooth muscle responsiveness was studied by measuring cumulative ACH dose responses (10(-10) to 10(-4) M ACH). Basal ACH dose-response curves, measured twice in intervals of 1 h using tracheal preparations of 11 guinea pigs previously not exposed to IC, were reproducible. Subchronic in vivo exposures to TDI, HDI, and MDI atmospheres of 10 and 20 parts per billion (ppb) on 5 consecutive days led to significantly (p < 0.05) increased ACH responsiveness of tracheal smooth muscle, whereas concentrations of 2.5 and 5 ppb were not effective. Exposure to HDI atmospheres of 10 ppb for 1, 2, 4, or 8 weeks resulted in a time-dependent increase in ACH responses (p < 0.05) of guinea pig tracheal smooth muscle. Increased tracheal muscle responses to ACH were transient since tracheal preparations from animals exposed to 10 and 20 ppb MDI for 4 weeks and with an exposure-free interval of 8 weeks before preparation did not show enlarged ACH responses, which were present in preparations at the end of the exposure period (p < 0.05). Exposure to low IC concentrations as present in workplaces cause increased ACH responsiveness of guinea pig tracheal smooth muscle. The increased responsiveness of the airways seems to be largely reversible, since normal responses were found after 8 weeks of IC avoidance. Reversibility of IC-induced airway hyperresponsiveness is of great occupational and preventive medical importance. Workers with acquired airway hyperresponsiveness might escape lung damage if the changes are detected in an early stage before alterations in lung function are in a chronic stage.

  13. Paraoxon attenuates vascular smooth muscle contraction through inhibiting Ca2+ influx in the rabbit thoracic aorta.

    PubMed

    Zhou, Shouhong; Liu, Liying; Yang, Xuhong; Wu, Shujin; Chen, Gengrong

    2010-01-01

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

  14. Fraktalkine produced by airway smooth muscle cells contributes to mast cell recruitment in asthma.

    PubMed

    El-Shazly, Amr; Berger, Patrick; Girodet, Pierre-Olivier; Ousova, Olga; Fayon, Michael; Vernejoux, Jean-Marc; Marthan, Roger; Tunon-de-Lara, J Manuel

    2006-02-01

    Human airway smooth muscle cells (HASMC) secrete fractalkine (FKN), a chemokine the concentration of which is increased in asthmatic patients. HASMC also induce mast cell chemotaxis, as a component of asthma inflammation. We therefore evaluated the role of smooth muscle-derived FKN in mast cell migration. We assessed the capacity of recombinant FKN to induce human mast cell chemotaxis. This effect implicates a calcium-independent pathway involving actin reorganization and protein kinase C-delta. We found that HASMC constitutively produce FKN, the synthesis of which is reinforced upon proinflammatory stimulation. Under basal experimental conditions, FKN production by HASMC is not sufficient to induce mast cell chemotaxis. However, pretreatment of mast cells with the neuropeptide vasoactive intestinal peptide (VIP) increases FKN potency to attract mast cells. Since we observed, in asthmatic patients, an increase in both FKN and VIP expression by airway smooth muscle and a positive correlation between VIP staining and mast cell infiltration of the smooth muscle layer, we conclude that HASMC-derived FKN may contribute to mast cell recruitment in asthma.

  15. Electrically Stimulated Adipose Stem Cells on Polypyrrole-Coated Scaffolds for Smooth Muscle Tissue Engineering.

    PubMed

    Björninen, Miina; Gilmore, Kerry; Pelto, Jani; Seppänen-Kaijansinkko, Riitta; Kellomäki, Minna; Miettinen, Susanna; Wallace, Gordon; Grijpma, Dirk; Haimi, Suvi

    2017-04-01

    We investigated the use of polypyrrole (PPy)-coated polymer scaffolds and electrical stimulation (ES) to differentiate adipose stem cells (ASCs) towards smooth muscle cells (SMCs). Since tissue engineering lacks robust and reusable 3D ES devices we developed a device that can deliver ES in a reliable, repeatable, and cost-efficient way in a 3D environment. Long pulse (1 ms) or short pulse (0.25 ms) biphasic electric current at a frequency of 10 Hz was applied to ASCs to study the effects of ES on ASC viability and differentiation towards SMCs on the PPy-coated scaffolds. PPy-coated scaffolds promoted proliferation and induced stronger calponin, myosin heavy chain (MHC) and smooth muscle actin (SMA) expression in ASCs compared to uncoated scaffolds. ES with 1 ms pulse width increased the number of viable cells by day 7 compared to controls and remained at similar levels to controls by day 14, whereas shorter pulses significantly decreased viability compared to the other groups. Both ES protocols supported smooth muscle expression markers. Our results indicate that electrical stimulation on PPy-coated scaffolds applied through the novel 3D ES device is a valid approach for vascular smooth muscle tissue engineering.

  16. Changes in Inhibitory Control of Circular Smooth Muscle During Coilitis in the Rat

    DTIC Science & Technology

    1998-03-25

    cyclic guanosine monophosphate Concentration at which 50% of the maximum response occurs Enteric Nervous System Gastrointestinal Giant Migrating Complex...communication throughout the gastrointestinal tract. Acetylcholine (Ach) 1 2 is the preganglionic neurotransmitter at nicotinic receptors and is also the...properties of intestinal smooth muscle, are responsible for the motility and digestive processes that occur throughout the gastrointestinal tract

  17. 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.

  18. Intestinal smooth muscle response to chronic obstruction : possible applications in jejunoileal atresia.

    PubMed

    Cloutier, R

    1975-02-01

    Hyperplasia is the main change occurring in intestinal smooth muscle above a chronic obstruction and explains the functional obstruction seen in the proximal bowel of a jejunoileal atresia. With an experimental model in dogs, this hyperplasia has been shown to be reversible. However, changes are extreme in atresia, and experiments in animals with induced atresia will best evaluate various kinds of treatment.

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

    SciTech Connect

    Costa-Silva, Bruno; Programa de Pos-graduacao em Neurociencias, Centro de Ciencias Biologicas, Universidade Federal de Santa Catarina, Campus Universitario - Trindade, 88040-900, Florianopolis, S.C.; Coelho da Costa, Meline

    2009-04-01

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

  20. Sodium spirulan as a potent inhibitor of arterial smooth muscle cell proliferation in vitro.

    PubMed

    Kaji, Toshiyuki; Okabe, Maiko; Shimada, Satomi; Yamamoto, Chika; Fujiwara, Yasuyuki; Lee, Jung-Bum; Hayashi, Toshimitsu

    2004-03-26

    Sodium spirulan (Na-SP) is a sulfated polysaccharide with M(r) approximately 220,000 isolated from the blue-green alga Spirulina platensis. The polysaccharide consists of two types of disaccharide repeating units, O-hexuronosyl-rhamnose (aldobiuronic acid) and O-rhamnosyl-3-O-methylrhamnose (acofriose) with sulfate groups, other minor saccharides and sodium ion. Since vascular smooth muscle cell proliferation is a crucial event in the progression of atherosclerosis, we investigated the effect of Na-SP on the proliferation of bovine arterial smooth muscle cells in culture. It was found that Na-SP markedly inhibits the proliferation without nonspecific cell damage. Either replacement of sodium ion with calcium ion or depolymerization of the Na-SP molecule to M(r) approximately 14,700 maintained the inhibitory activity, however, removal of sodium ion or desulfation markedly reduced the activity. Heparin and heparan sulfate also inhibited vascular smooth muscle cell growth but their effect was weaker than that of Na-SP; dextran sulfate, chondroitin sulfate, dermatan sulfate and hyaluronan failed to inhibit the cell growth. The present data suggest that Na-SP is a potent inhibitor of arterial smooth muscle cell proliferation, and the inhibitory effect requires a certain minimum sequence of polysaccharide structure whose molecular conformation is maintained by sodium ion bound to sulfate group.

  1. Arterial smooth muscle cells express nitric oxide synthase in response to endothelial injury.

    PubMed

    Hansson, G K; Geng, Y J; Holm, J; Hårdhammar, P; Wennmalm, A; Jennische, E

    1994-08-01

    Endothelial cells regulate vascular tone by secreting paracrine mediators that control the contractility of arterial smooth muscle cells. Nitric oxide (NO) is an important vasodilating agent that is generated from L-arginine by the enzyme nitric oxide synthase (NOS), which is expressed constitutively by the endothelium. NO also inhibits platelet aggregation, contributing to the antithrombotic properties of the endothelial surface. It would therefore be expected that loss of the endothelium during arterial injury would lead to vasospasm and thrombosis but instead, the neointima formed after injury has a nonthrombogenic surface and a maintained vascular patency. We report here that arterial smooth muscle cells in the neointima formed after a deendothelializing balloon injury to the rat carotid artery express the cytokine-inducible isoform of NOS. Expression was detectable by reverse transcription-polymerase chain reaction from day 1-14 after injury and in situ hybridization showed expression of NOS mRNA by neointimal smooth muscle cells, particularly at the surface of the lesion. This was associated with systemically detectable NO production as revealed by electron paramagnetic resonance spectroscopic analysis of nitrosylated red cell hemoglobin. Local NO production by intimal smooth muscle cells after endothelial injury could represent an important mechanism for the maintenance of arterial patency and nonthrombogenicity in the injured artery.

  2. Reciprocal Regulation of Syndecan-2 and Notch Signaling in Vascular Smooth Muscle Cells*

    PubMed Central

    Zhao, Ning; Liu, Hua; Lilly, Brenda

    2012-01-01

    Vascular cell interactions mediated through cell surface receptors play a critical role in the assembly and maintenance of blood vessels. These signaling interactions transmit important information that alters cell function through changes in protein dynamics and gene expression. Here, we identify syndecan-2 (SDC2) as a gene whose expression is induced in smooth muscle cells upon physical contact with endothelial cells. Syndecan-2 is a heparan sulfate proteoglycan that is known to be important for developmental processes, including angiogenesis. Our results show that endothelial cells induce mRNA expression of syndecan-2 in smooth muscle cells by activating Notch receptor signaling. Both NOTCH2 and NOTCH3 contribute to the increased expression of syndecan-2 and are themselves sufficient to promote its expression independent of endothelial cells. Syndecan family members serve as coreceptors for signaling molecules, and interestingly, our data show that syndecan-2 regulates Notch signaling and physically interacts with NOTCH3. Notch activity is attenuated in smooth muscle cells made deficient in syndecan-2, and this specifically prevents expression of the differentiation marker smooth muscle α-actin. These results show a novel mechanism in which Notch receptors control their own activity by inducing the expression of syndecan-2, which then acts to propagate Notch signaling by direct receptor interaction. PMID:22437834

  3. Endothelial Cells Direct Mesenchymal Stem Cells Toward a Smooth Muscle Cell Fate

    PubMed Central

    Lin, Cho-Hao

    2014-01-01

    Under defined conditions, mesenchymal stem cells can differentiate into unique cell types, making them attractive candidates for cell-based disease therapies. Ischemic diseases would greatly benefit from treatments that include the formation of new blood vessels from mesenchymal stem cells. However, blood vessels are complex structures composed of endothelial cells and smooth muscle cells, and their assembly and function in a diseased environment is reliant upon joining with the pre-existing vasculature. Although endothelial cell/smooth muscle cell interactions are well known, how endothelial cells may influence mesenchymal stem cells and facilitate their differentiation has not been defined. Therefore, we sought to explore how endothelial cells might drive mesenchymal stem cells toward a smooth muscle fate. Our data show that cocultured endothelial cells induce smooth muscle cell differentiation in mesenchymal stem cells. Endothelial cells can promote a contractile phenotype, reduce proliferation, and enhance collagen synthesis and secretion. Our data show that Notch signaling is essential for endothelial cell-dependent differentiation, and this differentiation pathway is largely independent of growth factor signaling mechanisms. PMID:24914692

  4. Electrical activity from smooth muscle of the anal sphincteric area of the cat.

    PubMed Central

    Bouvier, M; Gonella, J

    1981-01-01

    1. The electrical activities of longitudinal and circular smooth muscle of the anal sphincteric area have been studied in the cat. 2. Electromyographic recordings were achieved with extracellular electrodes, in vivo on acute and chronic animals, and in vitro on the isolated organ. In addition, electrical and mechanical activities were recorded from muscle strips with the sucrose gap technique. 3. Circular muscle coat electrical activity consisted exclusively of slow variations of the membrane potential of the smooth muscle cells. Each slow potential variation was followed by a contraction. 4. The electrical activity and the concomitant contractions were tetrodotoxin resistant (10(-6) g/ml.). Both disappeared in Ca-free solution or in the presence of Mn ions (10(-3) M). 5. On circular muscle, noradrenaline (10(-8)-10(-7) g/ml. in vitro, or 0.1-0.15 mg/kg in vivo) had an excitatory effect consisting in an increase of slow potential frequency. The action of noradrenaline was antagonized by phentolamine (10(-6)-10(-5) g/ml. in vitro, or 0.2 mg/kg in vivo). 6. On circular muscle, acetylcholine (10(-8)-10(-6) g/ml.), used exclusively on muscle strips, did never produce any clear cut effect. 7. Longitudinal muscle coat electrical activity consisted of spike potentials superimposed on slow time course depolarizations which were never observed alone. Each spike was followed by a contraction. This electrical activity was tetrodotoxin resistant (10(-6) g/ml.). 8. Longitudinal muscle activity was abolished by noradrenaline (10(-6) g/ml.) and enhanced by acetylcholine (10(-8)-10(-6) g/ml.). The action of noradrenaline was antagonized by propranolol (0.2 mg/kg I.V.; 10(-6) g/ml.) and that of acetylcholine by atropine (10(-7) g/ml.). 9. Electrophysiological and pharmacological data indicate that electromechanical coupling is achieved (1) in circular muscle, through Ca dependent slow variations in membrane potential of the muscle cells and (2) in longitudinal muscle, through spike

  5. Smooth Muscle LDL Receptor-related Protein-1 Inactivation Reduces Vascular Reactivity and Promotes Injury-induced Neointima Formation

    PubMed Central

    Basford, Joshua E.; Moore, Zachary W.Q.; Zhou, Li; Herz, Joachim; Hui, David Y.

    2009-01-01

    Objective Defective smooth muscle expression of LDL receptor-related protein-1 (Lrp1) increases atherosclerosis in hypercholesterolemic mice. This study explored the importance of smooth muscle Lrp1 expression under normolipidemic conditions. Methods and Results Smooth muscle cells isolated from control (smLrp1+/+) and smooth muscle-specific Lrp1 knockout (smLrp1−/−) mice were characterized based on morphology, smooth muscle marker protein expression levels, and growth rates in vitro. Vascular functions were assessed by aortic constrictive response to agonist stimulation in situ and neointimal hyperplasia to carotid arterial injury in vivo. The smLrp1−/− smooth muscle cells displayed reduced α-actin and calponin expression and an accelerated growth rate due to sustained phosphorylation of platelet derived growth factor receptor (PRGFR) and protein kinase B/Akt. Vasoconstrictive response to agonist stimulation was impaired in aortic rings isolated from smLrp1−/− mice. Injury-induced neointimal hyperplasia was significantly increased in smLrp1−/−mice. The increase in neointima was associated with corresponding elevated activation of PDGFR signaling pathway. Conclusions Smooth muscle expression of Lrp1 is important in maintaining normal vascular functions under normolipidemic conditions. The absence of Lrp1 expression results in greater smooth muscle cell proliferation, deficient contractile protein expression, impairment of vascular contractility, and promotion of denudation-induced neointimal hyperplasia. PMID:19729608

  6. Smooth muscle myosin regulation by serum and cell density in cultured rat lung connective tissue cells.

    PubMed

    Babij, P; Zhao, J; White, S; Woodcock-Mitchell, J; Mitchell, J; Absher, M; Baldor, L; Periasamy, M; Low, R B

    1993-08-01

    RNA and protein analyses were used to detect expression of SM1 and SM2 smooth muscle myosin heavy chain (MHC) in cultured adult rat lung connective tissue cells (RL-90). Smooth muscle MHC mRNA expression in confluent cells grown in 10% serum was approximately 50% of the level in adult stomach. Similar results were obtained in cells cultured at low density (25% confluency) in 1% serum. However, in low-density cultures transferred to 10% serum for 24 h, the level of MHC mRNA decreased to approximately 20% of that in adult stomach. Smooth muscle alpha-actin showed a pattern of expression similar to that for smooth muscle MHC. Expression of nonmuscle MHC-A mRNA was higher in all culture conditions compared to stomach. MHC-A mRNA expression was less in low-density cultures in low serum and increased when low-density cultures were transferred to 10% serum for 24 h. MHC-B mRNA expression was less in low- vs. high-density cultures. In contrast to MHC-A, however, MHC-B mRNA expression in low-density cultures was higher in low serum. Immunofluorescence and immunoblotting with SM1-specific antibody demonstrated the presence of the SM1 protein isoform as well as reactivity to a protein band migrating slightly faster than SM2. These results demonstrate that cultured rat lung connective tissue cells express smooth muscle MHC and that expression is modulated by culture conditions.

  7. ADAR1-Mediated RNA Editing, A Novel Mechanism Controlling Phenotypic Modulation of Vascular Smooth Muscle Cells.

    PubMed

    Fei, Jia; Cui, Xiao-Bing; Wang, Jia-Ning; Dong, Kun; Chen, Shi-You

    2016-07-22

    Vascular smooth muscle cell (SMC) phenotypic modulation is characterized by the downregulation of SMC contractile genes. Platelet-derived growth factor-BB, a well-known stimulator of SMC phenotypic modulation, downregulates SMC genes via posttranscriptional regulation. The underlying mechanisms, however, remain largely unknown. To establish RNA editing as a novel mechanism controlling SMC phenotypic modulation. Precursor mRNAs (pre-mRNA) of SMC myosin heavy chain and smooth muscle α-actin were accumulated while their mature mRNAs were downregulated during SMC phenotypic modulation, suggesting an abnormal splicing of the pre-mRNAs. The abnormal splicing resulted from SMC marker pre-mRNA editing that was facilitated by adenosine deaminase acting on RNA 1 (ADAR1), an enzyme converting adenosines to inosines (A→I editing) in RNA sequences. ADAR1 expression inversely correlated with SMC myosin heavy chain and smooth muscle α-actin levels; knockdown of ADAR1 restored SMC myosin heavy chain and smooth muscle α-actin expression in phenotypically modulated SMC, and editase domain mutation diminished the ADAR1-mediated abnormal splicing of SMC marker pre-mRNAs. Moreover, the abnormal splicing/editing of SMC myosin heavy chain and smooth muscle α-actin pre-mRNAs occurred during injury-induced vascular remodeling. Importantly, heterozygous knockout of ADAR1 dramatically inhibited injury-induced neointima formation and restored SMC marker expression, demonstrating a critical role of ADAR1 in SMC phenotypic modulation and vascular remodeling in vivo. Our results unraveled a novel molecular mechanism, that is, pre-mRNA editing, governing SMC phenotypic modulation. © 2016 American Heart Association, Inc.

  8. [Comparison of electrophysiological properties of vascular smooth muscle cells in different arterioles in guinea pig].

    PubMed

    Ma, Ke-Tao; Li, Xin-Zhi; Li, Li; Zhang, Zhi-Ping; Zhao, Lei; Zhu, He; Si, Jun-Qiang

    2010-10-25

    Arterioles are major contributors to the control of systemic blood pressure and local blood flow. In this study, we compared electrophysiological properties of vascular smooth muscle cells (VSMCs) in anterior inferior cerebellar artery (AICA), mesenteric artery (MA) and spiral modiolar artery (SMA) by intracellular microelectrode recording and whole-cell patch clamp recording techniques. Results were shown as below: (1) Intracellular microelectrode recordings were made from VSMCs in AICA, MA and SMA with resting potentials of (-68±1.8) (n=65), (-71±2.4) (n=80) and (-66±2.9) mV (n=58), respectively. There was no significant difference in resting potentials among arterioles. (2) The membrane capacitance and membrane conductance in situ cells were much larger than those in dispersed smooth muscle cells by whole-cell recording techniques, and there was significant difference among arterioles, which were in the order: MA>AICA>SMA. After application of gap junction blocker 2-APB (100 μmol/L), the membrane capacitance and membrane conductance in situ cells were very close with those in single smooth muscle cells. (3) The I/V relation of whole-cell current of dissociated smooth muscle cells (AICA, MA and SMA) showed a prominent outward rectification, and the currents were substantially inhibited by 1 mmol/L 4-AP or 10 mmol/L TEA. When the command voltage was +40 mV, the current densities of VSMCs in AICA, MA and SMA were (26±2.0), (24±1.7) and (18±1.3) pA/pF respectively. SMA showed significant difference in the current density from AICA and MA respectively. These results suggest that the electrophysiological properties of coupling strength of gap junction and current density of smooth muscle cells are different among arterioles in the guinea pig.

  9. Testosterone induces hyporesponsiveness by interfering with IP3receptors in guinea pig airway smooth muscle.

    PubMed

    Montaño, Luis M; Flores-Soto, Edgar; Reyes-García, Jorge; Díaz-Hernández, Verónica; Carbajal-García, Abril; Campuzano-González, Elías; Ramírez-Salinas, G Lizbeth; Velasco-Velázquez, Marco A; Sommer, Bettina

    2017-12-21

    Asthma symptoms have been associated with sex steroids. During childhood, this illness seems more frequent in boys than in girls and this tendency reverts in puberty when it is more severe in women. Testosterone (TES), at supraphysiological concentrations, relaxed pre-contracted airway smooth muscle, but its effects at physiological concentrations have not been thoroughly studied. We explored this possibility in guinea pig tracheal smooth muscle. In myocytes TES (10 nM) abolished carbachol (CCh)-induced intracellular Ca 2+ concentration ([Ca 2+ ]i) increment. Ca 2+ responses to ATP were partially modified by TES while histamine's were not. These results indicate that inositol 1,4,5-trisphosphate (IP 3 ) signaling pathway might be involved. Photolysis of caged-IP 3 increased [Ca 2+ ]i and TES abolished this effect. TES diminished reactivity of the smooth muscle to CCh and this effect was non-genomic since it was unchanged by flutamide. In tracheal smooth muscle, mRNA for each IP 3 receptor (ITPR) isoform was found and, by immunofluorescence, ITPR1 and ITPR3 seems to be the main isoforms observed while ITPR2 was less prominent. Comparing the amino acid sequence of ITPR1 and the sequence of the TES binding site on the androgen receptor, we found that they share a short sequence. This domain could be responsible for the TES binding to the ITPR1 and probably for its blocking effect. We conclude that TES modifies ITPR1 function in airway smooth muscle, turning this tissue less reactive to contractile agonists that act through PLCβ-IP 3 signaling cascade. These results might be related to the low asthma prevalence in males from puberty to adulthood. Copyright © 2017 Elsevier B.V. All rights reserved.

  10. Electrical properties of purinergic transmission in smooth muscle of the guinea-pig prostate.

    PubMed

    Lam, Michelle; Mitsui, Retsu; Hashitani, Hikaru

    2016-01-01

    Prostatic smooth muscle develops spontaneous myogenic tone which is modulated by autonomic neuromuscular transmission. This study aimed to investigate the role of purinergic transmission in regulating electrical activity of prostate smooth muscle and whether its contribution may be altered with age. Intracellular recordings were simultaneously made with isometric tension recordings in smooth muscle preparations of the guinea-pig prostate. Immunostaining for P2X1 receptors on whole mount preparations was also performed. In prostate preparations which generated spontaneous slow waves, electrical field stimulation (EFS)-evoked excitatory junction potentials (EJPs) which were abolished by guanethidine (10 μM), α-β-methylene ATP (10 μM) or pyridoxal phosphate-6-azophenyl-2,4-disulfonic acid (PPADS, 10 μM) but not phentolamine (1 μM). Consistently, immunostaining revealed the expression of P2X1 receptors on prostatic smooth muscle. EJPs themselves did not cause contractions, but EJPs could sum to trigger a slow wave and associated contraction. Yohimbine (1 μM) and 3,7-dimethyl-1-propargylxanthine (DMPX, 10 μM) but not propranolol (1 μM) potentiated EJPs. Although properties of EJPs were not different between young and aging guinea-pig prostates, ectoATPase inhibitor ARL 67156 (100 μM) augmented EJP amplitudes by 64.2 ± 29.6% in aging animals, compared to 22.1 ± 19.9% in young animals. These results suggest that ATP released from sympathetic nerves acts on P2X1 purinoceptors located on prostate smooth muscle to evoke EJPs, while pre-junctional α2-adrenergic and adenosine A2 receptors may play a role in preventing excessive transmitter release. Age-related up-regulation of enzymatic ATP breakdown may be a compensatory mechanism for the enhanced purinergic transmission which would cause hypercontractility arising from increased ATP release in older animals. Copyright © 2015 Elsevier B.V. All rights reserved.

  11. Titanium Dioxide Modulation of the Contractibility of Visceral Smooth Muscles In Vivo

    NASA Astrophysics Data System (ADS)

    Tsymbalyuk, Olga V.; Naumenko, Anna M.; Rohovtsov, Oleksandr O.; Skoryk, Mykola A.; Voiteshenko, Ivan S.; Skryshevsky, Valeriy A.; Davydovska, Tamara L.

    2017-02-01

    Electronic scanning microscopy was used in the work to obtain the image and to identify the sizes of titanium dioxide (TiO2) nanoparticles 21 ± 5 nm. The qualitative and quantitative elemental analysis of the preparations of the caecum, antrum, myometrium, kidneys, and lungs of the rats, burdened with titanium dioxide, was also performed. It was established using the tenzometric method in the isometric mode that the accumulation of titanium dioxide in smooth muscles of the caecum resulted in the considerable, compared to the control, increase in the frequency of their spontaneous contractions, the decrease in the duration of the contraction-relaxation cycle, and the decrease in the indices of muscle functioning efficiency (the index of contractions in Montevideo units (MU) and the index of contractions in Alexandria units (AU)). In the same experimental conditions, there was not the increase, but the decrease in the frequency of spontaneous contractions, the duration of the contraction-relaxation cycle, and the increase in MU and AU indices in the smooth muscles of myometrium (in the group of rats, burdened with TiO2 for 30 days). It was also determined that TiO2 modulates both the mechanisms of the input of extracellular Ca2+ ions and the mechanisms of decreasing the concentration of these cations in smooth muscle cells of the caecum during the generation of the high potassium contraction. In these conditions, there is a considerable increase in the normalized maximal velocity of the contraction phase and the relaxation phase. It was demonstrated in the work that titanium dioxide also changes the cholinergic excitation in these muscles. The impact of titanium dioxide in the group of rats, burdened with TiO2, was accompanied with a considerable impairment of the kinetics of forming the tonic component of the oxytocin-induced contraction of the smooth muscles of myometrium.

  12. Myosin light chain kinase is central to smooth muscle contraction and required for gastrointestinal motility in mice.

    PubMed

    He, Wei-Qi; Peng, Ya-Jing; Zhang, Wen-Cheng; Lv, Ning; Tang, Jing; Chen, Chen; Zhang, Cheng-Hai; Gao, Song; Chen, Hua-Qun; Zhi, Gang; Feil, Robert; Kamm, Kristine E; Stull, James T; Gao, Xiang; Zhu, Min-Sheng

    2008-08-01

    Smooth muscle is essential for maintaining homeostasis for many body functions and provides adaptive responses to stresses imposed by pathologic disorders. Identified cell signaling networks have defined many potential mechanisms for initiating smooth muscle contraction with or without myosin regulatory light chain (RLC) phosphorylation by myosin light chain kinase (MLCK). We generated tamoxifen-inducible and smooth muscle-specific MLCK knockout (KO) mice and provide direct loss-of-function evidence that shows the primary importance of MLCK in phasic smooth muscle contractions. We used the Cre-loxP system to establish Mlck floxed mice in which exons 23, 24, and 25 were flanked by 2 loxP sites. Smooth muscle-specific MLCK KO mice were generated by crossing Mlck floxed mice with SM-CreER(T2) (ki) mice followed by tamoxifen treatment. The phenotype was assessed by histologic, biochemical, molecular, cell biological, and physiologic analyses. Targeted deletion of MLCK in adult mouse smooth muscle resulted in severe gut dysmotility characterized by weak peristalsis, dilation of the digestive tract, and reduction of feces excretion and food intake. There was also abnormal urinary bladder function and lower blood pressure. Isolated muscles showed a loss of RLC phosphorylation and force development induced by K(+)-depolarization. The kinase knockout also markedly reduced RLC phosphorylation and force development with acetylcholine which activates Ca(2+)-sensitizing signaling pathways. MLCK and its phosphorylation of RLC are required physiologically for smooth muscle contraction and are essential for normal gastrointestinal motility.

  13. Ranolazine inhibits voltage-gated mechanosensitive sodium channels in human colon circular smooth muscle cells.

    PubMed

    Neshatian, Leila; Strege, Peter R; Rhee, Poong-Lyul; Kraichely, Robert E; Mazzone, Amelia; Bernard, Cheryl E; Cima, Robert R; Larson, David W; Dozois, Eric J; Kline, Crystal F; Mohler, Peter J; Beyder, Arthur; Farrugia, Gianrico

    2015-09-15

    Human jejunum smooth muscle cells (SMCs) and interstitial cells of Cajal (ICCs) express the SCN5A-encoded voltage-gated, mechanosensitive sodium channel NaV1.5. NaV1.5 contributes to small bowel excitability, and NaV1.5 inhibitor ranolazine produces constipation by an unknown mechanism. We aimed to determine the presence and molecular identity of Na(+) current in the human colon smooth muscle and to examine the effects of ranolazine on Na(+) current, mechanosensitivity, and smooth muscle contractility. Inward currents were recorded by whole cell voltage clamp from freshly dissociated human colon SMCs at rest and with shear stress. SCN5A mRNA and NaV1.5 protein were examined by RT-PCR and Western blots, respectively. Ascending human colon strip contractility was examined in a muscle bath preparation. SCN5A mRNA and NaV1.5 protein were identified in human colon circular muscle. Freshly dissociated human colon SMCs had Na(+) currents (-1.36 ± 0.36 pA/pF), shear stress increased Na(+) peaks by 17.8 ± 1.8% and accelerated the time to peak activation by 0.7 ± 0.3 ms. Ranolazine (50 μM) blocked peak Na(+) current by 43.2 ± 9.3% and inhibited shear sensitivity by 25.2 ± 3.2%. In human ascending colon strips, ranolazine decreased resting tension (31%), reduced the frequency of spontaneous events (68%), and decreased the response to smooth muscle electrical field stimulation (61%). In conclusion, SCN5A-encoded NaV1.5 is found in human colonic circular smooth muscle. Ranolazine blocks both peak amplitude and mechanosensitivity of Na(+) current in human colon SMCs and decreases contractility of human colon muscle strips. Our data provide a likely mechanistic explanation for constipation induced by ranolazine. Copyright © 2015 the American Physiological Society.

  14. A new level of plasticity: Drosophila smooth-like testes muscles compensate failure of myoblast fusion

    PubMed Central

    Kuckwa, Jessica; Fritzen, Katharina; Buttgereit, Detlev; Rothenbusch-Fender, Silke; Renkawitz-Pohl, Renate

    2016-01-01

    The testis of Drosophila resembles an individual testis tubule of mammals. Both are surrounded by a sheath of smooth muscles, which in Drosophila are multinuclear and originate from a pool of myoblasts that are set aside in the embryo and accumulate on the genital disc later in development. These muscle stem cells start to differentiate early during metamorphosis and give rise to all muscles of the inner male reproductive system. Shortly before the genital disc and the developing testes connect, multinuclear nascent myotubes appear on the anterior tips of the seminal vesicles. Here, we show that adhesion molecules are distinctly localized on the seminal vesicles; founder cell (FC)-like myoblasts express Dumbfounded (Duf) and Roughest (Rst), and fusion-competent myoblast (FCM)-like cells mainly express Sticks and stones (Sns). The smooth but multinuclear myotubes of the testes arose by myoblast fusion. RNAi-mediated attenuation of Sns or both Duf and Rst severely reduced the number of nuclei in the testes muscles. Duf and Rst probably act independently in this context. Despite reduced fusion in all of these RNAi-treated animals, myotubes migrated onto the testes, testes were shaped and coiled, muscle filaments were arranged as in the wild type and spermatogenesis proceeded normally. Hence, the testes muscles compensate for fusion defects so that the myofibres encircling the adult testes are indistinguishable from those of the wild type and male fertility is guaranteed. PMID:26657767

  15. α-smooth muscle actin is not a marker of fibrogenic cell activity in skeletal muscle fibrosis.

    PubMed

    Zhao, Wanming; Wang, Xingyu; Sun, Kai-Hui; Zhou, Lan

    2018-01-01

    α-Smooth muscle actin (α-SMA) is used as a marker for a subset of activated fibrogenic cells, myofibroblasts, which are regarded as important effector cells of tissue fibrogenesis. We address whether α-SMA-expressing myofibroblasts are detectable in fibrotic muscles of mdx5cv mice, a mouse model for Duchenne muscular dystrophy (DMD), and whether the α-SMA expression correlates with the fibrogenic function of intramuscular fibrogenic cells. α-SMA immunostaining signal was not detected in collagen I (GFP)-expressing cells in fibrotic muscles of ColI-GFP/mdx5cv mice, but it was readily detected in smooth muscle cells lining intramuscular blood vessel walls. α-SMA expression was detected by quantitative RT-PCR and Western blot in fibrogenic cells sorted from diaphragm and quadriceps muscles of the ColI-GFP/mdx5cv mice. Consistent with the more severe fibrosis in the ColI-GFP/mdx5cv diaphragm, the fibrogenic cells in the diaphragm exerted a stronger fibrogenic function than the fibrogenic cells in the quadriceps as gauged by their extracellular matrix gene expression. However, both gene and protein expression of α-SMA was lower in the diaphragm fibrogenic cells than in the quadriceps fibrogenic cells in the ColI-GFP/mdx5cv mice. We conclude that myofibroblasts are present in fibrotic skeletal muscles, but their expression of α-SMA is not detectable by immunostaining. The level of α-SMA expression by intramuscular fibrogenic cells does not correlate positively with the level of collagen gene expression or the severity of skeletal muscle fibrosis in the mdx5cv mice. α-SMA is not a functional marker of fibrogenic cells in skeletal muscle fibrosis associated with muscular dystrophy.

  16. Hypotension due to Kir6.1 gain-of-function in vascular smooth muscle.

    PubMed

    Li, Anlong; Knutsen, Russell H; Zhang, Haixia; Osei-Owusu, Patrick; Moreno-Dominguez, Alex; Harter, Theresa M; Uchida, Keita; Remedi, Maria S; Dietrich, Hans H; Bernal-Mizrachi, Carlos; Blumer, Kendall J; Mecham, Robert P; Koster, Joseph C; Nichols, Colin G

    2013-08-23

    KATP channels, assembled from pore-forming (Kir6.1 or Kir6.2) and regulatory (SUR1 or SUR2) subunits, link metabolism to excitability. Loss of Kir6.2 results in hypoglycemia and hyperinsulinemia, whereas loss of Kir6.1 causes Prinzmetal angina-like symptoms in mice. Conversely, overactivity of Kir6.2 induces neonatal diabetes in mice and humans, but consequences of Kir6.1 overactivity are unknown. We generated transgenic mice expressing wild-type (WT), ATP-insensitive Kir6.1 [Gly343Asp] (GD), and ATP-insensitive Kir6.1 [Gly343Asp,Gln53Arg] (GD-QR) subunits, under Cre-recombinase control. Expression was induced in smooth muscle cells by crossing with smooth muscle myosin heavy chain promoter-driven tamoxifen-inducible Cre-recombinase (SMMHC-Cre-ER) mice. Three weeks after tamoxifen induction, we assessed blood pressure in anesthetized and conscious animals, as well as contractility of mesenteric artery smooth muscle and KATP currents in isolated mesenteric artery myocytes. Both systolic and diastolic blood pressures were significantly reduced in GD and GD-QR mice but normal in mice expressing the WT transgene and elevated in Kir6.1 knockout mice as well as in mice expressing dominant-negative Kir6.1 [AAA] in smooth muscle. Contractile response of isolated GD-QR mesenteric arteries was blunted relative to WT controls, but nitroprusside relaxation was unaffected. Basal KATP conductance and pinacidil-activated conductance were elevated in GD but not in WT myocytes. KATP overactivity in vascular muscle can lead directly to reduced vascular contractility and lower blood pressure. We predict that gain of vascular KATP function in humans would lead to a chronic vasodilatory phenotype, as indeed has recently been demonstrated in Cantu syndrome.

  17. Hypotension Due to Kir6.1 Gain‐of‐Function in Vascular Smooth Muscle

    PubMed Central

    Li, Anlong; Knutsen, Russell H.; Zhang, Haixia; Osei‐Owusu, Patrick; Moreno‐Dominguez, Alex; Harter, Theresa M.; Uchida, Keita; Remedi, Maria S.; Dietrich, Hans H.; Bernal‐Mizrachi, Carlos; Blumer, Kendall J.; Mecham, Robert P.; Koster, Joseph C.; Nichols, Colin G.

    2013-01-01

    Background KATP channels, assembled from pore‐forming (Kir6.1 or Kir6.2) and regulatory (SUR1 or SUR2) subunits, link metabolism to excitability. Loss of Kir6.2 results in hypoglycemia and hyperinsulinemia, whereas loss of Kir6.1 causes Prinzmetal angina–like symptoms in mice. Conversely, overactivity of Kir6.2 induces neonatal diabetes in mice and humans, but consequences of Kir6.1 overactivity are unknown. Methods and Results We generated transgenic mice expressing wild‐type (WT), ATP‐insensitive Kir6.1 [Gly343Asp] (GD), and ATP‐insensitive Kir6.1 [Gly343Asp,Gln53Arg] (GD‐QR) subunits, under Cre‐recombinase control. Expression was induced in smooth muscle cells by crossing with smooth muscle myosin heavy chain promoter–driven tamoxifen‐inducible Cre‐recombinase (SMMHC‐Cre‐ER) mice. Three weeks after tamoxifen induction, we assessed blood pressure in anesthetized and conscious animals, as well as contractility of mesenteric artery smooth muscle and KATP currents in isolated mesenteric artery myocytes. Both systolic and diastolic blood pressures were significantly reduced in GD and GD‐QR mice but normal in mice expressing the WT transgene and elevated in Kir6.1 knockout mice as well as in mice expressing dominant‐negative Kir6.1 [AAA] in smooth muscle. Contractile response of isolated GD‐QR mesenteric arteries was blunted relative to WT controls, but nitroprusside relaxation was unaffected. Basal KATP conductance and pinacidil‐activated conductance were elevated in GD but not in WT myocytes. Conclusions KATP overactivity in vascular muscle can lead directly to reduced vascular contractility and lower blood pressure. We predict that gain of vascular KATP function in humans would lead to a chronic vasodilatory phenotype, as indeed has recently been demonstrated in Cantu syndrome. PMID:23974906

  18. Investigating the role of smooth muscle cells in large elastic arteries: a finite element analysis.

    PubMed

    Murtada, Sae-Il; Holzapfel, Gerhard A

    2014-10-07

    Physiological loading in large elastic arteries is considered to be mainly carried by the passive components of the media but it is not known how much the contraction of the smooth muscle cells is actually involved in the load carrying. Smooth muscle contraction is considered to occur in a relatively slow time domain but the contraction is able to produce significant tension. In the present work the role of smooth muscle contraction in large elastic arteries is investigated by analyzing how changes in the intracellular calcium, and thereby the active tone of smooth muscle cells, influence the deformation and stress behavior; different intracellular calcium functions and medial wall thicknesses with cycling internal pressure are studied. In particular, a recently proposed mechanochemical model (Murtada et al., 2012. J. Theor. Biol. 297, 176-186), which links intracellular calcium with mechanical contraction and an anisotropic model representing the elastin/collagen composite, was implemented into a 3D finite element framework. Details of the implementation procedure are described and a verification of the model implementation is provided by means of the isometric contraction/relaxation analysis of a medial strip at optimal muscle length. In addition, numerically obtained pressure-radius relationships of arterial rings modeled with one and two layers are analyzed with different geometries and at different calcium levels; a comparison with the Laplace equation is provided. Finally, a two-layer arterial ring is loaded with a realistic pressure wave and with various intracellular calcium functions (different amplitudes and mean values) and medial wall thicknesses; residual stresses are considered. The finite element results show that changes in the calcium amplitudes hardly have an influence on the current inner ring radius and the circumferential stress. However, an increase in the mean intracellular calcium value and the medial wall thickness leads to a clear

  19. Smooth muscle myosin light chain kinase efficiently phosphorylates serine 15 of cardiac myosin regulatory light chain

    SciTech Connect

    Josephson, Matthew P.; Sikkink, Laura A.; Penheiter, Alan R.

    2011-12-16

    Highlights: Black-Right-Pointing-Pointer Cardiac myosin regulatory light chain (MYL2) is phosphorylated at S15. Black-Right-Pointing-Pointer Smooth muscle myosin light chain kinase (smMLCK) is a ubiquitous kinase. Black-Right-Pointing-Pointer It is a widely believed that MYL2 is a poor substrate for smMLCK. Black-Right-Pointing-Pointer In fact, smMLCK efficiently and rapidly phosphorylates S15 in MYL2. Black-Right-Pointing-Pointer Phosphorylation kinetics measured by novel fluorescence method without radioactivity. -- Abstract: Specific phosphorylation of the human ventricular cardiac myosin regulatory light chain (MYL2) modifies the protein at S15. This modification affects MYL2 secondary structure and modulates the Ca{sup 2+} sensitivity of contraction in cardiac tissue. Smooth muscle myosin light chainmore » kinase (smMLCK) is a ubiquitous kinase prevalent in uterus and present in other contracting tissues including cardiac muscle. The recombinant 130 kDa (short) smMLCK phosphorylated S15 in MYL2 in vitro. Specific modification of S15 was verified using the direct detection of the phospho group on S15 with mass spectrometry. SmMLCK also specifically phosphorylated myosin regulatory light chain S15 in porcine ventricular myosin and chicken gizzard smooth muscle myosin (S20 in smooth muscle) but failed to phosphorylate the myosin regulatory light chain in rabbit skeletal myosin. Phosphorylation kinetics, measured using a novel fluorescence method eliminating the use of radioactive isotopes, indicates similar Michaelis-Menten V{sub max} and K{sub M} for regulatory light chain S15 phosphorylation rates in MYL2, porcine ventricular myosin, and chicken gizzard myosin. These data demonstrate that smMLCK is a specific and efficient kinase for the in vitro phosphorylation of MYL2, cardiac, and smooth muscle myosin. Whether smMLCK plays a role in cardiac muscle regulation or response to a disease causing stimulus is unclear but it should be considered a potentially

  20. Increased IGF-IEc expression and mechano-growth factor production in intestinal muscle of fibrostenotic Crohn's disease and smooth muscle hypertrophy.

    PubMed

    Li, Chao; Vu, Kent; Hazelgrove, Krystina; Kuemmerle, John F

    2015-12-01

    The igf1 gene is alternatively spliced as IGF-IEa and IGF-IEc variants in humans. In fibrostenotic Crohn's disease, the fibrogenic cytokine TGF-β1 induces IGF-IEa expression and IGF-I production in intestinal smooth muscle and results in muscle hyperplasia and collagen I production that contribute to stricture formation. Mechano-growth factor (MGF) derived from IGF-IEc induces skeletal and cardiac muscle hypertrophy following stress. We hypothesized that increased IGF-IEc expression and MGF production mediated smooth muscle hypertrophy also characteristic of fibrostenotic Crohn's disease. IGF-IEc transcripts and MGF protein were increased in muscle cells isolated from fibrostenotic intestine under regulation by endogenous TGF-β1. Erk5 and MEF2C were phosphorylated in vivo in fibrostenotic muscle; both were phosphorylated and colocalized to nucleus in response to synthetic MGF in vitro. Smooth muscle-specific protein expression of α-smooth muscle actin, γ-smooth muscle actin, and smoothelin was increased in affected intestine. Erk5 inhibition or MEF2C siRNA blocked smooth muscle-specific gene expression and hypertrophy induced by synthetic MGF. Conditioned media of cultured fibrostenotic muscle induced muscle hypertrophy that was inhibited by immunoneutralization of endogenous MGF or pro-IGF-IEc. The results indicate that TGF-β1-dependent IGF-IEc expression and MGF production in patients with fibrostenotic Crohn's disease regulates smooth muscle cell hypertrophy a critical factor that contributes to intestinal stricture formation. Copyright © 2015 the American Physiological Society.

  1. Chitosan-based scaffolds for the support of smooth muscle constructs in intestinal tissue engineering

    PubMed Central

    Zakhem, Elie; Raghavan, Shreya; Gilmont, Robert R; Bitar, Khalil N

    2012-01-01

    Intestinal tissue engineering is an emerging field due to a growing demand for intestinal lengthening and replacement procedures secondary to massive resections of the bowel. Here, we demonstrate the potential use of a chitosan/collagen scaffold as a 3D matrix to support the bioengineered circular muscle constructs maintain their physiological functionality. We investigated the biocompatibility of chitosan by growing rabbit colonic circular smooth muscle cells (RCSMCs) on chitosan-coated plates. The cells maintained their spindle-like morphology and preserved their smooth muscle phenotypic markers. We manufactured tubular scaffolds with central openings composed of chitosan and collagen in a 1:1 ratio. Concentrically-aligned 3D circular muscle constructs were bioengineered using fibrin-based hydrogel seeded with RCSMCs. The constructs were placed around the scaffold for 2 weeks, after which they were taken off and tested for their physiological functionality. The muscle constructs contracted in response to Acetylcholine (Ach) and potassium chloride (KCl) and they relaxed in response to vasoactive intestinal peptide (VIP). These results demonstrate that chitosan is a biomaterial possibly suitable for intestinal tissue engineering applications. PMID:22483012

  2. Airway smooth muscle responsiveness from dogs with airway hyperresponsiveness after O/sub 3/ inhalation

    SciTech Connect

    Jones, G.L.; O'Byrne, P.M.; Pashley, M.

