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Sample records for bladder smooth muscle

  1. Involvement of STAT3 in Bladder Smooth Muscle Hypertrophy Following Bladder Outlet Obstruction

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    Ogawa,Norio

    2006-12-01

    Full Text Available We examined the involvement of the signal transducer and activator of transcription 3 (STAT3 in bladder outlet obstruction (BOO-induced bladder smooth muscle hypertrophy using a rat in vivo and in vitro study. BOO induced increases in bladder weight and bladder smooth muscle thickness 1 week after the operation. By using antibody microarrays, 64 of 389 proteins blotted on the array met our selection criteria of an INR value between > or = 2.0 and < or = 0.5. This result revealed up-regulation of transcription factors, cell cycle regulatory proteins, apoptosis-associated proteins and so on. On the other hand, down-regulation (INR value < or = 0.5 of proteins was not found. In a profiling study, we found an increase in the expression of STAT3. A significant increase in nuclear phosphorylated STAT3 expression was confirmed in bladder smooth muscle tissue by immunohistochemistry and Western blot analysis. Cyclical stretch-relaxation (1 Hz at 120% elongation significantly increased the expression of STAT3 and of alpha-smooth muscle actin in primary cultured bladder smooth muscle cells. Furthermore, the blockade of STAT3 expression by the transfection of STAT3 small interfering RNA (siRNA significantly prevented the stretch-induced increase in alpha-smooth muscle actin expression. These results suggest that STAT3 has an important role in the induction of bladder smooth muscle hypertrophy.

  2. TGF-β1 inhibits connexin-43 expression in cultured smooth muscle cells of human bladder

    Institute of Scientific and Technical Information of China (English)

    Chi Qiang; Zhou Fenghai; Wang Yangmin

    2009-01-01

    Objective: In this research, we studied the TGF-β1 effects on connexin-43 expression in cultured human bladder smooth muscle cells. Methods: Human bladder smooth muscle cells primary cultures, with bladder tissue obtained from patients undergoing cystectomy, were intervened by recombinant human TGF-β1. Connexin-43 expression in human bladder smooth muscle cells was then examined by Western blotting and immunocytochemistry. Results: Stimulation with TGF-β1 led to significant reduction of cormexin-43 immunoreactivity and coupling (P<0.0001). Connexin-43 protein expression was significantly downregnlated (P<0.05). Simultaneously, low phosphorylation species of connexin-43 were particularly affected. Conclusion: Our experiments demonstrated a significant downregulation of connexin-43 by TGF-β1 in cultured human bladder smooth muscle cells. These findings support the view that TGF-β1 is involved in the pathophysiology of urinary bladder dysfunction.

  3. Cell length measurements in longitudinal smooth muscle strips of the pig urinary bladder

    NARCIS (Netherlands)

    E. van Asselt (Els); R. Schot; R. van Mastrigt (Ron)

    1993-01-01

    textabstractIn this study the length of smooth muscle cells in muscle bundles of pig urinary bladder wall was determined after dissection in Tyrode buffers with different calcium concentrations ([Ca2+]). Previous studies have shown that the length of isolated smooth muscle cells decreases with an in

  4. Original Research: Combined model of bladder detrusor smooth muscle and interstitial cells.

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    Rosenberg, Josef; Byrtus, Miroslav; Stengl, Milan

    2016-10-01

    Although patients with lower urinary tract symptoms constitute a large and still growing population, understanding of bladder detrusor muscle physiology remains limited. Understanding the interactions between the detrusor smooth muscle cells and other bladder cell types (e.g. interstitial cells, IC) that may significantly contribute to coordinating and modulating detrusor contractions represents a considerable challenge. Computer modeling could help to elucidate some properties that are difficult to address experimentally; therefore, we developed in silico models of detrusor smooth muscle cell and interstitial cells, coupled through gap junctions. The models include all of the major ion conductances and transporters described in smooth muscle cell and interstitial cells in the literature. The model of normal detrusor muscle (smooth muscle cell and interstitial cells coupled through gap junctions) completely reproduced the experimental results obtained with detrusor strips in the presence of several pharmacological interventions (ryanodine, caffeine, nimodipine), whereas the model of smooth muscle cell alone (without interstitial cells) failed to reproduce the experimental results. Next, a model of overactive bladder, a highly prevalent clinical condition in both men and women with increasing incidence at older ages, was produced by modifying several processes as reported previously: a reduction of Ca(2+)-release through ryanodine receptors and a reduction of Ca(2+)-dependent K(+)-conductance with augmented gap junctional coupling. This model was also able to reproduce the pharmacological modulation of overactive bladder. In conclusion, a model of bladder detrusor muscle was developed that reproduced experimental results obtained in both normal and overactive bladder preparations. The results indicate that the non-smooth muscle cells of the detrusor (interstitial cells) contribute significantly to the contractile behavior of bladder detrusor muscle and should not be

  5. The force recovery following repeated quick releases applied to pig urinary bladder smooth muscle

    NARCIS (Netherlands)

    R. van Mastrigt (Ron)

    1991-01-01

    textabstractA method for measuring several quick-releases during one contraction of a pig urinary bladder smooth muscle preparation was developed. The force recovery following quick release in this muscle type was studied by fitting a multiexponential model to 926 responses measured during the first

  6. Strain history and TGF-β1 induce urinary bladder wall smooth muscle remodeling and elastogenesis

    OpenAIRE

    Heise, Rebecca L.; Parekh, Aron; Joyce, Erinn M.; Michael B. Chancellor; Sacks, Michael S.

    2011-01-01

    Mechanical cues that trigger pathological remodeling in smooth muscle tissues remain largely unknown and are thought to be pivotal triggers for strain-induced remodeling. Thus, an understanding of the effects mechanical stimulation is important to elucidate underlying mechanisms of disease states and in the development of methods for smooth muscle tissue regeneration. For example, the urinary bladder wall (UBW) adaptation to spinal cord injury (SCI) includes extensive hypertrophy as well as i...

  7. Intracellular electrical activity in human urinary bladder smooth muscle: the effect of high sucrose medium

    NARCIS (Netherlands)

    A.J. Visser (Anna); R. van Mastrigt (Ron)

    2001-01-01

    textabstractIntroduction: The primary key to pharmacotherapy of bladder instability is in the excitation-contraction coupling of detrusor smooth muscle cells. To study this process, simultaneous recordings of mechanical and electrical activity are required. However, recording of mechanical activity

  8. Supramaximal stimuli do not evoke a maximal contraction in urinary bladder smooth muscle fibers

    NARCIS (Netherlands)

    J. Minekus (Joanne); A.J. Visser (Anna); R. van Mastrigt (Ron)

    2001-01-01

    textabstractBACKGROUND: Smooth muscle fibers can be stimulated with an electrical field, high potassium or carbachol. We studied the effect of combined, supramaximal stimulation on the isometric force and the maximum shortening velocity of the pig urinary bladder. MATERIALS AND METHODS: After determ

  9. The length dependence of the series elasticity of pig bladder smooth muscle

    NARCIS (Netherlands)

    R. van Mastrigt (Ron)

    1988-01-01

    textabstractStrips of urinary bladder smooth muscle were subjected to a series of quick release measurements. Each measurement consisted of several releases and resets to the original length, made during one contraction. The complete length-force characteristic of series elasticity was quantified by

  10. Strain history and TGF-β1 induce urinary bladder wall smooth muscle remodeling and elastogenesis.

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    Heise, Rebecca L; Parekh, Aron; Joyce, Erinn M; Chancellor, Michael B; Sacks, Michael S

    2012-01-01

    Mechanical cues that trigger pathological remodeling in smooth muscle tissues remain largely unknown and are thought to be pivotal triggers for strain-induced remodeling. Thus, an understanding of the effects mechanical stimulation is important to elucidate underlying mechanisms of disease states and in the development of methods for smooth muscle tissue regeneration. For example, the urinary bladder wall (UBW) adaptation to spinal cord injury (SCI) includes extensive hypertrophy as well as increased collagen and elastin, all of which profoundly alter its mechanical response. In addition, the pro-fibrotic growth factor TGF-β1 is upregulated in pathologies of other smooth muscle tissues and may contribute to pathological remodeling outcomes. In the present study, we utilized an ex vivo organ culture system to investigate the response of UBW tissue under various strain-based mechanical stimuli and exogenous TGF-β1 to assess extracellular matrix (ECM) synthesis, mechanical responses, and bladder smooth muscle cell (BSMC) phenotype. Results indicated that a 0.5-Hz strain frequency triangular waveform stimulation at 15% strain resulted in fibrillar elastin production, collagen turnover, and a more compliant ECM. Further, this stretch regime induced changes in cell phenotype while the addition of TGF-β1 altered this phenotype. This phenotypic shift was further confirmed by passive strip biomechanical testing, whereby the bladder groups treated with TGF-β1 were more compliant than all other groups. TGF-β1 increased soluble collagen production in the cultured bladders. Overall, the 0.5-Hz strain-induced remodeling caused increased compliance due to elastogenesis, similar to that seen in early SCI bladders. Thus, organ culture of bladder strips can be used as an experimental model to examine ECM remodeling and cellular phenotypic shift and potentially elucidate BMSCs ability to produce fibrillar elastin using mechanical stretch either alone or in combination with

  11. Mechanical stretch upregulates proteins involved in Ca2+ sensitization in urinary bladder smooth muscle hypertrophy.

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    Boopathi, Ettickan; Gomes, Cristiano; Zderic, Stephen A; Malkowicz, Bruce; Chakrabarti, Ranjita; Patel, Darshan P; Wein, Alan J; Chacko, Samuel

    2014-09-15

    Partial bladder outlet obstruction (pBOO)-induced remodeling of bladder detrusor smooth muscle (DSM) is associated with the modulation of cell signals regulating contraction. We analyzed the DSM from obstructed murine urinary bladders for the temporal regulation of RhoA GTPase and Rho-activated kinase (ROCK), which are linked to Ca(2+) sensitization. In addition, the effects of equibiaxial cell stretch, a condition thought to be associated with pBOO-induced bladder wall smooth muscle hypertrophy and voiding frequency, on the expression of RhoA, ROCK, and C-kinase-activated protein phosphatase I inhibitor (CPI-17) were investigated. DSM from 1-, 3-, 7-, and 14-day obstructed male mice bladders and benign prostatic hyperplasia (BPH)-induced obstructed human bladders revealed overexpression of RhoA and ROCK-β at the mRNA and protein levels compared with control. Primary human bladder myocytes seeded onto type I collagen-coated elastic silicone membranes were subjected to cyclic equibiaxial stretch, mimicking the cellular mechanical stretch in the bladder in vivo, and analyzed for the expression of RhoA, ROCK-β, and CPI-17. Stretch caused a significant increase of RhoA, ROCKβ, and CPI-17 expression. The stretch-induced increase in CPI-17 expression occurs at the transcriptional level and is associated with CPI-17 promoter binding by GATA-6 and NF-κB, the transcription factors responsible for CPI-17 gene transcription. Cell stretch caused by bladder overdistension in pBOO is the likely mechanism for initiating overexpression of the signaling proteins regulating DSM tone.

  12. [The effect of prostatic peptides on the contractile activity of smooth-muscle cells from the bladder].

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    Barabanova, V V; Gorbachev, A G; Parastaeva, M M; Khavinson, V Kh

    1993-02-01

    Prostatilene (PST) enhanced the functional activity of the bladder smooth-muscle cells (SMC). The possibility of activation of the SMC contractility by the PST through pharmacomechanical associations, is discussed.

  13. [The effect of prostatilen on the contractile activity of the smooth-muscle cells of the blood vessels and bladder in cats].

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    al-Shchukri, S Kh; Barabanov, S V; Barabanova, V V; Bobkov, Iu A; Gorbachev, A G; Parastaeva, M M

    1996-07-01

    Prostatilene enhanced the functional activity of the bladder and blood vessels' smooth muscle cells. A possibility of activation of the smooth muscle cells contractility with prostatilene by a pharmaco-mechanical association, is discussed.

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

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    Juliane Brun

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

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

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

    2015-01-01

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

  16. Effects of hypoxia and glucose-removal condition on muscle contraction of the smooth muscles of porcine urinary bladder.

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    Nagai, Yuta; Kaneda, Takeharu; Miyamoto, Yasuyuki; Nuruki, Takaomi; Kanda, Hidenori; Urakawa, Norimoto; Shimizu, Kazumasa

    2016-01-01

    To elucidate the dependence of aerobic energy metabolism and utilization of glucose in contraction of urinary bladder smooth muscle, we investigated the changes in the reduced pyridine nucleotide (PNred) fluorescence, representing glycolysis activity, and determined the phosphocreatine (PCr) and ATP contents of the porcine urinary bladder during contractions induced by high K(+) or carbachol (CCh) and with and without hypoxia (achieved by bubbling N2 instead of O2) or in a glucose-free condition. Hyperosmotic addition of 65 mM KCl (H-65K(+)) and 1 µM CCh induced a phasic contraction followed by a tonic contraction. A glucose-free physiological salt solution (PSS) did not change the subsequent contractile responses to H-65K(+) and CCh. However, hypoxia significantly attenuated H-65K(+)- and CCh-induced contraction. H-65K(+) and CCh induced a sustained increase in PNred fluorescence, representing glycolysis activity. Hypoxia enhanced H-65K(+)- and CCh-induced increases in PNred fluorescence, whereas glucose-free PSS decreased these increases, significantly. In the presence of H-65K(+), hypoxia decreased the PCr and ATP contents; however, the glucose-free PSS did not change the PCr contents. In conclusion, we demonstrated that high K(+)- and CCh-induced contractions depend on aerobic metabolism and that an endogenous substrate may be utilized to maintain muscle contraction in a glucose-free PSS in the porcine urinary bladder.

  17. Free fatty acid palmitate impairs the vitality and function of cultured human bladder smooth muscle cells.

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    Andreas Oberbach

    Full Text Available BACKGROUND: Incidence of urinary tract infections is elevated in patients with diabetes mellitus. Those patients show increased levels of the saturated free fatty acid palmitate. As recently shown metabolic alterations induced by palmitate include production and secretion of the pro-inflammatory cytokine interleukine-6 (IL-6 in cultured human bladder smooth muscle cells (hBSMC. Here we studied the influence of palmitate on vital cell properties, for example, regulation of cell proliferation, mitochondrial enzyme activity and antioxidant capacity in hBSMC, and analyzed the involvement of major cytokine signaling pathways. METHODOLOGY/PRINCIPAL FINDINGS: HBSMC cultures were set up from bladder tissue of patients undergoing cystectomy and stimulated with palmitate. We analyzed cell proliferation, mitochondrial enzyme activity, and antioxidant capacity by ELISA and confocal immunofluorescence. In signal transduction inhibition experiments we evaluated the involvement of NF-κB, JAK/STAT, MEK1, PI3K, and JNK in major cytokine signaling pathway regulation. We found: (i palmitate decreased cell proliferation, increased mitochondrial enzyme activity and antioxidant capacity; (ii direct inhibition of cytokine receptor by AG490 even more strongly suppressed cell proliferation in palmitate-stimulated cells, while counteracting palmitate-induced increase of antioxidant capacity; (iii in contrast knockdown of the STAT3 inhibitor SOCS3 increased cell proliferation and antioxidant capacity; (iv further downstream JAK/STAT3 signaling cascade the inhibition of PI3K or JNK enhanced palmitate induced suppression of cell proliferation; (v increase of mitochondrial enzyme activity by palmitate was enhanced by inhibition of PI3K but counteracted by inhibition of MEK1. CONCLUSIONS/SIGNIFICANCE: Saturated free fatty acids (e.g., palmitate cause massive alterations in vital cell functions of cultured hBSMC involving distinct major cytokine signaling pathways. Thereby

  18. Expression and function of K(V)2-containing channels in human urinary bladder smooth muscle.

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    Hristov, Kiril L; Chen, Muyan; Afeli, Serge A Y; Cheng, Qiuping; Rovner, Eric S; Petkov, Georgi V

    2012-06-01

    The functional role of the voltage-gated K(+) (K(V)) channels in human detrusor smooth muscle (DSM) is largely unexplored. Here, we provide molecular, electrophysiological, and functional evidence for the expression of K(V)2.1, K(V)2.2, and the electrically silent K(V)9.3 subunits in human DSM. Stromatoxin-1 (ScTx1), a selective inhibitor of K(V)2.1, K(V)2.2, and K(V)4.2 homotetrameric channels and of K(V)2.1/9.3 heterotetrameric channels, was used to examine the role of these channels in human DSM function. Human DSM tissues were obtained during open bladder surgeries from patients without a history of overactive bladder. Freshly isolated human DSM cells were studied using RT-PCR, immunocytochemistry, live-cell Ca(2+) imaging, and the perforated whole cell patch-clamp technique. Isometric DSM tension recordings of human DSM isolated strips were conducted using tissue baths. RT-PCR experiments showed mRNA expression of K(V)2.1, K(V)2.2, and K(V)9.3 (but not K(V)4.2) channel subunits in human isolated DSM cells. K(V)2.1 and K(V)2.2 protein expression was confirmed by Western blot analysis and immunocytochemistry. Perforated whole cell patch-clamp experiments revealed that ScTx1 (100 nM) inhibited the amplitude of the voltage step-induced K(V) current in freshly isolated human DSM cells. ScTx1 (100 nM) significantly increased the intracellular Ca(2+) level in DSM cells. In human DSM isolated strips, ScTx1 (100 nM) increased the spontaneous phasic contraction amplitude and muscle force, and enhanced the amplitude of the electrical field stimulation-induced contractions within the range of 3.5-30 Hz stimulation frequencies. These findings reveal that ScTx1-sensitive K(V)2-containing channels are key regulators of human DSM excitability and contractility and may represent new targets for pharmacological or genetic intervention for bladder dysfunction.