    1988-07-01

    Airway hyperresponsiveness occurs after inhalation of O3 in dogs. The purpose of this study was to examine the responsiveness of trachealis smooth muscle in vitro to electrical field stimulation, exogenous acetylcholine, and potassium chloride from dogs with airway hyperresponsiveness after inhaled O3 in vivo and to compare this with the responsiveness of trachealis muscle from control dogs. In addition, excitatory junction potentials were measured with the use of single and double sucrose gap techniques in both groups of dogs to determine whether inhaled O3 affects the release of acetylcholine from parasympathetic nerves in trachealis muscle. Airway hyperresponsiveness developed in allmore » dogs after inhaled O3 (3 ppm for 30 min). The acetylcholine provocative concentration decreased from 4.11 mg/ml before O3 inhalation to 0.66 mg/ml after O3 (P less than 0.0001). The acetylcholine provocative concentration increased slightly after control inhalation of dry room air. Airway smooth muscle showed increased responses to both electrical field stimulation and exogenous acetylcholine but not to potassium chloride in preparations from dogs with airway hyperresponsiveness in vivo. The increased response to electrical field stimulation was not associated with a change in excitatory junctional potentials. These results suggest that a postjunctional alteration in trachealis muscle function occurs after inhaled O3 in dogs, which may account for airway hyperresponsiveness after O3 in vivo.« less

  3. Telocytes in skeletal, cardiac and smooth muscle interstitium: morphological and functional aspects.

    PubMed

    Marini, Mirca; Rosa, Irene; Ibba-Manneschi, Lidia; Manetti, Mirko

    2018-04-25

    Telocytes (TCs) represent a new distinct type of cells found in the stromal compartment of many organs, including the skeletal, cardiac and smooth muscles. TCs are morphologically defined as interstitial cells with a small cellular body from which arise very long (up to hundreds of micrometers) and thin moniliform processes (named telopodes) featuring the alternation of slender segments (called podomers) and small dilated portions (called podoms) accommodating some organelles. Although these stromal cells are mainly characterized by their ultrastructural traits, in the last few years TCs have been increasingly studied for their immunophenotypes, microRNA profiles, and gene expression and proteomic signatures. By their long-distance spreading telopodes, TCs build a three-dimensional network throughout the whole stromal space and communicate with each other and neighboring cells through homocellular and heterocellular junctions, respectively. Moreover, increasing evidence suggests that TCs may exert paracrine functions being able to transfer genetic information and signaling molecules to other cells via the release of different types of extracellular vesicles. A close relationship between TCs and stem/progenitor cell niches has also been described in several organs. However, the specific functions of TCs located in the muscle interstitium remain to be unraveled. Here, we review the morphological and possible functional aspects of TCs in skeletal, cardiac and smooth muscle tissues. The potential involvement of TCs in muscle tissue pathological changes and future possibilities for targeting TCs as a novel promising therapeutic strategy to foster muscle tissue regeneration and repair are also discussed.

  4. The relaxant effect of Nigella sativa on smooth muscles, its possible mechanisms and clinical applications

    PubMed Central

    Keyhanmanesh, Rana; Gholamnezhad, Zahra; Boskabady, Mohammad Hossien

    2014-01-01

    Nigella sativa (N. sativa) is a spice plant which has been traditionally used for culinary and medicinal purposes. Different therapeutic properties including the beneficial effects on asthma and dyspnea, digestive and gynecology disorders have been described for the seeds of N. sativa. There is evidence of the relaxant effects of this plant and some of its constituents on different types of smooth muscle including rabbit aorta, rabbit jejunum and trachea. The relaxant effect of N. sativa could be of therapeutic importance such as bronchodilation in asthma, vasodilation in hypertension and therapeutic effect on digestive or urogenital disorders. Therefore in the present article, the relaxant effects of N. sativa and its constituents on smooth muscles and its possible mechanisms as well as clinical application of this effect were reviewed. PMID:25859297

  5. Effects of Gymnodinium breve toxin on the smooth muscle preparation of guinea-pig ileum

    PubMed Central

    Grunfeld, Y.; Spiegelstein, M.Y.

    1974-01-01

    1 The effects of Gymnodinium breve neurotoxin (GT) on smooth muscles were studied using the guinea-pig isolated ileum. 2 The toxin caused strong spasmogenic effects at 1-4 μg/ml, characterized by prolonged tonic contraction with superimposed pronounced pendular movements. Tachyphylaxis was observed upon administration of successive doses. 3 Atropine blocked the contractile response elicited by GT, whereas mepyramine and hexamethonium failed to do so. These findings tentatively suggested a cholinergic involvement at a post-ganglionic site of action. 4 In the presence of tetrodotoxin the effects of GT were abolished, excluding direct action of the toxin on the smooth muscle. 5 It is concluded that GT exerts its spasmogenic effects through stimulation of the post-ganglionic cholinergic nerve fibres. PMID:4155337

  6. [Physiological and pathophysiological meanings of gastrointestinal smooth muscle motor unit SIP syncytium].

    PubMed

    Song, Ni-Na; Xu, Wen-Xie

    2016-10-25

    Gastrointestinal smooth muscle layer contains two kinds of interstitial cells with special differentiation, i.e., interstitial cells of Cajal (ICC) and platelet-derived growth factor receptor α-positive (PDGFRα + ) cells. The ICC and PDGFRα + cells contact with smooth muscle cells (SMCs) by gap junctions and regulate contractive function of the SMCs. Therefore, these three kinds of cells constitute a functional syncytium, i.e., the SMC, ICC and PDGFRα + cells syncytium (SIP syncytium). Various neurotransmitters, humoral factors, endogenous bioactive molecules, as well as drugs regulate gastrointestinal motility through the SIP syncytium. In this review, we introduce the concept of SIP syncytium and summarize functions of the syncytium, as well as its physiological and pathological significances.

  7. Characterization of bacterial artificial chromosome transgenic mice expressing mCherry fluorescent protein substituted for the murine smooth muscle-alpha-actin gene

    USDA-ARS?s Scientific Manuscript database

    Smooth muscle a actin (SMA) is a cytoskeletal protein expressed by mesenchymal and smooth muscle cell types, including mural cells(vascular smooth muscle cells and pericytes). Using Bacterial Artificial Chromosome (BAC) recombineering technology, we generated transgenic reporter mice that express a ...

  8. Inhibitory mechanism of xestospongin-C on contraction and ion channels in the intestinal smooth muscle.

    PubMed

    Ozaki, Hiroshi; Hori, Masatoshi; Kim, Yoon-Sun; Kwon, Seong-Chun; Ahn, Duck-Sun; Nakazawa, Hiroshi; Kobayashi, Motomasa; Karaki, Hideaki

    2002-12-01

    1. Xestospongin-C isolated from a marine sponge, Xestospongia sp., has recently been shown to be a membrane-permeable IP(3) receptor inhibitor. In this study we examined the effects of this compound on smooth muscle from guinea-pig ileum. 2. In guinea-pig ileum permeabilized with alpha-toxin, xestospongin-C (3 microM) inhibited contractions induced by Ca(2+) mobilized from sarcoplasmic reticulum (SR) with IP(3) or carbachol with GTP, but not with caffeine. 3. In intact smooth muscle tissue, xestospongin-C (3-10 microM) inhibited carbachol- and high-K+-induced increases in [Ca(2+)](i) and contractions at sustained phase. 4. It also inhibited voltage-dependent inward Ba(2+) currents in a concentration-dependent manner with an IC(50) of 0.63 microM. Xestospongin-C (3-10 microM) had no effect on carbachol-induced inward Ca(2+) currents via non-selective cation channels; but it did reduce voltage-dependent K+ currents in a concentration-dependent manner with an IC(50) of 0.13 microM. 5. These results suggest that xestospongin-C inhibits the IP(3) receptor but not the ryanodine receptor in smooth muscle SR membrane. In intact smooth muscle cells, however, xestospongin-C appears to inhibit voltage-dependent Ca(2+) and K+ currents at a concentration range similar to that at which it inhibits the IP(3) receptor. Xestospongin-C is a selective blocker of the IP(3) receptor in permeabilised cells but not in cells with intact plasma membrane.

  9. [Relaxation mechanism of smooth muscle cells and its relationship with penile erection].

    PubMed

    Zhao, Hu; Jiang, Rui

    2016-09-01

    The contractile and diastolic function of smooth muscle cells (SMCs) is closely related to penile erection and erectile dysfunction (ED). In addition to nitric oxide (NO), carbon monoxide (CO), and hydrogen sulfide (H2S), sulfur dioxide (SO2), estrogen receptor (ER), P2Y receptor, perivascular tissue (PVT), and calcium activated potassium channel (Kca) are found to be involved in the relaxation of SMCs. This review updates the mechanisms of the relaxation of SMCs and its relationship with ED.

  10. Rho-kinase inhibitors augment the inhibitory effect of propofol on rat bronchial smooth muscle contraction.

    PubMed

    Hanazaki, Motohiko; Yokoyama, Masataka; Morita, Kiyoshi; Kohjitani, Atsushi; Sakai, Hiroyasu; Chiba, Yoshihiko; Misawa, Miwa

    2008-06-01

    Airway smooth muscle contraction is not caused by the increase in intracellular Ca(2+) ([Ca(2+)](i)) alone because agonist stimulation increases tension at the same [Ca(2+)](i) (increase in Ca(2+) sensitivity). The small G protein Rho A and Rho-kinase (ROCK) play important roles in the regulation of Ca(2+) sensitivity. In this study, we investigated the effects of three ROCK inhibitors (fasudil, Y-27632, and H-1152) on rat airway smooth muscle contraction and the effects of ROCK inhibitors on propofol-induced bronchodilatory effects. Ring strips from intrapulmonary bronchus of male Wistar rats were placed in 400-microL organ baths containing Krebs-Henseleit solution. After obtaining stable contraction with 30 microM acetylcholine, (1) propofol (1 microM-1 mM) was cumulatively applied; (2) cumulative doses of Y-27632 (0.01-300 microM), fasudil (0.01-100 microM), or H-1152 (0.01-100 microM) were applied; (3) propofol (1 microM-1 mM), with Y-27632, fasudil or H-1152 (0.03 microM or 0.1 microM), was cumulatively applied. (1) Propofol produced concentration-dependent relaxation of rat bronchial smooth muscle. (2) All ROCK inhibitors produced concentration-dependent relaxation. (3) 0.03 microM Y-27632 and fasudil had no significant effect on the concentration-response curve for propofol, while 0.1 microM of both agents significantly shifted concentration-response curves to the left and decreased EC(50). H-1152 (both 0.03 microM and 0.1 microM) significantly sifted the concentration-response curve for propofol to the left and decreased EC(50). ROCK inhibitors, especially H-1152, can attenuate the contraction of rat airway smooth muscle. The combined use of ROCK inhibitors and propofol causes greater relaxation.

  11. Neuropilin 1 Is Essential for Gastrointestinal Smooth Muscle Contractility and Motility in Aged Mice

    PubMed Central

    Yamaji, Maiko; Mahmoud, Marwa; Evans, Ian M.; Zachary, Ian C.

    2015-01-01

    Background and Aims Neuropilin 1 (NRP1) is a non-tyrosine kinase receptor for vascular endothelial growth factor (VEGF) and class 3 semaphorins, playing a role in angiogenesis and neuronal axon guidance, respectively. NRP1 is expressed in smooth muscle cells (SMC) but the functional role of NRP1 in SMC has not been elucidated. We therefore investigated the biological relevance of NRP1 in SMC in vivo by generating mice with SMC-specific Nrp1 deficiency. Methods Conditional gene targeting generated SMC-specific Nrp1 knockout mice (Nrp1SMKO) in which Cre recombinase is driven by the smooth muscle-specific myosin heavy chain (smMHC) promoter. Results SMC-specific Nrp1 deficiency resulted in a significant reduction in intestinal length by 6 months, and, by 18 months, in severe constipation, and enlargement of the intestine consistent with chronic intestinal pseudo-obstruction. These effects were associated with significant thinning of the intestinal smooth muscle, and decreased intestinal contractility. Expression of contractile proteins was reduced in Nrp1SMKO mice, including the smMHC isoform, SMB, whereas we observed a significant increase in the expression of the small-conductance calcium-activated potassium channel 3 (SK3/KCa2.3), implicated in negative regulation of smooth muscle contraction. Conclusions Nrp1 deficiency in visceral SMC results in adult-onset defects in gastrointestinal contractility and motility and causes a shift to a less contractile SMC phenotype. These findings indicate a new role for Nrp1 in the maintenance of the visceral SMC contractile phenotype required for normal GI motility in aged mice. PMID:25659123

  12. Neuropilin 1 is essential for gastrointestinal smooth muscle contractility and motility in aged mice.

    PubMed

    Yamaji, Maiko; Mahmoud, Marwa; Evans, Ian M; Zachary, Ian C

    2015-01-01

    Neuropilin 1 (NRP1) is a non-tyrosine kinase receptor for vascular endothelial growth factor (VEGF) and class 3 semaphorins, playing a role in angiogenesis and neuronal axon guidance, respectively. NRP1 is expressed in smooth muscle cells (SMC) but the functional role of NRP1 in SMC has not been elucidated. We therefore investigated the biological relevance of NRP1 in SMC in vivo by generating mice with SMC-specific Nrp1 deficiency. Conditional gene targeting generated SMC-specific Nrp1 knockout mice (Nrp1SMKO) in which Cre recombinase is driven by the smooth muscle-specific myosin heavy chain (smMHC) promoter. SMC-specific Nrp1 deficiency resulted in a significant reduction in intestinal length by 6 months, and, by 18 months, in severe constipation, and enlargement of the intestine consistent with chronic intestinal pseudo-obstruction. These effects were associated with significant thinning of the intestinal smooth muscle, and decreased intestinal contractility. Expression of contractile proteins was reduced in Nrp1SMKO mice, including the smMHC isoform, SMB, whereas we observed a significant increase in the expression of the small-conductance calcium-activated potassium channel 3 (SK3/KCa2.3), implicated in negative regulation of smooth muscle contraction. Nrp1 deficiency in visceral SMC results in adult-onset defects in gastrointestinal contractility and motility and causes a shift to a less contractile SMC phenotype. These findings indicate a new role for Nrp1 in the maintenance of the visceral SMC contractile phenotype required for normal GI motility in aged mice.

  13. Smooth muscle caldesmon modulates peristalsis in the wild type and non-innervated zebrafish intestine

    PubMed Central

    ABRAMS, J.; DAVULURI, G.; SEILER, C.; PACK, M.

    2013-01-01

    Background The high molecular weight isoform of the actin-binding protein Caldesmon (h-CaD) regulates smooth muscle contractile function by modulating cross-bridge cycling of myosin heads. The normal inhibitory activity of h-CaD is regulated by the enteric nervous system; however, the role of h-CaD during intestinal peristalsis has never been studied. Methods We identified a zebrafish paralog of the human CALD1 gene that encodes an h-CaD isoform expressed in intestinal smooth muscle. We examined the role of h-CaD during intestinal peristalsis in zebrafish larvae by knocking down the h-CaD protein using an antisense morpholino oligonucleotide. We also developed transgenic zebrafish that express inhibitory peptides derived from the h-CaD myosin and actin-binding domains, and examined their effect on peristalsis in wild-type zebrafish larvae and sox10colourless mutant larvae that lack enteric nerves. Key Results Genomic analyses identified two zebrafish Caldesmon paralogs. The cald1a ortholog encoded a high molecular weight isoform generated by alternative splicing whose intestinal expression was restricted to smooth muscle. Propulsive intestinal peristalsis was increased in wild-type zebrafish larvae by h-CaD knockdown and by expression of transgenes encoding inhibitory myosin and actin-binding domain peptides. Peristalsis in the non-innervated intestine of sox10colourless larvae was partially restored by h-CaD knockdown and expression of the myosin-binding peptide. Conclusions & Inferences Disruption of the normal inhibitory function of h-CaD enhances intestinal peristalsis in both wild-type zebrafish larvae and mutant larvae that lack enteric nerves, thus confirming a physiologic role for regulation of smooth muscle contraction at the actin filament. PMID:22316291

  14. Generalized smooth muscle hamartoma with multiple congenital anomalies without the "Michelin tire baby" phenotype.

    PubMed

    Janicke, Elise C; Nazareth, Michael R; Rothman, Ilene L

    2014-01-01

    We report a patient with generalized smooth muscle hamartoma who presented with many of the variety of congenital anomalies that have been reported in babies with multiple symmetric circumferential rings of folded skin known as Michelin tire baby (MTB) syndrome, but our patient did not show the MTB phenotype. This constellation of findings in the absence of the MTB phenotype has not been previously reported. © 2014 Wiley Periodicals, Inc.

  15. Altered cytoskeletal structure of smooth muscle cells in ureteropelvic junction obstruction.

    PubMed

    Cutroneo, Giuseppina; Arena, Salvatore; Anastasi, Giuseppe; Cervellione, Raimondo M; Grimaldi, Silvia; Di Mauro, Debora; Speciale, Francesco; Arena, Francesco; Di Benedetto, Vincenzo; Favaloro, Angelo; Magno, Carlo

    2011-06-01

    Ureteropelvic junction obstruction is one of the most common causes of hydronephrosis in children. A malfunction of smooth muscle cells is believed to be the underlying mechanism causing obstruction. We investigated the expression of some integrins, talin and β-dystroglycan, considered the main compound of smooth muscle cell cytoskeleton, and active caspase 3 at the level of the ureteropelvic junction obstruction. Specimens were obtained at pyeloplasty in 12 children with ureteropelvic junction obstruction. Six control specimens were obtained during organ explantation. Specimens were divided into renal pelvis, ureteropelvic junction and ureter below the obstruction. Western blot analysis of active caspase 3, and immunofluorescence and polymerase chain reaction analysis were performed for α7A, β1A, α7B and β1D integrins, talin and β-dystroglycan. Talin and β-dystroglycan were slightly impaired in ureteropelvic junction obstruction, while α7B and β1D integrins were severely reduced, and α7A, β1A and active caspase 3 were significantly enhanced compared to controls. We demonstrated activation of apoptosis and a critical alteration of cytoskeleton that might explain the altered function and the increased apoptosis in smooth muscle cells in ureteropelvic junction obstruction. The delayed rearrangement of the cytoskeleton of smooth muscle cells in ureteropelvic junction obstruction might be linked to a postnatal splicing from α7A and β1A to α7B and β1D integrins, respectively. This relationship could explain the common clinical scenario of spontaneous improvement of hydronephrosis in children with suspected ureteropelvic junction obstruction. Copyright © 2011 American Urological Association Education and Research, Inc. Published by Elsevier Inc. All rights reserved.

  16. Smooth muscle progenitor cells from peripheral blood promote the neovascularization of endothelial colony-forming cells

    SciTech Connect

    Joo, Hyung Joon; Seo, Ha-Rim; Jeong, Hyo Eun

    2014-07-11

    Highlights: • Two distinct vascular progenitor cells are induced from adult peripheral blood. • ECFCs induce vascular structures in vitro and in vivo. • SMPCs augment the in vitro and in vivo angiogenic potential of ECFCs. • Both cell types have synergistic therapeutic potential in ischemic hindlimb model. - Abstract: Proangiogenic cell therapy using autologous progenitors is a promising strategy for treating ischemic disease. Considering that neovascularization is a harmonized cellular process that involves both endothelial cells and vascular smooth muscle cells, peripheral blood-originating endothelial colony-forming cells (ECFCs) and smooth muscle progenitor cells (SMPCs), which are similar to mature endothelialmore » cells and vascular smooth muscle cells, could be attractive cellular candidates to achieve therapeutic neovascularization. We successfully induced populations of two different vascular progenitor cells (ECFCs and SMPCs) from adult peripheral blood. Both progenitor cell types expressed endothelial-specific or smooth muscle-specific genes and markers, respectively. In a protein array focused on angiogenic cytokines, SMPCs demonstrated significantly higher expression of bFGF, EGF, TIMP2, ENA78, and TIMP1 compared to ECFCs. Conditioned medium from SMPCs and co-culture with SMPCs revealed that SMPCs promoted cell proliferation, migration, and the in vitro angiogenesis of ECFCs. Finally, co-transplantation of ECFCs and SMPCs induced robust in vivo neovascularization, as well as improved blood perfusion and tissue repair, in a mouse ischemic hindlimb model. Taken together, we have provided the first evidence of a cell therapy strategy for therapeutic neovascularization using two different types of autologous progenitors (ECFCs and SMPCs) derived from adult peripheral blood.« less

  17. Tra2β as a novel mediator of vascular smooth muscle diversification

    PubMed Central

    Shukla, Supriya; Fisher, Steven A.

    2009-01-01

    Transformer splicing regulatory proteins determine the sexually dimorphic traits of Drosophila. The role of the vertebrate homologues of Tra-2 in phenotypic specification is undefined. We are using the alternative splicing of the MYPT1 E23 exon as a model for the study of smooth muscle diversification into fast and slow contractile phenotypes. Tra2β mRNA and protein is expressed at up to 10-fold higher levels in fast smooth muscle tissues such as the rat portal vein (PV) and small mesenteric artery (MA), in which E23 is spliced, as compared to the slow smooth muscle tissues of the large arteries and veins, in which E23 is skipped. Tra2β is up-regulated up to 10-fold concordant with the initiation of E23 splicing as the rat PV and avian gizzard implement the fast program of gene expression in the peri-natal period. In disease models such as portal hypertension and MA high/low flow, the PV and MA1 dynamically down-regulate Tra2β concordant with a shift to E23 skipping and the slow program of gene expression. Tra2β binds to a highly conserved sequence within E23 and trans-activates its splicing in vitro and in vivo; this is abolished with mutation or deletion of this sequence. RNAi mediated knock-down of Tra2β markedly reduces E23 splicing. We propose that Tra2β has been conserved through evolution and re-deployed for the specification of the fast smooth muscle phenotype, and may serve as a novel nodal point for the investigation of this process in developmental and disease models. PMID:18669920

  18. Bladder Smooth Muscle Cells Differentiation from Dental Pulp Stem Cells: Future Potential for Bladder Tissue Engineering.

    PubMed

    Song, Bing; Jiang, Wenkai; Alraies, Amr; Liu, Qian; Gudla, Vijay; Oni, Julia; Wei, Xiaoqing; Sloan, Alastair; Ni, Longxing; Agarwal, Meena

    2016-01-01

    Dental pulp stem cells (DPSCs) are multipotent cells capable of differentiating into multiple cell lines, thus providing an alternative source of cell for tissue engineering. Smooth muscle cell (SMC) regeneration is a crucial step in tissue engineering of the urinary bladder. It is known that DPSCs have the potential to differentiate into a smooth muscle phenotype in vitro with differentiation agents. However, most of these studies are focused on the vascular SMCs. The optimal approaches to induce human DPSCs to differentiate into bladder SMCs are still under investigation. We demonstrate in this study the ability of human DPSCs to differentiate into bladder SMCs in a growth environment containing bladder SMCs-conditioned medium with the addition of the transforming growth factor beta 1 (TGF-β1). After 14 days of exposure to this medium, the gene and protein expression of SMC-specific marker (α-SMA, desmin, and calponin) increased over time. In particular, myosin was present in differentiated cells after 11 days of induction, which indicated that the cells differentiated into the mature SMCs. These data suggested that human DPSCs could be used as an alternative and less invasive source of stem cells for smooth muscle regeneration, a technology that has applications for bladder tissue engineering.

  19. Synergistic interaction between PPAR ligands and salbutamol on human bronchial smooth muscle cell proliferation

    PubMed Central

    Fogli, S; Stefanelli, F; Picchianti, L; Del Re, M; Mey, V; Bardelli, C; Danesi, R; Breschi, MC

    2013-01-01

    Background and Purpose An important objective in asthma therapy is to prevent the accelerated growth of airway smooth muscle cells which leads to hyperplasia and bronchial hyperreactivity. We investigated the effect of combination of salbutamol and PPARγ agonists on growth factor-stimulated human bronchial smooth muscle cell (BSMC) proliferation. Experimental Approach Synergism was quantified by the combination index-isobologram method. Assays used here included analyses of growth inhibition, cell viability, DNA fragmentation, gene transcription, cell cycle and protein expression. Key Results The PPARγ gene was highly expressed in BSMC and the protein was identified in cell nuclei. Single-agent salbutamol or PPARγ agonists prevented growth factor-induced human BSMC proliferation within a micromolar range of concentrations through their specific receptor subtypes. Sub-micromolar levels of combined salbutamol-PPARγ agonist inhibited growth by 50% at concentrations from ∼2 to 12-fold lower than those required for each drug alone, without induction of apoptosis or necrosis. Combination treatments also promoted cell cycle arrest at the G1/S transition phase and inhibition of ERK phosphorylation. Conclusions and Implications The synergistic interaction between PPARγ agonists and β2-adrenoceptor agonists on airway smooth muscle cell proliferation highlights the anti-remodelling potential of this combination in chronic lung diseases. PMID:22924744

  20. Immunohistochemical characterization of endometriosis-associated smooth muscle cells in human peritoneal endometriotic lesions.

    PubMed

    Barcena de Arellano, Maria L; Gericke, Jessica; Reichelt, Uta; Okuducu, Ali Fuat; Ebert, Andreas D; Chiantera, Vito; Schneider, Achim; Mechsner, Sylvia

    2011-10-01

    Smooth muscle cells (SMC) are common components of endometriotic lesions. SMC have been characterized previously in peritoneal, ovarian and deep infiltrating endometriotic lesions and adenomyosis. The aim of this retrospective study was to investigate the extent of differentiation in endometriosis-associated SMC (EMaSMC) in peritoneal endometriotic lesions. We obtained biopsies from peritoneal endometriotic lesions (n = 60) and peritoneal sites distant from the endometriotic lesion (n = 60), as well as healthy peritoneum from patients without endometriosis (control tissue, n = 10). These controls were hysterectomy specimens from patients without endometriosis or adenomyosis. Histopathological examination of peritoneal specimens using antibodies against oxytocin receptor (OTR), vasopressin receptor (VPR), smooth muscle myosin heavy chain (SM-MHC), estrogen receptor (ER) or progesterone receptor (PR) was performed. To identify SMC and their level of differentiation, antibodies for smooth muscle actin desmin and caldesmon were used. SMC were detected in all endometriotic lesions. SMC were more abundant in unaffected peritoneum of women with endometriosis (38%) compared with women without endometriosis (6%; P < 0.0001). Depending on the level of differentiation, SMC stained for SM-MHC, OTR, VPR, ER and PR. OTR was only detected in fully differentiated SMC. Identification of OTR, VPR, ER and PR leads to the hypothesis that the EMaSMC might be functionally active and possibly involved in the generation of pain associated with endometriosis.

  1. mTOR pathway and Ca2+ stores mobilization in aged smooth muscle cells

    PubMed Central

    Martín-Cano, Francisco E; Camello-Almaraz, Cristina; Hernandez, David; Pozo, Maria J; Camello, Pedro J

    2013-01-01

    Aging is considered to be driven by the so called senescence pathways, especially the mTOR route, although there is almost no information on its activity in aged tissues. Aging also induces Ca2+ signal alterations, but information regarding the mechanisms for these changes is almost inexistent. We investigated the possible involvement of the mTOR pathway in the age-dependent changes on Ca2+ stores mobilization in colonic smooth muscle cells of young (4 month old) and aged (24 month old) guinea pigs. mTORC1 activity was enhanced in aged smooth muscle, as revealed by phosphorylation of mTOR and its direct substrates S6K1 and 4E-BP1. Mobilization of intracellular Ca2+ stores through IP3R or RyR channels was impaired in aged cells, and it was facilitated by mTOR and by FKBP12, as indicated by the inhibitory effects of KU0063794 (a direct mTOR inhibitor), rapamycin (a FKBP12-mediated mTOR inhibitor) and FK506 (an FKBP12 binding immunosuppressant). Aging suppressed the facilitation of the Ca2+ mobilization by FKBP12 but not by mTOR, without changing the total expression of FKBP12 protein. In conclusion, or study shows that in smooth muscle aging enhances the constitutive activity of mTORC1 pathway and impairs Ca2+ stores mobilization by suppression of the FKBP12-induced facilitation of Ca2+ release. PMID:23661091

  2. Advanced glycation endproducts regulate smooth muscle cells calcification in cultured HSMCs.

    PubMed

    He, Hu-Qiang; Liu, Yong; Zeng, Hong; Sun, Xiao-Lei; Zhang, Lei; Zhang, Xue-Lin; Liao, Wen-Jun; Zhou, Xiang-Yu; He, Yan-Zheng

    2015-01-01

    To investigate the mechanism of Advanced glycation end products (AGEs) promoting the calcification of smooth muscle cells. The successfully cultured smooth muscle cells were divided into three groups: normal culture group (group A), calcified culture group (group B), calcification + AGEs group (group C); the concentration of intracellular calcium ion was detected in each group; the promotion of AGEs on the calcification of HSMCs was confirmed by VON KOSSA staining; and the expressions of β-catenin, RAGE, β-catenin, OPG and E-cadherin protein were detected by immunofluorescence and western blot. The morphology of the cells in each group showed that the amount of calcified plaques in calcification + AGES group were significantly higher than the calcification group. VON KOSSA staining showed that with increasing concentrations of AGE-BSA, the amount of its calcification gradually increased. Calcium concentration in Calcification + 20 mg/L AGEs group was significantly higher, followed by 40 mg/L AGEs group. The expression of β-catenin increased with the increasing concentrations of AGEs. AGEs can promote the calcification of human femoral artery smooth muscle cells, with a concentration gradient effect. With increasing concentrations of AGEs, the expression of RAGE increased, indicating that AGEs-induced HSMCs proliferation was correlated with RAGE expression.

  3. Smooth muscle cells of penis in the rat: noninvasive quantification with shear wave elastography.

    PubMed

    Zhang, Jia-Jie; Qiao, Xiao-Hui; Gao, Feng; Bai, Ming; Li, Fan; Du, Lian-Fang; Xing, Jin-Fang

    2015-01-01

    Smooth muscle cells (SMCs) of cavernosum play an important role in erection. It is of great significance to quantitatively analyze the level of SMCs in penis. In this study, we investigated the feasibility of shear wave elastography (SWE) on evaluating the level of SMCs in penis quantitatively. Twenty healthy male rats were selected. The SWE imaging of penis was carried out and then immunohistochemistry analysis of penis was performed to analyze the expression of alpha smooth muscle actin in penis. The measurement index of SWE examination was tissue stiffness (TS). The measurement index of immunohistochemistry analysis was positive area percentage of alpha smooth muscle actin (AP). Sixty sets of data of TS and AP were obtained. The results showed that TS was significantly correlated with AP and the correlation coefficient was -0.618 (p < 0.001). The result of TS had been plotted against the AP measurements. The relation between the two results has been fitted with quadric curve; the goodness-of-fit index was 0.364 (p < 0.001). The level of SMCs in penis was successfully quantified in vivo with SWE. SWE can be used clinically for evaluating the level of SMCs in penis quantitatively.

  4. Overexpression of functional TrkA receptors after internalisation in human airway smooth muscle cells.

    PubMed

    Freund-Michel, Véronique; Frossard, Nelly

    2008-10-01

    Trafficking of the TrkA receptor after stimulation by NGF is of emerging importance in structural cells in the context of airway inflammatory diseases. We have recently reported the expression of functional TrkA receptors in human airway smooth muscle cells (HASMC). We have here studied the TrkA trafficking mechanisms in these cells. TrkA disappearance from the cell membrane was induced within 5 min of NGF (3pM) stimulation. Co-immunoprecipitation of clathrin-TrkA was revealed, and TrkA internalisation inhibited either by clathrin inhibitors or by siRNA inducing downregulation of endogenous clathrin. TrkA internalised receptors were totally degraded in lysosomes, with no recycling phenomenon. Newly synthesized TrkA receptors were thereafter re-expressed at the cell membrane within 10 h. TrkA re-synthesis was inhibited by blockade of clathrin-dependent internalisation, but not of TrkA receptors lysosomal degradation. Finally, we observed that NGF multiple stimulations progressively increased TrkA expression in HASMC, which was associated with an increase in NGF/TrkA-dependent proliferation. In conclusion, we show here the occurrence of clathrin-dependent TrkA internalisation and lysosomal degradation in the airway smooth muscle, followed by upregulated re-synthesis of functional TrkA receptors and increased proliferative effect in the human airway smooth muscle. This may have pathophysiological consequences in airway inflammatory diseases.

  5. Smooth muscle metaplasia and innervation in interstitium of endometriotic lesions related to pain.

    PubMed

    Odagiri, Kohei; Konno, Ryo; Fujiwara, Hiroyuki; Netsu, Sachiho; Yang, Chenghui; Suzuki, Mitsuaki

    2009-11-01

    To investigate the pathogenesis of endometriotic pain. Retrospective nonrandomized immunohistochemical study. A university hospital, Department of Gynecology. Twenty human endometriotic specimens were selected from different lesions including ovarian endometrioma, peritoneal lesion, and deep infiltrating lesion. Premenopausal women with histologically diagnosed endometriosis were selected (mean age 39 years; range, 25-53 years). The chief complaint was dysmenorrhea, dyschezia, and dyspareunia. A rat endometriosis model was induced in 10 SLC-Sprague-Dawley rats (8 weeks old) by surgical autotransplantation of the uterus. Immunohistochemical staining of endometriotic specimens for alpha-smooth muscle actin (ASMA), neural cell adhesion molecule (NCAM), and nerve growth factor (NGF) expression. Comparison of the immunoreactive staining of ASMA, NCAM, and NGF expression in human endometriosis and a rat endometriosis model. Morphological analysis revealed thick interstitium in both human and rat endometriotic lesions. The major components of fibrotic interstitium are smooth muscle cells, stained by anti-ASMA antibody, and nerve cells, stained by anti-NCAM antibody. Inflammatory cells are also present (e.g., macrophages and lymphocytes) as revealed by anti-NGF antibody staining. These results suggest that the contraction of smooth muscle cells and the hyperalgia derived from innervation in the interstitial area is related to pain in endometriosis.

  6. Potentiation of contraction of rabbit airway smooth muscle by some cyclooxygenase products.

    PubMed

    Armour, C L; Johnson, P R; Black, J L

    1988-06-01

    An alteration in smooth muscle sensitivity may be one of the mechanisms of the airway hyperresponsiveness observed in asthma. Indomethacin inhibits experimentally induced airway hyperresponsiveness. We thus examined the effects of the cyclooxygenase products PGD2, PGF2 alpha and a thromboxane A2 analogue U46619 on contractile responses of rabbit airway smooth muscle to histamine, carbachol and electrical field stimulation (EFS). PGD2 did not potentiate any contractile responses. When PGF2 alpha (1 microM) was administered 30 min before cumulative concentration-response curves to histamine and carbachol, no potentiation was observed. However, PGF2 alpha (1 microM) added immediately before EFS and bolus doses of histamine potentiated the contractile responses. U46619 increased the cumulative concentration-responses to both histamine and carbachol. The fact that we could alter smooth muscle sensitivity in vitro with PGF2 alpha and a thromboxane analogue suggests that these mediators may be involved in the airway hyperresponsiveness observed in asthma.

  7. Functional Constituents of a Local Serotonergic System, Intrinsic to the Human Coronary Artery Smooth Muscle Cells

    PubMed Central

    Baskar, Kannan; Sur, Swastika; Selvaraj, Vithyalakashmi; Agrawal, Devendra K.

    2015-01-01

    Human coronary artery smooth muscle cells (HCASMCs) play an important role in the pathogenesis of coronary atherosclerosis and coronary artery diseases (CAD). Serotonin is a mediator known to produce vascular smooth muscle cell (VSMC) mitogenesis and contribute to coronary atherosclerosis. We hypothesize that the human coronary artery smooth muscle cell possesses certain functional constituents of the serotonergic system such as: tryptophan hydroxylase and serotonin transporter. Our aim was to examine the presence of functional tryptophan hydroxylase-1 (TPH1) and serotonin transporter (SERT) in HCASMCs. The mRNA transcripts by qPCR and protein expression by Western blot of TPH1 and SERT were examined. The specificity and accuracy of the primers were verified using DNA gel electrophoresis and sequencing of qPCR products. The functionality of SERT was examined using a fluorescence dye-based serotonin transporter assay. The enzymatic activity of TPH was evaluated using UPLC. The HCASMCs expressed both mRNA transcripts and protein of SERT and TPH. The qPCR showed a single melt curve peak for both transcripts and in sequence analysis the amplicons were aligned with the respective genes. SERT and TPH enzymatic activity was present in the HCASMCs. Taken together, both TPH and SERT are functionally expressed in HCASMCs. These findings are novel and represent an initial step in examining the clinical relevance of the serotonergic system in HCASMCs and its role in the pathogenesis of coronary atherosclerosis and CAD. PMID:25861735

  8. Non-receptor tyrosine kinases and the actin cytoskeleton in contractile vascular smooth muscle.

    PubMed

    Ohanian, Jacqueline; Pieri, Maria; Ohanian, Vasken

    2015-09-01

    The contractility of vascular smooth muscle cells within the walls of arteries is regulated by mechanical stresses and vasoactive signals. Transduction of these diverse stimuli into a cellular response occurs through many different mechanisms, one being reorganisation of the actin cytoskeleton. In addition to a structural role in maintaining cellular architecture it is now clear that the actin cytoskeleton of contractile vascular smooth muscle cells is a dynamic structure reacting to changes in the cellular environment. Equally clear is that disrupting the cytoskeleton or interfering with its rearrangement, has profound effects on artery contractility. The actin cytoskeleton associates with dense plaques, also called focal adhesions, at the plasma membrane of smooth muscle cells. Vasoconstrictors and mechanical stress induce remodelling of the focal adhesions, concomitant with cytoskeletal reorganisation. Recent work has shown that non-receptor tyrosine kinases and tyrosine phosphorylation of focal adhesion proteins such as paxillin and Hic-5 are important for actin cytoskeleton and focal adhesion remodelling and contraction. © 2014 The Authors. The Journal of Physiology © 2014 The Physiological Society.

  9. Gastric mucosal smooth muscles may explain oscillations in glandular pressure: role of vasoactive intestinal peptide.

    PubMed

    Synnerstad, I; Ekblad, E; Sundler, F; Holm, L

    1998-02-01

    Oscillating (3-7 cycles/min) high pressures in gastric glands during acid secretion suggest the existence of rhythmically contracting mucosal muscles. The aim of this study was to study vasoactive intestinal peptide (VIP), an inhibitory neurotransmitter in the gastrointestinal tract, in relation to mucosal muscles, glandular pressure, and blood flow. Rat, dog, and human mucosae were examined immunocytochemically for smooth muscle actin and VIP. Glandular pressure was measured using microelectrodes, red blood cell velocity (V[RBC]) was measured using a cross-correlation technique, and blood flow was measured using laser Doppler flowmetry in exposed gastric mucosa of thiobutabarbital sodium-anesthetized rats. Actin immunostaining showed muscle strands arising from muscularis mucosae, extending toward the gastric pits. VIP-immunoreactive nerve fibers were found in close relation to these muscles. VIP, administered intra-arterially close to the stomach (2 microg/kg bolus, followed by 10 microg x kg[-1] x h[-1]), significantly decreased glandular pressure from 18.2 +/- 1.6 to 8.9 +/- 1.6 mm Hg and almost eliminated the pressure oscillations. VIP infusion also abolished the oscillations in V(RBC) and significantly increased blood flow by approximately 35%. Contracting mucosal muscles may be responsible for oscillations in glandular pressure and possibly also in V(RBC). VIP probably relaxes these muscles.

  10. Comparison of ionic currents from interstitial cells and smooth muscle cells of canine colon.

    PubMed Central

    Lee, H K; Sanders, K M

    1993-01-01

    1. Voltage-dependent ionic currents of isolated interstitial cells were characterized using the whole-cell voltage clamp technique, and compared with currents recorded from circular muscle cells. Both cell types were isolated from the submucosal pacemaking region in the canine distal colon. 2. Upon depolarization, interstitial cells and smooth muscle cells generated transient inward, followed by slowly inactivating outward, currents. 3. After blocking inward current and much of the Ca(2+)-dependent outward current, interstitial cells displayed voltage-dependent outward current that rapidly activated, reached a peak, and then inactivated. This current was resistant to 4-aminopyridine(4-AP; 1 mM). Smooth muscle cells expressed a similar current but it was reduced by about 40% at a test potential of +20 mV by 4-AP (1 mM). 4. The inactivation characteristics of the voltage-dependent outward currents of interstitial cells and smooth muscle cells were compared. The outward current of interstitial cells inactivated at more negative potentials; half-inactivation occurred at -53 mV, whereas half-inactivation occurred at -20 mV in smooth muscle cells. 5. Inward currents were not strikingly different in the two cell types when dialysing pipettes were used. When the perforated patch technique (using Amphotericin-B) was used, a negatively activating inward current was observed in interstitial cells that had a resolution threshold of -70 to -60 mV. This current peaked at -10 mV. Inward currents in smooth muscle cells were resolved at test potentials positive to -50 mV and peaked at 0 to +10 mV. 6. When interstitial cells were held at -40 mV, inward current could not be resolved with test depolarization negative to -30 mV. From this holding potential, peak amplitude was reduced by 85% with test depolarizations to -10 mV. Holding smooth muscle cells at -40 mV also reduced inward current, but the peak current in these cells was reduced by only 39% at 0 mV. 7. Ni2+ partially

  11. Altered Expression of Human Smooth Muscle Myosin Phosphatase Targeting (MYPT) Isovariants with Pregnancy and Labor

    PubMed Central

    Taggart, Julie; Robson, Stephen; Taggart, Michael

    2016-01-01

    Background Myosin light-chain phosphatase is a trimeric protein that hydrolyses phosphorylated myosin II light chains (MYLII) to cause relaxation in smooth muscle cells including those of the uterus. A major component of the phosphatase is the myosin targeting subunit (MYPT), which directs a catalytic subunit to dephosphorylate MYLII. There are 5 main MYPT family members (MYPT1 (PPP1R12A), MYPT2 (PPP1R12B), MYPT3 (PPP1R16A), myosin binding subunit 85 MBS85 (PPP1R12C) and TIMAP (TGF-beta-inhibited membrane-associated protein (PPP1R16B)). Nitric oxide (NO)-mediated smooth muscle relaxation has in part been attributed to activation of the phosphatase by PKG binding to a leucine zipper (LZ) dimerization domain located at the carboxyl-terminus of PPP1R12A. In animal studies, alternative splicing of PPP1R12A can lead to the inclusion of a 31-nucleotide exonic segment that generates a LZ negative (LZ-) isovariant rendering the phosphatase less sensitive to NO vasodilators and alterations in PPP1R12ALZ- and LZ+ expression have been linked to phenotypic changes in smooth muscle function. Moreover, PPP1R12B and PPP1R12C, but not PPP1R16A or PPP1R16B, have the potential for LZ+/LZ- alternative splicing. Yet, by comparison to animal studies, the information on human MYPT genomic sequences/mRNA expressions is scant. As uterine smooth muscle undergoes substantial remodeling during pregnancy we were interested in establishing the patterns of expression of human MYPT isovariants during this process and also following labor onset as this could have important implications for determining successful pregnancy outcome. Objectives We used cross-species genome alignment, to infer putative human sequences not available in the public domain, and isovariant-specific quantitative PCR, to analyse the expression of mRNA encoding putative LZ+ and LZ- forms of PPP1R12A, PPP1R12B and PPP1R12C as well as canonical PPP1R16A and PPP1R16B genes in human uterine smooth muscle from non

  12. Effects of Ginkgo biloba extracts with mirodenafil on the relaxation of corpus cavernosal smooth muscle and the potassium channel activity of corporal smooth muscle cells.

    PubMed

    Kim, Jung Jun; Han, Deok Hyun; Lim, Soo Hyun; Kim, Tae Hun; Chae, Mee Ree; Chung, Kyung Jin; Kam, Sung Chul; Jeon, Ju-Hong; Park, Jong Kwan; Lee, Sung Won

    2011-09-01

    In this study, we investigated the effects of a combination of Ginkgo biloba extracts (GBE) and phosphodiesterase type 5 (PDE-5) inhibitors on the muscular tone of the corpus cavernosum and potassium channel activity of corporal smooth muscle cells. Strips of corpus cavernosum from male New Zealand white rabbits were mounted in organ baths for isometric tension studies. After contraction with 1×10⁻⁵ mol l⁻¹ norepinephrine, GBE (0.01-1 mg ml⁻¹) and mirodenafil (0.01-100 nmol l⁻¹) were added together into the organ bath. In electrophysiological studies, whole-cell currents were recorded by the conventional patch-clamp technique in cultured smooth muscle cells of the human corpus cavernosum. The corpus cavernosum was relaxed in response to GBE in a dose-dependent manner (from 0.64%±8.35% at 0.01 mg ml⁻¹ to 52.28% ± 11.42% at 1 mg ml⁻¹). After pre-treatment with 0.03 mg ml⁻¹ of GBE, the relaxant effects of mirodenafil were increased at all concentrations. After tetraethylammonium (TEA) (1 mmol l⁻¹) administration, the increased effects were inhibited (P<0.01). Extracellular administration of GBE increased the whole-cell K(+) outward currents in a dose-dependent fashion. The increase of the outward current was inhibited by 1 mmol l⁻¹ TEA. These results suggest that GBE could increase the relaxant potency of mirodenafil even at a minimally effective dose. The K(+) flow through potassium channels might be one of the mechanisms involved in this synergistic relaxation.

  13. Pathway of programmed cell death and oxidative stress induced by β-hydroxybutyrate in dairy cow abomasum smooth muscle cells and in mouse gastric smooth muscle.