  19. Eviprostat Activates cAMP Signaling Pathway and Suppresses Bladder Smooth Muscle Cell Proliferation

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    Masayuki Takeda

    2013-06-01

    Full Text Available Eviprostat is a popular phytotherapeutic agent for the treatment of lower urinary tract symptoms (LUTS. At present, the signaling mechanisms underlying its therapeutic effects are still poorly understood. Given that cAMP has been reported to suppress cell hyperplasia and hypertrophy in various pathological situations, we asked whether the effect of Eviprostat could be ascribed to the activation of the cAMP signaling pathway. In the study, exposure of cAMP response element (CRE-secreted alkaline phosphatase (SEAP (CRE-SEAP-reporter cells to Eviprostat elevated SEAP secretion, which was associated with an increased phosphorylation of vasodilator-stimulated phosphoprotein (VASP and cAMP-response element-binding protein (CREB, as well as enhanced expression of CRE-regulated protein connexin43, indicating an activation of the cAMP signaling pathway. Consistent with these observations, Eviprostat-induced expression of Cx43 was abolished in the presence of adenylyl cyclase inhibitor SQ22536 or PKA inhibitor H89, whereas it was mimicked by adenylyl cyclase activator, forskolin. Further analysis demonstrated that Eviprostat significantly potentiated the effect of phosphodiesterase 3 (PDE3 inhibitor, but not that of PDE4 inhibitor, on CRE activation. Moreover, Eviprostat suppressed PDGF-induced activation of ERK and Akt and inhibited cell proliferation and hillock formation in both mesangial cells and bladder smooth muscle cells. Collectively, activation of the cAMP signaling pathway could be an important mechanism by which Eviprostat exerts its therapeutic effects for LUTS.

  20. Cytokine effects on gap junction communication and connexin expression in human bladder smooth muscle cells and suburothelial myofibroblasts.

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    Marco Heinrich

    Full Text Available BACKGROUND: The last decade identified cytokines as one group of major local cell signaling molecules related to bladder dysfunction like interstitial cystitis (IC and overactive bladder syndrome (OAB. Gap junctional intercellular communication (GJIC is essential for the coordination of normal bladder function and has been found to be altered in bladder dysfunction. Connexin (Cx 43 and Cx45 are the most important gap junction proteins in bladder smooth muscle cells (hBSMC and suburothelial myofibroblasts (hsMF. Modulation of connexin expression by cytokines has been demonstrated in various tissues. Therefore, we investigate the effect of interleukin (IL 4, IL6, IL10, tumor necrosis factor-alpha (TNFα and transforming growth factor-beta1 (TGFβ1 on GJIC, and Cx43 and Cx45 expression in cultured human bladder smooth muscle cells (hBSMC and human suburothelial myofibroblasts (hsMF. METHODOLOGY/PRINCIPAL FINDINGS: HBSMC and hsMF cultures were set up from bladder tissue of patients undergoing cystectomy. In cytokine stimulated cultured hBSMC and hsMF GJIC was analyzed via Fluorescence Recovery after Photo-bleaching (FRAP. Cx43 and Cx45 expression was assessed by quantitative PCR and confocal immunofluorescence. Membrane protein fraction of Cx43 and Cx45 was quantified by Dot Blot. Upregulation of cell-cell-communication was found after IL6 stimulation in both cell types. In hBSMC IL4 and TGFβ1 decreased both, GJIC and Cx43 protein expression, while TNFα did not alter communication in FRAP-experiments but increased Cx43 expression. GJ plaques size correlated with coupling efficacy measured, while Cx45 expression did not correlate with modulation of GJIC. CONCLUSIONS/SIGNIFICANCE: Our finding of specific cytokine effects on GJIC support the notion that cytokines play a pivotal role for pathophysiology of OAB and IC. Interestingly, the effects were independent from the classical definition of pro- and antiinflammatory cytokines. We conclude, that

  1. NS19504: a novel BK channel activator with relaxing effect on bladder smooth muscle spontaneous phasic contractions.

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    Nausch, Bernhard; Rode, Frederik; Jørgensen, Susanne; Nardi, Antonio; Korsgaard, Mads P G; Hougaard, Charlotte; Bonev, Adrian D; Brown, William D; Dyhring, Tino; Strøbæk, Dorte; Olesen, Søren-Peter; Christophersen, Palle; Grunnet, Morten; Nelson, Mark T; Rønn, Lars C B

    2014-09-01

    Large-conductance Ca(2+)-activated K(+) channels (BK, KCa1.1, MaxiK) are important regulators of urinary bladder function and may be an attractive therapeutic target in bladder disorders. In this study, we established a high-throughput fluorometric imaging plate reader-based screening assay for BK channel activators and identified a small-molecule positive modulator, NS19504 (5-[(4-bromophenyl)methyl]-1,3-thiazol-2-amine), which activated the BK channel with an EC50 value of 11.0 ± 1.4 µM. Hit validation was performed using high-throughput electrophysiology (QPatch), and further characterization was achieved in manual whole-cell and inside-out patch-clamp studies in human embryonic kidney 293 cells expressing hBK channels: NS19504 caused distinct activation from a concentration of 0.3 and 10 µM NS19504 left-shifted the voltage activation curve by 60 mV. Furthermore, whole-cell recording showed that NS19504 activated BK channels in native smooth muscle cells from guinea pig urinary bladder. In guinea pig urinary bladder strips, NS19504 (1 µM) reduced spontaneous phasic contractions, an effect that was significantly inhibited by the specific BK channel blocker iberiotoxin. In contrast, NS19504 (1 µM) only modestly inhibited nerve-evoked contractions and had no effect on contractions induced by a high K(+) concentration consistent with a K(+) channel-mediated action. Collectively, these results show that NS19504 is a positive modulator of BK channels and provide support for the role of BK channels in urinary bladder function. The pharmacologic profile of NS19504 indicates that this compound may have the potential to reduce nonvoiding contractions associated with spontaneous bladder overactivity while having a minimal effect on normal voiding.

  2. Amino acid mutations in the caldesmon COOH-terminal functional domain increase force generation in bladder smooth muscle.

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    Deng, Maoxian; Boopathi, Ettickan; Hypolite, Joseph A; Raabe, Tobias; Chang, Shaohua; Zderic, Stephen; Wein, Alan J; Chacko, Samuel

    2013-11-15

    Caldesmon (CaD), a component of smooth muscle thin filaments, binds actin, tropomyosin, calmodulin, and myosin and inhibits actin-activated ATP hydrolysis by smooth muscle myosin. Internal deletions of the chicken CaD functional domain that spans from amino acids (aa) 718 to 731, which corresponds to aa 512-530 including the adjacent aa sequence in mouse CaD, lead to diminished CaD-induced inhibition of actin-activated ATP hydrolysis by myosin. Transgenic mice with mutations of five aa residues (Lys(523) to Gln, Val(524) to Leu, Ser(526) to Thr, Pro(527) to Cys, and Lys(529) to Ser), which encompass the ATPase inhibitory determinants located in exon 12, were generated by homologous recombination. Homozygous (-/-) animals did not develop, but heterozygous (+/-) mice carrying the expected mutations in the CaD ATPase inhibitory domain (CaD mutant) matured and reproduced normally. The peak force produced in response to KCl and electrical field stimulation by the detrusor smooth muscle from the CaD mutant was high compared with that of the wild type. CaD mutant mice revealed nonvoiding contractions during bladder filling on awake cystometry, suggesting that the CaD ATPase inhibitory domain suppresses force generation during the filling phase and this suppression is partially released by mutations in 50% of CaD in heterozygous. Our data show for the first time a functional phenotype, at the intact smooth muscle tissue and in vivo organ levels, following mutation of a functional domain at the COOH-terminal region of CaD.

  3. Hypoxia-increased expression of genes involved in inflammation, dedifferentiation, pro-fibrosis, and extracellular matrix remodeling of human bladder smooth muscle cells.

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    Wiafe, Bridget; Adesida, Adetola; Churchill, Thomas; Adewuyi, Esther Ekpe; Li, Zack; Metcalfe, Peter

    2017-01-01

    Partial bladder outlet obstruction (pBOO) is characterized by exaggerated stretch, hydrodynamic pressure, and inflammation which cause significant damage and fibrosis to the bladder wall. Several studies have implicated hypoxia in its pathophysiology. However, the isolated progressive effects of hypoxia on bladder cells are not yet defined. Sub-confluent normal human bladder smooth muscle cells (hbSMC) were cultured in 3% O2 tension for 2, 24, 48, and 72 h. RNA, cellular proteins, and secreted proteins were used for gene expression analysis, immunoblotting, and ELISA, respectively. Transcription of hypoxia-inducible factor (HIF)1α and HIF2α were transiently induced after 2 h of hypoxia (p inflammation, de-differentiation, pro-fibrotic changes, and increased extracellular matrix expression. This elucidates mechanisms of hypoxia-driven bladder deterioration in bladder cells, which is important in tailoring in vivo experiments and may ultimately translate into improved clinical outcomes.

  4. The purinergic component of human bladder smooth muscle cells’ proliferation and contraction under physiological stretch

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    Wazir, Romel; Luo, De-Yi; Tian, Ye; Yue, Xuan; Li, Hong; Wang, Kun-Jie, E-mail: kunjiewangatscu@163.com

    2013-07-26

    Highlights: •Stretch induces proliferation and contraction. •Optimum applied stretch in vitro is 5% and 10% equibiaxial stretching respectively. •Expression of P2X1 and P2X2 is upregulated after application of stretch. •P2X2 is possibly more susceptible to stretch related changes. •Purinoceptors functioning may explain conditions with atropine resistance. -- Abstract: Objective: To investigate whether cyclic stretch induces proliferation and contraction of human smooth muscle cells (HBSMCs), mediated by P2X purinoceptor 1 and 2 and the signal transduction mechanisms of this process. Methods: HBSMCs were seeded on silicone membrane and stretched under varying parameters; (equibiaxial elongation: 2.5%, 5%, 10%, 15%, 20%, 25%), (Frequency: 0.05 Hz, 0.1 Hz, 0.2 Hz, 0.5 Hz, 1 Hz). 5-Bromo-2-deoxyuridine assay was employed for proliferative studies. Contractility of the cells was determined using collagen gel contraction assay. After optimal physiological stretch was established; P2X1 and P2X2 were analyzed by real time polymerase chain reaction and Western Blot. Specificity of purinoceptors was maintained by employing specific inhibitors; (NF023 for P2X1, and A317491for P2X2), in some experiments. Results: Optimum proliferation and contractility were observed at 5% and 10% equibiaxial stretching respectively, applied at a frequency of 0.1 Hz; At 5% stretch, proliferation increased from 0.837 ± 0.026 (control) to 1.462 ± 0.023%, p < 0.05. Mean contraction at 10% stretching increased from 31.7 ± 2.3%, (control) to 78.28 ±1.45%, p < 0.05. Expression of P2X1 and P2X2 was upregulated after application of stretch. Inhibition had effects on proliferation (1.232 ± 0.051, p < 0.05 NF023) and (1.302 ± 0.021, p < 0.05 A314791) while contractility was markedly reduced (68.24 ± 2.31, p < 0.05 NF023) and (73.2 ± 2.87, p < 0.05 A314791). These findings shows that mechanical stretch can promote magnitude-dependent proliferative and contractile modulation of HBSMCs in

  5. Isolation and individual electrical stimulation of single smooth-muscle cells from the urinary bladder of the pig

    NARCIS (Netherlands)

    J.J. Glerum (Jacobus); R. van Mastrigt (Ron); J.C. Romijn (Johannes); D.J. Griffiths (Derek)

    1987-01-01

    textabstractIn contrast to striated muscle, measurements on strips of smooth muscle cannot be uniquely interpreted in terms of an array of contractile units. Therefore scaling down to the single-cell level is necessary to gain detailed understanding of the contractile process in this type of muscle.

  6. Transcriptional repression of Caveolin-1 (CAV1) gene expression by GATA-6 in bladder smooth muscle hypertrophy in mice and human beings.

    Science.gov (United States)

    Boopathi, Ettickan; Gomes, Cristiano Mendes; Goldfarb, Robert; John, Mary; Srinivasan, Vittala Gopal; Alanzi, Jaber; Malkowicz, S Bruce; Kathuria, Hasmeena; Zderic, Stephen A; Wein, Alan J; Chacko, Samuel

    2011-05-01

    Hypertrophy occurs in urinary bladder wall smooth muscle (BSM) in men with partial bladder outlet obstruction (PBOO) caused by benign prostatic hyperplasia (BPH) and in animal models of PBOO. Hypertrophied BSM from the rabbit model exhibits down-regulation of caveolin-1, a structural and functional protein of caveolae that function as signaling platforms to mediate interaction between receptor proteins and adaptor and effector molecules to regulate signal generation, amplification, and diversification. Caveolin-1 expression is diminished in PBOO-induced BSM hypertrophy in mice and in men with BPH. The proximal promoter of the human and mouse caveolin-1 (CAV1) gene was characterized, and it was observed that the transcription factor GATA-6 binds this promoter, causing reduced expression of caveolin-1. Furthermore, caveolin-1 expression levels inversely correlate with the abundance of GATA-6 in BSM hypertrophy in mice and human beings. Silencing of GATA6 gene expression up-regulates caveolin-1 expression, whereas overexpression of GATA-6 protein sustains the transcriptional repression of caveolin-1 in bladder smooth muscle cells. Together, these data suggest that GATA-6 acts as a transcriptional repressor of CAV1 gene expression in PBOO-induced BSM hypertrophy in men and mice. GATA-6-induced transcriptional repression represents a new regulatory mechanism of CAV1 gene expression in pathologic BSM, and may serve as a target for new therapy for BPH-induced bladder dysfunction in aging men.

  7. A droplet-based building block approach for bladder smooth muscle cell (SMC) proliferation.

    Science.gov (United States)

    Xu, F; Moon, S J; Emre, A E; Turali, E S; Song, Y S; Hacking, S A; Nagatomi, J; Demirci, U

    2010-03-01

    Tissue engineering based on building blocks is an emerging method to fabricate 3D tissue constructs. This method requires depositing and assembling building blocks (cell-laden microgels) at high throughput. The current technologies (e.g., molding and photolithography) to fabricate microgels have throughput challenges and provide limited control over building block properties (e.g., cell density). The cell-encapsulating droplet generation technique has potential to address these challenges. In this study, we monitored individual building blocks for viability, proliferation and cell density. The results showed that (i) SMCs can be encapsulated in collagen droplets with high viability (>94.2 +/- 3.2%) for four cases of initial number of cells per building block (i.e. 7 +/- 2, 16 +/- 2, 26 +/- 3 and 37 +/- 3 cells/building block). (ii) Encapsulated SMCs can proliferate in building blocks at rates that are consistent (1.49 +/- 0.29) across all four cases, compared to that of the controls. (iii) By assembling these building blocks, we created an SMC patch (5 mm x 5 mm x 20 microm), which was cultured for 51 days forming a 3D tissue-like construct. The histology of the cultured patch was compared to that of a native rat bladder. These results indicate the potential of creating 3D tissue models at high throughput in vitro using building blocks.

  8. An aryloxypropanolamine hβ3-adrenoceptor agonist as bladder smooth muscle relaxant.

    Science.gov (United States)

    Tasler, Stefan; Baumgartner, Roland; Behr-Roussel, Delphine; Oger-Roussel, Stephanie; Gorny, Diane; Giuliano, Francois; Ney, Peter

    2012-08-15

    The relaxant effect of an aryloxypropanolamine β3-adrenoceptor agonist on carbachol pre-contracted human detrusor muscle strips was evaluated and compared with literature results from reference compounds of similar mode of action, including mirabegron. A significant relaxation was observed for rac-4-{2-hydroxy-3-[1-(5-phenylthieno[2,3-d]pyrimidin-4-yl)piperidin-4-ylamino]propoxy}-2-(hydroxymethyl)phenol which was similar to that exerted by mirabegron. In order to allow for a thorough discussion of results in comparison to reference compounds, their affinity, selectivity and efficacy as hβ3-AR agonists have been evaluated and discussed thoroughly. A ranking of hβ3-AR agonists by relative efficacy resulted in the closest analogy to the order of relaxation potential, with only the relaxant effect of mirabegron not reflecting its excellent relative efficacy as such.

  9. Voltage-gated K(+) channels sensitive to stromatoxin-1 regulate myogenic and neurogenic contractions of rat urinary bladder smooth muscle.