    PubMed

    Tian, Wulin; Wei, Teng; Li, Bin; Wang, Zhe; Zhang, Naisheng; Xie, Guanghong

    2014-01-01

    The administration of exogenous β-hydroxybutyrate (β-HB), as well as fasting and caloric restriction, is a condition associated with β-HB abundance and decreased appetite in animals. Increased β-HB and decreased appetite exist simultaneously in some diseases, such as bovine left displaced abomasums (LDA) and human chronic gastritis. However, the effects of β-HB on stomach injuries have not been explored. To elucidate the possible effects of exogenous β-HB on the stomach, mice were injected intraperitoneally with β-HB, and bovine abomasum smooth muscle cells (BSMCs) were treated with different concentrations of β-HB. We found that β-HB induced BSMCs endoplasmic reticulum- and mitochondria-mediated apoptotic cell death. β-HB promoted Bax expression and caspase-12, -9, and -3 activation while blocking Bcl-2 expression. β-HB also promoted AIF, EndoG release and p53 expression. β-HB acted on key molecules in the apoptotic cell death pathway and increased p38 and c-June NH2-terminal kinase phosphorylation while inhibiting ERK phosphorylation and PCNA expression. β-HB upregulated P27 and P21 mRNA levels while downregulating cyclin and CDK mRNA levels, arresting the cell cycle. These results suggest that BSMCs treated with β-HB can induce oxidative stress, which can be prevented by intracellular calcium chelators BAPTA/AM but not antioxidant NAC. Additionally, these results suggest that β-HB causes ROS generation through a Ca2+-dependent mechanism and that intracellular Ca2+ levels play a critical role in β-HB -induced apoptotic cell death. The impact of β-HB on programmed cell death and oxidative stress in vivo was confirmed in murine experiments. For the first time, we show oxidative stress effects of β-HB on smooth muscle. We propose that β-HB is a possible cause of some stomach diseases, including bovine LDA.

  14. Whole animal knockout of smooth muscle alpha-actin does not alter excisional wound healing or the fibroblast-to-myofibroblast transition.

    PubMed

    Tomasek, James J; Haaksma, Carol J; Schwartz, Robert J; Howard, Eric W

    2013-01-01

    The contractile phenotype and function of myofibroblasts have been proposed to play a critical role in wound closure. It has been hypothesized that smooth muscle α-actin expressed in myofibroblasts is critical for its formation and function. We have used smooth muscle α-actin-null mice to test this hypothesis. Full-thickness excisional wounds closed at a similar rate in smooth muscle α-actin-null and wild-type mice. In addition, fibroblasts in smooth muscle α-actin-null granulation tissue when immunostained with a monoclonal antibody that recognizes all muscle actin isoforms exhibited a myofibroblast-like distribution and a stress fiber-like pattern, showing that these cells acquired the myofibroblast phenotype. Dermal fibroblasts from smooth muscle α-actin-null and wild-type mice formed stress fibers and supermature focal adhesions, and generated similar amounts of contractile force in response to transforming growth factor-β1. Smooth muscle γ-actin and skeletal muscle α-actin were expressed in smooth muscle α-actin-null myofibroblasts, as shown by immunostaining, real-time polymerase chain reaction, and mass spectrometry. These results show that smooth muscle α-actin is not necessary for myofibroblast formation and function and for wound closure, and that smooth muscle γ-actin and skeletal muscle α-actin may be able to functionally compensate for the lack of smooth muscle α-actin in myofibroblasts. © 2012 by the Wound Healing Society.

  15. Identification of G-Protein-Coupled Receptors (GPCRs) in Pulmonary Artery Smooth Muscle Cells as Novel Therapeutic Targets

    DTIC Science & Technology

    2017-10-01

    AWARD NUMBER: W81XWH-14-1-0372 TITLE: Identification of G- Protein -Coupled Receptors (GPCRs) in Pulmonary Artery Smooth Muscle Cells as Novel...DATES COVERED 2 Sep 2016 - 1 Sep 2017 4. TITLE AND SUBTITLE Identification of G- Protein -Coupled Receptors (GPCRs) in Pulmonary Artery Smooth Muscle...recognized) G- protein -coupled receptors (GPCRs) and their functional activity in the PASMCs from subjects with PAH can reveal new insights into

  16. A network of 2-4 nm filaments found in sea urchin smooth muscle. Protein constituents and in situ localization.

    PubMed

    Pureur, R P; Coffe, G; Soyer-Gobillard, M O; de Billy, F; Pudles, J

    1986-01-01

    In this report the coisolation of two proteins from sea urchin smooth muscle of apparent molecular weights (Mr) 54 and 56 kD respectively, as determined on SDS-PAGE, is described. Like the intermediate filament proteins, these two proteins are insoluble in high ionic strength buffer solution. On two-dimensional gel electrophoresis and by immunological methods it is shown that these proteins are not related (by these criteria) to rat smooth muscle desmin (54 kD) or vimentin (56 kD). Furthermore, in conditions where both desmin and vimentin assemble in vitro into 10 nm filaments, the sea urchin smooth muscle proteins do not assemble into filaments. Ultrastructural studies on the sea urchin smooth muscle cell show that the thin and thick filaments organization resembles that described in the vertebrate smooth muscle. However, instead of 10 nm filaments, a network of filaments, 2-4 nm in diameter, is revealed, upon removal of the thin and thick filaments by 0.6 M KCl treatment. By indirect immunofluorescence microscopy, and in particular by immunocytochemical electron microscopy studies on the sea urchin smooth muscle cell, it is shown that the antibodies raised against both 54 and 56 kD proteins appear to specifically label these 2-4 nm filaments. These findings indicate that both the 54 and 56 kD proteins might be constituents of this category of filaments. The possible significance of this new cytoskeletal element, that we have named echinonematin filaments, is discussed.

  17. Phenotypic heterogeneity influences the behavior of rat aortic smooth muscle cells in collagen lattice

    SciTech Connect

    Orlandi, Augusto; Ferlosio, Amedeo; Gabbiani, Giulio

    2005-12-10

    Phenotypic modulation of vascular smooth muscle cells (SMCs) in atherosclerosis and restenosis involves responses to the surrounding microenvironment. SMCs obtained by enzymatic digestion from tunica media of newborn, young adult (YA) and old rats and from the thickened intima (TI) and underlying media of young adult rat aortas 15 days after ballooning were entrapped in floating populated collagen lattice (PCL). TI-SMCs elongated but were poor at PCL contraction and remodeling and expressed less {alpha}2 integrin compared to other SMCs that appeared more dendritic. During early phases of PCL contraction, SMCs showed a marked decrease in the expression of {alpha}-smooth musclemore » actin and myosin. SMCs other than TI-SMCs required 7 days to re-express {alpha}-smooth muscle actin and myosin. Only TI-SMCs in PCL were able to divide in 48 h, with a greater proportion in S and G2-M cell cycle phases compared to other SMCs. Anti-{alpha}2 integrin antibody markedly inhibited contraction but not proliferation in YA-SMC-PLCs; anti-{alpha}1 and anti-{alpha}2 integrin antibodies induced a similar slight inhibition in TI-SMC-PCLs. Finally, TI-SMCs rapidly migrated from PCL on plastic reacquiring their epithelioid phenotype. Heterogeneity in proliferation and cytoskeleton as well the capacity to remodel the extracellular matrix are maintained, when SMCs are suspended in PCLs.« less

  18. A role of stretch-activated potassium currents in the regulation of uterine smooth muscle contraction

    PubMed Central

    Buxton, Iain L O; Heyman, Nathanael; Wu, Yi-ying; Barnett, Scott; Ulrich, Craig

    2011-01-01

    Rates of premature birth are alarming and threaten societies and healthcare systems worldwide. Premature labor results in premature birth in over 50% of cases. Preterm birth accounts for three-quarters of infant morbidity and mortality. Children that survive birth before 34 weeks gestation often face life-long disability. Current treatments for preterm labor are wanting. No treatment has been found to be generally effective and none are systematically evaluated beyond 48 h. New approaches to the treatment of preterm labor are desperately needed. Recent studies from our laboratory suggest that the uterine muscle is a unique compartment with regulation of uterine relaxation unlike that of other smooth muscles. Here we discuss recent evidence that the mechanically activated 2-pore potassium channel, TREK-1, may contribute to contraction-relaxation signaling in uterine smooth muscle and that TREK-1 gene variants associated with human labor and preterm labor may lead to a better understanding of preterm labor and its possible prevention. PMID:21642947

  19. Relaxing Effect of TSU-68, an Antiangiogenic Agent, on Mouse Airway Smooth Muscle.

    PubMed

    Tan, Honghao; Lei, Jun; Xue, Lu; Cai, Congli; Liu, Qing-Hua; Shen, Jinhua

    2017-01-01

    Recently, some small-molecule compounds that were designed for cancer therapy have acquired new roles in the treatment of pulmonary diseases. However, drug screening aimed at abnormal muscle contraction is still limited. TSU-68 is a potent, orally administered, small-molecule agent that can reduce the vascular endothelial growth factor (VEGF)-induced Ca2+ increase in endothelial cells. We questioned whether TSU-68 could also affect calcium influx and relax airway smooth muscle (ASM) cells. The current study aimed to investigate these effects and to explore the underlying mechanisms. The effects of TSU-68 on ASM cells were studied in mice using a series of biophysiological techniques, including force measurement and patch-clamp experiments. TSU-68 inhibited high K+ or acetylcholine chloride (ACh)-induced pre-contracted mouse tracheal rings in a concentration-dependent manner. Further research demonstrated that the TSU-68-induced ASM relaxation was mediated by calcium, which was decreased by blocking voltage-dependent Ca2+ channels (VDCCs) and non-selective cation channels (NSCCs). Our data indicated that TSU-68 relaxes tense ASM by reducing the intracellular Ca2+ concentration through blocking VDCCs and NSCCs, which suggested that this small molecule might be useful in the treatment of abnormal smooth muscle. © 2017 The Author(s). Published by S. Karger AG, Basel.

  20. The impact of prolapse mesh on vaginal smooth muscle structure and function

    PubMed Central

    Jallah, Z; Liang, R; Feola, A; Barone, W; Palcsey, S; Abramowitch, SD; Yoshimura, N; Moalli, P

    2016-01-01

    Objective To evaluate the impact of prolapse meshes on vaginal smooth muscle structure (VaSM) and function, and to evaluate these outcomes in the context of the mechanical and textile properties of the mesh. Design Three months following the implantation of three polypropylene prolapse meshes with distinct textile and mechanical properties, mesh tissue explants were evaluated for smooth muscle contraction, innervation, receptor function, and innervation density. Setting Magee-Womens Research Institute at the University of Pittsburgh. Population Thirty-four parous rhesus macaques of similar age, parity, and pelvic organ prolapse quantification (POP–Q) scores. Methods Macaques were implanted with mesh via sacrocolpopexy. The impact of Gynemesh™ PS (Ethicon; n = 7), Restorelle® (Coloplast; n = 7), UltraPro™ parallel and UltraPro™ perpendicular (Ethicon; n = 6 and 7, respectively) were compared with sham-operated controls (n = 7). Outcomes were analysed by Kruskal–Wallis ANOVA, Mann–Whitney U–tests and multiple regression analysis (P < 0.05). Mean outcome measures Vaginal tissue explants were evaluated for the maximum contractile force generated following muscle, nerve, and receptor stimulation, and for peripheral nerve density. Results Muscle myofibre, nerve, and receptor-mediated contractions were negatively affected by mesh only in the grafted region (P < 0.001, P = 0.002, and P = 0.008, respectively), whereas cholinergic and adrenergic nerve densities were affected in the grafted (P = 0.090 and P = 0.008, respectively) and non-grafted (P = 0.009 and P = 0.005, respectively) regions. The impact varied by mesh property, as mesh stiffness was a significant predictor of the negative affect on muscle function and nerve density (P < 0.001 and P = 0.013, respectively), whereas mesh and weight was a predictor of receptor function (P < 0.001). Conclusions Mesh has an overall negative impact on VaSM, and the effects are a function of mesh properties, most

  1. Force maintenance and myosin filament assembly regulated by Rho-kinase in airway smooth muscle.

    PubMed

    Lan, Bo; Deng, Linhong; Donovan, Graham M; Chin, Leslie Y M; Syyong, Harley T; Wang, Lu; Zhang, Jenny; Pascoe, Christopher D; Norris, Brandon A; Liu, Jeffrey C-Y; Swyngedouw, Nicholas E; Banaem, Saleha M; Paré, Peter D; Seow, Chun Y

    2015-01-01

    Smooth muscle contraction can be divided into two phases: the initial contraction determines the amount of developed force and the second phase determines how well the force is maintained. The initial phase is primarily due to activation of actomyosin interaction and is relatively well understood, whereas the second phase remains poorly understood. Force maintenance in the sustained phase can be disrupted by strains applied to the muscle; the strain causes actomyosin cross-bridges to detach and also the cytoskeletal structure to disassemble in a process known as fluidization, for which the underlying mechanism is largely unknown. In the present study we investigated the ability of airway smooth muscle to maintain force after the initial phase of contraction. Specifically, we examined the roles of Rho-kinase and protein kinase C (PKC) in force maintenance. We found that for the same degree of initial force inhibition, Rho-kinase substantially reduced the muscle's ability to sustain force under static conditions, whereas inhibition of PKC had a minimal effect on sustaining force. Under oscillatory strain, Rho-kinase inhibition caused further decline in force, but again, PKC inhibition had a minimal effect. We also found that Rho-kinase inhibition led to a decrease in the myosin filament mass in the muscle cells, suggesting that one of the functions of Rho-kinase is to stabilize myosin filaments. The results also suggest that dissolution of myosin filaments may be one of the mechanisms underlying the phenomenon of fluidization. These findings can shed light on the mechanism underlying deep inspiration induced bronchodilation. Copyright © 2015 the American Physiological Society.

  2. The impact of prolapse mesh on vaginal smooth muscle structure and function.

    PubMed

    Jallah, Z; Liang, R; Feola, A; Barone, W; Palcsey, S; Abramowitch, S D; Yoshimura, N; Moalli, P

    2016-06-01

    To evaluate the impact of prolapse meshes on vaginal smooth muscle structure (VaSM) and function, and to evaluate these outcomes in the context of the mechanical and textile properties of the mesh. Three months following the implantation of three polypropylene prolapse meshes with distinct textile and mechanical properties, mesh tissue explants were evaluated for smooth muscle contraction, innervation, receptor function, and innervation density. Magee-Womens Research Institute at the University of Pittsburgh. Thirty-four parous rhesus macaques of similar age, parity, and pelvic organ prolapse quantification (POP-Q) scores. Macaques were implanted with mesh via sacrocolpopexy. The impact of Gynemesh(™)  PS (Ethicon; n = 7), Restorelle(®) (Coloplast; n = 7), UltraPro(™) parallel and UltraPro(™) perpendicular (Ethicon; n = 6 and 7, respectively) were compared with sham-operated controls (n = 7). Outcomes were analysed by Kruskal-Wallis ANOVA, Mann-Whitney U-tests and multiple regression analysis (P < 0.05). Vaginal tissue explants were evaluated for the maximum contractile force generated following muscle, nerve, and receptor stimulation, and for peripheral nerve density. Muscle myofibre, nerve, and receptor-mediated contractions were negatively affected by mesh only in the grafted region (P < 0.001, P = 0.002, and P = 0.008, respectively), whereas cholinergic and adrenergic nerve densities were affected in the grafted (P = 0.090 and P = 0.008, respectively) and non-grafted (P = 0.009 and P = 0.005, respectively) regions. The impact varied by mesh property, as mesh stiffness was a significant predictor of the negative affect on muscle function and nerve density (P < 0.001 and P = 0.013, respectively), whereas mesh and weight was a predictor of receptor function (P < 0.001). Mesh has an overall negative impact on VaSM, and the effects are a function of mesh properties, most notably, mesh stiffness. Prolapse mesh affects vaginal

  3. Testosterone replacement maintains smooth muscle content in the corpus cavernosum of orchiectomized rats.

    PubMed

    Halmenschlager, Graziele; Rhoden, Ernani Luis; Motta, Gabriela Almeida; Sagrillo Fagundes, Lucas; Medeiros, Jorge Luiz; Meurer, Rosalva; Rhoden, Cláudia Ramos

    2017-10-01

    To evaluate the effects of testosterone (T) on the maintenance of corpus cavernosum (CC) structure and apoptosis. Animals were divided into three groups: sham operation group ( n  = 8) underwent sham operation; Orchiectomized (Orchiec)+ oily vehicle group ( n  = 8) underwent bilateral orchiectomy and received a single dose of oily vehicle by intramuscular injection (i.m.) 30 days after orchiectomy; and Orchiec + T group ( n  = 8) underwent bilateral orchiectomy and received a single dose of T undecanoate 100 mg/kg i.m. 30 days after the surgery. Animals were euthanized 60 days after the beginning of the experiment with an anesthetic overdose of ketamine and xylazine. Blood samples and penile tissue were collected on euthanasia. Azan's trichrome staining was used to evaluate smooth muscle, Weigert's Fucsin-Resorcin staining was used to evaluate elastic fibers and Picrosirius red staining was used to evaluate collagen. Apoptosis was evaluated using TUNEL technique. T levels decreased in Orchiec + oily vehicle when compared to sham operation and Orchiec + T groups ( p  < 0.001). T deprivation reduced trabecular smooth muscle content and penile diameter and T replacement maintained both parameters ( p  = 0.005 and p  = 0.001, respectively). No difference was observed in the content of sinusoidal space ( p  = 0.207), elastic fibers ( p  = 0.849), collagen ( p  = 0.216) and in apoptosis ( p  = 0.095). Normal testosterone levels maintain CC smooth muscle content and do not influence elastic fibers, collagen content and apoptotic index. Further studies should be performed in order to investigate the mechanisms by which androgen mediates its effects on CC structure.

  4. Rosiglitazone reverses salbutamol-induced β2-adrenoceptor tolerance in airway smooth muscle

    PubMed Central

    Fogli, Stefano; Pellegrini, Silvia; Adinolfi, Barbara; Mariotti, Veronica; Melissari, Erika; Betti, Laura; Fabbrini, Laura; Giannaccini, Gino; Lucacchini, Antonio; Bardelli, Claudio; Stefanelli, Fabio; Brunelleschi, Sandra; Breschi, Maria Cristina

    2011-01-01

    BACKGROUND AND PURPOSE β2-Adrenoceptor agonists are important therapeutic agents in the treatment of asthma and chronic obstructive pulmonary disease. The regular use of these drugs has been associated with proasthmatic-like changes that limit their efficacy and increase the risk of severe adverse reactions. We investigated whether the peroxisome-proliferator-activated receptor (PPAR)γ agonist rosiglitazone modulated salbutamol-induced β2-adrenoceptor desensitization in vivo and in vitro. EXPERIMENTAL APPROACH An in vivo model of homologous β2-adrenoceptor desensitization, established in guinea-pigs by administering salbutamol continuously, was used to study the ability of rosiglitazone to prevent β2-adrenoceptor tolerance. In vitro experiments on human bronchial smooth muscle cells were performed to increase the clinical relevance of the study. KEY RESULTS In tracheal smooth muscle tissues from desensitized animals, we observed a decrease in the protective effect of salbutamol on carbachol-induced contraction, a hyperresponsiveness to cholinergic stimuli, a modest underexpression of β2-adrenoceptor gene and a marked decrease in β-adrenoceptor number, relative to control values. Treatment with rosiglitazone preserved salbutamol relaxant activity, mitigated carbachol hyperresponsiveness and partially restored β2-adrenoceptor binding sites in tracheal tissues from homologously desensitized animals. The highly selective PPARγ agonist, GW1929, reproduced the effect of rosiglitazone, in vivo. In vitroβ2-adrenoceptor desensitization decreased salbutamol-mediated cAMP production, without affecting forskolin responses and β2-adrenoceptor expression. Rosiglitazone and 15-deoxy-Δ12,14-prostaglandin J2 restored salbutamol sensitivity in homologously desensitized cells. CONCLUSIONS AND IMPLICATIONS These data suggest a potential pharmacodynamic interaction between PPARγ agonists and salbutamol on airway smooth muscle responsiveness, supporting the therapeutic

  5. Smooth muscle cell differentiation in the processus vaginalis of children with hernia or hydrocele.

    PubMed

    Mouravas, V K; Koletsa, T; Sfougaris, D K; Philippopoulos, A; Petropoulos, A S; Zavitsanakis, A; Kostopoulos, I

    2010-04-01

    Incomplete obliteration of the processus vaginalis (PV) in children with inguinal hernia or hydrocele has recently been proposed to relate to smooth muscle cell (SMC) persistence. The aim of this study was to evaluate the diversity and differentiation of smooth muscle phenotypes in sacs associated with inguinal hernia and hydrocele through the expression of alpha-smooth muscle actin (SMA), h-caldesmon, desmin, and vimentin. Sacs associated with male hernia (n = 22), female hernia (n = 8), and hydrocele (n = 10) were immunohistochemically evaluated using monoclonal antibodies against SMA, h-caldesmon, desmin, and vimentin. Peritoneal samples (male, 4; female, 3) and obliterated PV (male, 3) obtained from age-matched patients served as controls. Expressions according to the groups were compared through chi-squared test, and P values less than 0.05 were considered to be statistically significant. Immunohistochemistry did not shown the presence of SMCs in control samples. The expression of SMA, desmin, and h-caldesmon did not differ among sacs obtained from patients with inguinal hernia and hydrocele. However, strong expression of vimentin in SMCs within sacs obtained from patients with hydrocele in comparison with sacs from male patients with inguinal hernia were observed. Our results indicate that sacs from patients with inguinal hernias and especially from male inguinal hernias have fully differentiated SMCs. On the other hand SMCs in sacs obtained from boys with hydrocele are in an intermediate state of differentiation-dedifferentiation. This phenotypic modulation may represent attempted apoptosis of SMCs, since sacs more sensitive to apoptosis appeared to have more dedifferentiated SMCs. It also probably depicts the differing influence of sympathetic and parasympathetic tonuses during the descent of the testis and the obliteration of PV.

  6. Exaggerated smooth muscle contraction segments on esophageal high-resolution manometry: prevalence and clinical relevance.

    PubMed

    Mello, M D; Duraiswamy, S; Price, L H; Li, Y; Patel, A; Gyawali, C P

    2015-02-01

    Two smooth muscle contraction segments (S2, S3) on esophageal high-resolution manometry (HRM) demonstrate varying contraction vigor in symptomatic patients. Significance of isolated exaggerated smooth muscle contraction remains unclear. High-resolution manometry studies were reviewed in 272 consecutive patients (56.4 ± 0.8 years, 62% F) and compared to 21 healthy controls (27.6 ± 0.6 years, 52% F), using HRM tools (distal contractile integral, DCI; distal latency, DL; integrated relaxation pressure, IRP), Chicago Classification (CC) and multiple rapid swallows (MRS). Segments were designated merged when the trough between S2 and S3 was ≥150 mmHg, and exaggerated S3 when peak S3 amplitude was ≥150 mmHg without merging with S2. Presenting symptoms and global symptom severity (on 100 mm visual analog scale) were recorded. Prevalence of merged and exaggerated segments was determined, and characteristics compared to symptomatic patients with normal HRM, and to healthy controls. Merged segments were identified in 5.6%, and exaggerated S3 in another 12.5%, but only 17-50% had a CC diagnosis; one healthy control had merged segments. DCI with wet swallows was similar in cohorts with merged and exaggerated segments (p = 0.7), significantly higher than symptomatic patients with normal HRM and healthy controls (p ≤ 0.003 for each comparison). Incomplete inhibition and prominent DCI augmentation on MRS (p ≤ 0.01), and presenting symptoms (chest pain and dysphagia, p = 0.04) characterized exaggerated segments, but not demographics or symptom burden. Merged esophageal smooth muscle segments and exaggerated S3 may represent hypermotility phenomena from abnormal inhibition and/or excitation, and are not uniformly identified by the CC algorithm. © 2014 John Wiley & Sons Ltd.

  7. Unique properties of muscularis mucosae smooth muscle in guinea pig urinary bladder

    PubMed Central

    Layne, Jeffrey J.; Pearson, Jessica M.; Sarkissian, Hagop; Nelson, Mark T.

    2011-01-01

    The muscularis mucosae, a type of smooth muscle located between the urothelium and the urinary bladder detrusor, has been described, although its properties and role in bladder function have not been characterized. Here, using mucosal tissue strips isolated from guinea pig urinary bladders, we identified spontaneous phasic contractions (SPCs) that appear to originate in the muscularis mucosae. This smooth muscle layer exhibited Ca2+ waves and flashes, but localized Ca2+ events (Ca2+ sparks, purinergic receptor-mediated transients) were not detected. Ca2+ flashes, often in bursts, occurred with a frequency (∼5.7/min) similar to that of SPCs (∼4/min), suggesting that SPCs are triggered by bursts of Ca2+ flashes. The force generated by a single mucosal SPC represented the maximal force of the strip, whereas a single detrusor SPC was ∼3% of maximal force of the detrusor strip. Electrical field stimulation (0.5–50 Hz) evoked force transients in isolated detrusor and mucosal strips. Inhibition of cholinergic receptors significantly decreased force in detrusor and mucosal strips (at higher frequencies). Concurrent inhibition of purinergic and cholinergic receptors nearly abolished evoked responses in detrusor and mucosae. Mucosal SPCs were unaffected by blocking small-conductance Ca2+-activated K+ (SK) channels with apamin and were unchanged by blocking large-conductance Ca2+-activated K+ (BK) channels with iberiotoxin (IbTX), indicating that SK and BK channels play a much smaller role in regulating muscularis mucosae SPCs than they do in regulating detrusor SPCs. Consistent with this, BK channel current density in myocytes from muscularis mucosae was ∼20% of that in detrusor myocytes. These findings indicate that the muscularis mucosae in guinea pig represents a second smooth muscle compartment that is physiologically and pharmacologically distinct from the detrusor and may contribute to the overall contractile properties of the urinary bladder. PMID:21632849

  8. Non-selective cationic channels of smooth muscle and the mammalian homologues of Drosophila TRP

    PubMed Central

    Beech, D J; Muraki, K; Flemming, R

    2004-01-01

    Throughout the body there are smooth muscle cells controlling a myriad of tubes and reservoirs. The cells show enormous diversity and complexity compounded by a plasticity that is critical in physiology and disease. Over the past quarter of a century we have seen that smooth muscle cells contain – as part of a gamut of ion-handling mechanisms – a family of cationic channels with significant permeability to calcium, potassium and sodium. Several of these channels are sensors of calcium store depletion, G-protein-coupled receptor activation, membrane stretch, intracellular Ca2+, pH, phospholipid signals and other factors. Progress in understanding the channels has, however, been hampered by a paucity of specific pharmacological agents and difficulty in identifying the underlying genes. In this review we summarize current knowledge of these smooth muscle cationic channels and evaluate the hypothesis that the underlying genes are homologues of Drosophila TRP (transient receptor potential). Direct evidence exists for roles of TRPC1, TRPC4/5, TRPC6, TRPV2, TRPP1 and TRPP2, and more are likely to be added soon. Some of these TRP proteins respond to a multiplicity of activation signals – promiscuity of gating that could enable a variety of context-dependent functions. We would seem to be witnessing the first phase of the molecular delineation of these cationic channels, something that should prove a leap forward for strategies aimed at developing new selective pharmacological agents and understanding the activation mechanisms and functions of these channels in physiological systems. PMID:15272031

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

    PubMed

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

    2015-04-01

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

  10. Loss of neurotrophin-3 from smooth muscle disrupts vagal gastrointestinal afferent signaling and satiation.

    PubMed

    Fox, Edward A; Biddinger, Jessica E; Baquet, Zachary C; Jones, Kevin R; McAdams, Jennifer

    2013-12-01

    A large proportion of vagal afferents are dependent on neurotrophin-3 (NT-3) for survival. NT-3 is expressed in developing gastrointestinal (GI) smooth muscle, a tissue densely innervated by vagal mechanoreceptors, and thus could regulate their survival. We genetically ablated NT-3 from developing GI smooth muscle and examined the pattern of loss of NT-3 expression in the GI tract and whether this loss altered vagal afferent signaling or feeding behavior. Meal-induced c-Fos activation was reduced in the solitary tract nucleus and area postrema in mice with a smooth muscle-specific NT-3 knockout (SM-NT-3(KO)) compared with controls, suggesting a decrease in vagal afferent signaling. Daily food intake and body weight of SM-NT-3(KO) mice and controls were similar. Meal pattern analysis revealed that mutants, however, had increases in average and total daily meal duration compared with controls. Mutants maintained normal meal size by decreasing eating rate compared with controls. Although microstructural analysis did not reveal a decrease in the rate of decay of eating in SM-NT-3(KO) mice, they ate continuously during the 30-min meal, whereas controls terminated feeding after 22 min. This led to a 74% increase in first daily meal size of SM-NT-3(KO) mice compared with controls. The increases in meal duration and first meal size of SM-NT-3(KO) mice are consistent with reduced satiation signaling by vagal afferents. This is the first demonstration of a role for GI NT-3 in short-term controls of feeding, most likely involving effects on development of vagal GI afferents that regulate satiation.

  11. miR-125b Regulates Calcification of Vascular Smooth Muscle Cells

    PubMed Central

    Goettsch, Claudia; Rauner, Martina; Pacyna, Nicole; Hempel, Ute; Bornstein, Stefan R.; Hofbauer, Lorenz C.

    2011-01-01

    Vascular calcification is a prominent feature of atherosclerosis and is closely linked to osteoporosis. Cellular differentiation is regulated by various microRNAs (miRs), including miR-125b, which is known to be involved in osteoblast differentiation. However, no specific miR has been defined that modulates vascular calcification. Herein, we assessed the impact of miR-125b in osteogenic transformation of vascular smooth muscle cells. Osteogenic transdifferentiation of human coronary artery smooth muscle cells was induced by osteogenic medium and enhanced the formation of mineralized matrix, resulting in a significantly higher mineral deposition after 21 days. Increased expression of miR-125b was time-dependent in human coronary artery smooth muscle cells and diminished during osteogenic transdifferentiation. At day 21, miR-125b was significantly reduced (−42%) compared with that in the untreated control. The expression of miR-processing enzymes, RNase III endonucleases DICER1 and DROSHA, was also decreased. Furthermore, inhibition of endogenous miR-125b promoted osteogenic transdifferentiation, as measured by increased alkaline phosphatase activity and matrix mineralization. Expression analysis revealed the osteoblast transcription factor SP7 (osterix) as a target of miR-125b. In vivo, miR-125b was decreased in calcified aortas of apolipoprotein E knockout mice. In conclusion, our results suggest that miR-125b is involved in vascular calcification in vitro and in vivo, at least partially by targeting SP7. Evaluating the role of miRs in arterial calcification in vivo may have important therapeutic implications. PMID:21806957

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

    PubMed

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

    2016-08-01

    Hypertension, which is a risk factor of heart failure, provokes adaptive changes at the vasculature and cardiac levels. Notch3 signaling plays an important role in resistance arteries by controlling the maturation of vascular smooth muscle cells. Notch3 deletion is protective in pulmonary hypertension while deleterious in arterial hypertension. Although this latter phenotype was attributed to renal and cardiac alterations, the underlying mechanisms remained unknown. To investigate the role of Notch3 signaling in the cardiac adaptation to hypertension, we used mice with either constitutive Notch3 or smooth muscle cell-specific conditional RBPJκ knockout. At baseline, both genotypes exhibited a cardiac arteriolar rarefaction associated with oxidative stress. In response to angiotensin II-induced hypertension, the heart of Notch3 knockout and SM-RBPJκ knockout mice did not adapt to pressure overload and developed heart failure, which could lead to an early and fatal acute decompensation of heart failure. This cardiac maladaptation was characterized by an absence of media hypertrophy of the media arteries, the transition of smooth muscle cells toward a synthetic phenotype, and an alteration of angiogenic pathways. A subset of mice exhibited an early fatal acute decompensated heart failure, in which the same alterations were observed, although in a more rapid timeframe. Altogether, these observations indicate that Notch3 plays a major role in coronary adaptation to pressure overload. These data also show that the hypertrophy of coronary arterial media on pressure overload is mandatory to initially maintain a normal cardiac function and is regulated by the Notch3/RBPJκ pathway. © 2016 American Heart Association, Inc.

  13. Fluid flow releases fibroblast growth factor-2 from human aortic smooth muscle cells

    NASA Technical Reports Server (NTRS)

    Rhoads, D. N.; Eskin, S. G.; McIntire, L. V.

    2000-01-01

    This study tested the hypothesis that fluid shear stress regulates the release of fibroblast growth factor (FGF)-2 from human aortic smooth muscle cells. FGF-2 is a potent mitogen that is involved in the response to vascular injury and is expressed in a wide variety of cell types. FGF-2 is found in the cytoplasm of cells and outside cells, where it associates with extracellular proteoglycans. To test the hypothesis that shear stress regulates FGF-2 release, cells were exposed to flow, and FGF-2 amounts were measured from the conditioned medium, pericellular fraction (extracted by heparin treatment), and cell lysate. Results from the present study show that after 15 minutes of shear stress at 25 dyne/cm(2) in a parallel-plate flow system, a small but significant fraction (17%) of the total FGF-2 was released from human aortic smooth muscle cells. FGF-2 levels in the circulating medium increased 10-fold over medium from static controls (P<0.01). A 50% increase in FGF-2 content versus control (P<0.01) was found in the pericellular fraction (extracted by heparin treatment). Furthermore, a significant decrease in FGF-2 was detected in the cell lysate, indicating that FGF-2 was released from inside the cell. Cell permeability studies with fluorescent dextran were performed to examine whether transient membrane disruption caused FGF-2 release. Flow cytometry detected a 50% increase in mean fluorescence of cells exposed to 25 dyne/cm(2) versus control cells. This indicates that the observed FGF-2 release from human aortic smooth muscle cells is likely due to transient membrane disruption on initiation of flow.

  14. Mechanisms underlying the pathogenesis of hyper-contractility of bronchial smooth muscle in allergic asthma.

    PubMed

    Sakai, Hiroyasu; Suto, Wataru; Kai, Yuki; Chiba, Yoshihiko

    2017-01-01

    Airway hyperresponsiveness (AHR) and inflammation are key pathophysiological features of asthma. Enhanced contraction of bronchial smooth muscle (BSM) is one of the causes of the AHR. It is thus important for development of asthma therapy to understand the change in the contractile signaling of airway smooth muscle cells associated with the AHR. In addition to the Ca 2+ -mediated phosphorylation of myosin light chain (MLC), contractile agonists also enhance MLC phosphorylation level, Ca 2+ -independently, by inactivating MLC phosphatase (MLCP), called Ca 2+ sensitization of contraction, in smooth muscle cells including airways. To date, involvements of RhoA/ROCKs and PKC/Ppp1r14a (also called as CPI-17) pathways in the Ca 2+ sensitization have been identified. Our previous studies revealed that the agonist-induced Ca 2+ sensitization of contraction is markedly augmented in BSMs of animal models of allergen-induced AHR. In BSMs of these animal models, the expression of RhoA and CPI-17 proteins were significantly increased, indicating that both the Ca 2+ sensitizing pathways are augmented. Interestingly, incubation of BSM cells with asthma-associated cytokines, such as interleukin-13 (IL-13), IL-17, and tumor necrosis factor-α (TNF-α), caused up-regulations of RhoA and CPI-17 in BSM cells of naive animals and cultured human BSM cells. In addition to the transcription factors such as STAT6 and NF-κB activated by these inflammatory cytokines, an involvement of down-regulation of miR-133a, a microRNA that negatively regulates RhoA translation, has also been suggested in the IL-13- and IL-17-induced up-regulation of RhoA. Thus, the Ca 2+ sensitizing pathways and the cytokine-mediated signaling including microRNAs in BSMs might be potential targets for treatment of allergic asthma, especially the AHR.

  15. Developmental origins of colon smooth muscle dysfunction in IBS-like rats.

    PubMed

    Li, Qingjie; Winston, John H; Sarna, Sushil K

    2013-10-01

    Epidemiological studies show that subsets of adult and pediatric patients with irritable bowel syndrome (IBS) have prior exposures to psychological or inflammatory stress. We investigated the cellular mechanisms of colonic smooth muscle dysfunction in adult rats subjected to neonatal inflammation. Ten-day-old male rat pups received 2,4,6-trinitrobenzene sulfonic acid to induce colonic inflammation. Colonic circular smooth muscle strips were obtained 6 to 8 wk later. We found that about half of the neonate pups subjected to inflammatory insult showed a significant increase in expression of the pore-forming α1C-subunit of Cav1.2b channels in adult life. These were the same rats in whom Vip mRNA increased in the colon muscularis externae. Additional experiments showed reduced interaction of histone deacetylase (HDAC) 3 with α1C1b promoter that increased the acetylation of histone H3 lysine 9 (H3K9) in the core promoter region. Vasoactive intestinal peptide (VIP) treatment of naïve muscularis externae swiftly recruited CREB-binding protein (CBP) to the α1C1b promoter and dissociated HDAC3 from this region to initiate transcription. The CBP interaction with the α1C1b promoter was transient, but the dissociation of HDAC3 persisted to sustain H3K9 hyperacetylation and increase in transcription. Intraperitoneal treatment of adult naïve rats with butyrate mimicked the effects of neonatal colon inflammation. We concluded that neonatal inflammation upregulates VIP in the colon muscularis externae, which modulates epigenetic events at the α1C1b promoter to activate α1C1b gene transcription. Inflammatory insult in early life may be one of the etiologies of smooth muscle dysfunction in adult life, which contributes to the altered motility function in patients with diarrhea-predominant IBS.

  16. Developmental origins of colon smooth muscle dysfunction in IBS-like rats

    PubMed Central

    Li, Qingjie; Winston, John H.

    2013-01-01

    Epidemiological studies show that subsets of adult and pediatric patients with irritable bowel syndrome (IBS) have prior exposures to psychological or inflammatory stress. We investigated the cellular mechanisms of colonic smooth muscle dysfunction in adult rats subjected to neonatal inflammation. Ten-day-old male rat pups received 2,4,6-trinitrobenzene sulfonic acid to induce colonic inflammation. Colonic circular smooth muscle strips were obtained 6 to 8 wk later. We found that about half of the neonate pups subjected to inflammatory insult showed a significant increase in expression of the pore-forming α1C-subunit of Cav1.2b channels in adult life. These were the same rats in whom Vip mRNA increased in the colon muscularis externae. Additional experiments showed reduced interaction of histone deacetylase (HDAC) 3 with α1C1b promoter that increased the acetylation of histone H3 lysine 9 (H3K9) in the core promoter region. Vasoactive intestinal peptide (VIP) treatment of naïve muscularis externae swiftly recruited CREB-binding protein (CBP) to the α1C1b promoter and dissociated HDAC3 from this region to initiate transcription. The CBP interaction with the α1C1b promoter was transient, but the dissociation of HDAC3 persisted to sustain H3K9 hyperacetylation and increase in transcription. Intraperitoneal treatment of adult naïve rats with butyrate mimicked the effects of neonatal colon inflammation. We concluded that neonatal inflammation upregulates VIP in the colon muscularis externae, which modulates epigenetic events at the α1C1b promoter to activate α1C1b gene transcription. Inflammatory insult in early life may be one of the etiologies of smooth muscle dysfunction in adult life, which contributes to the altered motility function in patients with diarrhea-predominant IBS. PMID:23886858

  17. Acrolein relaxes mouse isolated tracheal smooth muscle via a TRPA1-dependent mechanism.

    PubMed

    Cheah, Esther Y; Burcham, Philip C; Mann, Tracy S; Henry, Peter J

    2014-05-01

    Airway sensory C-fibres express TRPA1 channels which have recently been identified as a key chemosensory receptor for acrolein, a toxic and highly prevalent component of smoke. TRPA1 likely plays an intermediary role in eliciting a range of effects induced by acrolein including cough and neurogenic inflammation. Currently, it is not known whether acrolein-induced activation of TRPA1 produces other airway effects including relaxation of mouse airway smooth muscle. The aims of this study were to examine the effects of acrolein on airway smooth muscle tone in mouse isolated trachea, and to characterise the cellular and molecular mechanisms underpinning the effects of acrolein. Isometric tension recording studies were conducted on mouse isolated tracheal segments to characterise acrolein-induced relaxation responses. Release of the relaxant PGE₂ was measured by EIA to examine its role in the response. Use of selective antagonists/inhibitors permitted pharmacological characterisation of the molecular and cellular mechanisms underlying this relaxation response. Acrolein induced dose-dependent relaxation responses in mouse isolated tracheal segments. Importantly, these relaxation responses were significantly inhibited by the TRPA1 antagonists AP-18 and HC-030031, an NK₁ receptor antagonist RP-67580, and the EP₂ receptor antagonist PF-04418948, whilst completely abolished by the non-selective COX inhibitor indomethacin. Acrolein also caused rapid PGE₂ release which was suppressed by HC-030031. In summary, acrolein induced a novel bronchodilator response in mouse airways. Pharmacologic studies indicate that acrolein-induced relaxation likely involves interplay between TRPA1-expressing airway sensory C-fibres, NK₁ receptor-expressing epithelial cells, and EP₂-receptor expressing airway smooth muscle cells. Copyright © 2014 Elsevier Inc. All rights reserved.

  18. High pulsatility flow stimulates smooth muscle cell hypertrophy and contractile protein expression

    PubMed Central

    Scott, Devon; Tan, Yan; Shandas, Robin; Stenmark, Kurt R.

    2013-01-01

    Proximal arterial stiffening is an important predictor of events in systemic and pulmonary hypertension, partly through its contribution to downstream vascular abnormalities. However, much remains undetermined regarding the mechanisms involved in the vascular changes induced by arterial stiffening. We therefore addressed the hypothesis that high pulsatility flow, caused by proximal arterial stiffening, induces downstream pulmonary artery endothelial cell (EC) dysfunction that in turn leads to phenotypic change of smooth muscle cells (SMCs). To test the hypothesis, we employed a model pulmonary circulation in which upstream compliance regulates the pulsatility of flow waves imposed onto a downstream vascular mimetic coculture composed of pulmonary ECs and SMCs. The effects of high pulsatility flow on SMCs were determined both in the presence and absence of ECs. In the presence of ECs, high pulsatility flow increased SMC size and expression of the contractile proteins, smooth muscle α-actin (SMA) and smooth muscle myosin heavy chain (SM-MHC), without affecting proliferation. In the absence of ECs, high pulsatility flow decreased SMC expression of SMA and SM-MHC, without affecting SMC size or proliferation. To identify the molecular signals involved in the EC-mediated SMC responses, mRNA and/or protein expression of vasoconstrictors [angiotensin-converting enzyme (ACE) and endothelin (ET)-1], vasodilator (eNOS), and growth factor (TGF-β1) in EC were examined. Results showed high pulsatility flow decreased eNOS and increased ACE, ET-1, and TGF-β1 expression. ACE inhibition with ramiprilat, ET-1 receptor inhibition with bosentan, and treatment with the vasodilator bradykinin prevented flow-induced, EC-dependent SMC changes. In conclusion, high pulsatility flow stimulated SMC hypertrophy and contractile protein expression by altering EC production of vasoactive mediators and cytokines, supporting the idea of a coupling between proximal vascular stiffening, flow

  19. A multiscale active structural model of the arterial wall accounting for smooth muscle dynamics.

    PubMed

    Coccarelli, Alberto; Edwards, David Hughes; Aggarwal, Ankush; Nithiarasu, Perumal; Parthimos, Dimitris

    2018-02-01

    Arterial wall dynamics arise from the synergy of passive mechano-elastic properties of the vascular tissue and the active contractile behaviour of smooth muscle cells (SMCs) that form the media layer of vessels. We have developed a computational framework that incorporates both these components to account for vascular responses to mechanical and pharmacological stimuli. To validate the proposed framework and demonstrate its potential for testing hypotheses on the pathogenesis of vascular disease, we have employed a number of pharmacological probes that modulate the arterial wall contractile machinery by selectively inhibiting a range of intracellular signalling pathways. Experimental probes used on ring segments from the rabbit central ear artery are: phenylephrine, a selective α 1-adrenergic receptor agonist that induces vasoconstriction; cyclopiazonic acid (CPA), a specific inhibitor of sarcoplasmic/endoplasmic reticulum Ca 2+ -ATPase; and ryanodine, a diterpenoid that modulates Ca 2+ release from the sarcoplasmic reticulum. These interventions were able to delineate the role of membrane versus intracellular signalling, previously identified as main factors in smooth muscle contraction and the generation of vessel tone. Each SMC was modelled by a system of nonlinear differential equations that account for intracellular ionic signalling, and in particular Ca 2+ dynamics. Cytosolic Ca 2+ concentrations formed the catalytic input to a cross-bridge kinetics model. Contractile output from these cellular components forms the input to the finite-element model of the arterial rings under isometric conditions that reproduces the experimental conditions. The model does not account for the role of the endothelium, as the nitric oxide production was suppressed by the action of L-NAME, and also due to the absence of shear stress on the arterial ring, as the experimental set-up did not involve flow. Simulations generated by the integrated model closely matched experimental

  20. Developmental differences in the contractile response of isolated ovine tracheal smooth muscle cells

    PubMed Central

    Laudadio, Rachel E; Wolfson, Marla R; Shaffer, Thomas H; Driska, Steven P

    2015-01-01

    Previous studies have shown developmental differences in smooth muscle tone of the airways. Differences in airway mechanics may be based upon cellular differences between animals of different ages. We developed a method [JAP 86(1): 427–35, 1999] for isolating ovine tracheal smooth muscle cells and for measuring shortening velocity. This technique was used to study the change in contractile response of the airway smooth muscle cell during development. Here we report differences between preterm (110–124 or 125–140 days post-conception), newborn (3–7 days postnatal) and adult (9–36 months) cells. These cells were compared with respect to morphometry, shortening velocity, and percent shortening. The neonatal cells were shorter and narrower than the adult cells. Maximum shortening velocity was faster for adult (45.1 μm/sec) than for neonatal cells (range 11.1 to 25.1 μm/s). When velocity was normalized to the cell length, there was no difference between the adult and preterm cells, but there was a significant difference between the newborn (0.30 sec−1) and adult (0.54 sec−1) cells. The percent shortening did not show any significant difference with age. Within the neonatal groups, there were no significant differences in morphometry, shortening or velocity. To facilitate comparison between ASM tissues of different sizes with different sized cells, we also expressed percent shortening and velocities relative to a hypothetical 1mm segment of tissue. Represented this way, the amount of shortening for all age groups was the same, but the predicted maximum velocity of the hypothetical preterm tissue (125–140 days) was significantly greater than for newborn. PMID:19434687

  1. The expression of functional postsynaptic α2-adrenoceptors in the corpus cavernosum smooth muscle

    PubMed Central

    Gupta, Sandeep; Moreland, Robert B; Yang, Stone; Gallant, Cynthia M; Goldstein, Irwin; Traish, Abdulmaged

    1998-01-01

    The purpose of this study was to determine if corpus cavernosum smooth muscle expresses functional postsynaptic α2-adrenoceptors (AR).The α2-adrenoceptor agonist UK 14,304 elicited concentration-dependent contractions in rabbit corpus cavernosum smooth muscle (CCSM). The half-maximal response occurred at 0.32±0.03 μM and the maximum contraction at 10 μM UK 14,304.Pretreatment of CCSM strips with selective α2-adrenoceptor antagonists, rauwolscine and RS-15385, produced rightward shifts in the dose-response curves to UK 14,304 (pA2 values 7.1 and 8.5, respectively). In contrast, these antagonists did not alter contraction induced by the α1-adrenoceptor agonist phenylephrine (PE) or oxymetazoline. UK 14,304-induced contractions were also inhibited by prazosin (pA2=9.08).UK 14,304-induced contractions, unlike those to PE, were highly dependent on the presence of extracellular Ca2+.[3H]-rauwolscine bound to CCSM membranes with high affinity (Kd=1.5 nM). [3H]-rauwolscine binding was displaced by unlabelled rauwolscine, RS-15385, UK 14,304 and prazosin, but not by PE.UK 14,304 inhibited forskolin and prostaglandin E1 (PGE1)-induced increases in intracellular cyclic AMP concentration in primary cultures of rabbit CCSM cells.These results demonstrate that CCSM expresses Gi-coupled postsynaptic α2-adrenoceptors, and activation of these receptors causes contraction of trabecular smooth muscle. PMID:9559910

  2. Effect of periurethral denervation on smooth muscles of the lower urinary tract.

    PubMed

    Wai, Clifford Y; Liehr, Peter; Boreham, Muriel K; Schaffer, Joseph I; Word, R Ann

    2004-12-01

    This study was undertaken to determine the effect of periurethral denervation on contractile function of the smooth muscle of the lower urinary tract of the female rat. Periurethral nerve transection or sham operation was performed in 35 young female rats. Contractile function of the bladder dome and base was determined as a function of time after surgery. Statistical analysis was conducted by Student t test. Periurethral denervation resulted in impaired contractile responses to electrical field stimulation in the bladder base (nerve-transected 45 +/- 11 g/cm 2 ; sham 84 +/- 10 g/cm 2 , P < .05) and dome (nerve-transected 179 +/- 16 g/cm 2 ; sham 334 +/- 29 g/cm 2 , P < .05) 2 weeks after nerve transection. The ability to respond to potassium chloride and the muscarinic agonist, carbachol, and the rates of contraction and relaxation, however, remained intact. Baseline phasic contractile activity was increased significantly in bladders from nerve-transected animals. Maximal field-stimulated contractions of the longitudinal urethra smooth muscle were not altered by periurethral denervation (sham 21 +/- 6 g/cm 2 , nerve-transected 19 +/- 5 g/cm 2 , P = .4). Compromised nerve-mediated contractions of the bladder dome and base improved significantly by 21 days. Periurethral nerve transection results in transient impairment of neurogenic contractile responses in the bladder base and dome, though the intrinsic ability of the bladder to contract remains intact. This compromised response of the dome, in conjunction with previous results demonstrating impaired urethral smooth muscle relaxation, suggests that transection of periurethral neurons may have complex effects on the entire lower urinary tract.