    Science.gov (United States)

    Chen, Muyan; Kellett, Whitney F; Petkov, Georgi V

    2010-07-01

    Members of the voltage-gated K(+) (K(V)) channel family are suggested to control the resting membrane potential and the repolarization phase of the action potential in urinary bladder smooth muscle (UBSM). Recent studies report that stromatoxin-1, a peptide isolated from tarantulas, selectively inhibits K(V)2.1, K(V)2.2, K(V)4.2, and K(V)2.1/9.3 channels. The objective of this study was to investigate whether K(V) channels sensitive to stromatoxin-1 participate in the regulation of rat UBSM contractility and to identify their molecular fingerprints. Stromatoxin-1 (100 nM) increased the spontaneous phasic contraction amplitude, muscle force, and tone in isolated UBSM strips. However, stromatoxin-1 (100 nM) had no effect on the UBSM contractions induced by depolarizing agents such as KCl (20 mM) or carbachol (1 microM). This indicates that, under conditions of sustained membrane depolarization, the K(V) channels sensitive to stromatoxin-1 have no further contribution to the membrane excitability and contractility. Stromatoxin-1 (100 nM) increased the amplitude of the electrical field stimulation-induced contractions, suggesting also a role for these channels in neurogenic contractions. RT-PCR experiments on freshly isolated UBSM cells showed mRNA expression of K(V)2.1, K(V)2.2, and K(V)9.3, but not K(V)4.2 channel subunits. Protein expression of K(V)2.1 and K(V)2.2 channels was detected using Western blot and was further confirmed by immunocytochemical detection in freshly isolated UBSM cells. These novel findings indicate that K(V)2.1 and K(V)2.2, but not K(V)4.2, channel subunits are expressed in rat UBSM and play a key role in opposing both myogenic and neurogenic UBSM contractions.

  10. Novel mechanism of hydrogen sulfide-induced guinea pig urinary bladder smooth muscle contraction: role of BK channels and cholinergic neurotransmission.

    Science.gov (United States)

    Fernandes, Vítor S; Xin, Wenkuan; Petkov, Georgi V

    2015-07-15

    Hydrogen sulfide (H2S) is a key signaling molecule regulating important physiological processes, including smooth muscle function. However, the mechanisms underlying H2S-induced detrusor smooth muscle (DSM) contractions are not well understood. This study investigates the cellular and tissue mechanisms by which H2S regulates DSM contractility, excitatory neurotransmission, and large-conductance voltage- and Ca(2+)-activated K(+) (BK) channels in freshly isolated guinea pig DSM. We used a multidisciplinary experimental approach including isometric DSM tension recordings, colorimetric ACh measurement, Ca(2+) imaging, and patch-clamp electrophysiology. In isolated DSM strips, the novel slow release H2S donor, P-(4-methoxyphenyl)-p-4-morpholinylphosphinodithioic acid morpholine salt (GYY4137), significantly increased the spontaneous phasic and nerve-evoked DSM contractions. The blockade of neuronal voltage-gated Na(+) channels or muscarinic ACh receptors with tetrodotoxin or atropine, respectively, reduced the stimulatory effect of GYY4137 on DSM contractility. GYY4137 increased ACh release from bladder nerves, which was inhibited upon blockade of L-type voltage-gated Ca(2+) channels with nifedipine. Furthermore, GYY4137 increased the amplitude of the Ca(2+) transients and basal Ca(2+) levels in isolated DSM strips. GYY4137 reduced the DSM relaxation induced by the BK channel opener, NS11021. In freshly isolated DSM cells, GYY4137 decreased the amplitude and frequency of transient BK currents recorded in a perforated whole cell configuration and reduced the single BK channel open probability measured in excised inside-out patches. GYY4137 inhibited spontaneous transient hyperpolarizations and depolarized the DSM cell membrane potential. Our results reveal the novel findings that H2S increases spontaneous phasic and nerve-evoked DSM contractions by activating ACh release from bladder nerves in combination with a direct inhibition of DSM BK channels.

  11. Anti-smooth muscle antibody

    Science.gov (United States)

    ... gov/ency/article/003531.htm Anti-smooth muscle antibody To use the sharing features on this page, please enable JavaScript. Anti-smooth muscle antibody is a blood test that detects the presence ...

  12. Functional BK channels facilitate the β3-adrenoceptor agonist-mediated relaxation of nerve-evoked contractions in rat urinary bladder smooth muscle isolated strips.

    Science.gov (United States)

    Afeli, Serge A Y; Petkov, Georgi V

    2013-07-05

    The large-conductance voltage- and Ca(2+)-activated K(+) (BK) channel is a major regulator of detrusor smooth muscle (DSM) contractility thus facilitating urinary bladder function. Recent findings suggest that activation of β3-adrenoceptors causes DSM relaxation. However, it is unknown whether the β3-adrenoceptor-mediated DSM relaxation is BK channel-dependent during nerve-evoked contractions. To test this hypothesis, we induced nerve-evoked contractions in rat DSM isolated strips by using a tissue bath system equipped with platinum electrodes for electrical field stimulation (EFS). (±)-(R(*),R(*))-[4-[2-[[2-(3-chlorophenyl)-2-hydroxyethyl]amino]propyl]phenoxy] acetic acid sodium hydrate (BRL37344), a β3-adrenoceptor agonist, significantly decreased the amplitude and muscle force of the 20 Hz EFS-induced DSM contractions in a concentration-dependent manner. The BRL37344 inhibitory effect was significantly antagonized by 1-(2-ethylphenoxy)-3-[[(1S)-1,2,3,4-tetrahydro-1-naphthalenyl]amino]-(2S)-2-propanol hydrochloride (SR59230A), a β3-adrenoceptor antagonist. We further isolated the cholinergic from the purinergic component of the 0.5-50 Hz EFS-induced DSM contractions by using selective inhibitors, atropine as well as suramin and α,β-methylene-ATP. We found that BRL37344 inhibited both the purinergic and cholinergic components of the nerve-evoked contractions in rat DSM isolated strips. The pharmacological blockade of the BK channels with iberiotoxin, a selective BK channel inhibitor, increased the amplitude and muscle force of the 20 Hz EFS-induced contractions in rat DSM isolated strips. In the presence of iberiotoxin, there was a significant reduction in the BRL37344-induced inhibition of the 20 Hz EFS-induced contractions in rat DSM isolated strips. These latter findings suggest that BK channels play a critical role in the β3-adrenoceptor-mediated inhibition of rat DSM nerve-evoked contractions.

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

    Science.gov (United States)

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

    2012-03-01

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

  14. Regeneration and Maintenance of Intestinal Smooth Muscle Phenotypes

    Science.gov (United States)

    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

  15. Pharmacology of airway smooth muscle proliferation

    NARCIS (Netherlands)

    Gosens, Reinoud; Roscioni, Sara S.; Dekkers, Bart G. J.; Pera, Tonio; Schmidt, Martina; Schaafsma, Dedmer; Zaagsma, Johan; Meurs, Herman

    2008-01-01

    Airway smooth muscle thickening is a pathological feature that contributes significantly to airflow limitation and airway hyperresponsiveness in asthma. Ongoing research efforts aimed at identifying the mechanisms responsible for the increased airway smooth muscle mass have indicated that hyperplasi

  16. Large-conductance voltage- and Ca2+-activated K+ channel regulation by protein kinase C in guinea pig urinary bladder smooth muscle.

    Science.gov (United States)

    Hristov, Kiril L; Smith, Amy C; Parajuli, Shankar P; Malysz, John; Petkov, Georgi V

    2014-03-01

    Large-conductance voltage- and Ca(2+)-activated K(+) (BK) channels are critical regulators of detrusor smooth muscle (DSM) excitability and contractility. PKC modulates the contraction of DSM and BK channel activity in non-DSM cells; however, the cellular mechanism regulating the PKC-BK channel interaction in DSM remains unknown. We provide a novel mechanistic insight into BK channel regulation by PKC in DSM. We used patch-clamp electrophysiology, live-cell Ca(2+) imaging, and functional studies of DSM contractility to elucidate BK channel regulation by PKC at cellular and tissue levels. Voltage-clamp experiments showed that pharmacological activation of PKC with PMA inhibited the spontaneous transient BK currents in native freshly isolated guinea pig DSM cells. Current-clamp recordings revealed that PMA significantly depolarized DSM membrane potential and inhibited the spontaneous transient hyperpolarizations in DSM cells. The PMA inhibitory effects on DSM membrane potential were completely abolished by the selective BK channel inhibitor paxilline. Activation of PKC with PMA did not affect the amplitude of the voltage-step-induced whole cell steady-state BK current or the single BK channel open probability (recorded in cell-attached mode) upon inhibition of all major Ca(2+) sources for BK channel activation with thapsigargin, ryanodine, and nifedipine. PKC activation with PMA elevated intracellular Ca(2+) levels in DSM cells and increased spontaneous phasic and nerve-evoked contractions of DSM isolated strips. Our results support the concept that PKC activation leads to a reduction of BK channel activity in DSM via a Ca(2+)-dependent mechanism, thus increasing DSM contractility.

  17. Airway Epithelium Stimulates Smooth Muscle Proliferation

    OpenAIRE

    Malavia, Nikita K.; Raub, Christopher B.; Mahon, Sari B.; Brenner, Matthew; Reynold A Panettieri; George, Steven C.

    2009-01-01

    Communication between the airway epithelium and stroma is evident during embryogenesis, and both epithelial shedding and increased smooth muscle proliferation are features of airway remodeling. Hence, we hypothesized that after injury the airway epithelium could modulate airway smooth muscle proliferation. Fully differentiated primary normal human bronchial epithelial (NHBE) cells at an air–liquid interface were co-cultured with serum-deprived normal primary human airway smooth muscle cells (...

  18. [Influence of prostatilen on smooth muscle organs functional activity in surgical patients (clinical and experimental study)].

    Science.gov (United States)

    Al'-Shukri, S Kh; Aĭvazian, A I; Barabanov, S V; Barabanova, V V; Bobkov, Iu A; Gorbachev, A G; Parastaeva, M M

    1999-01-01

    The action of prostatilen on contractile activity of smooth muscles of isolated line slices of urine bladder of Wistar rats (myography) and arterial vessels of cat kidneys (resistography) was studied. On the basis of clinical cases effectiveness of prostatilen was analysed as a treatment restorting urine bladder function in acute reflex urinary retention after operations in the area of rectal sphincter, as well as in treatment of patients with chronic prostatitis. It is shown, that prostatilen produces contractile action on smooth muscles of renal blood vessels in cats and urine bladder walls in rats and it raises contractile activity of smooth muscles of human urine bladder. The results of experimental and clinical investigations make it possible to recommend the application of this bioregulating preparation for treatment and prophylaxis of disturbances in urination.

  19. Cardiac, Skeletal, and smooth muscle mitochondrial respiration

    DEFF Research Database (Denmark)

    Park, Song-Young; Gifford, Jayson R; Andtbacka, Robert H I

    2014-01-01

    Unlike cardiac and skeletal muscle, little is known about vascular smooth muscle mitochondrial function. Therefore, this study examined mitochondrial respiratory rates in the smooth muscle of healthy human feed arteries and compared with that of healthy cardiac and skeletal muscle. Cardiac......, skeletal, and smooth muscle was harvested from a total of 22 subjects (53±6 yrs) and mitochondrial respiration assessed in permeabilized fibers. Complex I+II, state 3 respiration, an index of oxidative phosphorylation capacity, fell progressively from cardiac, skeletal, to smooth muscle (54±1; 39±4; 15......±1 pmol•s(-1)•mg (-1), psmooth muscle (222±13; 115±2; 48±2 umol•g(-1)•min(-1), p

  20. Novel treatment strategies for smooth muscle disorders: Targeting Kv7 potassium channels.

    Science.gov (United States)

    Haick, Jennifer M; Byron, Kenneth L

    2016-09-01

    Smooth muscle cells provide crucial contractile functions in visceral, vascular, and lung tissues. The contractile state of smooth muscle is largely determined by their electrical excitability, which is in turn influenced by the activity of potassium channels. The activity of potassium channels sustains smooth muscle cell membrane hyperpolarization, reducing cellular excitability and thereby promoting smooth muscle relaxation. Research over the past decade has indicated an important role for Kv7 (KCNQ) voltage-gated potassium channels in the regulation of the excitability of smooth muscle cells. Expression of multiple Kv7 channel subtypes has been demonstrated in smooth muscle cells from viscera (gastrointestinal, bladder, myometrial), from the systemic and pulmonary vasculature, and from the airways of the lung, from multiple species, including humans. A number of clinically used drugs, some of which were developed to target Kv7 channels in other tissues, have been found to exert robust effects on smooth muscle Kv7 channels. Functional studies have indicated that Kv7 channel activators and inhibitors have the ability to relax and contact smooth muscle preparations, respectively, suggesting a wide range of novel applications for the pharmacological tool set. This review summarizes recent findings regarding the physiological functions of Kv7 channels in smooth muscle, and highlights potential therapeutic applications based on pharmacological targeting of smooth muscle Kv7 channels throughout the body.

  1. Role of Smooth Muscle in Intestinal Inflammation

    Directory of Open Access Journals (Sweden)

    Stephen M Collins

    1996-01-01

    Full Text Available The notion that smooth muscle function is altered in inflammation is prompted by clinical observations of altered motility in patients with inflammatory bowel disease (IBD. While altered motility may reflect inflammation-induced changes in intrinsic or extrinsic nerves to the gut, changes in gut hormone release and changes in muscle function, recent studies have provided in vitro evidence of altered muscle contractility in muscle resected from patients with ulcerative colitis or Crohn’s disease. In addition, the observation that smooth muscle cells are more numerous and prominent in the strictured bowel of IBD patients compared with controls suggests that inflammation may alter the growth of intestinal smooth muscle. Thus, inflammation is associated with changes in smooth muscle growth and contractility that, in turn, contribute to important symptoms of IBD including diarrhea (from altered motility and pain (via either altered motility or stricture formation. The involvement of smooth muscle in this context may be as an innocent bystander, where cells and products of the inflammatory process induce alterations in muscle contractility and growth. However, it is likely that intestinal muscle cells play a more active role in the inflammatory process via the elaboration of mediators and trophic factors, including cytokines, and via the production of collagen. The concept of muscle cells as active participants in the intestinal inflammatory process is a new concept that is under intense study. This report summarizes current knowledge as it relates to these two aspects of altered muscle function (growth and contractility in the inflamed intestine, and will focus on mechanisms underlying these changes, based on data obtained from animal models of intestinal inflammation.

  2. Autonomic Modification of Intestinal Smooth Muscle Contractility

    Science.gov (United States)

    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…

  3. The small GTPase Rac1 is required for smooth muscle contraction

    DEFF Research Database (Denmark)

    Rahman, Awahan; Davis, Benjamin; Lövdahl, Cecilia

    2014-01-01

    The role of the small GTP-binding protein Rac1 in smooth muscle contraction was examined using small molecule inhibitors (EHT1864, NSC23766) and a novel smooth muscle-specific, conditional, Rac1 knockout mouse strain. EHT1864, which affects nucleotide binding and inhibits Rac1 activity...... at lowered intracellular [Ca2+]. These results show that Rac1 activity is required for active contraction in smooth muscle, probably via enabling an adequate Ca2+ transient. At the same time, specific agonists recruit Rac1 signalling via upstream modulators, resulting in either a potentiation of contraction......, aorta) smooth muscle tissues. This contractile inhibition was associated with inhibition of the Ca2+ transient. Knockout of Rac1 (with a 50% loss of Rac1 protein) lowered active stress in the urinary bladder and the saphenous artery consistent with a role of Rac1 in facilitating smooth muscle...

  4. 生理范围内静水压力对体外培养的人膀胱平滑肌细胞的影响%Effects of Hydrostatic Pressure in Physiological Range on Bladder Smooth Muscle Cells in Vitro

    Institute of Scientific and Technical Information of China (English)

    韦堂墙; 陈林; 王彦; 许锋; 王坤杰

    2012-01-01

    为探索生理范围内的静水压对体外培养的人膀胱平滑肌细胞(HBSMCs)的影响,实验采用自行设计的静水压加压装置并按压力梯度分组,用HE染色观察细胞形态,用免疫荧光、流式细胞仪、逆转录聚合酶链式反应(RT-PCR)分别检测细胞骨架F-actin、细胞周期、增殖细胞核抗原(PCNA)与基质金属蛋白酶7(MMP-7).发现各组的细胞形态、骨架、细胞周期和增殖无明显差异,而各组间MMP-7 mRNA的表达呈现出随静水压力的升高而增加随后逐渐下降的趋势.因此,我们得出生理范围内持续静水压可能对体外培养的HBSMCs的形态、骨架、周期、增殖无明显影响,而对MMP-7 mRNA的表达具有一定的影响.%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 exper-iments,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 metalloprotein-ase 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 sig-nificant influence on proliferation,morphology,skeleton,and cell cycle of HBSMCs; but it may have great effect on the expression of MMP-7.

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

    Science.gov (United States)

    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.

  6. Mechanical properties of mammalian single smooth muscle cells. I. A low cost large range microforce transducer.

    NARCIS (Netherlands)

    J.J. Glerum (Jacobus); R. van Mastrigt (Ron)

    1990-01-01

    textabstractA transducer has been developed for measuring the minute forces generated during isometric contractions (1.0-10.0 microN) of single smooth muscle cells from the pig urinary bladder and the human uterus. In addition to its high sensitivity, resolution and stability (100 mV microN-1, and l

  7. Autophagic regulation of smooth muscle cell biology

    Science.gov (United States)

    Salabei, Joshua K.; Hill, Bradford G.