  3. Phenotypic modulation of corpus cavernosum smooth muscle cells in a rat model of cavernous neurectomy.

    PubMed

    Yang, Fan; Zhao, Jian F; Shou, Qi Y; Huang, Xiao J; Chen, Gang; Yang, Ke B; Zhang, Shi G; Lv, Bo D; Fu, Hui Y

    2014-01-01

    Patients undergoing radical prostatectomy (RP) are at high risk for erectile dysfunction (ED) due to potential cavernous nerve (CN) damage during surgery. Penile hypoxia after RP is thought to significantly contribute to ED pathogenesis. We previously showed that corpora cavernosum smooth muscle cells (CCSMCs) undergo phenotypic modulation under hypoxic conditions in vitro. Here, we studied such changes in an in vivo post-RP ED model by investigating CCSMCs in bilateral cavernous neurectomy (BCN) rats. Sprague-Dawley rats underwent sham (n = 12) or BCN (n = 12) surgery. After 12 weeks, they were injected with apomorphine to determine erectile function. The penile tissues were harvested and assessed for fibrosis using Masson trichrome staining and for molecular markers of phenotypic modulation using immunohistochemistry and western blotting. CCSMC morphological structure was evaluated by hematoxylin-eosin (H&E) staining and transmission electron microscopy (TEM). Erectile function was significantly lower in BCN rats than in sham rats. BCN increased hypoxia-inducible factor-1α and collagen protein expression in corpora cavernous tissue. H&E staining and TEM showed that CCSMCs in BCN rats underwent hypertrophy and showed rough endoplasmic reticulum formation. The expression of CCSMC phenotypic markers, such as smooth muscle α-actin, smooth muscle myosin heavy chain, and desmin, was markedly lower, whereas vimentin protein expression was significantly higher in BCN rats than in control rats. CCSMCs undergo phenotype modulation in rats with cavernous neurectomy. The results have unveiled physiological transformations that occur at the cellular and molecular levels and have helped characterize CN injury-induced ED.

  4. Control of stomach smooth muscle development and intestinal rotation by transcription factor BARX1

    PubMed Central

    Jayewickreme, Chenura D.; Shivdasani, Ramesh A.

    2015-01-01

    Diverse functions of the homeodomain transcription factor BARX1 include Wnt-dependent, non-cell autonomous specification of the stomach epithelium, tracheo-bronchial septation, and Wnt-independent expansion of the spleen primordium. Tight spatio-temporal regulation of Barx1 levels in the mesentery and stomach mesenchyme suggests additional roles. To determine these functions, we forced constitutive BARX1 expression in the Bapx1 expression domain, which includes the mesentery and intestinal mesenchyme, and also examined Barx1−/− embryos in further detail. Transgenic embryos invariably showed intestinal truncation and malrotation, in part reflecting abnormal left-right patterning. Ectopic BARX1 expression did not affect intestinal epithelium, but intestinal smooth muscle developed with features typical of the stomach wall. BARX1, which is normally restricted to the developing stomach, drives robust smooth muscle expansion in this organ by promoting proliferation of myogenic progenitors at the expense of other sub-epithelial cells. Undifferentiated embryonic stomach and intestinal mesenchyme showed modest differences in mRNA expression and BARX1 was sufficient to induce much of the stomach profile in intestinal cells. However, limited binding at cis-regulatory sites implies that BARX1 may act principally through other transcription factors. Genes expressed ectopically in BARX1+ intestinal mesenchyme and reduced in Barx1−/− stomach mesenchyme include Isl1, Pitx1, Six2 and Pitx2, transcription factors known to control left-right patterning and influence smooth muscle development. The sum of evidence suggests that potent BARX1 functions in intestinal rotation and stomach myogenesis occur through this small group of intermediary transcription factors. PMID:26057579

  5. Control of stomach smooth muscle development and intestinal rotation by transcription factor BARX1.

    PubMed

    Jayewickreme, Chenura D; Shivdasani, Ramesh A

    2015-09-01

    Diverse functions of the homeodomain transcription factor BARX1 include Wnt-dependent, non-cell autonomous specification of the stomach epithelium, tracheo-bronchial septation, and Wnt-independent expansion of the spleen primordium. Tight spatio-temporal regulation of Barx1 levels in the mesentery and stomach mesenchyme suggests additional roles. To determine these functions, we forced constitutive BARX1 expression in the Bapx1 expression domain, which includes the mesentery and intestinal mesenchyme, and also examined Barx1(-/)(-) embryos in further detail. Transgenic embryos invariably showed intestinal truncation and malrotation, in part reflecting abnormal left-right patterning. Ectopic BARX1 expression did not affect intestinal epithelium, but intestinal smooth muscle developed with features typical of the stomach wall. BARX1, which is normally restricted to the developing stomach, drives robust smooth muscle expansion in this organ by promoting proliferation of myogenic progenitors at the expense of other sub-epithelial cells. Undifferentiated embryonic stomach and intestinal mesenchyme showed modest differences in mRNA expression and BARX1 was sufficient to induce much of the stomach profile in intestinal cells. However, limited binding at cis-regulatory sites implies that BARX1 may act principally through other transcription factors. Genes expressed ectopically in BARX1(+) intestinal mesenchyme and reduced in Barx1(-/-) stomach mesenchyme include Isl1, Pitx1, Six2 and Pitx2, transcription factors known to control left-right patterning and influence smooth muscle development. The sum of evidence suggests that potent BARX1 functions in intestinal rotation and stomach myogenesis occur through this small group of intermediary transcription factors. Copyright © 2015 Elsevier Inc. All rights reserved.

  6. Loss of neurotrophin-3 from smooth muscle disrupts vagal gastrointestinal afferent signaling and satiation

    PubMed Central

    Biddinger, Jessica E.; Baquet, Zachary C.; Jones, Kevin R.; McAdams, Jennifer

    2013-01-01

    A large proportion of vagal afferents are dependent on neurotrophin-3 (NT-3) for survival. NT-3 is expressed in developing gastrointestinal (GI) smooth muscle, a tissue densely innervated by vagal mechanoreceptors, and thus could regulate their survival. We genetically ablated NT-3 from developing GI smooth muscle and examined the pattern of loss of NT-3 expression in the GI tract and whether this loss altered vagal afferent signaling or feeding behavior. Meal-induced c-Fos activation was reduced in the solitary tract nucleus and area postrema in mice with a smooth muscle-specific NT-3 knockout (SM-NT-3KO) compared with controls, suggesting a decrease in vagal afferent signaling. Daily food intake and body weight of SM-NT-3KO mice and controls were similar. Meal pattern analysis revealed that mutants, however, had increases in average and total daily meal duration compared with controls. Mutants maintained normal meal size by decreasing eating rate compared with controls. Although microstructural analysis did not reveal a decrease in the rate of decay of eating in SM-NT-3KO mice, they ate continuously during the 30-min meal, whereas controls terminated feeding after 22 min. This led to a 74% increase in first daily meal size of SM-NT-3KO mice compared with controls. The increases in meal duration and first meal size of SM-NT-3KO mice are consistent with reduced satiation signaling by vagal afferents. This is the first demonstration of a role for GI NT-3 in short-term controls of feeding, most likely involving effects on development of vagal GI afferents that regulate satiation. PMID:24068045

  7. Concomitant Epstein-Barr Virus-associated smooth muscle tumor and granulomatous inflammation of the liver.

    PubMed

    Can, Nhu Thuy; Grenert, James P; Vohra, Poonam

    2017-10-01

    Epstein-Barr Virus-associated smooth muscle tumor (EBV-SMT) is a rare mesenchymal tumor typically seen in immunocompromised patients. Here, we report a case of EBV-SMT and associated granulomatous inflammation in the liver of a 32-year-old man with history of human immunodeficiency virus/acquired immune deficiency syndrome (HIV/AIDS). To our knowledge, an association of these two lesions has not been previously reported. We review the literature and discuss pathogenesis, differential diagnosis and immunohistochemical (IHC) stains helpful for the diagnosis of this rare entity. Finally, we consider possible explanations for the concomitant presence of these lesions. Published by Elsevier GmbH.

  8. Resveratrol causes cell cycle arrest, decreased collagen synthesis, and apoptosis in rat intestinal smooth muscle cells.

    PubMed

    Garcia, Patricia; Schmiedlin-Ren, Phyllissa; Mathias, Jason S; Tang, Huaijing; Christman, Gregory M; Zimmermann, Ellen M

    2012-02-01

    One of the most difficult and treatment-resistant complications of Crohn's disease is the development of fibrotic intestinal strictures due to mesenchymal cell hyperplasia and collagen deposition. Resveratrol, a phytoalexin found in berries, peanuts, grapes, and red wine, has been shown to inhibit fibrosis in vasculature, heart, lung, kidney, liver, and esophagus in animal models. Resveratrol has also been shown to inhibit oxidation, inflammation, and cell proliferation and to decrease collagen synthesis in several cell types or animal models. The aim of this study was to determine whether resveratrol has antifibrotic effects on intestinal smooth muscle cells. Responses to resveratrol by cultured smooth muscle cells isolated from colons of untreated Lewis rats were examined; this rat strain is used in a model of Crohn's disease with prominent intestinal fibrosis. A relative decrease in cell numbers following treatment with 50 and 100 μM resveratrol was evident at 24 h (P ≤ 0.005). This effect was largely due to cell cycle arrest, with an increase in the percent of cells in S phase from 8 to 25-35% (P < 0.05). Cell viability was unchanged until 2-3 days of treatment when there was a 1.2- to 5.0-fold increase in the percent of apoptotic cells, depending on the assay (P < 0.05). Expression of collagen type I protein was decreased following treatment with resveratrol for 24 h (to 44 and 25% of control levels with 50 and 100 μM resveratrol, respectively; P < 0.05). Expression of procollagen types I and III mRNA was also decreased with resveratrol treatment. Resveratrol (50 μM) diminished the proliferative response to TGF-β₁ (P = 0.02) as well as IGF-I-stimulated collagen production (P = 0.02). Thus resveratrol decreases intestinal smooth muscle cell numbers through its effects on cell cycle arrest and apoptosis and also decreases collagen synthesis by the cells. These effects could be useful in preventing the smooth muscle cell hyperplasia and collagen

  9. Smooth muscle myopathy as an underrecognized manifestation of active systemic lupus erythematosus.

    PubMed

    Ngian, G S; Naidoo, P; Morand, E F; Hoi, A Y

    2011-06-01

    Systemic lupus erythematosus (SLE) is a systemic autoimmune disease with protean manifestations. We here present a case of unexplained diarrhoea and abdominal pain in a patient with SLE. Investigations revealed dilatation of stomach, small bowel and colonic wall, biliary and pancreatic ducts, renal collecting systems and ureters as well as thoracic aorta and major pulmonary arteries, as manifestations of a smooth muscle myopathy that was responsive to immunosuppressive therapy with cyclosporin A. © 2011 The Authors. Internal Medicine Journal © 2011 Royal Australasian College of Physicians.

  10. Effects of Scleroderma Antibodies and Pooled Human Immunoglobulin on Anal Sphincter and Colonic Smooth Muscle Function

    PubMed Central

    SINGH, JAGMOHAN; COHEN, SIDNEY; MEHENDIRATTA, VAIBHAV; MENDOZA, FABIAN; JIMENEZ, SERGIO A.; DIMARINO, ANTHONY J.; RATTAN, SATISH

    2012-01-01

    BACKGROUND & AIMS Patients with systemic sclerosis (SSc) have impairments in gastrointestinal smooth muscle function. The disorder has been associated with circulating antibodies to cholinergic muscarinic type-3 receptor (M3-R). We investigated whether it is possible to neutralize these antibodies with pooled human immunoglobulin (Ig)Gs (pooledhIgG). METHODS We studied the effects of IgGs purified from patients with SSc (SScIgGs) on cholinergic nerve stimulation in rat colon tissues. We also examined the effects of SScIgGs on M3-R activation by bethanechol (BeCh), M3-R occupancy, and receptor binding using mmunofluorescence, immunoblot, and ELISA analyses of human internal anal sphincter (IAS) smooth muscle cells (hSMCs), before and after administration of pooledhIgG. Functional displacement of M3-R occupancy by the SScIgGs was compared with that of other IgGs during the sustained phase of BeCh-induced contraction of intact smooth muscles from rats. RESULTS SScIgG significantly attenuated neutrally mediated contraction and acetylcholine release in rat colon as well as BeCh-induced sustained contraction of the IAS smooth muscle. In immunofluorescence analysis, SScIgG co-localized with M3-R. In immunoblot and ELISA analyses, M3-R loop-2 peptide and human IAS SMC membrane lysates bound significant amounts of SScIgG, compared with IgGs from healthy individuals and pooledhIgG. Binding was significantly attenuated by application of pooledhIgG, which by itself had no significant effect. Incubation of samples with pooledhIgG, or mixing pooledhIgG with SScIgG before administration to tissues, significantly reduced binding of SScIgG, indicating that pooledhIgG prevents SScIgG blockade of M3-R. CONCLUSIONS In studies of rat and human tissues, pooled human IgGs prevent and reverse the cholinergic dysfunctions associated with the progressive gastrointestinal manifestations of SSc, by neutralizing functional M3-R antibodies present in the circulation of patients with SSc. PMID

  11. [Effect of eosin Y on ATPase activity of actomyosine complex of the uterus smooth muscle].

    PubMed

    Labyntseva, R D; Kosterin, S O

    2005-01-01

    The effect of eosin Y (2,4,5,7 - tetrabromofluorescein; 0.1-100 microM) on ATPase activity smooth muscle actomyosine was studied. The inhibition coefficient i50 of ATPase activity with eosin Y was 0.74 +/- 0.07 microM. The inhibitor decreased V(max) of actomyosine ATPase for ATP, but no influence on affinity of actomyosine for ATP was observed. It is suggested that eosin Y inhibits actomyosine ATPase activity noncompetitively in respect of ATP.

  12. Salicylates and vascular smooth muscle cell proliferation: molecular mechanisms for cell cycle arrest.

    PubMed

    Marra, D E; Liao, J K

    2001-11-01

    Salicylates are effective prophylactic treatment strategies for myocardial infarction and ischemic strokes. Recent evidence suggests that high doses of salicylates may exert direct, platelet-independent effects on the vascular wall. Salicylate and aspirin, in concentrations between 1 and 5 mM, effectively inhibit vascular smooth muscle cell proliferation and DNA synthesis without inducing cellular toxicity or apoptosis. This inhibition is associated with effects on specific cell-cycle regulatory molecules, and may proceed via downregulation of the transcription factor, nuclear factor (NF)-kappaB. High-dose salicylates and selective NF-kappaB inhibitors may, therefore, play an important role in the management of vascular proliferative disorders.

  13. A smooth muscle inhibitory material from the bovine retractor penis and rat anococcygeus muscles.

    PubMed

    Gillespie, J S; Martin, W

    1980-12-01

    1. A material that powerfully inhibits the bovine retractor penis and rat anococcygeus muscles has been extracted from these muscles. The inhibitory activity is unaffected by atropine 10(-6) M, phentolamine 5 x 10(-6) M or propranolol 5 x 10(-6) M. 2. This inhibitory material exists in two forms, a stable but inactive form and an unstable inhibitory form. As isolated the material is in the stable, inactive form and is converted into the active form by a brief exposure to acid. The optimum for conversion is pH 2.0 and the active form, after neutralization, reverts with time to the inactive but can be reactivated by a further exposure to acid. The reversion to the inactive form is temperature sensitive, and is rapid at 37 degrees C. 3. The inhibitory material, both active and inactive, is irreversibly destroyed by 2 min in a boiling water bath or by exposure to U.V. irradiation. 4. The inhibitory material is not confined to tissues known to possess a non-adrenergic non-cholinergic innervation. Similar activity has been detected in extracts os skeletal and cardiac muscle and of the liver. The poorly innervated rat uterus and the non-innervated human umbilical artery, however, gave only small and variable amounts. The possible relationship of this material to non-adrenergic non-cholinergic nerves is discussed.

  14. Stretch-dependent potassium channels in murine colonic smooth muscle cells

    PubMed Central

    Koh, Sang Don; Sanders, Kenton M

    2001-01-01

    Gastrointestinal muscles are able to maintain negative resting membrane potentials in spite of stretch. We investigated whether stretch-dependent K+ channels might contribute to myogenic regulation of smooth muscle cells from the mouse colon. Negative pressure applied to on-cell membrane patches activated K+ channels that were voltage independent and had a slope conductance of 95 pS in symmetrical K+ gradients. The effects of negative pressure on open probability were graded as a function of pressure and reversible when atmospheric pressure was restored. Cell elongation activated K+ channels with the same properties as those activated by negative pressure, suggesting that the channels were stretch-dependent K+ (SDK) channels. Channels with the same properties were maximally activated by patch excision, suggesting that either an intracellular messenger or interactions with the cytoskeleton regulate open probability. Internal 4-aminopyridine, Ca2+ (10−8 to 10−6m), and tetraethylammonium (internal or external) were without effect on SDK channels. Nitric oxide donors (and cell-permeant cGMP analogues) activated SDK channels, suggesting that these channels may mediate a portion of the enteric inhibitory neural response in colonic muscles. In summary, SDK channels are an important conductance expressed by colonic muscle cells. SDK channels may stabilize membrane potential during dynamic changes in cell length and mediate responses to enteric neurotransmitters. PMID:11351024

  15. Gene transfer establishes primacy of striated vs. smooth muscle sarcoglycan complex in limb-girdle muscular dystrophy

    PubMed Central

    Durbeej, Madeleine; Sawatzki, Shanna M.; Barresi, Rita; Schmainda, Kathleen M.; Allamand, Valérie; Michele, Daniel E.; Campbell, Kevin P.

    2003-01-01

    Limb-girdle muscular dystrophy types 2E and F are characterized by skeletal muscle weakness and often cardiomyopathy and are due to mutations in the genes encoding β- and δ-sarcoglycan. We previously demonstrated that loss of sarcoglycans in smooth muscle leads to constrictions of the microvasculature that contributes to the cardiac phenotype. It is unclear how vasculature abnormalities affect skeletal muscle. We injected recombinant β- or δ-sarcoglycan adenoviruses into skeletal muscles of corresponding null mice. We hypothesized that the adenoviruses would not transduce vascular smooth muscle, and we would only target skeletal muscle. Indeed, sustained expression of intact sarcoglycan–sarcospan complex was noted at the sarcolemma, neuromuscular junction, myotendinous junction, and in peripheral nerve, but not in vascular smooth muscle. Gene transfer of the corresponding deleted sarcoglycan gene preserved sarcolemmal integrity, prevented pathological dystrophy and hypertrophy, and protected against exercised-induced damage. We conclude that vascular dysfunction is not a primary cause of β- and δ-sarcoglycan-deficient muscular dystrophy. In addition, we show successful functional rescue of entire muscles after adenovirus-mediated gene delivery. Thus, virus-mediated gene transfer of sarcoglycans to skeletal muscle in combination with pharmacological prevention of cardiomyopathy constitute promising therapeutic strategies for limb-girdle muscular dystrophies. PMID:12851463

  16. Identification of the endogenous smooth muscle myosin phosphatase-associated kinase

    PubMed Central

    MacDonald, Justin A.; Borman, Meredith A.; Murányi, Andrea; Somlyo, Avril V.; Hartshorne, David J.; Haystead, Timothy A. J.

    2001-01-01

    Ca2+ sensitization of smooth muscle contraction involves inhibition of myosin light chain phosphatase (SMPP-1M) and enhanced myosin light chain phosphorylation. Inhibition of SMPP-1M is modulated through phosphorylation of the myosin targeting subunit (MYPT1) by either Rho-associated kinase (ROK) or an unknown SMPP-1M-associated kinase. Activated ROK is predominantly membrane-associated and its putative substrate, SMPP-1M, is mainly myofibrillar-associated. This raises a conundrum about the mechanism of interaction between these enzymes. We present ZIP-like kinase, identified by “mixed-peptide” Edman sequencing after affinity purification, as the previously unidentified SMPP-1M-associated kinase. ZIP-like kinase was shown to associate with MYPT1 and phosphorylate the inhibitory site in intact smooth muscle. Phosphorylation of ZIP-like kinase was associated with an increase in kinase activity during carbachol stimulation, suggesting that the enzyme may be a terminal member of a Ca2+ sensitizing kinase cascade. PMID:11226254

  17. Initial characterization of a peripheral blood mononuclear cell chemoattractant derived from cultured arterial smooth muscle cells.

    PubMed Central

    Valente, A. J.; Fowler, S. R.; Sprague, E. A.; Kelley, J. L.; Suenram, C. A.; Schwartz, C. J.

    1984-01-01

    A mononuclear cell chemoattractant of high specific activity produced by baboon (Papio cynocephalus) aortic medial smooth-muscle cells (SMCs) in culture has been partially characterized. Smooth-muscle cells, between the third and eighth passage, were grown to confluence in Medium 199 containing 10% fetal calf serum and then incubated for 24 hours in either serumless medium (Neuman and Tytell) or Medium 199 containing 0.2% bovine serum albumin. The 24-hour SMC-conditioned medium was fractionated on Sephadex G100-Superfine and potent chemoattractant activity (SMC-CF) eluted in the 10,000-12,000 dalton region. SMC-CF displayed chemotactic and chemokinetic activity for peripheral blood mononuclear cells but not for polymorphonuclear leukocytes. Production of SMC-CF by the cells was significantly inhibited in the presence of cycloheximide, and its activity was abolished after incubation with the bacterial protease subtilisin. Chromatofocusing experiments indicate that SMC-CF is a cationic protein with a pI of greater than 10.5. The role of SMC-CF may play as an inflammatory mediator in monocyte recruitment to the arterial intima in atherogenesis is discussed. PMID:6391189

  18. Thin-film dielectric elastomer sensors to measure the contraction force of smooth muscle cells

    NASA Astrophysics Data System (ADS)

    Araromi, O.; Poulin, A.; Rosset, S.; Favre, M.; Giazzon, M.; Martin-Olmos, C.; Liley, M.; Shea, H.

    2015-04-01

    The development of thin-film dielectric elastomer strain sensors for the characterization of smooth muscle cell (SMC) contraction is presented here. Smooth muscle disorders are an integral part of diseases such as asthma and emphysema. Analytical tools enabling the characterization of SMC function i.e. contractile force and strain, in a low-cost and highly parallelized manner are necessary for toxicology screening and for the development of new and more effective drugs. The main challenge with the design of such tools is the accurate measurement of the extremely low contractile cell forces expected as a result of SMC monolayer contraction (as low as ~ 100 μN). Our approach utilizes ultrathin (~5 μm) and soft elastomer membranes patterned with elastomer-carbon composite electrodes, onto which the SMCs are cultured. The cell contraction induces an in-plane strain in the elastomer membrane, predicted to be in the order 1 %, which can be measured via the change in the membrane capacitance. The cell force can subsequently be deduced knowing the mechanical properties of the elastomer membrane. We discuss the materials and fabrication methods selected for our system and present preliminary results indicating their biocompatibility. We fabricate functional capacitive senor prototypes with good signal stability over the several hours (~ 0.5% variation). We succeed in measuring in-plane strains of 1 % with our fabricated devices with good repeatability and signal to noise ratio.

  19. Smooth muscle origin of postnatal 2nd CVP is pre-determined in early embryo

    PubMed Central

    Liu, Qiaozhen; Zhang, Hui; Tian, Xueying; He, Lingjuan; Huang, Xiuzhen; Tan, Zhen; Yan, Yan; Evans, Sylvia M.; Wythe, Joshua D.; Zhou, Bin

    2017-01-01

    Recent identification of the neonatal 2nd coronary vascular population (2nd CVP) suggests that a subset of these vessels form de novo and mature in the inner myocardial wall of the postnatal heart. However, the origin of smooth muscle cells (SMCs) in the postnatal 2nd CVP remains undetermined. Using a tamoxifen-inducible Wt1-CreER driver and a Rosa26-RFP reporter line, we traced the lineage of epicardial cells to determine if they contribute to SMCs of the 2nd CVP. Late embryonic and postnatal induction of Wt1-CreER activity demonstrated that at these stages Wt1-labeled epicardium does not significantly migrate into the myocardium to form SMCs. However, following tamoxifen treatment at an early embryonic stage (E10.5), we detected Wt1 descendants (epicardium-derived cells, or EPDCs) in the outer myocardial wall at E17.5. When the 2nd CVP forms and remodels at postnatal stage, these early labeled EDPCs re-migrate deep into the inner myocardial wall and contribute to 2nd CVP-SMCs in the adult heart. Our findings reveal that SMCs in the postnatal 2nd CVP are pre-specified as EPDCs from the earliest wave of epicardial cell migration. Rather than the re-activation and migration of epicardial cells at later stages, these resident EPDCs mobilize and contribute to smooth muscle of the 2nd CVP during postnatal development. PMID:26902114

  20. Receptor-based differences in human aortic smooth muscle cell membrane stiffness

    NASA Technical Reports Server (NTRS)

    Huang, H.; Kamm, R. D.; So, P. T.; Lee, R. T.

    2001-01-01

    Cells respond to mechanical stimuli with diverse molecular responses. The nature of the sensory mechanism involved in mechanotransduction is not known, but integrins may play an important role. The integrins are linked to both the cytoskeleton and extracellular matrix, suggesting that probing cells via integrins should yield different mechanical properties than probing cells via non-cytoskeleton-associated receptors. To test the hypothesis that the mechanical properties of a cell are dependent on the receptor on which the stress is applied, human aortic smooth muscle cells were plated, and magnetic beads, targeted either to the integrins via fibronectin or to the transferrin receptor by use of an IgG antibody, were attached to the cell surface. The resistance of the cell to deformation ("stiffness") was estimated by oscillating the magnetic beads at 1 Hz by use of single-pole magnetic tweezers at 2 different magnitudes. The ratio of bead displacements at different magnitudes was used to explore the mechanical properties of the cells. Cells stressed via the integrins required approximately 10-fold more force to obtain the same bead displacements as the cells stressed via the transferrin receptors. Cells stressed via integrins showed stiffening behavior as the force was increased, whereas this stiffening was significantly less for cells stressed via the transferrin receptor (P<0.001). Mechanical characteristics of vascular smooth muscle cells depend on the receptor by which the stress is applied, with integrin-based linkages demonstrating cell-stiffening behavior.

  1. Theoretical and experimental investigation of calcium-contraction coupling in airway smooth muscle.

    PubMed

    Mbikou, Prisca; Fajmut, Ales; Brumen, Milan; Roux, Etienne

    2006-01-01

    We investigated theoretically and experimentally the Ca2+-contraction coupling in rat tracheal smooth muscle. [Ca2+]i, isometric contraction and myosin light chain (MLC) phosphorylation were measured in response to 1 mM carbachol. Theoretical modeling consisted in coupling a model of Ca2+-dependent MLC kinase (MLCK) activation with a four-state model of smooth muscle contractile apparatus. Stimulation resulted in a short-time contraction obtained within 1 min, followed by a long-time contraction up to the maximal force obtained in 30 min. ML-7 and Wortmannin (MLCK inhibitors) abolished the contraction. Chelerythrine (PKC inhibitor) did not change the short-time, but reduced the long-time contraction. [Ca2+]i responses of isolated myocytes recorded during the first 90 s consisted in a fast peak, followed by a plateau phase and, in 28% of the cells, superimposed Ca2+ oscillations. MLC phosphorylation was maximal at 5 s and then decreased, whereas isometric contraction followed a Hill-shaped curve. The model properly predicts the time course of MLC phosphorylation and force of the short-time response. With oscillating Ca2+ signal, the predicted force does not oscillate. According to the model, the amplitude of the plateau and the frequency of oscillations encode for the amplitude of force, whereas the peak encodes for force velocity. The long-time phase of the contraction, associated with a second increase in MLC phosphorylation, may be explained, at least partially, by MLC phosphatase (MLCP) inhibition, possibly via PKC inhibition.

  2. Approaches to the mechanism of relaxing effect of vitamin K3 on smooth muscle.

    PubMed

    Altinkurt, O; Oztürk, Y

    1989-01-01

    Smooth muscle relaxing effect of vitamin K3 was tested by using isolated rat Vas deferens, normal and K+-depolarized rat duodenum. The contractions of the rat Vas deferens elicited by noradrenaline and phenylephrine were inhibited by vitamin K3, noncompetitively. Vitamin K3 inhibited the Ca2+-induced contractions of the K+-depolarized rat duodenum in a noncompetitive manner. The inhibitory effect of vitamin K3 on the K+-depolarized rat duodenum was evaluated by comparing it with that of verapamil, a well-known Ca2+-channel blocker and of trifluoperazine, a calmodulin inhibitor. Further, it was observed that vitamin K3 exerts a dose-dependent relaxing effect on the normal rat duodenum. The effect of vitamin K3 was compared with that of adrenaline, isoprenaline and papaverine. In addition, propranolol, a beta-adrenergic blocking agent, and nicotinic acid, an adenylate cyclase inhibitor, were used as tools in order to investigate the mechanism of the relaxing action of vitamin K3 on smooth muscle.

  3. Ethanol increases phosphate-mediated mineralization and osteoblastic transformation of vascular smooth muscle cells

    PubMed Central

    Oros, Melinda; Zavaczki, Erzsebet; Vadasz, Csaba; Jeney, Viktoria; Tosaki, Arpad; Lekli, Istvan; Balla, Gyorgy; Nagy, Laszlo; Balla, Jozsef

    2012-01-01

    Vascular calcification is implicated in the pathogenesis of atherosclerosis, diabetes and chronic kidney disease. Human vascular smooth muscle cells (HSMCs) undergo mineralization in response to elevated levels of inorganic phosphate (Pi) in an active and well-regulated process. This process involves increased activity of alkaline phosphatase and increased expression of core binding factor α-1 (CBF-α1), a bone-specific transcription factor, with the subsequent induction of osteocalcin. It has been shown that heavy alcohol consumption is associated with greater calcification in coronary arteries. The goal of our study was to examine whether ethanol alters mineralization of HSMCs provoked by high Pi. Exposure of HSMCs to ethanol increased extracellular matrix calcification in a dose responsive manner, providing a significant additional calcium deposition at concentrations of ≥60 mmol/l. HSMC calcification was accompanied by further enhancement in alkaline phosphatase activity. Ethanol also provoked a significant increase in the synthesis of osteocalcin. Moreover, in cells challenged with ethanol the expression of CBF-α1, a transcription factor involved in the regulation of osteoblastic transformation of HSMCs, was elevated. The observed effects of ethanol were not due to alterations of phosphate uptake by HSMCs. We conclude that ethanol enhances Pi-mediated human vascular smooth muscle calcification and transition of these cells into osteoblast-like cells. PMID:22260235

  4. Calcification of Vascular Smooth Muscle Cells and Imaging of Aortic Calcification and Inflammation

    PubMed Central

    Hutcheson, Joshua D.; Burke, Megan F.; Martyn, Trejeeve; Thayer, Timothy E.; Shakartzi, Hannah R.; Buswell, Mary D.; Tainsh, Robert E.; Yu, Binglan; Bagchi, Aranya; Rhee, David K.; Wu, Connie; Derwall, Matthias; Buys, Emmanuel S.; Yu, Paul B.; Bloch, Kenneth D.; Aikawa, Elena; Bloch, Donald B.; Malhotra, Rajeev

    2016-01-01

    Cardiovascular disease is the leading cause of morbidity and mortality in the world. Atherosclerotic plaques, consisting of lipid-laden macrophages and calcification, develop in the coronary arteries, aortic valve, aorta, and peripheral conduit arteries and are the hallmark of cardiovascular disease. In humans, imaging with computed tomography allows for the quantification of vascular calcification; the presence of vascular calcification is a strong predictor of future cardiovascular events. Development of novel therapies in cardiovascular disease relies critically on improving our understanding of the underlying molecular mechanisms of atherosclerosis. Advancing our knowledge of atherosclerotic mechanisms relies on murine and cell-based models. Here, a method for imaging aortic calcification and macrophage infiltration using two spectrally distinct near-infrared fluorescent imaging probes is detailed. Near-infrared fluorescent imaging allows for the ex vivo quantification of calcification and macrophage accumulation in the entire aorta and can be used to further our understanding of the mechanistic relationship between inflammation and calcification in atherosclerosis. Additionally, a method for isolating and culturing animal aortic vascular smooth muscle cells and a protocol for inducing calcification in cultured smooth muscle cells from either murine aortas or from human coronary arteries is described. This in vitro method of modeling vascular calcification can be used to identify and characterize the signaling pathways likely important for the development of vascular disease, in the hopes of discovering novel targets for therapy. PMID:27284788

  5. 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.

  6. Arginase I attenuates inflammatory cytokine secretion induced by lipopolysaccharide in vascular smooth muscle cells.

    PubMed

    Wang, Xu-ping; Chen, Yu-guo; Qin, Wei-dong; Zhang, Wei; Wei, Shu-jian; Wang, Juan; Liu, Fu Qiang; Gong, Lei; An, Feng Shuang; Zhang, Yun; Chen, Zhe-Yu; Zhang, Ming-Xiang

    2011-08-01

    Inflammation plays an important role in atherosclerosis. Arginase I (Arg I) promotes the proliferation of vascular smooth muscle cells; however, the effect of Arg I on inflammation remains unknown. The present study investigated the role of Arg I in inflammation in vitro and in vivo. Quantitative reverse transcription-polymerase chain reaction and Western blot analysis demonstrated that Arg I inhibited tumor necrosis factor-α production induced by lipopolysaccharide in human aortic smooth muscle cells. Inducible nitric oxide synthase substrate competition and nuclear factor-κB activation were main contributors to lipopolysaccharide-mediated inflammatory cytokine generation. However, Arg I could attenuate the function of inducible nitric oxide synthase and inhibit the subsequent nuclear factor-κB activation, leading to inhibition of tumor necrosis factor-α generation. Furthermore, upregulation of Arg I significantly decreased macrophage infiltration and inflammation in atherosclerotic plaque of rabbits, whereas downregulation of Arg I aggravated these adverse effects. The results indicate the antiinflammatory effects of Arg I and suggest an unexpected beneficial role of Arg I in inflammatory disease.

  7. Human Aortic Smooth Muscle Cells Promote Arteriole Formation by Coengrafted Endothelial Cells

    PubMed Central

    Shepherd, Benjamin R.; Jay, Steven M.; Saltzman, W. Mark; Tellides, George

    2009-01-01

    Collagen-fibronectin gels containing Bcl-2–transduced human umbilical vein endothelial cells (Bcl-2-HUVEC) implanted in the abdominal walls of immunodeficient mice form mature microvessels invested by host-derived smooth muscle cells (SMC) by 8 weeks. We tested the hypothesis that coengraftment of human aortic SMC (HASMC) could accelerate vessel maturation. To prevent SMC-mediated gel contraction, we polymerized the gel within a nonwoven poly(glycolic acid) (PGA) scaffold. Implanted grafts were evaluated at 15, 30, and 60 days. Acellular PGA-supported protein gels elicited a macrophage-rich foreign body reaction and transient host angiogenic response. When transplanted alone, HASMC tightly associated with the fibers of the scaffold and incorporated into the walls of angiogenic mouse microvessels, preventing their regression. When transplanted alone in PGA-supported gels, Bcl-2-HUVEC retained the ability to form microvessels invested by mouse SMC. Interestingly, grafts containing both Bcl-2-HUVEC and HASMC displayed greater numbers of smooth muscle α-actin–expressing cells associated with human EC-lined arteriole-like microvessels at all times examined and showed a significant increase in the number of larger caliber microvessels at 60 days. We conclude that SMC coengraftment can accelerate vessel development by EC and promote arteriolization. This strategy of EC-SMC coengraftment in PGA-supported protein gels may have broader application for perfusing bioengineered tissues. PMID:18620481

  8. [Migration and proliferation of smooth muscle cells in the vessel wall].

    PubMed

    Betz, E

    1990-03-01

    1. Intimal migration and proliferation causing artery stenoses in the course of atherogenesis can be inhibited by various drugs. 2. Secondary stenoses after ballooning of arteries are caused mainly by proliferation of smooth muscle cells. 3. Ballooning of arteries or repeated transmural electrical stimulations of artery walls with weak electrical current is followed by an increased mitotic activity of smooth muscle cells in the ballooned resp. stimulated area which reaches a maximal value about one week following the onset of the experiment. The mitotic activity returns then slowly to initial levels. 4. Adaptations to proliferation-inducing stimuli are possible. The experiments demonstrate that the proliferative phases in atherogenesis can be explained as a sequence of adaptations and deadaptations (= change of disposition) to the proliferation-inducing stimuli. 5. To select qualified drugs for an inhibition of the development of intimal proliferates in the course of atherogenesis makes it necessary to combine in vivo tests in animal experiments with tests on cell cultures of human cells from artery walls.

  9. TTX-sensitive voltage-gated Na+ channels are expressed in mesenteric artery smooth muscle cells.

    PubMed

    Berra-Romani, Roberto; Blaustein, Mordecai P; Matteson, Donald R

    2005-07-01

    The presence and properties of voltage-gated Na+ channels in mesenteric artery smooth muscle cells (SMCs) were studied using whole cell patch-clamp recording. SMCs from mouse and rat mesenteric arteries were enzymatically dissociated using two dissociation protocols with different enzyme combinations. Na+ and Ca2+ channel currents were present in myocytes isolated with collagenase and elastase. In contrast, Na+ currents were not detected, but Ca2+ currents were present in cells isolated with papain and collagenase. Ca2+ currents were blocked by nifedipine. The Na+ current was insensitive to nifedipine, sensitive to changes in the extracellular Na+ concentration, and blocked by tetrodotoxin with an IC50 at 4.3 nM. The Na+ conductance was half maximally activated at -16 mV, and steady-state inactivation was half-maximal at -53 mV. These values are similar to those reported in various SMC types. In the presence of 1 microM batrachotoxin, the Na+ conductance-voltage relationship was shifted by 27 mV in the hyperpolarizing direction, inactivation was almost completely eliminated, and the deactivation rate was decreased. The present study indicates that TTX-sensitive, voltage-gated Na+ channels are present in SMCs from the rat and mouse mesenteric artery. The presence of these channels in freshly isolated SMC depends critically on the enzymatic dissociation conditions. This could resolve controversy about the presence of Na+ channels in arterial smooth muscle.

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

    PubMed

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

    2000-04-27

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

  11. ROCK Isoform Regulation of Myosin Phosphatase and Contractility in Vascular Smooth Muscle Cells

    PubMed Central

    Wang, Yuepeng; Zheng, Xiaoyu Rayne; Riddick, Nadeene; Bryden, Meredith; Baur, Wendy; Zhang, Xin; Surks, Howard K.

    2009-01-01

    Abnormal VSMC contraction plays an important role in vascular diseases. The RhoA/ROCK signaling pathway is now well-recognized to mediate vascular smooth muscle contraction in response to vasoconstrictors by inhibiting myosin phosphatase (MLCP) activity and increasing myosin light chain (MLC) phosphorylation. Two ROCK isoforms, ROCK1 and ROCK2, are expressed in many tissues, yet the isoform specific roles of ROCK1 and ROCK2 in vascular smooth muscle (VSM) and the mechanism of ROCK-mediated regulation of MLCP are not well understood. In this study, ROCK2, but not ROCK1, bound directly to the myosin binding subunit (MBS) of MLCP, yet both ROCK isoforms regulated MLCP and MLC phosphorylation. Despite that both ROCK1 and ROCK2 regulated MLCP, the ROCK isoforms had distinct and opposing effects on VSMC morphology and ROCK2, but not ROCK1, had a predominant role in VSMC contractility. These data support that although the ROCK isoforms both regulate MLCP and MLC phosphorylation through different mechanisms, they have distinct roles in VSMC function. PMID:19131646

  12. Silencing miR-16 Expression Promotes Angiotensin II Stimulated Vascular Smooth Muscle Cell Growth

    PubMed Central

    Gu, Qingqing; Zhao, Guannan; Wang, Yinan; Xu, Biao; Yue, Junming

    2017-01-01

    miRNAs are a class of non-coding endogenous small RNAs that control gene expression at the posttranscriptional level and involved in cell proliferation, migration and differentiation. Dysregulation of miRNA expression is involved in a variety of human diseases including cardiovascular diseases. miRNAs have been shown to regulate vascular smooth muscle cell (VSMC) function and play vital roles in hypertension, restenosis and atherosclerosis. Here we reported that miR-16 as one of miRNAs in the miR-15 family was highly expressed in vascular smooth muscle cells (VSMCs) and involved in angiotensin II (Ang II) mediated VSMC signaling pathways. Ang II downregulated miR-16 expression in VSMCs. Lentiviral vector mediated miR-16 knockdown promoted Ang II-induced cell proliferation and migration. Moreover, silencing miR-16 enhanced Ang II induced cell cycle associated gene expression and promoted Ang II-activated cell proliferative pathways ERK1/2 and p38. Our finding demonstrated for the first time that miR-16 was a potential therapeutic target by participating in the Ang II-associated multiple signaling pathways in cardiovascular diseases. PMID:29104843

  13. Ayurvedic preparation of Zingiber officinale Roscoe: effects on cardiac and on smooth muscle parameters.

    PubMed

    Leoni, Alberto; Budriesi, Roberta; Poli, Ferruccio; Lianza, Mariacaterina; Graziadio, Alessandra; Venturini, Alice; Broccoli, Massimiliano; Micucci, Matteo

    2017-08-28

    The rhizome of the Zingiber officinale Roscoe, a biennial herb growing in South Asia, is commonly known as ginger. Ginger is used in clinical disorders, such as constipation, dyspepsia, diarrhoea, nausea and vomiting and its use is also recommended by the traditional medicine for cardiopathy, high blood pressure, palpitations and as a vasodilator to improve the circulation. The decoction of ginger rhizome is widely used in Ayurvedic medicine. In this papery by high-performance liquid chromatography, we have seen that its main phytomarkers were 6-gingerol, 8-gingerol and 6-shogaol and we report the effects of the decoction of ginger rhizome on cardiovascular parameters and on vascular and intestinal smooth muscle. In our experimental models, the decoction of ginger shows weak negative inotropic and chronotropic intrinsic activities but a significant intrinsic activity on smooth muscle with a potency on ileum is greater than on aorta: EC 50  = 0.66 mg/mL versus EC 50  = 1.45 mg/mL.

  14. Wood creosote inhibits calcium mobilization in Guinea pig colonic smooth muscle.

    PubMed

    Morino, Hirofumi; Ataka, Koji; Ito, Masafumi; Kuge, Tomoo

    2004-07-01

    Wood creosote, a mixture of simple phenolic compounds, has long been used as an herbal antidiarrheal medicine. Previous studies have shown that wood creosote has antimotility activity on the gastrointestinal (GI) tract, although its mechanism of action is not completely understood. The in vitro efficacy of wood creosote on calcium mobilization in guinea pig colonic smooth muscle was evaluated using a digital video camera system mounted on an inverted fluorescence microscope. The effects of wood creosote on spontaneous periodic increases in the free cytosolic calcium concentration ([Ca(2+)](i)), acetylcholine (ACh)-enhanced periodic increases in [Ca(2+)](i), and tetrodotoxin- or nifedipine-resistant spontaneous periodic increases in [Ca(2+)](i) were evaluated. Wood creosote decreased the amplitude of spontaneous (IC(50)=21 microg/ml) and ACh-enhanced (IC(50)=40 microg/ml) periodic increases in [Ca(2+)](i) in guinea pig colonic smooth muscle. Wood creosote also decreased the amplitude of both tetrodotoxin- and nifedipine-resistant spontaneous periodic increases in [Ca(2+)](i). These results suggest that antimotility activity through inhibition of Ca(2+) mobilization in the GI tract is at least partially responsible for the antidiarrheal activity of wood creosote. Wood creosote may exert its antimotility effect, at least in part, on network regions of interstitial cells of Cajal, which act as pacemaker cells and mediators of neurotransmission in the GI tract.

  15. Calcium-activated chloride channels anoctamin 1 and 2 promote murine uterine smooth muscle contractility

    PubMed Central

    Bernstein, Kyra; Vink, Joy Y; Fu, Xiao Wen; Wakita, Hiromi; Danielsson, Jennifer; Wapner, Ronald; Gallos, George

    2014-01-01

    Objective To determine the presence of calcium activated chloride channels anoctamin 1 and 2 in human and murine uterine smooth muscle and evaluate the physiologic role for these ion channels in murine myometrial contractility. Study Design We performed reverse transcription polymerase chain reaction to determine if anoctamin 1 and 2 are expressed in human and murine uterine tissue to validate the study of this protein in mouse models. Immunohistochemical staining of anoctamin 1 and 2 was then performed to determine protein expression in murine myometrial tissue. The function of anoctamin 1 and 2 in murine uterine tissue was evaluated using electrophysiological studies, organ bath, and calcium flux experiments. Results Anoctamin 1 and 2 are expressed in human and murine USM cells. Functional studies show that selective antagonism of these channels promotes relaxation of spontaneous murine uterine smooth muscle contractions. Blockade of anoctamin 1 and 2 inhibits both agonist-induced and spontaneous transient inward currents and abolishes G-protein coupled receptor (oxytocin) mediated elevations in intracellular calcium. Conclusion The calcium activated chloride channels ANO 1 and 2 are present in human and murine myometrial tissue and may provide novel potential therapeutic targets to achieve effective tocolysis. PMID:24928056

  16. A Potential Gravity-Sensing Role of Vascular Smooth Muscle Cell Glycocalyx in Altered Gravitational Stimulation

    PubMed Central

    Kang, Hongyan; Liu, Meili

    2013-01-01

    Abstract Previously, we have shown that vascular smooth muscle cells (VSMCs) exhibit varied physiological responses when exposed to altered gravitational conditions. In the present study, we focused on elucidating whether the cell surface glycocalyx could be a potential gravity sensor. For this purpose, a roller culture apparatus was used with the intent to provide altered gravitational conditions to cultured rat aortic smooth muscle cells (RASMCs). Heparinase III (Hep.III) was applied to degrade cell surface heparan sulfate proteoglycans (HSPG) selectively. Sodium chlorate was used to suppress new synthesis of HSPG. Glycocalyx remodeling, nitric oxide synthase (NOS) activation, and F-actin expression induced by gravity alteration were assessed by flow cytometry, reverse transcription polymerase chain reaction (RT-PCR), and Western blot. Results indicate that the exposure of cultured RASMCs to altered gravitational conditions led to a reduction in cell surface HSPG content and the activation of NOS. It also down-regulated the expression of glypican-1, constitutive NOS (NOSI and NOSIII), and F-actin. On the other hand, Hep.III followed by sodium chlorate treatment of HSPG attenuated the aforementioned NOS and F-actin modulation under altered gravitational conditions. All these findings suggest that the glycocalyx, and HSPG in particular, may be an important sensor of gravitational changes. This may play an important role in the regulation of NOS activation, F-actin modulation, and HSPG remodeling in VSMCs. Key Words: Glycocalyx—Gravity sensor—Gravity alteration—Roller culture apparatus. Astrobiology 13, 626–636. PMID:23848471

  17. Iptakalim inhibits PDGF-BB-induced human airway smooth muscle cells proliferation and migration

    SciTech Connect

    Liu, Wenrui; Kong, Hui; Zeng, Xiaoning

    2015-08-15

    Chronic airway diseases are characterized by airway remodeling which is attributed partly to the proliferation and migration of airway smooth muscle cells (ASMCs). ATP-sensitive potassium (K{sub ATP}) channels have been identified in ASMCs. Mount evidence has suggested that K{sub ATP} channel openers can reduce airway hyperresponsiveness and alleviate airway remodeling. Opening K{sup +} channels triggers K{sup +} efflux, which leading to membrane hyperpolarization, preventing Ca{sup 2+}entry through closing voltage-operated Ca{sup 2+} channels. Intracellular Ca{sup 2+} is the most important regulator of muscle contraction, cell proliferation and migration. K{sup +} efflux decreases Ca{sup 2+} influx, which consequently influences ASMCs proliferation andmore » migration. As a K{sub ATP} channel opener, iptakalim (Ipt) has been reported to restrain the proliferation of pulmonary arterial smooth muscle cells (PASMCs) involved in vascular remodeling, while little is known about its impact on ASMCs. The present study was designed to investigate the effects of Ipt on human ASMCs and the mechanisms underlying. Results obtained from cell counting kit-8 (CCK-8), flow cytometry and 5-ethynyl-2′-deoxyuridine (EdU) incorporation showed that Ipt significantly inhibited platelet-derived growth factor (PDGF)-BB-induced ASMCs proliferation. ASMCs migration induced by PDGF-BB was also suppressed by Ipt in transwell migration and scratch assay. Besides, the phosphorylation of Ca{sup 2+}/calmodulin-dependent kinase II (CaMKII), extracellular regulated protein kinases 1/2 (ERK1/2), protein kinase B (Akt), and cyclic adenosine monophosphate (cAMP) response element binding protein (CREB) were as well alleviated by Ipt administration. Furthermore, we found that the inhibition of Ipt on the PDGF-BB-induced proliferation and migration in human ASMCs was blocked by glibenclamide (Gli), a selective K{sub ATP} channel antagonist. These findings provide a strong evidence to support that

  18. Human airway smooth muscle maintain in situ cell orientation and phenotype when cultured on aligned electrospun scaffolds

    PubMed Central

    Morris, G. E.; Bridge, J. C.; Eltboli, O. M. I.; Lewis, M. P.; Knox, A. J.; Aylott, J. W.; Brightling, C. E.; Ghaemmaghami, A. M.

    2014-01-01

    Human airway smooth muscle (HASM) contraction plays a central role in regulating airway resistance in both healthy and asthmatic bronchioles. In vitro studies that investigate the intricate mechanisms that regulate this contractile process are predominantly conducted on tissue culture plastic, a rigid, 2D geometry, unlike the 3D microenvironment smooth muscle cells are exposed to in situ. It is increasingly apparent that cellular characteristics and responses are altered between cells cultured on 2D substrates compared with 3D topographies. Electrospinning is an attractive method to produce 3D topographies for cell culturing as the fibers produced have dimensions within the nanometer range, similar to cells' natural environment. We have developed an electrospun scaffold using the nondegradable, nontoxic, polymer polyethylene terephthalate (PET) composed of uniaxially orientated nanofibers and have evaluated this topography's effect on HASM cell adhesion, alignment, and morphology. The fibers orientation provided contact guidance enabling the formation of fully aligned sheets of smooth muscle. Moreover, smooth muscle cells cultured on the scaffold present an elongated cell phenotype with altered contractile protein levels and distribution. HASM cells cultured on this scaffold responded to the bronchoconstrictor bradykinin. The platform presented provides a novel in vitro model that promotes airway smooth muscle cell development toward a more in vivo-like phenotype while providing topological cues to ensure full cell alignment. PMID:24793171

  19. Leukotriene D4 receptor-mediated hydrolysis of phosphoinositide and mobilization of calcium in sheep tracheal smooth muscle cells

    SciTech Connect

    Mong, S.; Miller, J.; Wu, H.L.

    1988-02-01

    A sheep tracheal smooth muscle primary culture cell system was developed to characterize leukotriene D4 (LTD4) receptor-mediated biochemical and pharmacological effects. (/sup 3/H)LTD4 binding to the enriched plasma membrane receptor was specific, stereoselective and saturable. LTE4 and high affinity receptor antagonists bound to the receptors with a rank-order potency that was expected from previous smooth muscle contraction studies. In the (/sup 3/H)myoinositol labeled cells, LTD4 and LTE4 induced phosphoinositide hydrolysis. The biosynthesis of (/sup 3/H)inositol-trisphosphate was rapid and the induction of biosynthesis of (/sup 3/H)inositol-monophosphate by LTs was stereoselective and specific and was inhibited specifically by a receptor antagonist, SKFmore » 104353. In the fura-2 loaded smooth muscle cells, LTD4 and LTE4 induced transient intracellular Ca++ mobilization. The fura-2/Ca++ transient was stereoselective and specific and was inhibited by receptor antagonist, SKF 104353. These results suggest that the cultured sheep tracheal smooth muscle cells have plasma membrane receptors for LTD4. These receptors were coupled to a phospholipase C that, when activated by agonists, induced hydrolysis of inositol containing phospholipids. The hydrolysis products, e.g. diacylglycerol and inositol-trisphosphate, may serve as intracellular messengers that trigger or contribute to the contractile effect in sheep tracheal smooth muscle.« less

  20. Identification of a novel serum and growth factor-inducible gene in vascular smooth muscle cells.

    PubMed

    Feng, P; Liau, G

    1993-05-05

    We have used subtraction cloning to isolate a cDNA (PS4) that identified a serum-inducible mRNA of 1.9 kilobases in rabbit vascular smooth muscle cells. DNA sequence analysis revealed one major open reading frame encoding a 9,442 M(r) protein. Comparison of the DNA as well as the putative protein sequence with various data bases revealed no homology with other sequences. In vitro translation of synthesized PS4 mRNA generated a major polypeptide of 12 kDa. Serum stimulation of quiescent smooth muscle cells in culture induced a rapid increase in the level of PS4 mRNA. Expression of this message was detected by 1 h, peaked at approximately 4 h, and became undetectable by 12 h. The induction of PS4 by serum was completely blocked by cycloheximide, indicating its expression required prior protein synthesis. Epidermal growth factor, acidic fibroblast growth factor, and transforming growth factor-beta 1 also induced a strong increase in PS4 expression. By contrast, platelet-derived growth factor-BB was only able to mildly stimulate the level of PS4 mRNA and insulin-like growth factor-I was unable to enhance PS4 expression. There was a high level of PS4 mRNA in rabbit fetal muscle, esophagus, kidney, and lung, a low level in fetal aorta and heart, and an undetectable level in fetal liver, brain, as well as, in the placenta. The expression of PS4 in the corresponding adult tissues was low or undetectable. Our analysis indicate that PS4 expression is developmentally regulated and tightly controlled by growth factors, suggesting this novel gene has a role in cell growth and differentiation.

  1. Smooth muscle enfoldment internal sphincter construction after intersphincteric resection for rectal cancer.

    PubMed

    Jin, Heiying; Zhang, Bei; Yao, Hang; Du, Yonghong; Wang, Xiaofeng; Leng, Qiang

    2014-01-01

    To assess smooth muscle enfoldment and internal sphincter construction (SMESC) for improvement of continence after intersphincteric resection (ISR) for rectal cancer. Twenty-four Bama miniature pigs were randomly divided into a conventional ISR group and experimental SMESC group, with 12 pigs in each group. The proximal sigmoid colon was anastomosed directly to the anus in the ISR group. In the SMESC group, internal sphincter construction was performed. At 12 weeks before and after surgery, rectal resting pressure and anal canal length were assessed. Three-dimensional ultrasound was used to determine the thickness of the internal sphincter. After the animals were sacrificed, the rectum and anus were resected and pathological examinations were performed to evaluate the differences in sphincter thickness and muscle fibers. All 24 animals in the SMESC group and the ISR group survived the surgery. Twelve weeks post-surgery, the rectal resting pressure, length of the anal high-pressure zone and the postoperative internal sphincter thickness for the ISR group were significantly lower than for the SMESC group. There was a thickened area (about 2 cm) above the anastomotic stoma among animals from the SMESC group; in addition, the smooth muscles were significantly enlarged and enfolded when compared to the ISR group. This animal model study shows that the SMESC procedure achieved acceptable reconstruction of the internal anal neo-sphincter (IAN/S), without increasing surgical risk. However, the findings in this experimental animal model must be confirmed by clinical trials to determine the safety and efficacy of this procedure in clinical practice.

  2. Characterization of the effect of penehyclidine hydrochloride on muscarinic receptor subtypes mediating the contraction of guinea-pig isolated gastrointestinal smooth muscle.

    PubMed

    Xiao, Hong-Tao; Liao, Zhi; Meng, Xian-Min; Yan, Xiao-Yan; Chen, Shu-Jie; Mo, Zheng-Ji

    2009-07-01

    The aim was to characterize the effect of penehyclidine hydrochloride, which mediates the relaxation of guinea-pig isolated gastrointestinal smooth muscle, on muscarinic receptor subtypes. Radioimmune assay was used to determine cAMP levels in isolated guinea-pig gastrointestinal smooth muscle to compare the selective effects of penehyclidine hydrochloride on muscarinic receptor subtypes. The results indicated that the relaxing effect of penehyclidine hydrochloride on isolated gastrointestinal smooth muscle contraction induced by acetylcholine was stronger than that of atropine (based on PA2 values). In the radioimmune assay, penehyclidine hydrochloride increased the cAMP content in isolated guinea-pig stomach smooth muscle and decreased the cAMP content in isolated guinea-pig intestinal smooth muscle, but the difference was not statistically significant at a dose of 10 mumol/l. The results suggest that penehyclidine hydrochloride has little or no effect on M2 receptor subtypes in guinea-pig gastrointestinal smooth muscle.

  3. Development and maintenance of force and stiffness in airway smooth muscle.

    PubMed

    Lan, Bo; Norris, Brandon A; Liu, Jeffrey C-Y; Paré, Peter D; Seow, Chun Y; Deng, Linhong

    2015-03-01

    Airway smooth muscle (ASM) plays a central role in the excessive narrowing of the airway that characterizes the primary functional impairment in asthma. This phenomenon is known as airway hyper-responsiveness (AHR). Emerging evidence suggests that the development and maintenance of ASM force involves dynamic reorganization of the subcellular filament network in both the cytoskeleton and the contractile apparatus. In this review, evidence is presented to support the view that regulation of ASM contraction extends beyond the classical actomyosin interaction and involves processes within the cytoskeleton and at the interfaces between the cytoskeleton, the contractile apparatus, and the extracellular matrix. These processes are initiated when the muscle is activated, and collectively they cause the cytoskeleton and the contractile apparatus to undergo structural transformation, resulting in a more connected and solid state that allows force generated by the contractile apparatus to be transmitted to the extracellular domain. Solidification of the cytoskeleton also serves to stiffen the muscle and hence the airway. Oscillatory strain from tidal breathing and deep inspiration is believed to be the counter balance that prevents hypercontraction and stiffening of ASM in vivo. Dysregulation of this balance could lead to AHR seen in asthma.

  4. P53 Promotes Retinoid Acid-induced Smooth Muscle Cell Differentiation by Targeting Myocardin.

    PubMed

    Tan, Zhou; Li, Jingya; Zhang, Xuejing; Yang, Xueqin; Zhang, Zunyi; Yin, Ke-Jie; Huang, Huarong

    2018-04-15

    TP53 is a widely studied tumor suppressor gene that controls various cellular functions, including cell differentiation. However, little is known about its functional roles in smooth muscle cells (SMCs) differentiation from embryonic stem cells (ESCs). SMC differentiation is at the heart of our understanding of vascular development, normal blood pressure homeostasis, and the pathogenesis of vascular diseases such as atherosclerosis, hypertension, restenosis, as well as aneurysm. Using retinoid acid (RA)-induced SMC differentiation models, we observed that p53 expression is increased during in vitro differentiation of mouse ESCs into SMCs. Meanwhile, suppression of p53 by shRNA reduced RA-induced SMC differentiation. Mechanistically, we have identified for the first time that Myocardin, a transcription factor that induces muscle cell differentiation and muscle-specific gene expression, is the direct target of p53 by bioinformatic analysis, luciferase reporter assay, and chromatin immunoprecipitation approaches. Moreover, in vivo SMC-selective p53 transgenic overexpression inhibited injury-induced neointimal formation. Taken together, our data demonstrate that p53 and its target gene, Myocardin, play regulatory roles in SMC differentiation. This study may lead to the identification of novel target molecules that may, in turn, lead to novel drug discoveries for the treatment of vascular diseases.

  5. Heterogeneous CPA sensitivity of spontaneous excitation in smooth muscle of the rabbit urethra

    PubMed Central

    Hashitani, Hikaru; Yanai, Yoshimasa; Kohri, Kenjiro; Suzuki, Hikaru

    2006-01-01

    To investigate the role of intracellular Ca stores in generating spontaneous excitation of the urethra, the effects of cyclopiazonic acid (CPA) on spontaneous contractions, transient increases in intracellular calcium concentration ([Ca2+]i; Ca transients) and depolarizations were examined in smooth muscles of the rabbit urethra. In about 90% of circular smooth muscle (CSM) preparations, CPA (10 μM) increased the amplitude of spontaneous contractions by about 180% and reduced their frequency to some 25% of control values (CPA-resistant), while it readily abolished the contractions in the remaining preparations. In about 70% of CSM preparations, CPA prevented the generation of spontaneous depolarizations termed slow waves, but increased their amplitude and duration in the remainder. CPA also prevented the generation of spontaneous Ca transients in about 40% of CSM preparations, while increasing their amplitude and duration in the remaining preparations. In CPA-resistant preparations that had been exposed to nicardipine (1 μM), subsequent CPA invariably abolished residual spontaneous depolarizations or Ca transients. CPA abolished caffeine-induced Ca transients in Ca-free solutions, suggesting that it effectively depleted intracellular Ca stores. Longitudinal smooth muscles generated spontaneous action potentials, which had a shape distinct from that of slow waves in CSM. Spontaneous action potentials were abolished by nicardipine but not CPA. Transmural nerve stimulation increased the frequency of Ca transients to give a sustained rise in [Ca2+]i, but inhibited their generation after blocking α-adrenoceptors with phentolamine (1 μM). These nerve-evoked responses were preserved in preparations that had been exposed to CPA. Similarly, both in control and CPA-treated CSM preparations, spontaneous Ca transients were accelerated by noradrenaline (NAd, 1 μM) and were suppressed by 3-morpholino-sydnonimine (SIN-1, 10 μM), a nitric oxide (NO) donor. In

  6. Exploring smooth muscle phenotype and function in a bioreactor model of abdominal aortic aneurysm

    PubMed Central

    2013-01-01

    Background Vascular smooth muscle cells (SMC) are central to arterial structure and function yet their involvement in the progression of abdominal aortic aneurysm (AAA) disease is not well studied. The progressive and silent nature of AAA in man essentially restricts research to the use of “end-stage” tissue recovered during surgical repair. This study aimed to generate an ex vivo model of AAA using protease-treated porcine carotid arteries maintained in a novel bioreactor, and to compare the structural and functional changes in SMC cultured from the recovered vessels with those from human tissue acquired at elective surgical repair. Methods Freshly isolated porcine arteries were pretreated with collagenase and/or elastase before culturing under flow in a bioreactor for 12 days. Human end-stage aneurysmal tissue and saphenous veins from age-matched controls were collected from patients undergoing surgery. SMC were cultured and characterised (immunocytochemistry, measurement of spread cell area) and assessed functionally at the level of proliferation (cell-counting) and matrix-metalloproteinase (MMP) secretion (gelatin zymography). Cellular senescence was investigated using β-galactosidase staining and apoptosis was quantified using a fluorescence-based caspase 3 assay. Results Co-expression of alpha-smooth muscle actin and smooth muscle myosin heavy chain confirmed all cell populations as SMC. Porcine SMC harvested and cultivated after collagenase/elastase pretreatment displayed a prominent “rhomboid” morphology, increased spread area (32%, P < 0.01), impaired proliferation (47% reduction, P < 0.05), increased senescence (52%, P < 0.001), susceptibility to apoptosis and reduced MMP-2 secretion (60% decrease, P < 0.01) compared with SMC from vehicle, collagenase or elastase pre-treated vessels. Notably, these changes were comparable to those observed in human AAA SMC which were 2.4-fold larger than non-aneurysmal SMC (P < 0.001) and

  7. Calponin isoforms CNN1, CNN2 and CNN3: Regulators for actin cytoskeleton functions in smooth muscle and non-muscle cells.

    PubMed

    Liu, Rong; Jin, J-P

    2016-07-01

    Calponin is an actin filament-associated regulatory protein expressed in smooth muscle and many types of non-muscle cells. Three homologous genes, CNN1, CNN2 and CNN3, encoding calponin isoforms 1, 2, and 3, respectively, are present in vertebrate species. All three calponin isoforms are actin-binding proteins with functions in inhibiting actin-activated myosin ATPase and stabilizing the actin cytoskeleton, while each isoform executes different physiological roles based on their cell type-specific expressions. Calponin 1 is specifically expressed in smooth muscle cells and plays a role in fine-tuning smooth muscle contractility. Calponin 2 is expressed in both smooth muscle and non-muscle cells and regulates multiple actin cytoskeleton-based functions. Calponin 3 participates in actin cytoskeleton-based activities in embryonic development and myogenesis. Phosphorylation has been extensively studied for the regulation of calponin functions. Cytoskeleton tension regulates the transcription of CNN2 gene and the degradation of calponin 2 protein. This review summarizes our knowledge learned from studies over the past three decades, focusing on the evolutionary lineage of calponin isoform genes, their tissue- and cell type-specific expressions, structure-function relationships, and mechanoregulation. Copyright © 2016 Elsevier B.V. All rights reserved.

  8. Pro-elastogenic effects of bone marrow mesenchymal stem cell-derived smooth muscle cells on cultured aneurysmal smooth muscle cells.

    PubMed

    Swaminathan, Ganesh; Gadepalli, Venkat S; Stoilov, Ivan; Mecham, Robert P; Rao, Raj R; Ramamurthi, Anand

    2017-03-01

    Abdominal aortic aneurysms (AAAs) involve slow proteolysis and loss of structural matrix components (collagen and elastin), which lead to wall thinning, weakening and ultimate rupture. At this time, no established non-surgical therapy is available to slow or arrest AAA growth. Inhibiting matrix metalloproteases (MMPs; e.g. MMP2 and -9) overexpressed within AAAs is insufficient to arrest AAA growth, since resident smooth muscle cells (SMCs) are poorly elastogenic and cannot overcome elastolysis to reinstate a healthy elastic matrix. Towards overcoming this limitation, this first study sought to determine the utility of rat bone marrow mesenchymal stem cell (BM-MSC)-derived SMCs to stimulate elastin and elastic matrix synthesis and assembly by aneurysmal SMCs (EaRASMCs). BM-MSCs were successfully differentiated into cells of an SMC lineage (SMLCs). Our study indicates that BM-MSC-derived SMLCs secrete trophic factors, contained in conditioned medium (CM) from their cultures, that, when exposed to EaRASMC cultures in real time, stimulate elastin precursor and matrix deposition and crosslinking by these elastogenically deficient cells, with added benefits in terms of attenuating MMPs, specifically MMP9. The results thus lend support to a proposed cell therapy for AAAs, based on the use of BM-MSC-derived SMLCs. Although we observed no particular improvement in elastic fibre formation, no attenuation of MMP2 activity and increase in amounts of active MMP2 enzyme, we believe that this study justifies follow-up studies to improve upon these outcomes. Future studies will explore the effects of concentrated CM collected from long-term SMLC cultures on EaRASMCs and also investigate the elastogenic output of SMLCs themselves. Copyright © 2014 John Wiley & Sons, Ltd. Copyright © 2014 John Wiley & Sons, Ltd.

  9. A role for focal adhesion kinase in facilitating the contractile responses of murine gastric fundus smooth muscles.

    PubMed

    Xie, Yeming; Han, Koon Hee; Grainger, Nathan; Li, Wen; Corrigan, Robert D; Perrino, Brian A

    2018-03-12

    Smooth muscle contraction involves regulating myosin light chain phosphorylation and dephosphorylation by myosin light chain kinase and myosin light chain phosphatase. CPI-17 and MYPT1 are crucial for regulating gastrointestinal smooth muscle contraction by inhibiting myosin light chain phosphatase. Integrin signalling involves the dynamic recruitment of several proteins, including FAK, to focal adhesions. FAK tyrosine kinase activation is involved in cell adhesion to the extracellular matrix via integrin signalling. FAK participates in linking the force generated by myofilament activation to the extracellular matrix and throughout the smooth muscle tissue. Here, we show that cholinergic stimulation activates FAK in gastric fundus smooth muscles. Electrical field stimulation in the presence of L-NAME and MRS2500 contracted gastric fundus smooth muscle strips and increased FAK Y397 phosphorylation (pY397). Atropine blocked the contractions and prevented the increase in pY397. The FAK inhibitor PF-431396 inhibited the contractions and the increase in pY397. PF-431396 also inhibited the EFS-induced increase in CPI-17 T38 phosphorylation, and reduced MYPT1 T696 and T853, and myosin light chain S19 phosphorylation. Ca 2+ influx was unaffected by PF-431396. Nicardipine inhibited the contractions but had no effect on the increase in pY397. PDBu or calyculin A contracted gastric fundus smooth muscle strips Ca 2+ independently and increased pY397. Our findings suggest that FAK is activated by mechanical forces during contraction, and reveal a novel role of FAK in the regulation of CPI-17 phosphorylation. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.

  10. Myosin Va Plays a Role in Nitrergic Smooth Muscle Relaxation in Gastric Fundus and Corpora Cavernosa of Penis

    PubMed Central

    Carew, Josephine A.; Goyal, Raj K.; Sullivan, Maryrose P.

    2014-01-01

    The intracellular motor protein myosin Va is involved in nitrergic neurotransmission possibly by trafficking of neuronal nitric oxide synthase (nNOS) within the nerve terminals. In this study, we examined the role of myosin Va in the stomach and penis, proto-typical smooth muscle organs in which nitric oxide (NO) mediated relaxation is critical for function. We used confocal microscopy and co-immunoprecipitation of tissue from the gastric fundus (GF) and penile corpus cavernosum (CCP) to localize myosin Va with nNOS and demonstrate their molecular interaction. We utilized in vitro mechanical studies to test whether smooth muscle relaxations during nitrergic neuromuscular neurotransmission is altered in DBA (dilute, brown, non-agouti) mice which lack functional myosin Va. Myosin Va was localized in nNOS-positive nerve terminals and was co-immunoprecipitated with nNOS in both GF and CCP. In comparison to C57BL/6J wild type (WT) mice, electrical field stimulation (EFS) of precontracted smooth muscles of GF and CCP from DBA animals showed significant impairment of nitrergic relaxation. An NO donor, Sodium nitroprusside (SNP), caused comparable levels of relaxation in smooth muscles of WT and DBA mice. These normal postjunctional responses to SNP in DBA tissues suggest that impairment of smooth muscle relaxation resulted from inhibition of NO synthesis in prejunctional nerve terminals. Our results suggest that normal physiological processes of relaxation of gastric and cavernosal smooth muscles that facilitate food accommodation and penile erection, respectively, may be disrupted under conditions of myosin Va deficiency, resulting in complications like gastroparesis and erectile dysfunction. PMID:24516539

  11. Estradiol increases IP3 by a nongenomic mechanism in the smooth muscle cells from the rat oviduct.

    PubMed

    Reuquén, Patricia; Oróstica, María L; Rojas, Israel; Díaz, Patricia; Parada-Bustamante, Alexis; Orihuela, Pedro A

    2015-10-01

    Estradiol (E2) accelerates egg transport by a nongenomic action, requiring activation of estrogen receptor (ER) and successive cAMP and IP3 production in the rat oviduct. Furthermore, E2 increases IP3 production in primary cultures of oviductal smooth muscle cells. As smooth muscle cells are the mechanical effectors for the accelerated oocyte transport induced by E2 in the oviduct, herein we determined the mechanism by which E2 increases IP3 in these cells. Inhibition of protein synthesis by Actinomycin D did not affect the E2-induced IP3 increase, although this was blocked by the ER antagonist ICI182780 and the inhibitor of phospholipase C (PLC) ET-18-OCH3. Immunoelectron microscopy for ESR1 or ESR2 showed that these receptors were associated with the plasma membrane, indicating compatible localization with E2 nongenomic actions in the smooth muscle cells. Furthermore, ESR1 but not ESR2 agonist mimicked the effect of E2 on the IP3 level. Finally, E2 stimulated the activity of a protein associated with the contractile tone, calcium/calmodulin-dependent protein kinase II (CaMKII), in the smooth muscle cells. We conclude that E2 increases IP3 by a nongenomic action operated by ESR1 and that involves the activation of PLC in the smooth muscle cells of the rat oviduct. This E2 effect is associated with CaMKII activation in the smooth muscle cells, suggesting that IP3 and CaMKII are involved in the contractile activity necessary to accelerate oviductal egg transport. © 2015 Society for Reproduction and Fertility.

  12. Bronchodilatory and B-adrenergic effects of methanolic and aqueous extracts of Althaea root on isolated tracheobronchial smooth rat muscle.

    PubMed

    Alani, Behrang; Zare, Mohammad; Noureddini, Mahdi

    2015-01-01

    The smooth muscle contractions of the tracheobronchial airways are mediated through the balance of adrenergic, cholinergic and peptidergic nervous mechanisms. This research was designed to determine the bronchodilatory and B-adrenergic effects of methanolic and aqueous extracts of root Althaea on the isolated tracheobronchial smooth muscle of the rat. In this experimental study, 116 tracheobronchial sections (5 mm) from 58 healthy male Sprague-Dawley rats were dissected and divided into 23 groups. The effect of methanolic and aqueous extracts of the root Althaea was assayed at different concentrations (0.2, 0.6, 2.6, 6.6, 14.6 μg/ml) and epinephrine (5 μm) in the presence and absence of propranolol (1 μM) under one g tension based on the isometric method. This assay was recorded in an organ bath containing Krebs-Henseleit solution for tracheobronchial smooth muscle contractions using potassium chloride (KCl) (60 mM) induction. Epinephrine (5 μm) alone and root methanolic and aqueous extract concentrations (0.6-14.6 μg/ml) reduced tracheobronchial smooth muscle contractions induced using KCl (60 mM) in a dose dependent manner. Propranolol inhibited the antispasmodic effect of epinephrine on tracheobronchial smooth muscle contractions, but could not reduce the antispasmodic effect of the root extract concentrations. The methanolic and aqueous extracts of Althaea root inhibited the tracheobronchial smooth muscle contractions of rats in a dose dependent manner, but B-adrenergic receptors do not appear to engage in this process. Understanding the mechanism of this process can be useful in the treatment of pulmonary obstructive diseases like asthma.

  13. Bronchodilatory and B-adrenergic effects of methanolic and aqueous extracts of Althaea root on isolated tracheobronchial smooth rat muscle

    PubMed Central

    Alani, Behrang; Zare, Mohammad; Noureddini, Mahdi

    2015-01-01

    Background: The smooth muscle contractions of the tracheobronchial airways are mediated through the balance of adrenergic, cholinergic and peptidergic nervous mechanisms. This research was designed to determine the bronchodilatory and B-adrenergic effects of methanolic and aqueous extracts of root Althaea on the isolated tracheobronchial smooth muscle of the rat. Materials and Methods: In this experimental study, 116 tracheobronchial sections (5 mm) from 58 healthy male Sprague-Dawley rats were dissected and divided into 23 groups. The effect of methanolic and aqueous extracts of the root Althaea was assayed at different concentrations (0.2, 0.6, 2.6, 6.6, 14.6 μg/ml) and epinephrine (5 μm) in the presence and absence of propranolol (1 μM) under one g tension based on the isometric method. This assay was recorded in an organ bath containing Krebs-Henseleit solution for tracheobronchial smooth muscle contractions using potassium chloride (KCl) (60 mM) induction. Results: Epinephrine (5 μm) alone and root methanolic and aqueous extract concentrations (0.6-14.6 μg/ml) reduced tracheobronchial smooth muscle contractions induced using KCl (60 mM) in a dose dependent manner. Propranolol inhibited the antispasmodic effect of epinephrine on tracheobronchial smooth muscle contractions, but could not reduce the antispasmodic effect of the root extract concentrations. Conclusion: The methanolic and aqueous extracts of Althaea root inhibited the tracheobronchial smooth muscle contractions of rats in a dose dependent manner, but B-adrenergic receptors do not appear to engage in this process. Understanding the mechanism of this process can be useful in the treatment of pulmonary obstructive diseases like asthma. PMID:25879003

  14. Myosin Va plays a role in nitrergic smooth muscle relaxation in gastric fundus and corpora cavernosa of penis.

    PubMed

    Chaudhury, Arun; Cristofaro, Vivian; Carew, Josephine A; Goyal, Raj K; Sullivan, Maryrose P

    2014-01-01

    The intracellular motor protein myosin Va is involved in nitrergic neurotransmission possibly by trafficking of neuronal nitric oxide synthase (nNOS) within the nerve terminals. In this study, we examined the role of myosin Va in the stomach and penis, proto-typical smooth muscle organs in which nitric oxide (NO) mediated relaxation is critical for function. We used confocal microscopy and co-immunoprecipitation of tissue from the gastric fundus (GF) and penile corpus cavernosum (CCP) to localize myosin Va with nNOS and demonstrate their molecular interaction. We utilized in vitro mechanical studies to test whether smooth muscle relaxations during nitrergic neuromuscular neurotransmission is altered in DBA (dilute, brown, non-agouti) mice which lack functional myosin Va. Myosin Va was localized in nNOS-positive nerve terminals and was co-immunoprecipitated with nNOS in both GF and CCP. In comparison to C57BL/6J wild type (WT) mice, electrical field stimulation (EFS) of precontracted smooth muscles of GF and CCP from DBA animals showed significant impairment of nitrergic relaxation. An NO donor, Sodium nitroprusside (SNP), caused comparable levels of relaxation in smooth muscles of WT and DBA mice. These normal postjunctional responses to SNP in DBA tissues suggest that impairment of smooth muscle relaxation resulted from inhibition of NO synthesis in prejunctional nerve terminals. Our results suggest that normal physiological processes of relaxation of gastric and cavernosal smooth muscles that facilitate food accommodation and penile erection, respectively, may be disrupted under conditions of myosin Va deficiency, resulting in complications like gastroparesis and erectile dysfunction.

  15. Anti-actin antibodies revealed by counter-immunoelectrophoresis. Relation to smooth muscle antibodies and bile canalicular antibodies.

    PubMed Central

    Diederichsen, H; Riisom, K

    1980-01-01

    In investigations by counter-immunoelectrophoresis, anti-actin antibodies were found in 59% of patients with chronic hepatitis and in 8% of patients with non-hepatic diseases and normal blood donors. Anti-actin antibodies were found more frequently in patients with hepatitis and IgG smooth muscle antibodies than in other groups of diseases and normal subjects with IgG smooth muscle antibodies. Anti-actin antibodies showed no correlation with bile canalicular antibodies. Images Fig. 1 Fig. 2 PMID:6893598

  16. Logarithmic superposition of force response with rapid length changes in relaxed porcine airway smooth muscle

    PubMed Central

    Ijpma, G.; Cairns, S. P.; Sieck, G. C.

    2010-01-01

    We present a systematic quantitative analysis of power-law force relaxation and investigate logarithmic superposition of force response in relaxed porcine airway smooth muscle (ASM) strips in vitro. The term logarithmic superposition describes linear superposition on a logarithmic scale, which is equivalent to multiplication on a linear scale. Additionally, we examine whether the dynamic response of contracted and relaxed muscles is dominated by cross-bridge cycling or passive dynamics. The study shows the following main findings. For relaxed ASM, the force response to length steps of varying amplitude (0.25–4% of reference length, both lengthening and shortening) are well-fitted with power-law functions over several decades of time (10−2 to 103 s), and the force response after consecutive length changes is more accurately fitted assuming logarithmic superposition rather than linear superposition. Furthermore, for sinusoidal length oscillations in contracted and relaxed muscles, increasing the oscillation amplitude induces greater hysteresivity and asymmetry of force-length relationships, whereas increasing the frequency dampens hysteresivity but increases asymmetry. We conclude that logarithmic superposition is an important feature of relaxed ASM, which may facilitate a more accurate prediction of force responses in the continuous dynamic environment of the respiratory system. In addition, the single power-function response to length changes shows that the dynamics of cross-bridge cycling can be ignored in relaxed muscle. The similarity in response between relaxed and contracted states implies that the investigated passive dynamics play an important role in both states and should be taken into account. PMID:20817779

  17. Logarithmic superposition of force response with rapid length changes in relaxed porcine airway smooth muscle.

    PubMed

    Ijpma, G; Al-Jumaily, A M; Cairns, S P; Sieck, G C

    2010-12-01

    We present a systematic quantitative analysis of power-law force relaxation and investigate logarithmic superposition of force response in relaxed porcine airway smooth muscle (ASM) strips in vitro. The term logarithmic superposition describes linear superposition on a logarithmic scale, which is equivalent to multiplication on a linear scale. Additionally, we examine whether the dynamic response of contracted and relaxed muscles is dominated by cross-bridge cycling or passive dynamics. The study shows the following main findings. For relaxed ASM, the force response to length steps of varying amplitude (0.25-4% of reference length, both lengthening and shortening) are well-fitted with power-law functions over several decades of time (10⁻² to 10³ s), and the force response after consecutive length changes is more accurately fitted assuming logarithmic superposition rather than linear superposition. Furthermore, for sinusoidal length oscillations in contracted and relaxed muscles, increasing the oscillation amplitude induces greater hysteresivity and asymmetry of force-length relationships, whereas increasing the frequency dampens hysteresivity but increases asymmetry. We conclude that logarithmic superposition is an important feature of relaxed ASM, which may facilitate a more accurate prediction of force responses in the continuous dynamic environment of the respiratory system. In addition, the single power-function response to length changes shows that the dynamics of cross-bridge cycling can be ignored in relaxed muscle. The similarity in response between relaxed and contracted states implies that the investigated passive dynamics play an important role in both states and should be taken into account.

  18. SCA 40: studies of the relaxant effects on cryopreserved human airway and vascular smooth muscle.

    PubMed

    Müller-Schweinitzer, E; Fozard, J R

    1997-04-01

    1. 6-Bromo-8-methylaminoimidazol[1,2-a]pyrazine-2carbonitrile (SCA 40) has been claimed to induce relaxation in guinea-pig trachea by opening high conductance, calcium-activated potassium (BKCa) channels. The mechanism of action of SCA 40 has now been further investigated in ring preparations from cryopreserved human airway and vascular smooth muscle preparations in vitro. 2. Human bronchi with spontaneous tone relaxed in response to SCA 40 in a biphasic way. A high affinity component (pD2 8.61 +/- 0.21; mean +/- s.e.mean) accounted for 30% of the response and a low affinity component (pD2 6.53 +/- 0.14) for the remaining 70%. In contrast, in bronchi contracted with carbachol, 1 microM, the concentration-response curve to SCA 40 was monophasic and yielded a pD2 of 6.31 +/- 0.29. 3. SCA 40 relaxed pulmonary and mesenteric arteries and peripheral veins which had been precontracted by 10 nM U46619 nearly completely and in a monophasic way; the pD2 values were 6.37 +/- 0.08, 6.17 +/- 0.15 and 5.45 +/- 0.25, respectively. 4. Lemakalim, an opener of ATP-dependent potassium (KATP) channels, also relaxed human bronchi under spontaneous tone and the vascular tissues. NS 1619, a recognised opener of BKca channels, was inactive up to 10 microM on bronchial and vascular tissues. 5. The SCA 40-induced relaxation of human bronchi was reduced concentration-dependently in the presence of high potassium chloride (20 and 80 mM). However, in the presence of 80 mM KCl and nifedipine, 30 nM, SCA 40 fully relaxed the remaining contractile response with pD2 values of 8.08 +/- 0.13 and 5.27 +/- 0.13 for the high and low affinity component, respectively. 6. Relaxation responses to SCA 40 in human bronchi were resistant to blockade by glibenclamide at concentrations up to 10 microM (which blocked the relaxant response to lemakalim), quinine (30 microM), apamin (100 nM), tetraethylammonium (0.1-1 mM) and charybdotoxin (10-100 nM), thus excluding the involvement of a variety of K+ channels

  19. Rare Presentations of Epstein-Barr Virus--Associated Smooth Muscle Tumor in Children.

    PubMed

    Arva, Nicoleta C; Schafernak, Kristian T

    2016-01-01

    Epstein-Barr virus (EBV) has oncogenic potential and has been implicated in the etiology of a wide range of malignancies. Certain EBV-driven neoplasms, such as smooth muscle tumors (SMTs), manifest typically in immunocompromised patients. In children, these neoplasms have been encountered in the setting of primary immune disorders, specifically severe combined and common variable immunodeficiency syndromes. Human immunodeficiency virus infection and posttransplant immunosuppression, in particular liver and kidney transplantation, likewise increase the risk in the pediatric population. The location of these neoplasms appears related to the type of immunodeficiency: in acquired immunodeficiency syndrome they are frequently located intracranially or intraspinally, whereas after transplant they usually involve the liver or lung. We report 2 distinct cases of EBV-related SMT, unique through their coassociated immunosuppressive state or location: the 1st occurred in a patient with immunodeficiency secondary to NEMO gene mutation following hematopoietic stem cell transplantation; the 2nd developed in the orbit after heart transplant.

  20. Endothelium-dependent smooth muscle hyperpolarization: do gap junctions provide a unifying hypothesis?

    PubMed Central

    Griffith, Tudor M

    2004-01-01

    An endothelium-derived hyperpolarizing factor (EDHF) that is distinct from nitric oxide (NO) and prostanoids has been widely hypothesized to hyperpolarize and relax vascular smooth muscle following stimulation of the endothelium by agonists. Candidates as diverse as K+ ions, eicosanoids, hydrogen peroxide and C-type natriuretic peptide have been implicated as the putative mediator, but none has emerged as a ‘universal EDHF'. An alternative explanation for the EDHF phenomenon is that direct intercellular communication via gap junctions allows passive spread of agonist-induced endothelial hyperpolarization through the vessel wall. In some arteries, eicosanoids and K+ ions may themselves initiate a conducted endothelial hyperpolarization, thus suggesting that electrotonic signalling may represent a general mechanism through which the endothelium participates in the regulation of vascular tone. PMID:15028638

  1. Measurement of PLGA-NP interaction with single smooth muscle cells using optical tweezers

    NASA Astrophysics Data System (ADS)

    Gu, Ling; Mondal, Argha; Homayoni, Homa; Nguyen, Kytai; Mohanty, Samarendra

    2012-10-01

    For intervention of cardiovascular diseases, biodegradable and biocompatible, poly(lactic-co-glycolic acid) (PLGA) nanoparticles (NP) are emerging as agents of choice for controlled and targeted drug delivery. Therefore development of PLGA-NP with optimal physico-chemical properties will allow efficient binding and thus delivery of drug to targeted cells under various patho-physiological conditions. The force kinetics and its dependence on size of the NPs will be crucial for designing the NPs. Since optical tweezers allow non-contact, highly sensitive force measurement with high spatial and temporal resolution, we utilized it for studying interaction forces between magnetic PLGA nanoparticles with smooth muscle cells (SMC). In order to investigate effect of size, interaction force for 200 to 1100nm PLGA NP was measured. For similar interaction duration, the force was found to be higher with increase in size. The rupture force was found to depend on time of interaction of SMC with NPs.

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

    SciTech Connect

    Helkin, Alex; Maier, Kristopher G.; Department of Veterans Affairs VA Healthcare Network Upstate New York at Syracuse, Syracuse, NY

    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:more » 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

  3. Gastrointestinal peristalsis: joint action of enteric nerves, smooth muscle, and interstitial cells of Cajal.

    PubMed

    Huizinga, J D

    1999-11-15

    Peristalsis is a propulsive motor pattern orchestrated by neuronal excitation and inhibition in cooperation with intrinsic muscular control mechanisms, including those residing in interstitial cells of Cajal (ICC). Interstitial cells of Cajal form a network of cells in which electrical slow waves originate and then propagate into the musculature initiating rhythmic contractile activity upon excitaton by enteric nerves. Interstitial cells of Cajal have now been isolated and their intrinsic properties reveal the presence of rhythmic inward currents not found in smooth muscle cells. In tissues where classical slow waves are not present, enteric cholinergic excitation will evoke slow wave-like activity that forces action potentials to occur in a rhythmic manner. Intrinsic and induced slow wave activity directs many of the peristaltic motor patterns in the gut. Copyright 1999 Wiley-Liss, Inc.

  4. [Interactions between endothelial cells and smooth muscle cells of blood vessels in sepsis].