    2014-01-01

    Autophagy regulates the metabolism, survival, and function of numerous cell types, including those comprising the cardiovascular system. In the vasculature, changes in autophagy have been documented in atherosclerotic and restenotic lesions and in hypertensive vessels. The biology of vascular smooth muscle cells appears particularly sensitive to changes in the autophagic program. Recent evidence indicates that stimuli or stressors evoked during the course of vascular disease can regulate autophagic activity, resulting in modulation of VSMC phenotype and viability. In particular, certain growth factors and cytokines, oxygen tension, and pharmacological drugs have been shown to trigger autophagy in smooth muscle cells. Importantly, each of these stimuli has a redox component, typically associated with changes in the abundance of reactive oxygen, nitrogen, or lipid species. Collective findings support the hypothesis that autophagy plays a critical role in vascular remodeling by regulating smooth muscle cell phenotype transitions and by influencing the cellular response to stress. In this graphical review, we summarize current knowledge on the role of autophagy in the biology of the smooth muscle cell in (patho)physiology. PMID:25544597

  8. Autophagic regulation of smooth muscle cell biology

    Directory of Open Access Journals (Sweden)

    Joshua K. Salabei

    2015-04-01

    Full Text Available Autophagy regulates the metabolism, survival, and function of numerous cell types, including those comprising the cardiovascular system. In the vasculature, changes in autophagy have been documented in atherosclerotic and restenotic lesions and in hypertensive vessels. The biology of vascular smooth muscle cells appears particularly sensitive to changes in the autophagic program. Recent evidence indicates that stimuli or stressors evoked during the course of vascular disease can regulate autophagic activity, resulting in modulation of VSMC phenotype and viability. In particular, certain growth factors and cytokines, oxygen tension, and pharmacological drugs have been shown to trigger autophagy in smooth muscle cells. Importantly, each of these stimuli has a redox component, typically associated with changes in the abundance of reactive oxygen, nitrogen, or lipid species. Collective findings support the hypothesis that autophagy plays a critical role in vascular remodeling by regulating smooth muscle cell phenotype transitions and by influencing the cellular response to stress. In this graphical review, we summarize current knowledge on the role of autophagy in the biology of the smooth muscle cell in (pathophysiology.

  9. On the thermodynamics of smooth muscle contraction

    Science.gov (United States)

    Stålhand, Jonas; McMeeking, Robert M.; Holzapfel, Gerhard A.

    2016-09-01

    Cell function is based on many dynamically complex networks of interacting biochemical reactions. Enzymes may increase the rate of only those reactions that are thermodynamically consistent. In this paper we specifically treat the contraction of smooth muscle cells from the continuum thermodynamics point of view by considering them as an open system where matter passes through the cell membrane. We systematically set up a well-known four-state kinetic model for the cross-bridge interaction of actin and myosin in smooth muscle, where the transition between each state is driven by forward and reverse reactions. Chemical, mechanical and energy balance laws are provided in local forms, while energy balance is also formulated in the more convenient temperature form. We derive the local (non-negative) production of entropy from which we deduce the reduced entropy inequality and the constitutive equations for the first Piola-Kirchhoff stress tensor, the heat flux, the ion and molecular flux and the entropy. One example for smooth muscle contraction is analyzed in more detail in order to provide orientation within the established general thermodynamic framework. In particular the stress evolution, heat generation, muscle shorting rate and a condition for muscle cooling are derived.

  10. The Smooth Muscle of the Artery

    Science.gov (United States)

    1975-01-01

    ruling out the oossibility that depolarization is a junction potential due to rnovement. The low resting potential shown is indicative of the degree of...which is embryologically Soditm Pump in the Con- and functionally related to vascular trol ot %tscle Contrac- smooth muscle, many of the electrical...consideration a 5-hydroxytryptamine as well as histamine as being the factor that we are studying. This does not rule out the posni- bility that either

  11. Notch Signaling in Vascular Smooth Muscle Cells.

    Science.gov (United States)

    Baeten, J T; Lilly, B

    2017-01-01

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

  12. Length adaptation of smooth muscle contractile filaments in response to sustained activation.

    Science.gov (United States)

    Stålhand, Jonas; Holzapfel, Gerhard A

    2016-05-21

    Airway and bladder smooth muscles are known to undergo length adaptation under sustained contraction. This adaptation process entails a remodelling of the intracellular actin and myosin filaments which shifts the peak of the active force-length curve towards the current length. Smooth muscles are therefore able to generate the maximum force over a wide range of lengths. In contrast, length adaptation of vascular smooth muscle has attracted very little attention and only a handful of studies have been reported. Although their results are conflicting on the existence of a length adaptation process in vascular smooth muscle, it seems that, at least, peripheral arteries and arterioles undergo such adaptation. This is of interest since peripheral vessels are responsible for pressure regulation, and a length adaptation will affect the function of the cardiovascular system. It has, e.g., been suggested that the inward remodelling of resistance vessels associated with hypertension disorders may be related to smooth muscle adaptation. In this study we develop a continuum mechanical model for vascular smooth muscle length adaptation by assuming that the muscle cells remodel the actomyosin network such that the peak of the active stress-stretch curve is shifted towards the operating point. The model is specialised to hamster cheek pouch arterioles and the simulated response to stepwise length changes under contraction. The results show that the model is able to recover the salient features of length adaptation reported in the literature.

  13. Airway smooth muscle growth in asthma: proliferation, hypertrophy, and migration.

    Science.gov (United States)

    Bentley, J Kelley; Hershenson, Marc B

    2008-01-01

    Increased airway smooth muscle mass is present in fatal and non-fatal asthma. However, little information is available regarding the cellular mechanism (i.e., hyperplasia vs. hypertrophy). Even less information exists regarding the functional consequences of airway smooth muscle remodeling. It would appear that increased airway smooth muscle mass would tend to increase airway narrowing and airflow obstruction. However, the precise effects of increased airway smooth muscle mass on airway narrowing are not known. This review will consider the evidence for airway smooth muscle cell proliferation and hypertrophy in asthma, potential functional effects, and biochemical mechanisms.

  14. Deletion of Dicer in smooth muscle affects voiding pattern and reduces detrusor contractility and neuroeffector transmission.

    Directory of Open Access Journals (Sweden)

    Mardjaneh Karbalaei Sadegh

    Full Text Available MicroRNAs have emerged as important regulators of smooth muscle phenotype and may play important roles in pathogenesis of various smooth muscle related disease states. The aim of this study was to investigate the role of miRNAs for urinary bladder function. We used an inducible and smooth muscle specific Dicer knockout (KO mouse which resulted in significantly reduced levels of miRNAs, including miR-145, miR-143, miR-22, miR125b-5p and miR-27a, from detrusor preparations without mucosa. Deletion of Dicer resulted in a disturbed micturition pattern in vivo and reduced depolarization-induced pressure development in the isolated detrusor. Furthermore, electrical field stimulation revealed a decreased cholinergic but maintained purinergic component of neurogenic activation in Dicer KO bladder strips. The ultrastructure of detrusor smooth muscle cells was well maintained, and the density of nerve terminals was similar. Western blotting demonstrated reduced contents of calponin and desmin. Smooth muscle α-actin, SM22α and myocardin were unchanged. Activation of strips with exogenous agonists showed that depolarization-induced contraction was preferentially reduced; ATP- and calyculin A-induced contractions were unchanged. Quantitative real time PCR and western blotting demonstrated reduced expression of Cav1.2 (Cacna1c. It is concluded that smooth muscle miRNAs play an important role for detrusor contractility and voiding pattern of unrestrained mice. This is mediated in part via effects on expression of smooth muscle differentiation markers and L-type Ca(2+ channels in the detrusor.

  15. Pharmacological characterisation of the smooth muscle antispasmodic agent tiropramide.

    Science.gov (United States)

    Setnikar, I; Cereda, R; Pacini, M A; Revel, L; Makovec, F

    1989-09-01

    (+/-) Tiropramide hydrochloride, its D and L optical isomers and some of its metabolites were characterized in a number of in vitro pharmacological tests. Tiropramide showed broad spectrum antispasmodic activities on the isolated stomach of guinea pig electrically stimulated; on the longitudinal muscles of the ileum of guinea pig stimulated by electrical impulses, BaCl2, acetylcholine, histamine, serotonin, substance P and cholecystokinin octapeptide (CCK-8); on the spontaneous contractions and on the electrical inhibition of the jejunum of rabbit; on the spontaneous contractions and on the contractions provoked by BaCl2 and acetylcholine of the ascending colon of the rat; on the contractions provoked by BaCl2, acetylcholine, histamine and cerulein of the circular muscles of the gall bladder of the guinea pig; on the spontaneous contractions of the pyel-ureter preparation of the guinea pig; on the contractions of the uterus of the rat provoked by oxitocin, serotonin, acetylcholine, PGF2; on the spontaneous contraction of the portal vein of the rat; on the constriction of the tail artery of the rat provoked by electrical stimulation, epinephrine and ergotamine; on the contractions of the aortic strip of the rabbit stimulated by norepinephrine; on the contractions of the strip of bovine coronary artery depolarized by HCl. In general tiropramide had antispasmodic effect at 5-60 mumol/l concentration. It was more potent than papaverine on contractions provoked by electrical or chemical stimuli, and was less potent or ineffective on spontaneous and "physiological" contractions of the different smooth muscle preparations. Tiropramide had small effects on vascular smooth muscles and showed very small calcium channel blocking activity.