    PubMed

    Zhang, Yisen; Xu, Xiaohan; Sun, Bingwei

    2016-02-01

    Vascular endothelial cells (EC) and smooth muscle cells (SMC) are target and effecter cells of inflammation, and they play an important role in inflammatory responses. The abnormal structure and function of EC and SMC play a significant role in microcirculation disturbance in septic shock and multiple organ dysfunction. This review was meant to discuss the changes in structure and function of EC and SMC and their bidirectional regulation. The cellular linkage of EC and SMC is essential for the interactions between them, and it contributes to the course of sepsis. Paracrine and autocrine as produced by EC and SMC constitute a network for mutual adjustment. Replication of the interaction between EC and SMC facilitates the potential to support hemodynamics, tissue perfusion and cellular metabolism, thereby lower the mortality rate of sepsis. However, the detailed and specific mechanisms remain to be disclosed.

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

    PubMed

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

    2017-07-28

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

  6. ULTRASTRUCTURAL INTERRELATIONSHIPS OF NERVE PROCESSES AND SMOOTH MUSCLE CELLS IN THREE DIMENSIONS

    PubMed Central

    Thaemert, J. C.

    1966-01-01

    The muscularis externa of the intestinal wall of frogs was fixed in osmium tetroxide, embedded in Vestopal-W, serially sectioned for electron microscopy, and stained with uranyl acetate. A method to obtain individually mounted and properly positioned serial sections is described. The three-dimensional techniques used during the course of this investigation demonstrate that it is possible to examine carefully relatively large areas of tissue on individual serial sections with the electron microscope and subsequently to construct montages of electron micrographs of pertinent areas from each section. Several carefully rendered interrelationships of nerve processes and smooth muscle cells in three dimensions are exhibited and described. Recent studies of other neuro-effector relationships are discussed in relation to the present status of the nature and organization of the autonomic nervous system in visceral organs. PMID:5322460

  7. 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.

  8. Nicotinic acetylcholine receptor alpha7 subunit mediates migration of vascular smooth muscle cells toward nicotine.

    PubMed

    Li, Sheng; Zhao, Tiejun; Xin, Hong; Ye, Li-Hong; Zhang, Xiaodong; Tanaka, Hideyuki; Nakamura, Akio; Kohama, Kazuhiro

    2004-03-01

    GbaSM-4 cells, vascular smooth muscle cells (VSMCs) derived from brain basilar arteries, were shown to migrate toward d-nicotine by augmenting the actin cytoskeleton in their cell bodies and lamellipodia, and expression of nicotinic acetylcholine receptor (alpha7-nAChR) was detected in GbaSM-4 cells. Their chemotaxis was antagonized by an alpha7-nAChR antagonist of methyllycaconitine. It was also antagonized by inhibiting myosin light chain (MLC) kinase and by down-regulating MLC kinase. However, the changes in MLC phosphorylation were not associated with the nicotine treatment, suggesting the involvement of non-kinase activity of MLC kinase as reviewed by Gao et al. (IUBMB Life. 2001;51:337). This plot may work to induce arteriosclerosis during cigarette smoking.

  9. inhibits smooth muscle cell proliferation via the ERK1/2 MAPK signaling pathway.

    PubMed

    Jin, Enze; Han, Seongho; Son, Mina; Kim, Sung-Whan

    2016-01-01

    Cordyceps belongs to a genus of acormycete fungi and is known to exhibit various pharmacological effects. The aim of this study was to investigate the effect of Cordyceps species on the proliferation of vascular smooth muscle cells (VSMC) and their underlying molecular mechanism. A cell proliferation assay showed that Cordyceps bassiana ethanol extract (CBEE) significantly inhibited VSMC proliferation. In addition, neointimal formation was significantly reduced by treatment with CBEE in the carotid artery of balloon-injured rats. We also investigated the effects of CBEE on the extracellular signal-regulated kinase (ERK) signal pathway. Western blot analysis revealed increased ERK 1/2 phosphorylation in VSMCs treated with CBEE. Pretreatment with U0126 completely abrogated CBEE-induced ERK 1/2 phosphorylation. In conclusion, CBEE exhibited anti-proliferative properties that affected VSMCs through the ERK1/2 MAPK signaling pathway. Our data may elucidate the inhibitory mechanism of this natural product .

  10. Urokinase-induced signaling in human vascular smooth muscle cells is mediated by PDGFR-β

    PubMed Central

    Kiyan, Julia; Kiyan, Roman; Haller, Hermann; Dumler, Inna

    2005-01-01

    Urokinase (uPA)-induced signaling in human vascular smooth muscle cells (VSMC) elicits important cellular functional responses, such as cell migration and proliferation. However, how intracellular signaling is linked to glycolipid-anchored uPA receptor (uPAR) is unknown. We provide evidence that uPAR activation by uPA induces its association with platelet-derived growth factor receptor (PDGFR)-β. The interaction results in PDGF-independent PDGFR-β activation by phosphorylation of cytoplasmic tyrosine kinase domains and receptor dimerization. Association of the receptors as well as the tyrosine kinase activity of PDGFR-β are decisive in mediating uPA-induced downstream signaling that regulates VSMC migration and proliferation. These findings provide a molecular basis for mechanisms VSMC use to induce uPAR- and PDGFR-directed signaling. The processes may be relevant to VSMC function and vascular remodeling. PMID:15889147

  11. The intracranial injection of drug in goldfish. I: Hallucinogens and their antagonism to smooth muscle activity.

    PubMed

    Abramson, H A; Gettner, H H; Carone, P A; Rolo, A; Krinsky, L

    1979-01-01

    A simplified method of studying the surfacing reaction of goldfish to hallucinogens is described. Goldfish weighing up to three grams are injected intracranially. Employing this method, d-lysergic acid diethylamide (LSD-25), d-2-acetyl lysergic acid diethylamide (ALD-52), 1-methyl d-lysergic acid butano-lamide (UML-491), and 5-methoxy dimethyl tryptamine (5-MEO-DMT) were found to be as pharmacologically active as previously noted in fish and in man. The relationship of these drugs to their anti-serotonin activity is of particular interest to the allergist because of the way in which the congeners and derivatives of LSD block the action of serotonin on smooth muscle.

  12. [Effects of hydrostatic pressure in physiological range on bladder smooth muscle cells in vitro].

    PubMed

    Wei, Tangqiang; Chen, Lin; Wang, Yan; Xu, Feng; Wang, Kanjie

    2012-08-01

    To explore the effects of the physiological range of hydrostatic pressure on human bladder smooth muscle cells (HBSMCs) cultured in vitro, we used a hydrostatic compression device designed in our laboratory into the experiments, which were grouped by varied hydrostatic pressure gradients. Cellular morphology was observed with HE staining; cytoskeleton F-actin, cell cycle, both proliferating cell nuclear antigen (PCNA) and matrix metalloproteinase 7 (MMP-7) were detected respectively with immunofluorescence, flow cytometry and RT-PCR. We found that the proliferation, cytoskeleton and cycle distribution of HBSMCs were not obviously different among the groups of different hydrostatic pressure; however, the mRNA expression of MMP-7 exhibited a trend of first increasing and then declining as the pressure gradually rises. Thus the physiological range of hydrostatic pressure may not have significant influence on proliferation, morphology, skeleton, and cell cycle of HBSMCs, but it may have great effect on the expression of MMP-7.

  13. The effect of N-methyl-D-aspartate receptor antagonist (memantine) on esophageal and gastric smooth muscle: functional investigation in a rat hydrocephalus model.

    PubMed

    Bektaş, Arzu Ozyer; Tugay, Melih; Tugay, Sevinç; Göçmez, Semil Selcen; Etus, Volkan; Utkan, Tijen

    2008-09-01

    The purpose of this study was to investigate the effect of N-methyl-d-aspartate (NMDA) receptor antagonist on esophageal and gastric smooth muscle reactivity in a rat hydrocephalus model. Hydrocephalus was induced in rats by injection of kaolin into the cisterna magna. Two weeks after the procedure, memantine (20 mg/kg per day, 2 weeks) was given to rats with hydrocephalus in the memantine group (MG). The rest of the rats with hydrocephalus received serum physiologic (hydrocephalus group, HG). The control group (nonhydrocephalic rats, CG) was sham operated. The fourth group consisted of nonhydrocephalic rats with treated memantine (memantine control group, MC). Contractile (KCl, carbachol) and relaxant (isoprenaline, papaverine) esophageal and gastric smooth muscle reactivity were determined by in vitro muscle technique. No significant difference was found in the KCl (nonreceptor-mediated)-induced esophageal smooth muscle reactivity among the groups. Carbachol (receptor-mediated)-induced smooth muscle reactivity significantly decreased in HG compared to other groups. The isoprenaline (receptor-mediated)-induced smooth muscle reactivity significantly decreased in HG compared to other groups. No significant difference was found in smooth muscle reactivity to papaverine (nonreceptor-mediated) among the groups. Gastric smooth muscle reactivity to KCl significantly increased in HG compared to other groups. Also, KCl-induced smooth muscle reactivity significantly increased in MG compared to CG and MC. Carbachol-induced smooth muscle reactivity significantly decreased in HG compared to MG, CG, and MC. No significant difference was observed in isoprenaline- and papaverine-induced smooth muscle reactivity among the groups. Our findings suggest that memantine may influence esophageal and gastric smooth muscle reactivity in hydrocephalus.

  14. Involvement of vascular peroxidase 1 in angiotensin II-induced vascular smooth muscle cell proliferation

    PubMed Central

    Shi, Ruizheng; Hu, Changping; Yuan, Qiong; Yang, Tianlun; Peng, Jun; Li, Yuanjian; Bai, Yongping; Cao, Zehong; Cheng, Guangjie; Zhang, Guogang

    2011-01-01

    Aims Vascular peroxidase 1 (VPO1) is a newly identified haem-containing peroxidase that catalyses the oxidation of a variety of substrates by hydrogen peroxide (H2O2). Considering the well-defined effects of H2O2 on the vascular remodelling during hypertension, and that VPO1 can utilize H2O2 generated from co-expressed NADPH oxidases to catalyse peroxidative reactions, the aims of this study were to determine the potential role of VPO1 in vascular remodelling during hypertension. Methods and results The vascular morphology and the expression of VPO1 in arterial tissues of spontaneously hypertensive rats and Wistar–Kyoto rats were assessed. The VPO1 expression was significantly increased concomitantly with definite vascular remodelling assessed by evaluating the media thickness, lumen diameter, media thickness-to-lumen diameter ratio and mean nuclear area in artery media in spontaneously hypertensive rats. In addition, in cultured rat aortic smooth muscle cells we found that the angiotensin II-mediated cell proliferation was inhibited by knockdown of VPO1 using small hairpin RNA. Moreover, the NADPH oxidase inhibitor, apocynin, and the hydrogen peroxide scavenger, catalase, but not the ERK1/2 inhibitor, PD98059, attenuated angiotensin II-mediated up-regulation of VPO1 and generation of hypochlorous acid. Conclusion VPO1 is a novel regulator of vascular smooth muscle cell proliferation via NADPH oxidase–H2O2–VPO1–hypochlorous acid–ERK1/2 pathways, which may contribute to vascular remodelling in hypertension. PMID:21292788

  15. The inhibitory effect of Isoliquiritigenin on the proliferation of human arterial smooth muscle cell.

    PubMed

    Chen, Tianbao; Deng, Shaoxiong; Lin, Rong

    2017-07-17

    Isoliquiritigenin (ISL) has various biological activities including as antioxidant and an inhibitor of PI3K/AKT signaling pathway. However, both oxidative stress and activated PI3K/AKT signaling contribute to the aberrant proliferation of vascular smooth muscle cells (VSMCs). This study is aimed to explore the effect of ISL on the proliferation of human arterial smooth muscle cells (HASMCs) and to investigate the underlying mechanisms. BrdU incorporation, cell cycle and reactive oxygen species (ROS) in normal or ISL treated HASMCs were analyzed by flow cytometry. Cell viablity was measured by CCK-8. Protein expression levels were examined by Western blot, and superoxide dismutase (SOD) activity was detected by using commercial kit. We observed that ISL could inhibit the proliferation of HASMCs in a dose and time dependent manner. Cell cycle of ISL treated HASMCs arrested mainly in G1/S phase and accompanied with elevated expression of p27 and decreased expression of CyclinD1 and CyclinE. In addition, ISL could down-regulated the expression of p-PI3K and p-AKT, alleviated oxidative stress and enhanced the SOD activity in HASMCs. Furthermore, H 2 O 2 treatment partly improved cell viability and up regulated p-PI3K and p-AKT in HASMCs. Therefore, we concluded that ISL inhibited the proliferation of HASMCs via attenuating oxidative stress and suppressing PI3K/AKT signaling pathway. The inhibitory effect of ISL on PI3K/AKT signaling pathway, at least partly, was mediated by ROS.

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

    SciTech Connect

    Gao, Fu; Chambon, Pierre; Tellides, George

    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 studymore » 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.« less

  17. Essential role for calcium waves in migration of human vascular smooth muscle cells.

    PubMed

    Espinosa-Tanguma, Ricardo; O'Neil, Caroline; Chrones, Tom; Pickering, J Geoffrey; Sims, Stephen M

    2011-08-01

    Vascular smooth muscle cell (SMC) migration is characterized by extension of the lamellipodia at the leading edge, lamellipodial attachment to substrate, and release of the rear (uropod) of the cell, all of which enable forward movement. However, little is known regarding the role of intracellular cytosolic Ca(2+) concentration ([Ca(2+)](i)) in coordinating these distinct activities of migrating SMCs. The objective of our study was to determine whether regional changes of Ca(2+) orchestrate the migratory cycle in human vascular SMCs. We carried out Ca(2+) imaging using digital fluorescence microscopy of fura-2 loaded human smooth muscle cells. We found that motile SMCs exhibited Ca(2+) waves that characteristically swept from the rear of polarized cells toward the leading edge. Ca(2+) waves were less evident in nonpolarized, stationary cells, although acute stimulation of these SMCs with the agonists platelet-derived growth factor-BB or histamine could elicit transient rise of [Ca(2+)](i). To investigate a role for Ca(2+) waves in the migratory cycle, we loaded cells with the Ca(2+) chelator BAPTA, which abolished Ca(2+) waves and significantly reduced retraction, supporting a causal role for Ca(2+) in initiation of retraction. However, lamellipod motility was still evident in BAPTA-loaded cells. The incidence of Ca(2+) oscillations was reduced when Ca(2+) release from intracellular stores was disrupted with the sarcoplasmic reticulum Ca(2+)-ATPase inhibitor thapsigargin or by treatment with the inositol 1,4,5-trisphosphate receptor blocker 2-aminoethoxy-diphenyl borate or xestospongin C, implicating Ca(2+) stores in generation of waves. We conclude that Ca(2+) waves are essential for migration of human vascular SMCs and can encode cell polarity.

  18. Effects of hispidulin, a flavone isolated from Inula viscosa, on isolated guinea-pig smooth muscle.

    PubMed

    Abdalla, S; Abu-Zarga, M; Afifi, F; Al-Khalil, S; Sabri, S

    1988-01-01

    1. In small concentrations (10(-7)-3 X 10(-6) M), hispidulin caused concentration-dependent contraction of isolated guinea-pig ileum and only mild relaxation of guinea-pig tracheal rings. 2. Larger concentrations (up to 3 X 10(-4) M) caused concentration-dependent relaxation of the ileum and the trachea. All the effects on the ileum and the trachea are reversible upon removal of the compound. 3. In concentrations from 10(-7) to 3 X 10(-4) M, hispidulin had no effect on the tone of the epinephrine-contracted rings of the guinea-pig main pulmonary artery. 4. Hispidulin caused a shift to the right of the acetylcholine concentration-effect curves on ileum and trachea and significantly inhibited the maximum contractions induced by acetylcholine. 5. In Ca2+-free, depolarizing solution, hispidulin caused both a shift to the right, and an inhibition of the maximum contractions, of the CaCl2 concentration-effect curves on ileum, trachea and pulmonary artery. 6. In Ca2+-free, EGTA-containing solution, hispidulin caused concentration-dependent inhibition of the contractions induced in the pulmonary artery by epinephrine and in the ileum by histamine. 7. These observations suggest that hispidulin may interfere with Ca2+ binding to the Ca2+-receptor protein(s) in the smooth muscle cell and/or with the agonist-induced Ca2+-release from intracellular stores. Less likely, hispidulin may interfere with Ca2+ influx through smooth muscle cell membrane.

  19. Mesenchymal stem cells inhibit hypoxia-induced inflammatory and fibrotic pathways in bladder smooth muscle cells.

    PubMed

    Wiafe, Bridget; Adesida, Adetola; Churchill, Thomas; Metcalfe, Peter

    2018-03-02

    Partial bladder outlet obstruction is a multifactorial urological condition in which hypoxia plays a significant role. We recently investigated hypoxia's role as a single stressor and found that hypoxia induced an intense inflammatory and profibrotic switch in bladder smooth muscle cells (bSMCs). With the immunomodulatory capacity of mesenchymal stem cells (MSCs), we aimed to investigate if the hypoxia-signaling pathways can be mitigated using MSCs. Bladder smooth muscle cells were cultured in 3% oxygen tension for 72 h with either the direct or indirect co-culture with bone marrow derived MSCs. High pore density transwells were used for indirect co-cultures. Total RNA was extracted for gene expression analysis and the Mesoscale multiplex assay was used for secreted cytokines and growth factor measurements. Total collagen contents were determined using the Sirius Red collagen assay. Hypoxia induced increase of HIF3α, VEGF, TGFβ1, TNFα, IL-1β, IL-6, αSMA, and total collagen expression and decreased IL-10 levels in bSMCs. Both direct and indirect MSCs co-cultures inhibited > 50% of hypoxia-induced TGFβ1 and IL-6 expression (p < 0.005) in a HIF-independent manner. Also, both MSCs co-culture techniques induced > 200% increase in IL-10 protein (p < 0.005) and inhibited hypoxia-induced αSMA, collagen I and III transcripts as well as total collagen proteins (p < 0.0001). Contrastingly, the hypoxia-induced IL-1β and TNFα were inhibited by only the direct co-cultures (p < 0.05). MSCs co-culture with bSMCs potently mitigates hypoxia-induced inflammatory and profibrotic pathways. This work has elucidated the role of cell-cell contact and paracrine immunomodulatory mechanisms of MSCs action and opened avenues for therapeutic intervention.

  20. Nitric oxide-evoked transient kinetics of cyclic GMP in vascular smooth muscle cells.

    PubMed

    Cawley, Sharon M; Sawyer, Carolyn L; Brunelle, Kara F; van der Vliet, Albert; Dostmann, Wolfgang R

    2007-05-01

    Cyclic-3',5'-guanosine monophosphate (cGMP) mediates the intracellular signaling cascade responsible for the nitric oxide (NO) initiated relaxation of vascular smooth muscle (VSM). However, the temporal dynamics, including the regulation of cGMP turnover, are largely unknown. Here we report new mechanistic insights into the kinetics of cGMP synthesis and hydrolysis in primary VSM cells by utilizing FRET-based cGMP-indicators [A. Honda, S.R. Adams, C.L. Sawyer, V. Lev-Ram, R.Y. Tsien, W.R. Dostmann, Proc. Natl. Acad. Sci. U S A 98 (5) (2001) 2437.]. First, 2-(N,N-Diethylamino)-diazenolate 2-oxide (DEA/NO) and 2,2'-(Hydroxynitrosohydrazono)-bis-ethanimine (DETA/NO) induced NO-concentration dependent, transient cGMP responses ("peaks") irrespective of their rates of NO release. The kinetic characteristics of these cGMP peaks were governed by the concerted action of the NO-sensitive guanylyl cyclase (GC) and phosphodiesterase type V (PDE5) as shown by their respective inhibition using 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ) and Sildenafil. These responses occurred in the presence of moderately elevated cGMP (5-15% FRET ratio), and thus activated PKG and phosphorylated PDE5, suggesting a prominent role for GC in the maintenance and termination of cGMP peaks. Furthermore, cGMP transients could be elicited repeatedly without apparent desensitization of GC or by suppression of cGMP via long-term PDE5 activity. These results demonstrate a continuous sensitivity of the NO/cGMP signaling system, inherent to the phasic nature of smooth muscle physiology.

  1. Vascular smooth muscle modulates endothelial control of vasoreactivity via reactive oxygen species production through myoendothelial communications.

    PubMed

    Billaud, Marie; Marthan, Roger; Savineau, Jean-Pierre; Guibert, Christelle

    2009-07-30

    Endothelial control of vascular smooth muscle plays a major role in the resulting vasoreactivity implicated in physiological or pathological circulatory processes. However, a comprehensive understanding of endothelial (EC)/smooth muscle cells (SMC) crosstalk is far from complete. Here, we have examined the role of gap junctions and reactive oxygen species (ROS) in this crosstalk and we demonstrate an active contribution of SMC to endothelial control of vasomotor tone. In small intrapulmonary arteries, quantitative RT-PCR, Western Blot analyses and immunofluorescent labeling evidenced connexin (Cx) 37, 40 and 43 in EC and/or SMC. Functional experiments showed that the Cx-mimetic peptide targeted against Cx 37 and Cx 43 ((37,43)Gap27) (1) reduced contractile and calcium responses to serotonin (5-HT) simultaneously recorded in pulmonary arteries and (2) abolished the diffusion in SMC of carboxyfluorescein-AM loaded in EC. Similarly, contractile and calcium responses to 5-HT were decreased by superoxide dismutase and catalase which, catabolise superoxide anion and H(2)O(2), respectively. Both Cx- and ROS-mediated effects on the responses to 5-HT were reversed by L-NAME, a NO synthase inhibitor or endothelium removal. Electronic paramagnetic resonance directly demonstrated that 5-HT-induced superoxide anion production originated from the SMC. Finally, whereas 5-HT increased NO production, it also decreased cyclic GMP content in isolated intact arteries. These data demonstrate that agonist-induced ROS production in SMC targeting EC via myoendothelial gap junctions reduces endothelial NO-dependent control of pulmonary vasoreactivity. Such SMC modulation of endothelial control may represent a signaling pathway controlling vasoreactivity under not only physiological but also pathological conditions that often implicate excessive ROS production.

  2. 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. Copyright © 2013 American Society for Bone and Mineral Research.

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

    PubMed Central

    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, 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. PMID:22887744

  4. A Subpopulation of Smooth Muscle Cells, Derived from Melanocyte-Competent Precursors, Prevents Patent Ductus Arteriosus

    PubMed Central

    Puig, Isabel; Champeval, Delphine; Kumasaka, Mayuko; Belloir, Elodie; Bonaventure, Jacky; Mark, Manuel; Yamamoto, Hiroaki; Taketo, Mark M.; Choquet, Philippe; Etchevers, Heather C.; Beermann, Friedrich; Delmas, Véronique; Monassier, Laurent; Larue, Lionel

    2013-01-01

    Background Patent ductus arteriosus is a life-threatening condition frequent in premature newborns but also present in some term infants. Current mouse models of this malformation generally lead to perinatal death, not reproducing the full phenotypic spectrum in humans, in whom genetic inheritance appears complex. The ductus arteriosus (DA), a temporary fetal vessel that bypasses the lungs by shunting the aortic arch to the pulmonary artery, is constituted by smooth muscle cells of distinct origins (SMC1 and SMC2) and many fewer melanocytes. To understand novel mechanisms preventing DA closure at birth, we evaluated the importance of cell fate specification in SMC that form the DA during embryonic development. Upon specific Tyr::Cre-driven activation of Wnt/β-catenin signaling at the time of cell fate specification, melanocytes replaced the SMC2 population of the DA, suggesting that SMC2 and melanocytes have a common precursor. The number of SMC1 in the DA remained similar to that in controls, but insufficient to allow full DA closure at birth. Thus, there was no cellular compensation by SMC1 for the loss of SMC2. Mice in which only melanocytes were genetically ablated after specification from their potential common precursor with SMC2, demonstrated that differentiated melanocytes themselves do not affect DA closure. Loss of the SMC2 population, independent of the presence of melanocytes, is therefore a cause of patent ductus arteriosus and premature death in the first months of life. Our results indicate that patent ductus arteriosus can result from the insufficient differentiation, proliferation, or contractility of a specific smooth muscle subpopulation that shares a common neural crest precursor with cardiovascular melanocytes. PMID:23382837

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

    PubMed

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

    2009-11-01

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

  6. Immunohistochemistry with keratin and smooth muscle actin monoclonal antibodies in canine digestive tract and extramural glands.

    PubMed

    Vos, J H; van den Ingh, T S; de Neijs, M; van Mil, F N; Ivanyi, D; Ramaekers, F C

    1992-05-01

    The canine digestive system and its extramural glands (parotid gland, liver, pancreas) were immunohistochemically studied using a panel of twelve monoclonal antibodies (MoAbs) specific for human keratin proteins and for alpha-smooth muscle actin. Various epithelial tissues and cells were characterized by different keratin staining patterns. So, the epithelial lining of the upper alimentary tract was characterized by staining with the MoAb 6B10, specific for keratin-type (K) 4, and the absence of staining with the MoAbs directed against K 8 and 18 (CAM 5.2 and RGE 53, DE-K18 respectively), whereas the lower alimentary tract epithelium was not labeled by 6B10, but stained by the latter MoAbs. In the salivary glands the luminal and basal cells of the adenomeres as well as the different ductal structures could be immunohistochemically differentiated. The duct epithelium in liver and pancreas showed next to keratin staining characteristics in common with hepatocytes and exocrine pancreatic cells, additional staining by several keratin MoAbs. The keratin staining patterns in the canine tissues showed, in addition to similarities also distinct discrepancies when compared to the staining patterns in corresponding human tissues. Myoepithelial cells in salivary and oesophageal glands could be differentiated from other basally located epithelial cells by their exclusive immunoreactivity for alpha-smooth muscle actin. Canine pancreatic endocrine cells were not labeled by any of the keratin MoAbs. It is concluded that immunohistochemistry with polypeptide specific MoAbs specific for human keratin-types can be used to differentiate between different types of canine epithelial tissues and epithelial cells in the digestive tract. As a result such reagents may find their application in developmental biology and pathology of this species.

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

    PubMed

    Yin, Qianran; Jiang, Dehua; Li, Lei; Yang, Yu; Wu, Pei; Luo, Yuanyuan; Yang, Rongli; Li, Dongye

    2017-01-01

    Lipopolysaccharide (LPS) is a potent activator of vascular smooth muscle cells (VSMCs) proliferation, but the underlying mechanism remains unknown. In this study, we knocked down Toll-like receptor 4 (TLR4) and Ras-related C3 botulinum toxin substrate 1 (Rac1) expression using small interfering RNA (siRNA) in order to investigate the effects and possible mechanisms of LPS-induced VSMCs proliferation. VSMCs proliferation was monitored by 5-ethynyl-2'-deoxyuridine staining, and Rac1 activity was measured via Glutathione S-transferase pull-down assay. mRNAs encoding proliferating cell nuclear antigen (PCNA), smooth muscle 22α (SM22α), myosin heavy chain (MYH) and transient receptor potential channel 1 (TRPC1) were detected by qRT-PCR. The expression of total Akt, p-Akt (308), p-Akt (473), SM22α, MYH and TRPC1 protein was analysed by Western blot. Treatment with TLR4 siRNA (siTLR4) or Rac1 siRNA (siRac1) significantly decreased LPS-induced VSMCs proliferation. Moreover, LPS-induced activation of Rac1 through TLR4 was observed. Western blot analysis revealed that transfection with siTLR4 or siRac1 inhibited LPS-induced Akt phosphorylation. We discovered that LPS stimulated VSMCs proliferation via phenotypic modulation and that this effect was partially inhibited by pre-treatment with siTLR4 or siRac1. Further, TLR4 and Rac1 are involved in LPS-induced activation of TRPC1. This study suggests that LPS exerts an effect on VSMCs proliferation and that the TLR4/Rac1/Akt signalling pathway mediates this effect. © 2017 The Author(s). Published by S. Karger AG, Basel.

  8. Regulation of cyclo-oxygenase gene expression in rat smooth muscle cells by catalase.

    PubMed

    Chen, G; Kamal, M; Hannon, R; Warner, T D

    1998-05-15

    We have studied, in detail, the effect of catalase, one of the naturally occurring antioxidant enzymes, on the expression of cyclo-oxygenase (COX) mRNA and protein in rat aortic smooth muscle cells (RASMC). The activity of COX enzyme within the cells was also determined. Catalase either alone or in combination with interleukin-1beta (IL-1beta) enhanced mRNA and protein expression for cyclo-oxygenase 2 (COX-2) in a concentration-dependent manner. However, it did not affect the expression of mRNA or protein for cyclo-oxygenase 1 (COX-1). The expression of mRNA for COX-2 induced by catalase was blocked completely by actinomycin D (ACT) or cycloheximide (CHX). In comparison, expression of mRNA for COX-2 stimulated by IL-1beta was inhibited by actinomycin D, but not by cycloheximide. This suggests that induction of the synthesis of mRNA for COX-2 by catalase and IL-1beta involves different mechanisms. In particular, the induction of mRNA for COX-2 by catalase requires on-going protein and RNA synthesis, but the induction following exposure to IL-1beta does not. The increase in expression of mRNA for COX-2 induced by catalase may be related to the ability of catalase to stimulate cyclic AMP response element (CRE) and NF-IL6 transcription factors, but not nuclear factor kappa B (NF-kappaB), for electrophoretic mobility shift assays (EMSA) showed that catalase enhanced nuclear factor binding to cyclic AMP response element and NF-IL6 but not to NF-kappaB. Catalase exerted a biphasic effect on prostaglandin synthesis. At low concentrations it enhanced prostaglandin production, but at high concentrations it tended to inhibit it. These findings suggest that catalase has differential and multiple effects on COX expression and activity in rat aortic smooth muscle cells.

  9. Premature aortic smooth muscle cell differentiation contributes to matrix dysregulation in Marfan Syndrome.

    PubMed

    Dale, Matthew; Fitzgerald, Matthew P; Liu, Zhibo; Meisinger, Trevor; Karpisek, Andrew; Purcell, Laura N; Carson, Jeffrey S; Harding, Paul; Lang, Haili; Koutakis, Panagiotis; Batra, Rishi; Mietus, Constance J; Casale, George; Pipinos, Iraklis; Baxter, B Timothy; Xiong, Wanfen

    2017-01-01

    Thoracic aortic aneurysm and dissection are life-threatening complications of Marfan syndrome (MFS). Studies of human and mouse aortic samples from late stage MFS demonstrate increased TGF-β activation/signaling and diffuse matrix changes. However, the role of the aortic smooth muscle cell (SMC) phenotype in early aneurysm formation in MFS has yet to be fully elucidated. As our objective, we investigated whether an altered aortic SMC phenotype plays a role in aneurysm formation in MFS. We describe previously unrecognized concordant findings in the aortas of a murine model of MFS, mgR, during a critical and dynamic phase of early development. Using Western blot, gelatin zymography, and histological analysis, we demonstrated that at postnatal day (PD) 7, before aortic TGF-β levels are increased, there is elastic fiber fragmentation/disorganization and increased levels of MMP-2 and MMP-9. Compared to wild type (WT) littermates, aortic SMCs in mgR mice express higher levels of contractile proteins suggesting a switch to a more mature contractile phenotype. In addition, tropoelastin levels are decreased in mgR mice, a finding consistent with a premature switch to a contractile phenotype. Proliferation assays indicate a decrease in the proliferation rate of mgR cultured SMCs compared to WT SMCs. KLF4, a regulator of smooth muscle cell phenotype, was decreased in aortic tissue of mgR mice. Finally, overexpression of KLF4 partially reversed this phenotypic change in the Marfan SMCs. This study indicates that an early phenotypic switch appears to be associated with initiation of important metabolic changes in SMCs that contribute to subsequent pathology in MFS.

  10. Oxidized Low Density Lipoprotein (LDL) Affects Hyaluronan Synthesis in Human Aortic Smooth Muscle Cells*

    PubMed Central

    Viola, Manuela; Bartolini, Barbara; Vigetti, Davide; Karousou, Evgenia; Moretto, Paola; Deleonibus, Sara; Sawamura, Tatsuya; Wight, Thomas N.; Hascall, Vincent C.; De Luca, Giancarlo; Passi, Alberto

    2013-01-01

    Thickening of the vessel in response to high low density lipoprotein(s) (LDL) levels is a hallmark of atherosclerosis, characterized by increased hyaluronan (HA) deposition in the neointima. Human native LDL trapped within the arterial wall undergoes modifications such as oxidation (oxLDL). The aim of our study is to elucidate the link between internalization of oxLDL and HA production in vitro, using human aortic smooth muscle cells. LDL were used at an effective protein concentration of 20–50 μg/ml, which allowed 80% cell viability. HA content in the medium of untreated cells was 28.9 ± 3.7 nmol HA-disaccharide/cell and increased after oxLDL treatment to 53.9 ± 5.6. OxLDL treatments doubled the transcripts of HA synthase HAS2 and HAS3. Accumulated HA stimulated migration of aortic smooth muscle cells and monocyte adhesiveness to extracellular matrix. The effects induced by oxLDL were inhibited by blocking LOX-1 scavenger receptor with a specific antibody (10 μg/ml). The cholesterol moiety of LDL has an important role in HA accumulation because cholesterol-free oxLDL failed to induce HA synthesis. Nevertheless, cholesterol-free oxLDL and unmodified cholesterol (20 μg/ml) induce only HAS3 transcription, whereas 22,oxysterol affects both HAS2 and HAS3. Moreover, HA deposition was associated with higher expression of endoplasmic reticulum stress markers (CHOP and GRP78). Our data suggest that HA synthesis can be induced in response to specific oxidized sterol-related species delivered through oxLDL. PMID:23979132

  11. Oxidized low density lipoprotein (LDL) affects hyaluronan synthesis in human aortic smooth muscle cells.

    PubMed

    Viola, Manuela; Bartolini, Barbara; Vigetti, Davide; Karousou, Evgenia; Moretto, Paola; Deleonibus, Sara; Sawamura, Tatsuya; Wight, Thomas N; Hascall, Vincent C; De Luca, Giancarlo; Passi, Alberto

    2013-10-11

    Thickening of the vessel in response to high low density lipoprotein(s) (LDL) levels is a hallmark of atherosclerosis, characterized by increased hyaluronan (HA) deposition in the neointima. Human native LDL trapped within the arterial wall undergoes modifications such as oxidation (oxLDL). The aim of our study is to elucidate the link between internalization of oxLDL and HA production in vitro, using human aortic smooth muscle cells. LDL were used at an effective protein concentration of 20-50 μg/ml, which allowed 80% cell viability. HA content in the medium of untreated cells was 28.9 ± 3.7 nmol HA-disaccharide/cell and increased after oxLDL treatment to 53.9 ± 5.6. OxLDL treatments doubled the transcripts of HA synthase HAS2 and HAS3. Accumulated HA stimulated migration of aortic smooth muscle cells and monocyte adhesiveness to extracellular matrix. The effects induced by oxLDL were inhibited by blocking LOX-1 scavenger receptor with a specific antibody (10 μg/ml). The cholesterol moiety of LDL has an important role in HA accumulation because cholesterol-free oxLDL failed to induce HA synthesis. Nevertheless, cholesterol-free oxLDL and unmodified cholesterol (20 μg/ml) induce only HAS3 transcription, whereas 22,oxysterol affects both HAS2 and HAS3. Moreover, HA deposition was associated with higher expression of endoplasmic reticulum stress markers (CHOP and GRP78). Our data suggest that HA synthesis can be induced in response to specific oxidized sterol-related species delivered through oxLDL.

  12. Synergy between thrombin and serotonin in inducing vascular smooth muscle cell proliferation.

    PubMed

    Pakala, R; Benedict, C

    1999-12-01

    Previous studies have indicated that apart from playing an important role in hemostasis and thrombosis, thrombin may also contribute to the development of postangioplasty restenosis caused by the stimulation of vascular smooth muscle cell (VSMC) proliferation. Because thrombin generation in vivo is accompanied by platelet activation and release of smooth muscle cell (SMC) growth factors such as serotonin, we examined the possible interaction between these two compounds on VSMC proliferation. Thrombin (0.01 to 100 nmol/L), thrombin receptor-activating peptide (0.1 to 1000 micromol/L), and serotonin (5HT; 0.1 to 1000 micromol/L) increased tritiated thymidine incorporation into the DNA of canine aortic VSMCs in a dose-dependent manner. When thrombin and 5HT were added together at sub-threshold concentrations, they acted synergistically in inducing tritiated thymidine incorporation. These findings were paralleled by a 90%+/-5% increase in the cell number at 48 hours, as compared with a 37%+/-2% increase with 50 micromol/L serotonin and a 13%+/-3% increase with 0.1 nmol/L thrombin. We also demonstrated that a brief exposure to thrombin (1 hour) is sufficient to show its potentiating effect on serotonin. The mitogenic effect of serotonin and its synergistic interaction with thrombin on VSMC proliferation was abolished by serotonin type 2 receptor antagonist LY281067. Similarly, gamma-hirudin--a direct thrombin inhibitor--blocked the mitogenic effect of thrombin and its synergistic interaction with serotonin. When LY281067 and gamma-hirudin were used together, they abolished the mitogenic effects of both the agonists. Because clot-bound active thrombin can escape inactivation by anti-thrombin, this thrombin may potentiate the mitogenic effect of serotonin and keep the SMCs in a proliferative state for a long period of time. These findings support the use of 5HT2 receptor antagonists in combination with thrombin inhibitors in the prevention of SMC proliferation after

  13. Supercontracted state of vertebrate smooth muscle cell fragments reveals myofilament lengths

    PubMed Central

    1990-01-01

    Isolated cell preparations from chicken gizzard smooth muscle typically contain a mixture of cell fragments and whole cells. Both species are spontaneously permeable and may be preloaded with externally applied phalloidin and antibodies and then induced to contract with Mg ATP. Labeling with antibodies revealed that the cell fragments specifically lacked certain cytoskeletal proteins (vinculin, filamin) and were depleted to various degrees in others (desmin, alpha-actinin). The cell fragments showed a unique mode of supercontraction that involved the protrusion of actin filaments through the cell surface during the terminal phase of shortening. In the presence of dextran, to minimize protein loss, the supercontracted products were star-like in form, comprising long actin bundles radiating in all directions from a central core containing myosin, desmin, and alpha-actinin. It is concluded that supercontraction is facilitated by an effective uncoupling of the contractile apparatus from the cytoskeleton, due to partial degradation of the latter, which allows unhindered sliding of actin over myosin. Homogenization of the cell fragments before or after supercontraction produced linear bipolar dimer structures composed of two oppositely polarized bundles of actin flanking a central bundle of myosin filaments. Actin filaments were shown to extend the whole length of the bundles and their length averaged integral to 4.5 microns. Myosin filaments in the supercontracted dimers averaged 1.6 microns in length. The results, showing for the first time the high actin to myosin filament length ratio in smooth muscle are readily consistent with the slow speed of shortening of this tissue. Other implications of the results are also discussed. PMID:2277067

  14. Activation of hypoxia-inducible factor-1 in pulmonary arterial smooth muscle cells by endothelin-1

    PubMed Central

    Pisarcik, Sarah; Maylor, Julie; Lu, Wenju; Yun, Xin; Undem, Clark; Sylvester, J. T.; Semenza, Gregg L.

    2013-01-01

    Numerous cellular responses to hypoxia are mediated by the transcription factor hypoxia-inducible factor-1 (HIF-1). HIF-1 plays a central role in the pathogenesis of hypoxic pulmonary hypertension. Under certain conditions, HIF-1 may utilize feedforward mechanisms to amplify its activity. Since hypoxia increases endothelin-1 (ET-1) levels in the lung, we hypothesized that during moderate, prolonged hypoxia ET-1 might contribute to HIF-1 signaling in pulmonary arterial smooth muscle cells (PASMCs). Primary cultures of rat PASMCs were treated with ET-1 or exposed to moderate, prolonged hypoxia (4% O2 for 60 h). Levels of the oxygen-sensitive HIF-1α subunit and expression of HIF target genes were increased in both hypoxic cells and cells treated with ET-1. Both hypoxia and ET-1 also increased HIF-1α mRNA expression and decreased mRNA and protein expression of prolyl hydroxylase 2 (PHD2), which is the protein responsible for targeting HIF-1α for O2-dependent degradation. The induction of HIF-1α by moderate, prolonged hypoxia was blocked by BQ-123, an antagonist of ET-1 receptor subtype A. The effects of ET-1 were mediated by increased intracellular calcium, generation of reactive oxygen species, and ERK1/2 activation. Neither ET-1 nor moderate hypoxia induced the expression of HIF-1α or HIF target genes in aortic smooth muscle cells. These results suggest that ET-1 induces a PASMC-specific increase in HIF-1α levels by upregulation of HIF-1α synthesis and downregulation of PHD2-mediated degradation, thereby amplifying the induction of HIF-1α in PASMCs during moderate, prolonged hypoxia. PMID:23418090

  15. Impaired Integrity of DNA after Recovery from Inflammation Causes Persistent Dysfunction of Colonic Smooth Muscle

    PubMed Central

    Choi, Kuicheon; Chen, Jinghong; Mitra, Sankar; Sarna, Sushil K.

    2011-01-01

    Background & Aims Patients with inflammatory bowel disease who are in remission and those that developed inflammatory bowel syndrome after enteric infection continue to have symptoms of diarrhea or constipation in the absence of overt inflammation, indicating motility dysfunction. We investigated whether oxidative stress during inflammation impairs integrity of the promoter of Cacna1c, which encodes the pore-forming α1C subunit of Cav1.2b calcium channels. Methods We used long-extension PCR (LX-PCR) to evaluate DNA integrity in tissues from distal colons of rats; trinitrobenzene sulfonic acid (TNBS) was used to induce inflammation. Results H2O2 increased in the muscularis externa 1 to 7 days after inflammation was induced with TNBS. The oxidative stress significantly impaired DNA integrity in 2 specific segments of the Cacna1c promoter: −506 to −260 and −2,193 to −1,542. The impairment peaked at day 3 and recovered partially by day 7 after induction of inflammation; expression of the products of Cacna1c followed a similar time course. Oxidative stress suppressed the expression of Nrf2, an important regulator of anti-oxidant proteins. Intra-peritoneal administration of sulforaphane significantly reversed the suppression of Nrf2, oxidative damage in the promoter of Cacna1c, and suppression of Cacna1c on day 7 of inflammation. The inflammation subsided completely by 56 days after inflammation was induced; however, impairment of DNA integrity, expression of Nrf2 and Cacna1c, and smooth muscle reactivity to acetylcholine remained suppressed at this timepoint. Conclusion Oxidative stress during inflammation impairs the integrity of the promoter of Cacna1c; impairment persists partially after inflammation has subsided. Reduced transcription of Cacna1c contributes to smooth muscle dysfunction in the absence of inflammation. PMID:21745450

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

    PubMed

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

    2018-01-15

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

  17. [Venom of Latrodectus mactans from Chile (Araneae, Theridiidae): effect on smooth muscle].

    PubMed

    Romero, Fernando; Altieri, Elena; Urrutia, Mauricio; Jara, Jorge

    2003-06-01

    The venoms of Latrodectus sp. have been reported to induce contraction probably mediated by adrenergic and cholinergic transmitters. We have demonstrated that the venom of Chilean Latrodectus mactans contains neurotoxins that induce a contraction partially independent of transmitters release. Transmembrane mobility of Na+ and Ca2+ ions and more specifically, the increase of cytoplasmic calcium concentration are responsible for tonic contraction in smooth muscle. Calcium may enter the cell by several ways, such as the voltage-dependent Ca2+ L-type channels and the Na+/Ca2+ exchanger. This study aimed to examine the participation of this exchanger in the tonic contraction of smooth muscle in vas deferent of rat induced by the venom of the Chilean spider L. mactans. Blockers of Na+ channels (amiloride) and Ca2+ L-type channels (nifedipine), and a stimulator of the exchanger (modified Tyrode, Na+ 80 mM) were used. Simultaneously, variations of the cytoplasmic concentration of Ca2+ were registered by microfluorimetry (Fura-2 indicator) in the presence of nifedipine. In presence of amiloride, dose-dependent inhibition of venom-induced contraction was observed, suggesting the participation of voltage-dependent Ca2+ L-type channels. The contraction was only partially inhibited by nifedipine and the Ca2+ cytoplasmic concentration increased, as assessed by the microfluorimetric registration. Finally, the venom-induced contraction increased in the presence of modified Tyrode, probably due to the action of the Na+/Ca2+ exchanger. Taken together, our results support the idea that the Na+/Ca2+ exchanger is active and may be, at least in part, responsible for the contraction induced by the venom of Chilean L. mactans.

  18. Acute hypoxia selectively inhibits KCNA5 channels in pulmonary artery smooth muscle cells.

    PubMed

    Platoshyn, Oleksandr; Brevnova, Elena E; Burg, Elyssa D; Yu, Ying; Remillard, Carmelle V; Yuan, Jason X-J

    2006-03-01

    Acute hypoxia causes pulmonary vasoconstriction in part by inhibiting voltage-gated K(+) (Kv) channel activity in pulmonary artery smooth muscle cells (PASMC). The hypoxia-mediated decrease in Kv currents [I(K(V))] is selective to PASMC; hypoxia has little effect on I(K(V)) in mesenteric artery smooth muscle cells (MASMC). Functional Kv channels are homo- and/or heterotetramers of pore-forming alpha-subunits and regulatory beta-subunits. KCNA5 is a Kv channel alpha-subunit that forms functional Kv channels in PASMC and regulates resting membrane potential. We have shown that acute hypoxia selectively inhibits I(K(V)) through KCNA5 channels in PASMC. Overexpression of the human KCNA5 gene increased I(K(V)) and caused membrane hyperpolarization in HEK-293, COS-7, and rat MASMC and PASMC. Acute hypoxia did not affect I(K(V)) in KCNA5-transfected HEK-293 and COS-7 cells. However, overexpression of KCNA5 in PASMC conferred its sensitivity to hypoxia. Reduction of Po(2) from 145 to 35 mmHg reduced I(K(V)) by approximately 40% in rat PASMC transfected with human KCNA5 but had no effect on I(K(V)) in KCNA5-transfected rat MASMC (or HEK and COS cells). These results indicate that KCNA5 is an important Kv channel that regulates resting membrane potential and that acute hypoxia selectively reduces KCNA5 channel activity in PASMC relative to MASMC and other cell types. Because Kv channels (including KCNA5) are ubiquitously expressed in PASMC and MASMC, the observation from this study indicates that a hypoxia-sensitive mechanism essential for inhibiting KCNA5 channel activity is exclusively present in PASMC. The divergent effect of hypoxia on I(K(V)) in PASMC and MASMC also may be due to different expression levels of KCNA5 channels.

  19. Central role of the BK channel in urinary bladder smooth muscle physiology and pathophysiology

    PubMed Central

    2014-01-01

    The physiological functions of the urinary bladder are to store and periodically expel urine. These tasks are facilitated by the contraction and relaxation of the urinary bladder smooth muscle (UBSM), also known as detrusor smooth muscle, which comprises the bladder wall. The large-conductance voltage- and Ca2+-activated K+ (BK, BKCa, MaxiK, Slo1, or KCa1.1) channel is highly expressed in UBSM and is arguably the most important physiologically relevant K+ channel that regulates UBSM function. Its significance arises from the fact that the BK channel is the only K+ channel that is activated by increases in both voltage and intracellular Ca2+. The BK channels control UBSM excitability and contractility by maintaining the resting membrane potential and shaping the repolarization phase of the spontaneous action potentials that determine UBSM spontaneous rhythmic contractility. In UBSM, these channels have complex regulatory mechanisms involving integrated intracellular Ca2+ signals, protein kinases, phosphodiesterases, and close functional interactions with muscarinic and β-adrenergic receptors. BK channel dysfunction is implicated in some forms of bladder pathologies, such as detrusor overactivity, and related overactive bladder. This review article summarizes the current state of knowledge of the functional role of UBSM BK channels under normal and pathophysiological conditions and provides new insight toward the BK channels as targets for pharmacological or genetic control of UBSM function. Modulation of UBSM BK channels can occur by directly or indirectly targeting their regulatory mechanisms, which has the potential to provide novel therapeutic approaches for bladder dysfunction, such as overactive bladder and detrusor underactivity. PMID:24990859

  20. Differential transport kinetics of chondroitin sulfate and dermatan sulfate proteoglycan by monkey aorta smooth muscle cells.

    PubMed

    Yeo, T K; Yeo, K T; Wight, T N

    1992-04-01

    Pulse-chase studies were performed to study the kinetics of chondroitin sulfate proteoglycan (CSPG) and dermatan sulfate proteoglycan (DSPG) transport in monkey aorta smooth muscle cells. During a short pulse (5 min) with [35S]Na2SO4 (500 microCi/ml), the cells synthesized 59% DSPG, 38% CSPG, and 3% heparan sulfate proteoglycan. Both DSPG and CSPG were transported out of the cell very rapidly after sulfate incorporation. At various chase times, proteoglycans (PGs) were isolated from four cellular compartments: (a) medium, (b) total cell lysate, (c) intracellular pool, and (d) extracellular pool. The PGs from the different pools were analyzed by Sepharose CL-2B column chromatography. The data of intracellular DSPG loss fitted a double exponential decay model: approximately 90% was secreted quickly with a t1/2 of 7 min, and the remaining 10% had a dramatically slower rate of secretion (t1/2 of 130 min). DSPG was rapidly secreted into the medium without prior accumulation in the extracellular matrix. In contrast, the loss of intracellular CSPG fitted a single exponential decay model with a t1/2 of 8 min; however, there was a significant accumulation of CSPG in the extracellular matrix compartment before release into the medium, resulting in a relatively slower secretion of CSPG into the medium (t1/2 of about 31 min). This delay in CSPG secretion into the medium is probably due to aggregation in the extracellular matrix, since addition of short hyaluronan oligomers (8-14 oligosaccharides) to the medium during the chase increased the rate of CSPG being secreted into the medium. We concluded that in aortic smooth muscle cell cultures, CSPG and DSPG are secreted via two distinct pathways through the cellular compartments.

  1. Nelumbo nucifera leaf extract inhibits neointimal hyperplasia through modulation of smooth muscle cell proliferation and migration.

    PubMed

    Karki, Rajendra; Jeon, Eun-Raye; Kim, Dong-Wook

    2013-01-01

    Endovascular injury induced by balloon withdrawal leads to the increased activation of matrix metalloproteinases (MMPs) in the vascular wall allowing the proliferated smooth muscle cells (SMCs) to digest the surrounding extracellular matrix and migrate from the media into the intima leading to the intimal thickening. The objective of this study was to examine the effect of Nelumbo nucifera leaf extract (NL) on intimal thickening of rat carotid artery injured by balloon catheter and on the proliferation and migration of cultured vascular smooth muscle cells (VSMCs) induced by tumor necrosis factor-α. NL was administered orally using gastric sonde at three different doses, 100 mg kg(-1) (NL100), 400 mg kg(-1) (NL400), and 800 mg kg(-1) (NL800) for 4 wk from the day of balloon injury in the rats. VSMC proliferation and migration were assessed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide and Boyden chamber methods, whereas enzymatic action of matrix metalloproteinase-2 and -9 (MMP-2 and MMP-9) was carried out by gelatin zymography, and MMP-9 protein expression, extracellular signal-regulated kinase 1/2, and c-Jun N-terminal kinase phosphorylations were assessed by Western blot analyses. NL reduced the intimal thickening by suppressing VSMC's proliferation through inhibition of extracellular signal-regulated kinase 1/2 phosphorylation and their migration by reducing the expression of MMP-2 and -9 through inhibition of JNK1/2 phosphorylation. Thus, the results suggest that NL can be considered of therapeutic value in the prevention of atherosclerosis because restenosis after percutaneous transluminal coronary angioplasty can be considered a model of "accelerated atherosclerosis." Copyright © 2013 Elsevier Inc. All rights reserved.

  2. Facilitated hyperpolarization signaling in vascular smooth muscle-overexpressing TRIC-A channels.

    PubMed

    Tao, Shengchen; Yamazaki, Daiju; Komazaki, Shinji; Zhao, Chengzhu; Iida, Tsunaki; Kakizawa, Sho; Imaizumi, Yuji; Takeshima, Hiroshi

    2013-05-31

    The TRIC channel subtypes, namely TRIC-A and TRIC-B, are intracellular monovalent cation-specific channels and likely mediate counterion movements to support efficient Ca(2+) release from the sarco/endoplasmic reticulum. Vascular smooth muscle cells (VSMCs) contain both TRIC subtypes and two Ca(2+) release mechanisms; incidental opening of ryanodine receptors (RyRs) generates local Ca(2+) sparks to induce hyperpolarization and relaxation, whereas agonist-induced activation of inositol trisphosphate receptors produces global Ca(2+) transients causing contraction. Tric-a knock-out mice develop hypertension due to insufficient RyR-mediated Ca(2+) sparks in VSMCs. Here we describe transgenic mice overexpressing TRIC-A channels under the control of a smooth muscle cell-specific promoter. The transgenic mice developed congenital hypotension. In Tric-a-overexpressing VSMCs from the transgenic mice, the resting membrane potential decreased because RyR-mediated Ca(2+) sparks were facilitated and cell surface Ca(2+)-dependent K(+) channels were hyperactivated. Under such hyperpolarized conditions, L-type Ca(2+) channels were inactivated, and thus, the resting intracellular Ca(2+) levels were reduced in Tric-a-overexpressing VSMCs. Moreover, Tric-a overexpression impaired inositol trisphosphate-sensitive stores to diminish agonist-induced Ca(2+) signaling in VSMCs. These altered features likely reduced vascular tonus leading to the hypotensive phenotype. Our Tric-a-transgenic mice together with Tric-a knock-out mice indicate that TRIC-A channel density in VSMCs is responsible for controlling basal blood pressure at the whole-animal level.

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

    SciTech Connect

    Son, Dong Ju; Division of Cardiology, Department of Medicine, Emory University, Atlanta, GA; Kim, Soo Yeon

    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 murinemore » 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.« less

  4. Variable luminal sarcoplasmic reticulum Ca(2+) buffer capacity in smooth muscle cells.

    PubMed

    Dagnino-Acosta, Adán; Guerrero-Hernández, Agustín

    2009-09-01

    Sarcoplasmic reticulum contains the internal Ca(2+) store in smooth muscle cells and its lumen appears to be a continuum that lacks diffusion barriers. Accordingly, the free luminal Ca(2+) level is the same all throughout the SR; however, whether the Ca(2+) buffer capacity is the same in all the SR is unknown. We have estimated indirectly the luminal Ca(2+) buffer capacity of the SR by comparing the reduction in SR Ca(2+) levels with the corresponding increase in [Ca(2+)](i) during activation of either IP(3)Rs with carbachol or RyRs with caffeine, in smooth muscle cells from guinea pig urinary bladder. We have determined that carbachol-sensitive SR has a 2.4 times larger Ca(2+) buffer capacity than caffeine-sensitive SR. Rapid inhibition of SERCA pumps with thapsigargin revealed that this pump activity accounts for 80% and 60% of the Ca(2+) buffer capacities of carbachol- and caffeine-sensitive SR, respectively. Moreover, the Ca(2+) buffer capacity of carbachol-sensitive SR was similar to caffeine-sensitive SR when SERCA pumps were inhibited. Similar rates of Ca(2+) replenishments suggest similar levels of SERCA pump activities for either carbachol- or caffeine-sensitive SR. Paired pulses of caffeine, in conditions of low Ca(2+) influx, indicate the relevance of luminal SR Ca(2+) buffer capacity in the [Ca(2+)](i) response. To further study the importance of luminal SR Ca(2+) buffer capacity in the release process we used low levels of heparin to partially inhibit IP(3)Rs. This condition revealed carbachol-induced transient increase of luminal SR Ca(2+) levels provided that SERCA pumps were active. It thus appears that SERCA pump activity keeps the luminal SR Ca(2+)-binding proteins in the high-capacity, low-affinity conformation, particularly for IP(3)R-mediated Ca(2+) release.

  5. Opiorphin is a master regulator of the hypoxic response in corporal smooth muscle cells

    PubMed Central

    Fu, Shibo; Tar, Moses Tarndie; Melman, Arnold; Davies, Kelvin Paul

    2014-01-01

    Men with sickle cell disease (SCD) risk developing priapism. Recognizing that SCD is a disease of hypoxia, we investigated the effect of hypoxia on gene expression in corporal smooth muscle (CSM) cells. Rat CSM cells in vitro were treated with CoCl2 or low oxygen tension to mimic hypoxia. Hypoxic conditions increased expression of genes previously associated with priapism in animal models. Variable coding sequence a1 (Vcsa1; the rat opiorphin homologue, sialorphin), hypoxia-inducible factor 1a (Hif-1a), and A2B adenosine receptor (a2br) were increased by 10-, 4-, and 6-fold, respectively, by treatment with CoCl2, whereas low oxygen tension caused increases in expression of 3-, 4-, and 1.5-fold, respectively. Sialorphin-treated CSM cells increased expression of Hif-1a and a2br by 4-fold, and vcsa1-siRNA treatment reduced expression by ∼50%. Using a Hif-1a inhibitor, we demonstrated up-regulation of a2br by sialorphin is dependent on Hif-1a, and knockdown of vcsa1 expression with vcsa1-siRNA demonstrated that hypoxic-up-regulation of Hif-1a is dependent on vcsa1. In CSM from a SCD mouse, there was 15-fold up-regulation of opiorphin at a life stage prior to priapism. We conclude that in CSM, opiorphins are master regulators of the hypoxic response. Opiorphin up-regulation in response to SCD-associated hypoxia activates CSM “relaxant” pathways; excessive activation of these pathways results in priapism.—Fu, S., Tar, M. T., Melman, A., Davies, K. P. Opiorphin is a master regulator of the hypoxic response in corporal smooth muscle cells. PMID:24803544

  6. Calcium-dependent smooth muscle excitatory effect elicited by the venom of the hydrocoral Millepora complanata.

    PubMed

    Rojas, Alejandra; Torres, Mónica; Rojas, J Isela; Feregrino, Angélica; Heimer-de la Cotera, Edgar P

    2002-06-01

    In the present paper, we describe the results obtained from a preliminary pharmacological and biochemical study of the fire coral Millepora complanata, a regular component of coral reefs in the Mexican Caribbean. The protein-containing crude extract obtained from M. complanata (tested from 0.001 to 1000 microg protein/ml) caused a concentration-dependent stimulation of spontaneous contractions of the guinea pig ileum. The extract (EC(50)=11.55+/-2.36 microg/ml) was approximately 12-fold less potent than ionomycin (EC(50)=0.876+/-0.25 microg/ml) and its maximum induced contraction (1mg protein/ml) was equivalent to 68% of the response to 60mM KCl. FPLC size exclusion chromatography of the M. complanta extract afforded 12 primary fractions, of which only FV (containing proteins with molecular weights ranging from 17 to 44 kDa) and FVIII (consisting of peptides with molecular weights lesser than 1.8k Da) elicited an excitatory effect when tested at the EC(50) of the original extract. After incubation in Ca(2+)-free medium, the ileal response to FV and FVIII was significantly reduced. Blockage of L-type Ca(2+) channels with nifedipine (1 microM) inhibited FV and FVIII-evoked contractions. Cd(2+) (10 microM), an unspecific blocker of voltage-activated calcium channels, also antagonized FV and FVIII-induced effects, whereas the Na(+) channel blocker tetrodotoxin (10nM) did not significantly affect FV and FVIII responses. These results suggest that the contractions induced by the bioactive fractions obtained from the crude extract of M. complanata are caused mainly by a direct action on smooth muscle cells, via an increase in Ca(2+) permeability that occurs, at least partly, through L-type voltage-dependent Ca(2+) channels found in the cell membrane of smooth muscle. Copright 2002 Elsevier Science Ltd.

  7. The Kinetics Underlying the Velocity of Smooth Muscle Myosin Filament Sliding on Actin Filaments in Vitro*

    PubMed Central

    Haldeman, Brian D.; Brizendine, Richard K.; Facemyer, Kevin C.; Baker, Josh E.; Cremo, Christine R.

    2014-01-01

    Actin-myosin interactions are well studied using soluble myosin fragments, but little is known about effects of myosin filament structure on mechanochemistry. We stabilized unphosphorylated smooth muscle myosin (SMM) and phosphorylated smooth muscle myosin (pSMM) filaments against ATP-induced depolymerization using a cross-linker and attached fluorescent rhodamine (XL-Rh-SMM). Electron micrographs showed that these side polar filaments are very similar to unmodified filaments. They are ∼0.63 μm long and contain ∼176 molecules. Rate constants for ATP-induced dissociation and ADP release from acto-myosin for filaments and S1 heads were similar. Actin-activated ATPases of SMM and XL-Rh-SMM were similarly regulated. XL-Rh-pSMM filaments moved processively on F-actin that was bound to a PEG brush surface. ATP dependence of filament velocities was similar to that for solution ATPases at high [actin], suggesting that both processes are limited by the same kinetic step (weak to strong transition) and therefore are attachment-limited. This differs from actin sliding over myosin monomers, which is primarily detachment-limited. Fitting filament data to an attachment-limited model showed that approximately half of the heads are available to move the filament, consistent with a side polar structure. We suggest the low stiffness subfragment 2 (S2) domain remains unhindered during filament motion in our assay. Actin-bound negatively displaced heads will impart minimal drag force because of S2 buckling. Given the ADP release rate, the velocity, and the length of S2, these heads will detach from actin before slack is taken up into a backwardly displaced high stiffness position. This mechanism explains the lack of detachment-limited kinetics at physiological [ATP]. These findings address how nonlinear elasticity in assemblies of motors leads to efficient collective force generation. PMID:24907276

  8. The effect of Taraxacum officinale on gastric emptying and smooth muscle motility in Rodents.

    PubMed

    Jin, Y-R; Jin, J; Piao, X-X; Jin, N G

    2011-08-01

    Taraxacum officinale (TO) is a traditional herbal medicine that has been widely used for abdominal illnesses. However, the efficacy and the mechanism of TO on gastric emptying (GE) and smooth muscle motility are unknown. Ethyl acetate fraction (EA), n-butanol fraction (BF), and aqueous fraction (AF) were prepared in succession from 70% ethanol extract (EE) of TO using solvent polarity chromatography. Phenol red meal was adopted to estimate GE in mice. A polygraph was used to measure the smooth muscle motility in rats. The percentage of GE was 48.8 ± 6.1% (vehicle control), 75.3 ± 6.5% (cisapride positive control), 68.0±6.7% (EE), 53.3±6.0% (EA), 54.1±6.3% (AF), and 86.0±6.5% (BF). Thus, BF was determined to be most effective in accelerating GE. This stimulatory effect of BF on GE was also supported by the observation that BF increased spontaneous contraction of gastric fundus and antrum and decreased the spontaneous motility of pyloric sphincter in vitro. Atropine blocked the stimulatory effect of BF on GE, whereas phentolamine and propranolol had no effect. BF seems to be a promising prokinetic agent. BF-induced increase in the contraction of fundus and antrum contributes to an increase in the intra-gastric pressure. BF-induced decrease in the motility of pyloric sphincter contributes to a decrease in the resistance of food from the stomach to the small intestine. The acceleration of GE by BF is likely to be exerted through cholinergic stimulation. © 2011 Blackwell Publishing Ltd.

  9. Biomechanical effects of environmental and engineered particles on human airway smooth muscle cells.

    PubMed

    Berntsen, P; Park, C Y; Rothen-Rutishauser, B; Tsuda, A; Sager, T M; Molina, R M; Donaghey, T C; Alencar, A M; Kasahara, D I; Ericsson, T; Millet, E J; Swenson, J; Tschumperlin, D J; Butler, J P; Brain, J D; Fredberg, J J; Gehr, P; Zhou, E H

    2010-06-06

    The past decade has seen significant increases in combustion-generated ambient particles, which contain a nanosized fraction (less than 100 nm), and even greater increases have occurred in engineered nanoparticles (NPs) propelled by the booming nanotechnology industry. Although inhalation of these particulates has become a public health concern, human health effects and mechanisms of action for NPs are not well understood. Focusing on the human airway smooth muscle cell, here we show that the cellular mechanical function is altered by particulate exposure in a manner that is dependent upon particle material, size and dose. We used Alamar Blue assay to measure cell viability and optical magnetic twisting cytometry to measure cell stiffness and agonist-induced contractility. The eight particle species fell into four categories, based on their respective effect on cell viability and on mechanical function. Cell viability was impaired and cell contractility was decreased by (i) zinc oxide (40-100 nm and less than 44 microm) and copper(II) oxide (less than 50 nm); cell contractility was decreased by (ii) fluorescent polystyrene spheres (40 nm), increased by (iii) welding fumes and unchanged by (iv) diesel exhaust particles, titanium dioxide (25 nm) and copper(II) oxide (less than 5 microm), although in none of these cases was cell viability impaired. Treatment with hydrogen peroxide up to 500 microM did not alter viability or cell mechanics, suggesting that the particle effects are unlikely to be mediated by particle-generated reactive oxygen species. Our results highlight the susceptibility of cellular mechanical function to particulate exposures and suggest that direct exposure of the airway smooth muscle cells to particulates may initiate or aggravate respiratory diseases.

  10. THE STRUCTURE OF THE SMOOTH MUSCLE FIBRES IN THE BODY WALL OF THE EARTHWORM

    PubMed Central

    Hanson, Jean

    1957-01-01

    1. The structure of the smooth muscle fibres in the longitudinal muscle coat of the body wall of Lumbricus terrestris has been investigated by phase contrast light microscopy and electron microscopy. 2. The muscle fibre is ribbon-shaped, and attached to each of its two surfaces is a set of myofibrils. These are also ribbon-shaped, and they lie with their surfaces perpendicular to the surfaces of the fibre, and their inner edges nearly meeting in the middle of the fibre. These fibrils are oriented at an angle to the fibre axis, and diminish greatly in width as they approach the edge of the fibre. The orientation of the set of fibrils belonging to one surface of the fibre is the mirror image of that of the set belonging to the other surface; thus, when both sets are in view in a fibre lying flat on one face, the fibre exhibits double oblique striation. A comparison of extended and contracted fibres indicates that as the fibre contracts, the angle made between fibre and fibril axes increases (e.g. from 5 to 30°) and so does the angle made between the two sets of fibrils (e.g. from 10 to 60°). 3. The myofibril, throughout its length, contains irregularly packed filaments, commonly 250 A in diameter, which are parallel to its long axis and remain straight in contracted muscles. Between them is material which probably consists of much finer filaments. Thus A and I bands are absent. 4. Bound to one face of each fibril, but not penetrating inside it, is a regularly spaced series of transverse stripes. They are of two kinds, alternating along the length of the fibril, and it is suggested that they are comparable to the Z and M lines of a cross-striated fibril. The spacing of these stripes is about 0.5 µ ("Z" to "Z") in extended muscles, and 0.25 µ in contracted muscles. A bridge extends from each stripe across to the stripeless surface of the next fibril. PMID:13416316

  11. Cytogenetic analysis reveals clonal proliferation of smooth muscle cells in atherosclerotic plaques.

    PubMed

    Casalone, R; Granata, P; Minelli, E; Portentoso, P; Giudici, A; Righi, R; Castelli, P; Socrate, A; Frigerio, B

    1991-06-01

    Cytogenetic analysis of primary cell cultures from human atherosclerotic fibrous plaques revealed clonal chromosome abnormalities in 13 of the 18 cases studied. Loss of the Y chromosome and del(13)(q14) were present as single clonal abnormalities in eight cases; in five cases separate clones were found involving loss of the Y and a XXY karyotype, trisomy 10 and 18, loss of the Y and trisomy 7. A variety of single numerical and structural abnormalities were present in all but two of the 18 cases. Immunocytochemical studies were performed on cells from the same cultures used for cytogenetic analysis using monoclonal antibodies to human leucocyte common antigen, to human vimentin and to muscle actin. The immunoreactivity was positive for actin in 70-80% of the cells; 100% of the cells were positive for vimentin and all cells were ALC negative. These results indicated that the chromosomal abnormalities are present in the smooth muscle cells of the plaque. The hypothesis is proposed that the proliferation leading to the atherosclerotic lesion may primarily represent a hyperplastic response to mechanical and biological injuries and that this reactive proliferation is, in turn, associated with a tendency to chromosome instability.

  12. Membrane properties of smooth muscle cells in pulmonary arteries of the rat.

    PubMed

    Suzuki, H; Twarog, B M

    1982-05-01

    Electrical properties of the membrane of smooth muscle cells in the rat main pulmonary artery (MPA) and a small pulmonary artery (SPA) were compared. MPA and SPA differed in several important respects, suggesting characteristic quantitative and qualitative differences in membrane properties. 1) Resting membrane potentials were similar in both (MPA 52.2 +/- 1.3 mV; SPA 51.5 +/- 1.7 mV). The cells displayed no spontaneous electrical activity. The muscle layers in both MPA and SPA showed cablelike properties; a graded local response to outward current pulses was observed, but no action potentials were evoked. 2) Tetraethylammonium chloride (TEA, 1-5 mM) depolarized, increased membrane resistance, and suppressed rectification in MPA. TEA strongly depolarized SPA and contraction ensued. 3) The maximum membrane depolarization produced by a 10-fold increase in extracellular [K+] was 48 mV in MPA and 47 mV in SPA. In K+-free solution gradual depolarization was observed in SPA, but the membrane potential in MPA was not modified. Restoration of K+-containing solution produced equivalent hyperpolarization in both tissues, indicating a similar degree of stimulation of electrogenic Na+-K+ pumping. 4) A Na+-deficient solution did not affect the membrane potential in MPA but depolarized SPA.

  13. Deletion of BMAL1 in Smooth Muscle Cells Protects Mice From Abdominal Aortic Aneurysms.

    PubMed

    Lutshumba, Jenny; Liu, Shu; Zhong, Yu; Hou, Tianfei; Daugherty, Alan; Lu, Hong; Guo, Zhenheng; Gong, Ming C

    2018-05-01

    Abdominal aortic aneurysm (AAA) has high mortality rate when ruptured, but currently, there is no proven pharmacological therapy for AAA because of our poor understanding of its pathogenesis. The current study explored a novel role of smooth muscle cell (SMC) BMAL1 (brain and muscle Arnt-like protein-1)-a transcription factor known to regulate circadian rhythm-in AAA development. SMC-selective deletion of BMAL1 potently protected mice from AAA induced by (1) MR (mineralocorticoid receptor) agonist deoxycorticosterone acetate or aldosterone plus high salt intake and (2) angiotensin II infusion in hypercholesterolemia mice. Aortic BMAL1 was upregulated by deoxycorticosterone acetate-salt, and deletion of BMAL1 in SMCs selectively upregulated TIMP4 (tissue inhibitor of metalloproteinase 4) and suppressed deoxycorticosterone acetate-salt-induced MMP (matrix metalloproteinase) activation and elastin breakages. Moreover, BMAL1 bound to the Timp4 promoter and suppressed Timp4 transcription. These results reveal an important, but previously unexplored, role of SMC BMAL1 in AAA. Moreover, these results identify TIMP4 as a novel target of BMAL1, which may mediate the AAA protective effect of SMC BMAL1 deletion. © 2018 American Heart Association, Inc.

  14. Smooth Muscle Progenitor Cells Derived From Human Pluripotent Stem Cells Induce Histologic Changes in Injured Urethral Sphincter.

    PubMed

    Li, Yanhui; Wen, Yan; Wang, Zhe; Wei, Yi; Wani, Prachi; Green, Morgaine; Swaminathan, Ganesh; Ramamurthi, Anand; Pera, Renee Reijo; Chen, Bertha

    2016-12-01

    : Data suggest that myoblasts from various sources, including bone marrow, skeletal muscle, and adipose tissue, can restore muscle function in patients with urinary incontinence. Animal data have indicated that these progenitor cells exert mostly a paracrine effect on the native tissues rather than cell regeneration. Limited knowledge is available on the in vivo effect of human stem cells or muscle progenitors on injured muscles. We examined in vivo integration of smooth muscle progenitor cells (pSMCs) derived from human pluripotent stem cells (hPSCs). pSMCs were derived from a human embryonic stem cell line (H9-ESCs) and two induced pluripotent stem cell (iPSC) lines. pSMCs were injected periurethrally into urethral injury rat models (2 × 10 6 cells per rat) or intramuscularly into severe combined immunodeficiency mice. Histologic and quantitative image analysis revealed that the urethras in pSMC-treated rats contained abundant elastic fibers and thicker muscle layers compared with the control rats. Western blot confirmed increased elastin/collagen III content in the urethra and bladder of the H9-pSMC-treated rats compared with controls. iPSC-pSMC treatment also showed similar trends in elastin and collagen III. Human elastin gene expression was not detectable in rodent tissues, suggesting that the extracellular matrix synthesis resulted from the native rodent tissues rather than from the implanted human cells. Immunofluorescence staining and in vivo bioluminescence imaging confirmed long-term engraftment of pSMCs into the host urethra and the persistence of the smooth muscle phenotype. Taken together, the data suggest that hPSC-derived pSMCs facilitate restoration of urethral sphincter function by direct smooth muscle cell regeneration and by inducing native tissue elastin/collagen III remodeling. The present study provides evidence that a pure population of human smooth muscle progenitor cells (pSMCs) derived from human pluripotent stem cells (hPSCs) (human

  15. TRPM4 channel: a new player in urinary bladder smooth muscle function in rats

    PubMed Central

    Smith, Amy C.; Parajuli, Shankar P.; Hristov, Kiril L.; Cheng, Qiuping; Soder, Rupal P.; Afeli, Serge A. Y.; Earley, Scott; Xin, Wenkuan; Malysz, John

    2013-01-01

    The TRPM4 channel is a Ca2+-activated, monovalent cation-selective channel of the melastatin transient receptor potential (TRPM) family. The TRPM4 channel is implicated in the regulation of many cellular processes including the immune response, insulin secretion, and pressure-induced vasoconstriction of cerebral arteries. However, the expression and function of the TRPM4 channels in detrusor smooth muscle (DSM) have not yet been explored. Here, we provide the first molecular, electrophysiological, and functional evidence for the presence of TRPM4 channels in rat DSM. We detected the expression of TRPM4 channels at mRNA and protein levels in freshly isolated DSM single cells and DSM tissue using RT-PCR, Western blotting, immunohistochemistry, and immunocytochemistry. 9-Hydroxyphenanthrene (9-phenanthrol), a novel selective inhibitor of TRPM4 channels, was used to examine their role in DSM function. In perforated patch-clamp recordings using freshly isolated rat DSM cells, 9-phenanthrol (30 μM) decreased the spontaneous inward current activity at −70 mV. Real-time DSM live-cell Ca2+ imaging showed that selective inhibition of TRPM4 channels with 9-phenanthrol (30 μM) significantly reduced the intracellular Ca2+ levels. Isometric DSM tension recordings revealed that 9-phenanthrol (0.1–30 μM) significantly inhibited the amplitude, muscle force integral, and frequency of the spontaneous phasic and pharmacologically induced contractions of rat DSM isolated strips. 9-Phenanthrol also decreased the amplitude and muscle force integral of electrical field stimulation-induced contractions. In conclusion, this is the first study to examine the expression and provide evidence for TRPM4 channels as critical regulators of rat DSM excitability and contractility. PMID:23283997

  16. Triptolide inhibits TGF-β1-induced cell proliferation in rat airway smooth muscle cells by suppressing Smad signaling

    SciTech Connect

    Chen, Ming; Lv, Zhiqiang; Huang, Linjie

    2015-02-15

    Background: We have reported that triptolide can inhibit airway remodeling in a murine model of asthma via TGF-β1/Smad signaling. In the present study, we aimed to investigate the effect of triptolide on airway smooth muscle cells (ASMCs) proliferation and the possible mechanism. Methods: Rat airway smooth muscle cells were cultured and made synchronized, then pretreated with different concentration of triptolide before stimulated by TGF-β1. Cell proliferation was evaluated by MTT assay. Flow cytometry was used to study the influence of triptolide on cell cycle and apoptosis. Signal proteins (Smad2, Smad3 and Smad7) were detected by western blotting analysis. Results: Triptolidemore » significantly inhibited TGF-β1-induced ASMC proliferation (P<0.05). The cell cycle was blocked at G1/S-interphase by triptolide dose dependently. No pro-apoptotic effects were detected under the concentration of triptolide we used. Western blotting analysis showed TGF-β1 induced Smad2 and Smad3 phosphorylation was inhibited by triptolide pretreatment, and the level of Smad7 was increased by triptolide pretreatment. Conclusions: Triptolide may function as an inhibitor of asthma airway remodeling by suppressing ASMCs proliferation via negative regulation of Smad signaling pathway. - Highlights: • In this study, rat airway smooth muscle cells were cultured and made synchronized. • Triptolide inhibited TGF-β1-induced airway smooth muscle cells proliferation. • Triptolide inhibited ASMCs proliferation via negative regulation of Smad signaling pathway.« less

  17. Smooth Muscle LDL Receptor-Related Protein-1 Deletion Induces Aortic Insufficiency and Promotes Vascular Cardiomyopathy in Mice

    PubMed Central

    Basford, Joshua E.; Koch, Sheryl; Anjak, Ahmad; Singh, Vivek P.; Krause, Eric G.; Robbins, Nathan; Weintraub, Neal L.; Hui, David Y.; Rubinstein, Jack

    2013-01-01

    Valvular disease is common in patients with Marfan syndrome and can lead to cardiomyopathy. However, some patients develop cardiomyopathy in the absence of hemodynamically significant valve dysfunction, suggesting alternative mechanisms of disease progression. Disruption of LDL receptor-related protein-1 (Lrp1) in smooth muscle cells has been shown to cause vascular pathologies similar to Marfan syndrome, with activation of smooth muscle cells, vascular dysfunction and aortic aneurysms. This study used echocardiography and blood pressure monitoring in mouse models to determine whether inactivation of Lrp1 in vascular smooth muscle leads to cardiomyopathy, and if so, whether the mechanism is a consequence of valvular disease. Hemodynamic changes during treatment with captopril were also assessed. Dilation of aortic roots was observed in young Lrp1-knockout mice and progressed as they aged, whereas no significant aortic dilation was detected in wild type littermates. Diastolic blood pressure was lower and pulse pressure higher in Lrp1-knockout mice, which was normalized by treatment with captopril. Aortic dilation was followed by development of aortic insufficiency and subsequent dilated cardiomyopathy due to valvular disease. Thus, smooth muscle cell Lrp1 deficiency results in aortic dilation and insufficiency that causes secondary cardiomyopathy that can be improved by captopril. These findings provide novel insights into mechanisms of cardiomyopathy associated with vascular activation and offer a new model of valvular cardiomyopathy. PMID:24312398

  18. Hypoxia-Inducible Factor-1α in Smooth Muscle Cells Protects Against Aortic Aneurysms-Brief Report.

    PubMed

    Imanishi, Masaki; Chiba, Yoichi; Tomita, Noriko; Matsunaga, Shinji; Nakagawa, Toshitaka; Ueno, Masaki; Yamamoto, Kazuhiro; Tamaki, Toshiaki; Tomita, Shuhei

    2016-11-01

    The purpose of this study was to determine the role of smooth muscle cell-derived hypoxia-inducible factor-1α (Hif-1α) in the pathogenesis of aortic aneurysms. Control mice and smooth muscle cell-specific hypoxia-inducible factor-1α-deficient mice were infused with β-aminopropionitrile for 2 weeks and angiotensin II for 6 weeks to induce aortic aneurysm formation. Mutant mice experienced increased levels of aneurysm formation of the thoracic or abdominal aorta with more severe elastin disruption, compared with control mice. Smooth muscle cell-specific hypoxia-inducible factor-1α deficiency did not affect matrix metalloproteinase-2 activity; however, the activity of lysyl oxidase and the levels of tropoelastin mRNA in the angiotensin II- and β-aminopropionitrile-treated aortae, associated with elastin fiber formation, were suppressed. Furthermore, we observed reduced volumes of mature cross-linked elastin in the thoracoabdominal aorta after treatment with angiotensin II and β-aminopropionitrile. Deficiency of smooth muscle cell-derived hypoxia-inducible factor-1α augments aortic aneurysms, accompanied by disruption of elastin fiber formation, but not changes of elastin fiber degradation. © 2016 American Heart Association, Inc.

  19. Knockdown of parathyroid hormone related protein in smooth muscle cells alters renal hemodynamics but not blood pressure.

    PubMed

    Raison, Denis; Coquard, Catherine; Hochane, Mazène; Steger, Jacques; Massfelder, Thierry; Moulin, Bruno; Karaplis, Andrew C; Metzger, Daniel; Chambon, Pierre; Helwig, Jean-Jacques; Barthelmebs, Mariette

    2013-08-01

    Parathyroid hormone-related protein (PTHrP) belongs to vasoactive factors that regulate blood pressure and renal hemodynamics both by reducing vascular tone and raising renin release. PTHrP is expressed in systemic and renal vasculature. Here, we wanted to assess the contribution of vascular smooth muscle cell endogenous PTHrP to the regulation of cardiovascular and renal functions. We generated a mouse strain (SMA-CreERT2/PTHrPL2/L2 or premutant PTHrPSM-/-), which allows temporally controlled, smooth muscle-targeted PTHrP knockdown in adult mice. Tamoxifen treatment induced efficient recombination of PTHrP-floxed alleles and decreased PTHrP expression in vascular and visceral smooth muscle cells of PTHrPSM-/- mice. Blood pressure remained unchanged in PTHrPSM-/- mice, but plasma renin concentration and creatinine clearance were reduced. Renal hemodynamics were further analyzed during clearance measurements in anesthetized mice. Conditional knockdown of PTHrP decreased renal plasma flow and glomerular filtration rate with concomitant reduction in filtration fraction. Similar measurements were repeated during acute saline volume expansion. Saline volume expansion induced a rise in renal plasma flow and reduced filtration fraction; both were blunted in PTHrPSM-/- mice leading to impaired diuresis. These findings show that endogenous vascular smooth muscle PTHrP controls renal hemodynamics under basal conditions, and it is an essential factor in renal vasodilation elicited by saline volume expansion.

  20. Reconstitution of protein kinase C-induced contractile Ca2+ sensitization in Triton X-100-demembranated rabbit arterial smooth muscle

    PubMed Central

    Kitazawa, T; Takizawa, N; Ikebe, M; Eto, M

    1999-01-01

    Triton X-100-demembranated smooth muscle loses Ca2+-sensitizing responsiveness to protein kinase C (PKC) activators while intact and α-toxin-permeabilized smooth muscles remain responsive. We attempted to reconstitute the contractile Ca2+ sensitization by PKC in the demembranated preparations. Western blot analyses showed that the content of the PKC α-isoform (PKCα) was markedly reduced and that the smooth muscle-specific protein phosphatase-1 inhibitor protein CPI-17 was not detectable, while the amount of calponin and actin still remained similar to those of intact strips. Unphosphorylated recombinant CPI-17 alone induced a small but significant contraction at constant Ca2+. Isoform-selective PKC inhibitors inhibited unphosphorylated but not pre-thiophosphorylated CPI-17-induced contraction, suggesting that in situ conventional PKC isoform(s) can phosphorylate CPI-17. Exogenously replenishing PKCα alone did not induce potentiation of contraction and only slowly increased myosin light chain (MLC) phosphorylation at submaximal Ca2+. PKC in the presence of CPI-17, but not the [T38A]-CPI mutant, markedly induced potentiation of both contraction and MLC phosphorylation. CPI-17 itself was phosphorylated. In in vitro experiments, CPI-17 was a much better substrate for PKCα than calponin, caldesmon, MLC and myosin. Our results indicate that PKC requires CPI-17 phosphorylation at Thr-38 but not calponin for reconstitution of the contractile Ca2+ sensitization in the demembranated arterial smooth muscle. PMID:10517807

  1. Role of rho-kinase (ROCK) in tonic but not phasic contraction in the frog stomach smooth muscle.

    PubMed

    Sahin, Leyla; Cevik, Ozge Selin; Koyuncu, Dilan Deniz; Buyukafsar, Kansu

    2018-04-01

    Rho/Rho-kinase (ROCK) signaling has extensively been shown to take part in mammalian smooth muscle contractions in response to diverse agents yet its role in the contraction of amphibian smooth muscle has not been investigated. Therefore, we aimed to explore any role of this pathway in the contractions of frog stomach smooth. The strips were prepared and suspended in organ baths filled with Ringer solution. Changes in the circular strips of the frog stomach muscle length were recorded isotonically with a force transducer in organ baths. Carbachol (CCh) exerted both phasic and tonic contractions. In contrast, atropin abolished all types of contractions by CCh. The phasic contractions were suppressed by a Ca 2+ channel blocker, nifedipine but not by the ROCK inhibitor, Y-27632. However, the tonic contractions were markedly attenuated by Y-27632. Selective M 1 receptor blocker, pirenzepin, selective M 3 receptor blocker and DAMP had no effects on CCh-elicited contractions. On the other hand, selective M 2 receptor blocker, AF-DX suppressed all types of contractile activity by CCh. These data suggest that M 2 receptor activation could mainly mediate CCh-induced phasic and tonic contractions, and ROCK seems to be involved in the CCh-induced tonic but not phasic contractions of the frog stomach smooth muscle. Copyright © 2018 Elsevier Inc. All rights reserved.

  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., E-mail: jeffrey_alexis@urmc.rochester.edu

    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).more » 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.« less

  3. Endogenous γ-aminobutyric Acid Modulates Tonic Guinea Pig Airway Tone and Propofol-induced Airway Smooth Muscle Relaxation

    PubMed Central

    Gallos, George; Gleason, Neil R.; Virag, Laszlo; Zhang, Yi; Mizuta, Kentauro; Whittington, Robert A.; Emala, Charles W.

    2009-01-01

    Background Emerging evidence indicates that an endogenous autocrine/paracrine system involving γ-aminobutyric acid (GABA) is present in airways. GABAA channels, GABAB receptors and the enzyme that synthesizes GABA have been identified in airway epithelium and smooth muscle. However, the endogenous ligand itself, GABA, has not been measured in airway tissues. We sought to demonstrate that GABA is released in response to contractile agonists and tonically contributes a pro-relaxant component to contracted airway smooth muscle. Methods The amount and cellular localization of GABA in upper guinea pig airways under resting and contracted tone was determined by high pressure liquid chromatography and immunohistochemistry, respectively. The contribution that endogenous GABA imparts on the maintenance of airway smooth muscle acetylcholine-induced contraction was assessed in intact guinea pig airway tracheal rings using selective GABAA antagonism (gabazine) under resting or acetylcholine-contracted conditions. The ability of an allosteric agent (propofol) to relax a substance P-induced relaxation in an endogenous GABA-dependent manner was assessed. Results GABA levels increased and localized to airway smooth muscle following contractile stimuli in guinea pig upper airways. Acetylcholine-contracted guinea pig tracheal rings exhibited an increase in contracted force upon addition of the GABAA antagonist gabazine which was subsequently reversed by the addition of the GABAA agonist muscimol. Propofol dose-dependently relaxed a substance P contraction that was blocked by gabazine. Conclusion These studies demonstrate that GABA is endogenously present and increases following contractile stimuli in guinea pig upper airways and that endogenous GABA contributes a tonic pro-relaxant component in the maintenance of airway smooth muscle tone. PMID:19322939

  4. 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. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

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

    PubMed

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

    2016-05-01

    Smooth muscle contraction is controlled by the regulated activity of the myosin heavy chain ATPase (Myh11). Myh11 mutations have diverse effects in the cardiovascular, digestive and genitourinary systems in humans and animal models. We previously reported a recessive missense mutation, meltdown (mlt), which converts a highly conserved tryptophan to arginine (W512R) in the rigid relay loop of zebrafish Myh11. The mlt mutation disrupts myosin regulation and non-autonomously induces invasive expansion of the intestinal epithelium. Here, we report two newly identified missense mutations in the switch-1 (S237Y) and coil-coiled (L1287M) domains of Myh11 that fail to complement mlt Cell invasion was not detected in either homozygous mutant but could be induced by oxidative stress and activation of oncogenic signaling pathways. The smooth muscle defect imparted by the mlt and S237Y mutations also delayed intestinal transit, and altered vascular function, as measured by blood flow in the dorsal aorta. The cell-invasion phenotype induced by the three myh11 mutants correlated with the degree of myosin deregulation. These findings suggest that the vertebrate intestinal epithelium is tuned to the physical state of the surrounding stroma, which, in turn, governs its response to physiologic and pathologic stimuli. Genetic variants that alter the regulation of smooth muscle myosin might be risk factors for diseases affecting the intestine, vasculature, and other tissues that contain smooth muscle or contractile cells that express smooth muscle proteins, particularly in the setting of redox stress. © 2016. Published by The Company of Biologists Ltd.

  6. Affinities of pirenzepine for muscarinic cholinergic receptors in membranes isolated from bovine tracheal mucosa and smooth muscle

    SciTech Connect

    Madison, J.M.; Jones, C.A.; Tom-Moy, M.

    1987-03-01

    Muscarinic cholinergic receptors have been classified into subtypes based on their high (M-1 subtype) or low (M-2 subtype) affinities for the nonclassic antagonist pirenzepine, and this classification has important experimental and therapeutic implications. Because muscarinic receptors are abundant in the airways where they mediate several different cellular responses, the goal of this study was to characterize the affinities of pirenzepine for the muscarinic receptors in bovine tracheal mucosa and smooth muscle. After isolating membrane particulates from mucosa and smooth muscle, as well as from bovine cerebral cortex (a known source of M-1 receptors), we used /sup 3/H-quinuclidinyl benzilate to labelmore » muscarinic receptors in the particulates and performed competition radioligand binding assays in the presence of either atropine or pirenzepine. Receptors from all 3 tissues (mucosa, smooth muscle, and cerebral cortex) were of a relatively uniform affinity for atropine (range of KI values: 0.8 +/- 0.4 X 10(-9) to 2.4 +/- 1.7 X 10(-9) M), as would be predicted for this classic muscarinic antagonist. By contrast, affinities for pirenzepine differed depending on the tissue. In cerebral cortex, the majority of receptors were of high affinity for pirenzepine (KI = 1.8 +/- 1.4 X 10(-8) M). In both mucosa and smooth muscle, receptors were of low affinity for pirenzepine (Kl = 4.8 +/- 0.4 to 6.9 +/- 3.8 X 10(-7) M). We conclude that muscarinic cholinergic receptors in bovine tracheal mucosa and smooth muscle are predominantly of the M-2 subtype.« less

  7. Role of caveolae in cholesterol transport in arterial smooth muscle cells exposed to lipoproteins in vitro and in vivo.

    PubMed

    Thyberg, J; Calara, F; Dimayuga, P; Nilsson, J; Regnström, J

    1998-07-01

    Arterial smooth muscle cells are able to shift between two major differentiated states with distinct morphologic and functional properties, a contractile phenotype and a synthetic phenotype. Recently, it was demonstrated that contractile smooth muscle cells have numerous caveolae and that these specialized regions of the plasma membrane, to a large extent, are lost when the cells are modified into a synthetic phenotype. At the same time, the levels of the cholesterol-binding membrane protein caveolin remained unchanged and caveolin was redistributed from the cell surface to the perinuclear cytoplasm. In the present investigation, electron microscopy was used to study how smooth muscle cells of different phenotypes react to exposure to low-density lipoprotein and other lipoproteins both in vitro and in vivo. Our findings indicate that contractile cells (present early in primary culture and in the media of normal arterial walls) do not accumulate lipids in the cytoplasm and release excess cholesterol by means of plasma membrane caveolae. Extracellularly, the expelled lipids were built into membranous configurations and piled up as myelin-like deposits. In synthetic cells (formed after a few days in primary culture and as a response to arterial injury), lipids gathered in cytoplasmic droplets and increased amounts of membranous inclusions appeared in endosomes and lysosomes. On the other hand, no signs of extracellular discharge of lipids were detected. The results suggest that contractile smooth muscle cells use caveolin and caveolae to free themselves of excess lipoprotein-derived cholesterol and so manage to maintain a balance in the influx and efflux of cholesterol. Synthetic smooth muscle cells show a Golgi-like immunostaining for caveolin but have an insufficient capacity to use this protein to transport cholesterol to the plasma membrane and out of the cell. Cholesterol will then rather be esterified and collect in lipid droplets, eventually leading to foam

  8. Transforming growth factor-beta 3 alters intestinal smooth muscle function: implications for gastroschisis-related intestinal dysfunction.

    PubMed

    Moore-Olufemi, S D; Olsen, A B; Hook-Dufresne, D M; Bandla, V; Cox, C S

    2015-05-01

    Gastroschisis (GS) is a congenital abdominal wall defect that results in the development of GS-related intestinal dysfunction (GRID). Transforming growth factor-β, a pro-inflammatory cytokine, has been shown to cause organ dysfunction through alterations in vascular and airway smooth muscle. The purpose of this study was to evaluate the effects of TGF-β3 on intestinal smooth muscle function and contractile gene expression. Archived human intestinal tissue was analyzed using immunohistochemistry and RT-PCR for TGF-β isoforms and markers of smooth muscle gene and micro-RNA contractile phenotype. Intestinal motility was measured in neonatal rats ± TGF-β3 (0.2 and 1 mg/kg). Human intestinal smooth muscle cells (hiSMCs) were incubated with fetal bovine serum ± 100 ng/ml of TGF-β 3 isoforms for 6, 24 and 72 h. The effects of TGF-β3 on motility, hiSMC contractility and hiSMC contractile phenotype gene and micro-RNA expression were measured using transit, collagen gel contraction assay and RT-PCR analysis. Data are expressed as mean ± SEM, ANOVA (n = 6-7/group). GS infants had increased immunostaining of TGF-β3 and elevated levels of micro-RNA 143 & 145 in the intestinal smooth muscle. Rats had significantly decreased intestinal transit when exposed to TGF-β3 in a dose-dependent manner compared with Sham animals. TGF-β3 significantly increased hiSMC gel contraction and contractile protein gene and micro-RNA expression. TGF-β3 contributed to intestinal dysfunction at the organ level, increased contraction at the cellular level and elevated contractile gene expression at the molecular level. A hyper-contractile response may play a role in the persistent intestinal dysfunction seen in GRID.

  9. The effects of second messenger cAMP and its relative components on the contraction of uterine smooth muscle of rat.

    PubMed

    Shu, S-J; Lei, X-G; Liang, J-H; Song, Y-H; Xu, Q; Chen, X-D; Mao, L-G; Li, Z-G

    2017-04-01

    To investigate the effect of second messenger pathways on the uterine smooth muscle contraction and their associated mechanisms, and compare the evaluation methods. Preparation of uterine smooth muscle strips from healthy pregnant 18-21 d SD and non-pregnant rats. When the contraction of muscle strips was stable, we conducted gradient administration: PDE4 inhibitors (Z90), prostaglandin PGE2, adenylate cyclase inhibitor (SQ 22,530), cAMP analogs (dbcAMP) and AMPK agonists (AICAR), solvent dimethyl sulfoxide (DMSO) as controlled. Gradient administration of acetylcholine (Ach) and oxytocin (oxytocin) induced the contraction of muscle strips. The tension transducer and biological information collecting system were applied to record the changes, including duration, dilation tension, contraction tension, peak height, and mean tension, before and after different administration. Principal components analysis was adopted to evaluate the five changes. SQ 22,530, DMSO, cAMP alone had no significant effect on the contraction of uterine smooth muscle; Z90 can inhibit the spontaneous contraction of pregnant uterine smooth muscle strips; dbcAMP and AICAR can antagonize acetylcholine and oxytocin-induced the contraction of pregnant uterine smooth muscle strips. Z90, SQ 22,530 + Z90, dbcAMP, AICAR can inhibit the uterine contraction peak, diastolic amplitude, average muscle tone and contraction duration of the pregnant uterine smooth muscle in a concentration-dependent manners. At the same time, we compared the parameters, which reflect the contraction of uterine smooth muscle, and conduct main components analysis to determine the effect of the drugs. The second messenger cAMP and its related components ATP, 5'- AMP, AC, PDE, PKA, and AMPK can affect the uterine smooth muscle contraction via related signaling pathway in rats, and principal components analysis can be adopted to evaluate the smooth muscle relaxant.

  10. Effects of fetal hypothyroidism on uterine smooth muscle contraction and structure of offspring rats.

    PubMed

    Bagheripuor, Fatemeh; Ghanbari, Mahboubeh; Piryaei, Abbas; Ghasemi, Asghar

    2018-05-01

    What is the central question of this study? Does fetal hypothyroidism in rats alter uterine contractions and structure in the adult offspring? What is the main finding and its importance? Our study indicated that maternal hypothyroidism during pregnancy increased gestational length and decreased litter size. In addition, maternal hypothyroidism caused delayed puberty onset, irregular uterine contractions and histological changes in the uterus in the female offspring. This model might contribute to a better understanding of the cellular and molecular mechanisms involved in uterine contractions in fetal hypothyroidism, studies which are not possible in humans, and might help to establish therapeutic methods for these disorders observed in uterine contractions. Thyroid hormones play an essential role in fetal growth. Hypothyroidism impairs reproductive function in both humans and animals. The aim of this study was to assess the effects of fetal hypothyroidism on uterine smooth muscle contraction and structure in the adult offspring. The control group of female Wistar rats consumed tap water, whereas the hypothyroid group received water containing 0.025% of 6-propyl-2-thiouracial throughout gestation from mating until delivery. Isometric contractility and histological changes in uterine tissue were evaluated in the adult female offspring. We tested the effects of carbachol (10 -10 -10 -3  m) and oxytocin (10 -13 -10 -8  m) on uterine smooth muscle contraction in the fetal hypothyroid (FH) and control groups. Compared with control uteri, carbachol induced contractions with lower amplitude in the FH group (area under the curve: 1820.0 ± 250.0 versus 1370.0 ± 125.0 a.u., control versus FH group, respectively, P < 0.001) and frequency (86.4 ± 7.3 versus 37.0 ± 6.1 a.u., P < 0.001). Likewise, after exposure to oxytocin the amplitude (6614.0 ± 492.2 versus 4793.0 ± 735.2 a.u., P < 0.001) and frequency (367.4 ± 32.0 versus 167.0 ± 39.0

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

    SciTech Connect

    Salabei, Joshua K.; Department of Biochemistry and Molecular Biology, University of Louisville, Louisville, KY 40202; Balakumaran, Arun

    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 autophagymore » (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

  12. The dopamine D1 receptor is expressed and facilitates relaxation in airway smooth muscle

    PubMed Central

    2013-01-01

    Background Dopamine signaling is mediated by Gs protein-coupled “D1-like” receptors (D1 and D5) and Gi-coupled “D2-like” receptors (D2-4). In asthmatic patients, inhaled dopamine induces bronchodilation. Although the Gi-coupled dopamine D2 receptor is expressed and sensitizes adenylyl cyclase activity in airway smooth muscle (ASM) cells, the Gs-coupled dopamine D1-like receptor subtypes have never been identified on these cells. Activation of Gs-coupled receptors stimulates cyclic AMP (cAMP) production through the stimulation of adenylyl cyclase, which promotes ASM relaxation. We questioned whether the dopamine D1-like receptor is expressed on ASM, and modulates its function through Gs-coupling. Methods The mRNA and protein expression of dopamine D1-like receptor subtypes in both native human and guinea pig ASM tissue and cultured human ASM (HASM) cells was measured. To characterize the stimulation of cAMP through the dopamine D1 receptor, HASM cells were treated with dopamine or the dopamine D1-like receptor agonists (A68930 or SKF38393) before cAMP measurements. To evaluate whether the activation of dopamine D1 receptor induces ASM relaxation, guinea pig tracheal rings suspended under isometric tension in organ baths were treated with cumulatively increasing concentrations of dopamine or A68930, following an acetylcholine-induced contraction with or without the cAMP-dependent protein kinase (PKA) inhibitor Rp-cAMPS, the large-conductance calcium-activated potassium (BKCa) channel blocker iberiotoxin, or the exchange proteins directly activated by cAMP (Epac) antagonist NSC45576. Results Messenger RNA encoding the dopamine D1 and D5 receptors were detected in native human ASM tissue and cultured HASM cells. Immunoblots confirmed the protein expression of the dopamine D1 receptor in both native human and guinea pig ASM tissue and cultured HASM cells. The dopamine D1 receptor was also immunohistochemically localized to both human and guinea pig ASM. The

  13. Phenytoin inhibits contractions of rat gastrointestinal and portal vein smooth muscle by inhibiting calcium entry.

    PubMed

    Patejdl, R; Leroux, A-C; Noack, T

    2015-10-01

    Phenytoin is widely used as a second-line treatment for status epilepticus. Besides its well-known cardiac pro-arrhythmogenicity, side effects on other organ systems have received less attention. This study investigates the effects of phenytoin on gastrointestinal tissue function using an in vitro model of smooth muscle preparations from rats by combining registrations of pharmacological effects on mechanical contractions, electric field potentials, and dynamic intravital fluorescence microscopy. When added to the bathing solution at a concentration of 30 μM, phenytoin reduced the frequency of spontaneous activity significantly in antrum and portal vein preparations to 72.2 ± 36.5% (p = 0.022) and 80.7 ± 24.4% (p = 0.037) of control values, respectively. At a concentration of 100 μM, the height of spontaneous contractions declined to 9.8 ± 19.6% (p = 0.005) (antrum), 15.7 ± 28.2% (p = 0.004) (portal vein), and 31.8 ± 31.3% (p = 0.005) (colon) in comparison to the control conditions before the application of phenytoin. Depolarization triggered increases in calcium dependent fluorescence signals were reduced by 52.8 ± 39.1% (p = 0.012) The inhibition of spontaneous activity caused by phenytoin was reduced in the presence of the L-type calcium channel agonist BAY K8644(-). Phenytoin exerts strong inhibitory effects on the spontaneous and stimulated contractile activity of smooth muscles from both the upper and lower gastrointestinal tract. The mechanism underlying this effect is not related to the sodium channel blocking activity of phenytoin, but is rather caused by an inhibition of calcium entry through voltage dependent L-type calcium channels. The results of this study should raise vigilance to gastrointestinal complications in patients treated with phenytoin. © 2015 John Wiley & Sons Ltd.

  14. Heterogeneity of smooth muscle cells in atheromatous plaque of human aorta.

    PubMed Central

    Babaev, V. R.; Bobryshev, Y. V.; Stenina, O. V.; Tararak, E. M.; Gabbiani, G.

    1990-01-01

    This study was undertaken to investigate the expression of cytoskeletal proteins and the ultrastructure of cells in normal intima and atheromatous plaque of human aorta. It has been established, using double-labeling immunofluorescence, that smooth muscle cells (SMC) in normal aortic intima contain myosin, vimentin, and alpha-actin but do not react with antibodies against desmin. In contrast, 7 of 28 atherosclerotic plaques contained many cells expressing desmin in addition to the other cytoskeletal proteins characteristic of normal intima SMC. These cells were localized predominantly in the plaque cap and had the ultrastructural features of modulated SMC, ie, well-developed endoplasmic reticulum and Golgi apparatus. Besides, some cells in the 13 atherosclerotic plaques proved to be myosin, alpha actin, and desmin negative but contained vimentin and actin as revealed by fluorescent phalloidin. These cells were found in the immediate proximity of atheromatous material and reacted with a monoclonal antibody specific to SMC surface protein (11G10) but not with monoclonal anti-muscle actin (HHF35) and anti-macrophage (HAM56) antibodies. Electron microscopy of this plaque zone revealed that the cytoplasm of these cells was filled with rough endoplasmic reticulum and a developed Golgi complex. At the same time, a certain proportion of cells in this region retained morphologic features of differentiated SMC such as the presence of a basal lamina and myofilament bundles. The revealed peculiarities of cytoskeletal protein expression and the ultrastructure of cells in human aortic atherosclerotic plaques may be explained by a phenotypic modulation of vascular SMC. Images Figure 4 Figure 1 Figure 2 Figure 3 Figure 5 Figure 6 Figure 7 PMID:2190471

  15. Exogenous transforming growth factor-β1 enhances smooth muscle differentiation in embryonic mouse jejunal explants.

    PubMed

    Coletta, Riccardo; Roberts, Neil A; Randles, Michael J; Morabito, Antonino; Woolf, Adrian S

    2017-01-13

    An ex vivo experimental strategy that replicates in vivo intestinal development would in theory provide an accessible setting with which to study normal and dysmorphic gut biology. The current authors recently described a system in which mouse embryonic jejunal segments were explanted onto semipermeable platforms and fed with chemically defined serum-free media. Over 3 days in organ culture, explants formed villi and they began to undergo spontaneous peristalsis. As defined in the current study, the wall of the explanted gut failed to form a robust longitudinal smooth muscle (SM) layer as it would do in vivo over the same time period. Given the role of transforming growth factor β1 (TGFβ1) in SM differentiation in other organs, it was hypothesized that exogenous TGFβ1 would enhance SM differentiation in these explants. In vivo, TGFβ receptors I and II were both detected in embryonic longitudinal jejunal SM cells and, in organ culture, exogenous TGFβ1 induced robust differentiation of longitudinal SM. Microarray profiling showed that TGFβ1 increased SM specific transcripts in a dose dependent manner. TGFβ1 proteins were detected in amniotic fluid at a time when the intestine was physiologically herniated. By analogy with the requirement for exogenous TGFβ1 for SM differentiation in organ culture, the TGFβ1 protein that was demonstrated to be present in the amniotic fluid may enhance intestinal development when it is physiologically herniated in early gestation. Future studies of embryonic intestinal cultures should include TGFβ1 in the defined media to produce a more faithful model of in vivo muscle differentiation. Copyright © 2017 The Authors Journal of Tissue Engineering and Regenerative Medicine Published by John Wiley & Sons, Ltd. Copyright © 2017 The Authors Journal of Tissue Engineering and Regenerative Medicine Published by John Wiley & Sons, Ltd.

  16. Hypoxia inhibits human bladder smooth muscle cell proliferation: a potential mechanism of bladder dysfunction.

    PubMed

    Galvin, D J; Watson, R W G; O'Neill, A; Coffey, R N T; Taylor, C; Gillespie, J I; Fitzpatrick, J M

    2004-01-01

    Recent animal studies have suggested that bladder outflow obstruction causes bladder wall hypoxia during both the filling and the voiding phases of the micturition cycle. We have previously demonstrated that mechanical deformation of human detrusor leads to smooth muscle (SM) cell hypertrophy and hyperplasia, which may then contribute to hypoxia in the dysfunctional bladder. We hypothesise that the detrusor's response to a hypoxic environment contributes to bladder dysfunction. The aim of this study was to evaluate the effect of hypoxia on detrusor cell survival and growth. Normal human detrusor muscle was obtained at radical cystectomy and primary cultures were established. Cells were then cultured in the presence of 1% oxygen in a hypoxic chamber for different times. Apoptosis was assessed by propidium iodide DNA staining and flow cytometry. Proliferation was assessed by radiolabelled thymidine incorporation. Cell supernatants were retained for growth factor estimation by enzyme linked immuno-sorbent assay (ELISA), and total cell and nuclear extracts were isolated for Western blotting. SM cells responded to the presence of hypoxia through significant upregulation of survival factors hypoxia inducible factor (HIF 1alpha) and vascular endothelial growth factor (VEGF) in a time-dependent manner. Hypoxia did not induce cell death, but significantly reduced the rate of proliferation over time, associated with an increase in the cell cycle inhibitor p27kip1. In an in vitro human detrusor cell culture model, cells demonstrate a resistance to hypoxia-induced apoptosis but proliferation is inhibited. We suggest that the anti-proliferative effects of hypoxia may limit the ability of detrusor cells to respond to, and compensate for, alterations in their environment contributing to bladder dysfunction. Copyright 2004 Wiley-Liss, Inc.

  17. A functional study on small intestinal smooth muscles in jejunal atresia.

    PubMed

    Tyagi, Preeti; Mandal, Maloy B; Gangopadhyay, Ajay N; Patne, Shashikant C U

    2016-01-01

    The present study was aimed to assess the contractile status of neonatal small intestinal smooth muscle of dilated pre-atretic part of intestinal atresia to resolve debatable issues related to mechanisms of persistent dysmotility after surgical repair. A total of 34 longitudinally sectioned strips were prepared from pre-atretic dilated part of freshly excised 8 jejunal atresia type III a cases. Spontaneous as well as acetylcholine- and histamine-induced contractions were recorded in vitro by using organ bath preparations. Chemically evoked contractions were further evaluated after application of atropine (muscarinic blocker), pheniramine (H1 blocker), and lignocaine (neuronal blocker) to ascertain receptors and neuronal involvement. Histological examinations of strips were made by using Masson trichrome stain to assess the fibrotic changes. All 34 strips, except four showed spontaneous contractions with mean frequency and amplitude of 5.49 ± 0.26/min and 24.41 ± 5.26 g/g wet tissue respectively. The response to ACh was nearly twice as compared to histamine for equimolar concentrations (100 μM). ACh (100 μM) induced contractions were attenuated (by 60%) by atropine. Histamine (100 μM)-induced contractions was blocked by pheniramine (0.32 μM) and lignocaine (4 μM) by 74% and 78%, respectively. Histopathological examination showed varying degree of fibrotic changes in muscle layers. Pre-atretic dilated part of jejunal atresia retains functional activity but with definitive histopathologic abnormalities. It is suggested that excision of a length of pre-atretic part and early stimulation of peristalsis by locally acting cholinomimetic or H1 agonist may help in reducing postoperative motility problems in atresia patients.

  18. Transient contractions of urinary bladder smooth muscle are drivers of afferent nerve activity during filling

    PubMed Central

    Heppner, Thomas J.; Tykocki, Nathan R.; Hill-Eubanks, David

    2016-01-01

    Activation of afferent nerves during urinary bladder (UB) filling conveys the sensation of UB fullness to the central nervous system (CNS). Although this sensory outflow is presumed to reflect graded increases in pressure associated with filling, UBs also exhibit nonvoiding, transient contractions (TCs) that cause small, rapid increases in intravesical pressure. Here, using an ex vivo mouse bladder preparation, we explored the relative contributions of filling pressure and TC-induced pressure transients to sensory nerve stimulation. Continuous UB filling caused an increase in afferent nerve activity composed of a graded increase in baseline activity and activity associated with increases in intravesical pressure produced by TCs. For each ∼4-mmHg pressure increase, filling pressure increased baseline afferent activity by ∼60 action potentials per second. In contrast, a similar pressure elevation induced by a TC evoked an ∼10-fold greater increase in afferent activity. Filling pressure did not affect TC frequency but did increase the TC rate of rise, reflecting a change in the length-tension relationship of detrusor smooth muscle. The frequency of afferent bursts depended on the TC rate of rise and peaked before maximum pressure. Inhibition of small- and large-conductance Ca2+-activated K+ (SK and BK) channels increased TC amplitude and afferent nerve activity. After inhibiting detrusor muscle contractility, simulating the waveform of a TC by gently compressing the bladder evoked similar increases in afferent activity. Notably, afferent activity elicited by simulated TCs was augmented by SK channel inhibition. Our results show that afferent nerve activity evoked by TCs represents the majority of afferent outflow conveyed to the CNS during UB filling and suggest that the maximum TC rate of rise corresponds to an optimal length-tension relationship for efficient UB contraction. Furthermore, our findings implicate SK channels in controlling the gain of sensory

  19. Serum Response Factor Is Essential for Prenatal Gastrointestinal Smooth Muscle Development and Maintenance of Differentiated Phenotype

    PubMed Central

    Park, Chanjae; Lee, Moon Young; Park, Paul J; Ha, Se Eun; Berent, Robyn M; Fuchs, Robert; Miano, Joseph M; Becker, Laren S; Sanders, Kenton M; Ro, Seungil

    2015-01-01

    Background/Aims Smooth muscle cells (SMCs) characteristically express serum response factor (SRF), which regulates their development. The role of SRF in SMC plasticity in the pathophysiological conditions of gastrointestinal (GI) tract is less characterized. Methods We generated SMC-specific Srf knockout mice and characterized the prenatally lethal phenotype using ultrasound biomicroscopy and histological analysis. We used small bowel partial obstruction surgeries and primary cell culture using cell-specific enhanced green fluorescent protein (EGFP) mouse lines to study phenotypic and molecular changes of SMCs by immunofluorescence, Western blotting, and quantitative polymerase chain reaction. Finally we examined SRF change in human rectal prolapse tissue by immunofluorescence. Results Congenital SMC-specific Srf knockout mice died before birth and displayed severe GI and cardiac defects. Partial obstruction resulted in an overall increase in SRF protein expression. However, individual SMCs appeared to gradually lose SRF in the hypertrophic muscle. Cells expressing low levels of SRF also expressed low levels of platelet-derived growth factor receptor alpha (PDGFRαlow) and Ki67. SMCs grown in culture recaptured the phenotypic switch from differentiated SMCs to proliferative PDGFRαlow cells. The immediate and dramatic reduction of Srf and Myh11 mRNA expression confirmed the phenotypic change. Human rectal prolapse tissue also demonstrated significant loss of SRF expression. Conclusions SRF expression in SMCs is essential for prenatal development of the GI tract and heart. Following partial obstruction, SMCs down-regulate SRF to transition into proliferative PDGFRαlow cells that may represent a phenotype responsible for their plasticity. These findings demonstrate that SRF also plays a critical role in the remodeling process following GI injury. PMID:26424044

  20. Characterisation of K+ channels in human fetoplacental vascular smooth muscle cells.

    PubMed

    Brereton, Melissa F; Wareing, Mark; Jones, Rebecca L; Greenwood, Susan L

    2013-01-01

    Adequate blood flow through placental chorionic plate resistance arteries (CPAs) is necessary for oxygen and nutrient transfer to the fetus and a successful pregnancy. In non-placental vascular smooth muscle cells (SMCs), K(+) channels regulate contraction, vascular tone and blood flow. Previous studies showed that K(+) channel modulators alter CPA tone, but did not distinguish between effects on K(+) channels in endothelial cells and SMCs. In this study, we developed a preparation of freshly isolated CPASMCs of normal pregnancy and investigated K(+) channel expression and function. CPASMCs were isolated from normal human term placentas using enzymatic digestion. Purity and phenotype was confirmed with immunocytochemistry. Whole-cell patch clamp was used to assess K(+) channel currents, and mRNA and protein expression was determined in intact CPAs and isolated SMCs with RT-PCR and immunostaining. Isolated SMCs expressed α-actin but not CD31, a marker of endothelial cells. CPASMCs and intact CPAs expressed h-caldesmon and non-muscle myosin heavy chain-2; phenotypic markers of contractile and synthetic SMCs respectively. Whole-cell currents were inhibited by 4-AP, TEA, charybdotoxin and iberiotoxin implicating functional K(v) and BK(Ca) channels. 1-EBIO enhanced whole cell currents which were abolished by TRAM-34 and reduced by apamin indicating activation of IK(Ca) and SK(Ca) respectively. BK(Ca), IK(Ca) and SK(Ca)3 mRNA and/or protein were expressed in CPASMCs and intact CPAs. This study provides the first direct evidence for functional K(v), BK(Ca,) IK(Ca) and SK(Ca) channels in CPASMCs. These cells display a mixed phenotype implicating a dual role for CPASMCs in controlling both fetoplacental vascular resistance and vasculogenesis.

  1. Characterisation of K+ Channels in Human Fetoplacental Vascular Smooth Muscle Cells

    PubMed Central

    Brereton, Melissa F.; Wareing, Mark; Jones, Rebecca L.; Greenwood, Susan L.

    2013-01-01

    Adequate blood flow through placental chorionic plate resistance arteries (CPAs) is necessary for oxygen and nutrient transfer to the fetus and a successful pregnancy. In non-placental vascular smooth muscle cells (SMCs), K+ channels regulate contraction, vascular tone and blood flow. Previous studies showed that K+ channel modulators alter CPA tone, but did not distinguish between effects on K+ channels in endothelial cells and SMCs. In this study, we developed a preparation of freshly isolated CPASMCs of normal pregnancy and investigated K+ channel expression and function. CPASMCs were isolated from normal human term placentas using enzymatic digestion. Purity and phenotype was confirmed with immunocytochemistry. Whole-cell patch clamp was used to assess K+ channel currents, and mRNA and protein expression was determined in intact CPAs and isolated SMCs with RT-PCR and immunostaining. Isolated SMCs expressed α-actin but not CD31, a marker of endothelial cells. CPASMCs and intact CPAs expressed h-caldesmon and non-muscle myosin heavy chain-2; phenotypic markers of contractile and synthetic SMCs respectively. Whole-cell currents were inhibited by 4-AP, TEA, charybdotoxin and iberiotoxin implicating functional Kv and BKCa channels. 1-EBIO enhanced whole cell currents which were abolished by TRAM-34 and reduced by apamin indicating activation of IKCa and SKCa respectively. BKCa, IKCa and SKCa3 mRNA and/or protein were expressed in CPASMCs and intact CPAs. This study provides the first direct evidence for functional Kv, BKCa, IKCa and SKCa channels in CPASMCs. These cells display a mixed phenotype implicating a dual role for CPASMCs in controlling both fetoplacental vascular resistance and vasculogenesis. PMID:23437391

  2. Effect of Fructus Psoraleae on motility of gallbladder isolated smooth muscle strips from guinea pigs

    PubMed Central

    Jin, Shan; Li, Mei; Lin, Mei-Ling; Ding, Yong-Hui; Qu, Song-Yi; Li, Wei; Zheng, Tian-Zhen

    2006-01-01

    AIM: To observe the effect of Fructus Psoraleae on motility of isolated gallbladder muscle strips of guinea pigs and its mechanism. METHODS: Guinea pigs were hit to lose consciousness and the whole gallbladder was removed quickly. Two or three smooth muscle strips (8 mm × 3 mm) were cut along a longitudinal direction. The mucosa was gently removed. Every longitudinal muscle strip was suspended in a tissue chamber which was continuously perfused with 5 mL Krebs solution (37°C), pH 7.4, and aerated with 950 mL/L O2 and 50 mL /L CO2. The isometric response was recorded with an ink-writing recorder. After 2 h equilibration under 1 g-load, 50 μL Fructus Psoraleae (10, 20, 70, 200, 700, 1000 g/L) was added cumulatively into the tissue chamber in turn every 2 min to observe their effects on gallbladder muscle strips (cumulating final concentration of Fructus Psoraleae was 0.1, 0.3, 1.0, 3.0, 10.0, 20.0 g/L). The antagonists, including 4-DAMP, benzhydramine, hexamethonium, phentolamine, verapamil and idomethine were given 2 min before Fructus Psoraleae respectively to investigate the mechanisms involved. RESULTS: Fructus Psoraleae dose-dependently increased the resting tension (r = 0.992, P < 0.001), decreased the mean contractile amplitude (r = 0.970, P < 0.001) and meanwhile increased the contractile frequency of the gallbladder muscle strip in vitro (r = 0.965, P < 0.001). The exciting action of Fructus Psoraleae on the resting tension could be partially blocked by 4-DAMP (the resting tension decreased from 1.37 ± 0.41 to 0.70 ± 0.35, P < 0.001), benzhydramine (from 1.37 ± 0.41 to 0.45 ± 0.38, P < 0.001), hexamethonium (from 1.37 ± 0.41 to 0.94 ± 0.23, P < 0.05), phentolamine ( from 1.37 ± 0.41 to 0.89 ± 0.22, P < 0.01) and verapamil (from 1.37 ± 0.41 to 0.94 ± 0.26, P < 0.05). But the above antagonists had no significant effect on the action of Fructus Psoraleae–induced mean contractile amplitude (P > 0.05). Moreover, the increase of the contractile

  3. Role of Alpha-Smooth Muscle Actin and Fibroblast Activation Protein Alpha in Ovarian Neoplasms.

    PubMed

    da Silva, Ana Carolinne; Jammal, Millena Prata; Etchebehere, Renata Margarida; Murta, Eddie Fernando Candido; Nomelini, Rosekeila Simões

    2018-04-05

    Studies show that tumor growth is not just determined by the presence of malignant cells, since interactions between cancer cells and stromal microenvironment have important impacts on the cancer growth and progression. Cancer-associated fibroblasts play a prominent role in this process. The aims of the study were to investigate 2 cancer-associated fibroblasts markers, alpha-smooth muscle actin (α-SMA), and fibroblast activation protein alpha (FAP) in the stromal microenvironment of benign and malignant ovarian epithelial neoplasms, and to relate their tissue expression with prognostic factors in ovarian cancer. α-SMA and FAP were evaluated by immunohistochemistry in malignant (n = 28) and benign (n = 28) ovarian neoplasms. Fisher's exact test was used with a significance level lower than 0.05. FAP immunostaining was stronger in ovarian cancer when compared to benign neoplasms (p = 0.0366). There was no significant difference in relation to α-SMA expression between malignant and benign ovarian neoplasms as well as prognostic factors. In ovarian cancer, FAP stainings 2/3 was significantly related to histological grades 2 and 3 (p = 0.0183). FAP immunostaining is more intense in malignant neoplasms than in benign ovarian neoplasms, as well as in moderately differentiated and undifferentiated ovarian carcinomas compared to well-differentiated neoplasms, thus indicating that it can be used as a marker of worse prognosis. © 2018 S. Karger AG, Basel.

  4. Antiproliferative effect of UTP on human arterial and venous smooth muscle cells.

    PubMed

    White, P J; Kumari, R; Porter, K E; London, N J; Ng, L L; Boarder, M R

    2000-12-01

    We have investigated the hypothesis that responses associated with proliferation are regulated by extracellular nucleotides such as ATP and UTP in cultured human vascular smooth muscle cells (VSMC) derived from internal mammary artery (IMA) and saphenous vein (SV). Platelet-derived growth factor (PDGF), ATP, and UTP each generated an increase in cytosolic free Ca(2+) concentration ([Ca(2+)](i)) in both IMA- and SV-derived cells in the absence of detectable inositol 1,4,5-trisphosphate production. ATP alone had no effect on [(3)H]thymidine incorporation into DNA, but with a submaximal concentration of PDGF it raised [(3)H]thymidine incorporation in SV- but not IMA-derived cells. UTP alone also was without effect on [(3)H]thymidine incorporation or cell number. However, in both SV- and IMA-derived cells, UTP reduced the PDGF-stimulated [(3)H]thymidine response and PDGF-stimulated cell proliferation. This cannot be explained by an inhibitory effect on the p42/p44 mitogen-activated protein kinase (MAPK) cascade, since this response to PDGF was not attenuated by UTP. We conclude that, in human VSMC of both arterial and venous origin, UTP acts as an anti-proliferative regulator.

  5. Eugenol dilates rat cerebral arteries by inhibiting smooth muscle cell voltage-dependent calcium channels

    PubMed Central

    Peixoto-Neves, Dieniffer; Leal-Cardoso, Jose Henrique; Jaggar, Jonathan H.

    2014-01-01

    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 (Ca2+) currents, when using Ba2+ as a charge carrier, in isolated cerebral artery smooth muscle cells. Eugenol inhibition of voltage-dependent Ca2+ 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 inactivaiton rate. In summary, our data indicate that eugenol dilates cerebral arteries via multi-modal inhibition of voltage-dependent Ca2+ channels. PMID:24921632

  6. Antinociceptive and smooth muscle contracting activities of the methanolic extract of Cassia tora leaf.

    PubMed

    Chidume, F C; Kwanashie, H O; Adekeye, J O; Wambebe, C; Gamaniel, K S

    2002-07-01

    The leaves of Cassia tora Linn. (Family: Caesalpiniaceae) were soxhlet extracted with methanol. The spasmogenic effects of the extract were evaluated on guinea pig ileum, rabbit jejunum and mice intestinal transit. Antinociceptive activity of the extract was also evaluated in the mice. The LD(50) values of the extract in mice were >2000 mg/kg i.p. and p.o. The extract contracted smooth muscles of guinea pig ileum and rabbit jejunum in a concentration-dependent manner. Atropine reversibly blocked this activity. Mepyramine also reduced the contractile amplitude due to the extract in a concentration-dependent manner. The extract increased intestinal transit in mice dose dependently. C. tora extract significantly (P<0.05) reduced the number of acetic acid induced abdominal constrictions in mice and the effect was comparable to that of aspirin (150 mg/kg i.p.). The extract also significantly (P<0.05) reduced the nociceptive response of mice to increased force (g). The effects were dose-dependent. The studies suggest that the use of C. tora, traditionally, as a purgative and in the treatment of other ailments is justifiable.

  7. Relaxation of vascular smooth muscle induced by low-power laser radiation.

    PubMed

    Chaudhry, H; Lynch, M; Schomacker, K; Birngruber, R; Gregory, K; Kochevar, I

    1993-11-01

    The relaxation of rabbit aorta rings induced by low-power laser radiation was investigated in vitro to determine the location of the chromophore(s) responsible for this response and evaluate possible mechanisms. An action spectrum for relaxation was measured on rabbit thoracic aorta rings precontracted with norepinephrine. The decrease in isometric tension was measured during exposure to laser light (351-625 nm) delivered via a fiber optic to a small spot on the adventitial surface. The shortest UV wavelength (351 nm) was 35-fold more effective than 390 nm and 1700-fold more effective than 460 nm. Ultraviolet wavelengths also produced greater maximum relaxation (0.40-0.45) than visible wavelengths (0.20-0.25), suggesting that photovasorelaxation involves more than one chromophore. The adventitial layer was not necessary for photovasorelaxation, indicating that the light is absorbed by a chromophore in the medial layer. The same degree of relaxation was obtained on rings without adventitia when either one-half of the ring, or a small spot was irradiated indicating that communication between smooth muscle cells spreads a signal from the area illuminated to the entire ring. The mechanism for photovasorelaxation was investigated using potential inhibitors. N-monomethyl-L-arginine and N-amino-L-arginine, inhibitors of nitric oxide synthase, did not alter photovasorelaxation nor did indomethacin, an inhibitor of cyclooxygenase, and zinc protoporphyrin, an inhibitor of heme oxygenase.

  8. TRAIL-expressing T cells induce apoptosis of vascular smooth muscle cells in the atherosclerotic plaque

    PubMed Central

    Sato, Kayoko; Niessner, Alexander; Kopecky, Stephen L.; Frye, Robert L.; Goronzy, Jörg J.; Weyand, Cornelia M.

    2006-01-01

    Acute coronary syndromes (ACS) are precipitated by a rupture of the atherosclerotic plaque, often at the site of T cell and macrophage infiltration. Here, we show that plaque-infiltrating CD4 T cells effectively kill vascular smooth muscle cells (VSMC). VSMCs sensitive to T cell–mediated killing express the death receptor DR5 (TNF-related apoptosis-inducing ligand [TRAIL] receptor 2), and anti-TRAIL and anti-DR5 antibodies block T cell–mediated apoptosis. CD4 T cells that express TRAIL upon stimulation are expanded in patients with ACS and more effectively induce VSMC apoptosis. Adoptive transfer of plaque-derived CD4 T cells into immunodeficient mice that are engrafted with human atherosclerotic plaque results in apoptosis of VSMCs, which was prevented by coadministration of anti-TRAIL antibody. These data identify that the death pathway is triggered by TRAIL-producing CD4 T cells as a direct mechanism of VSMC apoptosis, a process which may lead to plaque destabilization. PMID:16418392

  9. Synthesis and in vitro safety assessment of magnetic bacterial cellulose with porcine aortic smooth muscle cells.

    PubMed

    Pastrana, Homero F; Cooper, Christy L; Alucozai, Milad; Reece, Lisa M; Avila, Alba G; Allain, Jean Paul

    2016-11-01

    Bacterial cellulose (BC) has been used as a scaffold for tissue regeneration (TR). Improving functional TR requires highly selective strategies for specific cell attraction. Embedding iron oxide nanoparticles into a BC matrix can drive magnetically labeled cells to specific tissues where they may begin to heal injured tissue. This article focuses on characterization and in vitro toxicity assessment of magnetic BC (MBC). We proposed to detect the production of radical oxygen species (ROS), esterase activity, and apoptosis to study cytotoxic interactions of MBC within its bioenvironment. Morphological characterization was performed using scanning electron microscopy where evidence shows that the diameter of MBC fibers compared to BC fibers was 33% smaller, and the pore areas were 25% bigger. Cytotoxicity assays in porcine aortic smooth muscle cells exposed for 24 hours to BC, MBC, and poly(ethylene glycol)-coated MBC (MBC-PEG) reveals 96% viability and 9% ROS production for MBC-PEG. In contrast, 25% of cells exposed to MBC were apoptotic, suggesting that even when the cells were metabolically active, MBC can induce damage. These outcomes support the need for more integral assessment in the hopes of assessing the potential biosafety and uses of nanocomposites for TR. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 104A: 2801-2809, 2016. © 2016 Wiley Periodicals, Inc.

  10. Tumstatin regulates the angiogenic and inflammatory potential of airway smooth muscle extracellular matrix.

    PubMed

    Harkness, Louise Margaret; Weckmann, Markus; Kopp, Matthias; Becker, Tim; Ashton, Anthony Wayne; Burgess, Janette Kay

    2017-12-01

    The extracellular matrix (ECM) creates the microenvironment of the tissue; an altered ECM in the asthmatic airway may be central in airway inflammation and remodelling. Tumstatin is a collagen IV-derived matrikine reduced in the asthmatic airway wall that reverses airway inflammation and remodelling in small and large animal models of asthma. This study hypothesized that the mechanisms underlying the broad asthma-resolving effects of tumstatin were due to autocrine remodelling of the ECM. Neutrophils and endothelial cells were seeded on decellularized ECM of non-asthmatic (NA) or asthmatic (A) airway smooth muscle (ASM) cells previously exposed to tumstatin in the presence or absence of a broad matrix metalloproteinase inhibitor, Marimastat. Gene expression in NA and A ASM induced by tumstatin was assessed using RT-PCR arrays. The presence of tumstatin during ECM deposition affected neutrophil and endothelial cell properties on both NA and A ASM-derived matrices and this was only partly due to MMP activity. Gene expression patterns in response to tumstatin in NA and A ASM cells were different. Tumstatin may foster an anti-inflammatory and anti-angiogenic microenvironment by modifying ASM-derived ECM. Further work is required to examine whether restoring tumstatin levels in the asthmatic airway represents a potential novel therapeutic approach. © 2017 The Authors. Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and Molecular Medicine.

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

    SciTech Connect

    Wada, Hiromichi; Abe, Mitsuru; Ono, Koh

    2008-10-03

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

  12. Angiotensin-Converting Enzyme in Smooth Muscle Cells Promotes Atherosclerosis-Brief Report.

    PubMed

    Chen, Xiaofeng; Howatt, Deborah A; Balakrishnan, Anju; Moorleghen, Jessica J; Wu, Congqing; Cassis, Lisa A; Daugherty, Alan; Lu, Hong

    2016-06-01

    Angiotensin-converting enzyme (ACE) is present in many cell types of atherosclerotic lesions. This study determined whether ACE activity in endothelial and smooth muscle cells (SMCs), 2 major resident cell types of the aorta, contributes to hypercholesterolemia-induced atherosclerosis. All study mice were in low-density lipoprotein receptor(-/-) background. To determine the contribution of ACE on endothelial cells to atherosclerosis, female ACE floxed mice were bred to male Tie2-Cre transgenic mice. Endothelial cell-specific deletion of ACE significantly decreased serum ACE activity, but had no effect on systolic blood pressure and atherosclerosis. Because ACE protein is present on SMCs, the most abundant cell type of the aorta, we then determined whether ACE on SMCs contributes to atherosclerosis. ACE was depleted from SMCs by breeding female ACE floxed mice with male SM22-Cre transgenic mice. SMC-specific deficiency of ACE did not affect ACE activity in serum, but ablated its presence and activity in the aortic media. Although SMC-specific deficiency of ACE had no effect on systolic blood pressure, it significantly attenuated hypercholesterolemia-induced atherosclerosis in both male and female mice. These studies provide direct evidence that ACE derived from endothelial cells does not play a critical role in atherosclerosis. Rather, SMC-derived ACE contributes to atherosclerosis, independent of circulating ACE activity and blood pressure. © 2016 American Heart Association, Inc.

  13. BMP-2 overexpression augments vascular smooth muscle cell motility by upregulating myosin Va via Erk signaling.

    PubMed

    Zhang, Ming; Yang, Min; Liu, Li-ping; Lau, Wayne Bond; Gao, Hai; Xin, Man-kun; Su, Li-Xiao; Wang, Jian; Cheng, Shu-Juan; Fan, Qian; Liu, Jing-Hua

    2014-01-01

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

  14. ADAMTS9 mediated extracellular matrix dynamics regulates umbilical cord vascular smooth muscle differentiation and rotation

    PubMed Central

    Nandadasa, Sumeda; Nelson, Courtney M.; Apte, Suneel S.

    2015-01-01

    Summary Despite the significance for fetal nourishment in mammals, mechanisms of umbilical cord vascular growth remain poorly understood. Here, the secreted metalloprotease ADAMTS9 is shown to be necessary for murine umbilical cord vascular development. Restricting it to the cell-surface using a gene trap allele, Adamts9Gt, impaired umbilical vessel elongation and radial growth, via reduced versican proteolysis and accumulation of extracellular matrix (ECM). Both Adamts9Gt and conditional Adamts9 deletion revealed that ADAMTS9 produced by mesenchymal cells acted non-autonomously to regulate smooth muscle cell (SMC) proliferation, differentiation and orthogonal reorientation during growth of the umbilical vasculature. In Adamts9Gt, we observed interference with PDGFRβ signaling via the MAPK/ERK pathway, which regulates cytoskeletal dynamics during SMC rotation. In addition, we observed disrupted Shh signaling and perturbed orientation of the mesenchymal primary cilium. Thus, ECM dynamics is a major influence on umbilical vascular SMC fate, with ADAMTS9 acting as its principal mediator. PMID:26027930

  15. Chloride channel blockade relaxes airway smooth muscle and potentiates relaxation by β-agonists

    PubMed Central

    Yim, Peter; Rinderspacher, Alison; Fu, Xiao Wen; Zhang, Yi; Landry, Donald W.; Emala, Charles W.

    2014-01-01

    Severe bronchospasm refractory to β-agonists continues to cause significant morbidity and mortality in asthmatic patients. We questioned whether chloride channels/transporters are novel targets for the relaxation of airway smooth muscle (ASM). We have screened a library of compounds, derivatives of anthranilic and indanyloxyacetic acid, that were originally developed to antagonize chloride channels in the kidney. We hypothesized that members of this library would be novel calcium-activated chloride channel blockers for the airway. The initial screen of this compound library identified 4 of 20 compounds that relaxed a tetraethylammonium chloride-induced contraction in guinea pig tracheal rings. The two most effective compounds, compounds 1 and 13, were further studied for their potential to either prevent the initiation of or relax the maintenance phase of an acetylcholine (ACh)-induced contraction or to potentiate β-agonist-mediated relaxation. Both relaxed an established ACh-induced contraction in human and guinea pig ex vivo ASM. In contrast, the prevention of an ACh-induced contraction required copretreatment with the sodium-potassium-chloride cotransporter blocker bumetanide. The combination of compound 13 and bumetanide also potentiated relaxation by the β-agonist isoproterenol in guinea pig tracheal rings. Compounds 1 and 13 hyperpolarized the plasma cell membrane of human ASM cells and blocked spontaneous transient inward currents, a measure of chloride currents in these cells. These functional and electrophysiological data suggest that modulating ASM chloride flux is a novel therapeutic target in asthma and other bronchoconstrictive diseases. PMID:24879056

  16. Interaction between aluminum and calcium ions in potassium chloride depolarized gastrointestinal smooth muscle.

    PubMed

    Hava, M; Hurwitz, A

    1975-04-01

    Aluminum chloride (5 times 10-minus 4 M) abolished the inhibitory effects of papaverine chloride (1.5 times 10- minus 5 M) on contractions of guinea-pig ileum and rat colon evoked by calcium in Tyrode's solution containing high KC1. The inhibitory effects of cocaine hydrochloride (10- minus 3 M) and MgSO4 (2 times 10- minus 2 M) were not antagonized. In isolated rat colon depolarized by partial exchange of NaCl by 40 mM KC1, AlC13 decreased the phasic contractions, slowed the onset of tonic contraction while increasing maximal tone and induced superimposed spontaneous activity during the tonic phase. Similar changes were evoked by increasing the CaCl2 concentration from 1.8 times 10- minus 3 M to 7.2 times 10- minus 3 M. The increase in spontaneous activity was not blocked by atropine sulfate (10- minus 7 minus 10- minus 4 M) but was partially antagonized by 10- minus 3 M caffeine and completely blocked by papaverine chloride (5 times 10- minus 4 M). More than one aluminum site of action is proposed for effects of aluminum on the gastro-intestinal smooth muscle.

  17. Smooth Muscle Ion Channels and Regulation of Vascular Tone in Resistance Arteries and Arterioles

    PubMed Central

    Tykocki, Nathan R.; Boerman, Erika M.; Jackson, William F.

    2017-01-01

    Vascular tone of resistance arteries and arterioles determines peripheral vascular resistance, contributing to the regulation of blood pressure and blood flow to, and within the body’s tissues and organs. Ion channels in the plasma membrane and endoplasmic reticulum of vascular smooth muscle cells (SMCs) in these blood vessels importantly contribute to the regulation of intracellular Ca2+ concentration, the primary determinant of SMC contractile activity and vascular tone. Ion channels provide the main source of activator Ca2+ that determines vascular tone, and strongly contribute to setting and regulating membrane potential, which, in turn, regulates the open-state-probability of voltage gated Ca2+ channels (VGCCs), the primary source of Ca2+ in resistance artery and arteriolar SMCs. Ion channel function is also modulated by vasoconstrictors and vasodilators, contributing to all aspects of the regulation of vascular tone. This review will focus on the physiology of VGCCs, voltage-gated K+ (KV) channels, large-conductance Ca2+-activated K+ (BKCa) channels, strong-inward-rectifier K+ (KIR) channels, ATP-sensitive K+ (KA