  16. De Novo ACTA2 Mutation Causes a Novel Syndrome of Multisystemic Smooth Muscle Dysfunction

    Science.gov (United States)

    Milewicz, Dianna M.; Østergaard, John R.; Ala-Kokko, Leena M.; Khan, Nadia; Grange, Dorothy K.; Mendoza-Londono, Roberto; Bradley, Timothy J.; Olney, Ann Haskins; Adès, Lesley; Maher, Joseph F.; Guo, Dongchuan; Buja, L. Maximilian; Kim, Dong; Hyland, James C.; Regalado, Ellen S.

    2011-01-01

    Smooth muscle cells (SMCs) contract to perform many physiological functions, including regulation of blood flow and pressure in arteries, contraction of the pupils, peristalsis of the gut and voiding of the bladder. SMC lineage in these organs is characterized by cellular expression of the SMC isoform of α-actin, encoded by the ACTA2 gene. We report here on a unique and de novo mutation in ACTA2, R179H, that causes a syndrome characterized by dysfunction of SMCs throughout the body, leading to aortic and cerebrovascular disease, fixed dilated pupils, hypotonic bladder, malrotation and hypoperistalsis of the gut and pulmonary hypertension. PMID:20734336

  17. Insulin induces a hypercontractile airway smooth muscle phenotype

    NARCIS (Netherlands)

    Gosens, R; Nelemans, SA; Bromhaar, MMG; Meurs, H; Zaagsma, J

    2003-01-01

    This study aims to investigate the effects of insulin on bovine tracheal smooth muscle phenotype in vitro. Contractility of muscle strips and DNA-synthesis ([3 H]thymidine incorporation) of isolated cells were used as parameters for smooth muscle phenotyping. Insulin (1 muM) was mitogenic for bovine

  18. Results of radiotherapy on ureteric obstruction in muscle-invasive bladder cancer

    DEFF Research Database (Denmark)

    Honnens De Lichtenberg, Mette; Miskowiak, J; Rolff, H

    1995-01-01

    To evaluate the effect of radiotherapy on ureteric obstruction due to muscle-invasive bladder cancer.......To evaluate the effect of radiotherapy on ureteric obstruction due to muscle-invasive bladder cancer....

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

    Directory of Open Access Journals (Sweden)

    Jorge L. Gutierrez-Pajares

    2015-06-01

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

  20. Changes of smooth muscle contractile filaments in small bowel atresia

    Institute of Scientific and Technical Information of China (English)

    Stefan Gfroerer; Henning Fiegel; Priya Ramachandran; Udo Rolle; Roman Metzger

    2012-01-01

    AIM:To investigate morphological changes of intestinal smooth muscle contractile fibres in small bowel atresia patients.METHODS:Resected small bowel specimens from small bowel atresia patients (n =12) were divided into three sections (proximal,atretic and distal).Standard histology hematoxylin-eosin staining and enzyme immunohistochemistry was performed to visualize smooth muscle contractile markers α-smooth muscle actin (SMA) and desmin using conventional paraffin sections of the proximal and distal bowel.Small bowel from agematched patients (n =2) undergoing Meckel's diverticulum resection served as controls.RESULTS:The smooth muscle coat in the proximal bowel of small bowel atresia patients was thickened compared with control tissue,but the distal bowel was unchanged.Expression of smooth muscle contractile fibres SMA and desmin within the proximal bowel was slightly reduced compared with the distal bowel and control tissue.There were no major differences in the architecture of the smooth muscle within the proximal bowel and the distal bowel.The proximal and distal bowel in small bowel atresia patients revealed only minimal differences regarding smooth muscle morphology and the presence of smooth muscle contractile filament markers.CONCLUSION:Changes in smooth muscle contractile filaments do not appear to play a major role in postoperative motility disorders in small bowel atresia.

  1. Neoadjuvant Chemotherapy in Muscle-Invasive Bladder Cancer

    Directory of Open Access Journals (Sweden)

    Kari T. Syvänen

    2014-05-01

    Full Text Available Neoadjuvant chemotherapy (NAC in muscle-invasive bladder cancer was introduced several years ago. Despite the evidence supporting its use in clinical practice, only a minority of patients who undergo radical cystectomy receive preoperative chemotherapy. In addition, recommendations and methods to detect patients who would benefit the most from NAC are still unclear. The European Association of Urology (EAU guidelines panel on muscle-invasive and metastatic bladder cancer recommends the use of cisplatin-based NAC for T2-T4a, cN0 M0 bladder cancer if the patient has a performance status ≥2 and if the renal function is not impaired, but the American Urological Association, for example, does not have any guideline recommendations on this topic at all. In this review we describe the current literature supporting NAC in association with radical cystectomy in muscle-invasive urothelial carcinoma of the bladder. Evidence acquisition was made searching the Medline database for original articles published before 1st February 2014, with search terms: “neoadjuvant chemotherapy”, “radical cystectomy”, and “invasive bladder cancer”.

  2. Smooth muscle cells largely develop independently of functional hemogenic endothelium

    Directory of Open Access Journals (Sweden)

    Monika Stefanska

    2014-01-01

    Full Text Available Vascular smooth muscle cells represent a major component of the cardiovascular system. In vitro studies have shown that FLK1+ cells derived from embryonic stem (ES cells can differentiate into both endothelial and smooth muscle cells. These FLK1+ cells also contain a mesodermal precursor, the hemangioblast, able to produce endothelial, blood and smooth muscle cells. The generation of blood precursors from the hemangioblast was recently shown to occur through a transient cell population of specialised endothelium, a hemogenic endothelium. To date, the lineage relationship between this cell population and smooth muscle cell progenitors has not been investigated. In this study, we generated a reporter ES cell line in which expression of the fluorescent protein H2B-VENUS is driven by the α-smooth muscle actin (α-SMA regulatory sequences. We demonstrated that this reporter cell line efficiently trace smooth muscle development during ES cell differentiation. Although some smooth muscle cells are associated with broad endothelial development, we established that smooth muscle cells are mostly generated independently from a specialised functional hemogenic endothelium. This study provides new and important insights into hematopoietic and vascular development, which may help in driving further progress towards the development of bioengineered vascular grafts for regenerative medicine.

  3. Treating asthma means treating airway smooth muscle cells

    NARCIS (Netherlands)

    Zuyderduyn, S; Sukkar, M B; Fust, A; Dhaliwal, S; Burgess, J K

    2008-01-01

    Asthma is characterised by airway hyperresponsiveness, airway inflammation and airway remodelling. Airway smooth muscle cells are known to be the main effector cells of airway narrowing. In the present paper, studies will be discussed that have led to a novel view of the role of airway smooth muscle

  4. Evidence of direct smooth muscle relaxant effects of the fibrate gemfibrozil.

    Science.gov (United States)

    Phelps, Laura E; Peuler, Jacob D

    2010-01-01

    Fibrates are commonly employed to treat abnormal lipid metabolism via their unique ability to stimulate peroxisome proliferator-activated receptor alpha (PPARalpha). Interestingly, they also decrease systemic arterial pressure, despite recent evidence that PPAR alpha may contribute to expression of renin and related hypertension. Yet, mechanisms responsible for their potential antihypertensive activity remain unresolved. Rapid decreases in arterial pressure following bolus intravenous injections of bezafibrate strongly suggest they may relax arterial smooth muscle directly. But since bezafibrate is highly susceptible to photodegradation in aqueous media, it has never been critically tested for this possibility in vitro with isolated arterial smooth muscle preparations. Accordingly, we tested gemfibrozil which is resistant to photodegradation. We examined it over a therapeutically-relevant range (50-400 microM) for both acute and delayed relaxant effects on contractions of the isolated rat tail artery; contractions induced by either depolarizing its smooth muscle cell membranes with high potassium or stimulating its membrane-bound receptors with norepinephrine and arginine-vasopressin. We also examined these same gemfibrozil levels for effects on spontaneously-occurring phasic rhythmic contractile activity, typically not seen in arteries under in vitro conditions but commonly exhibited by smooth muscle of uterus, duodenum and bladder. We found that gemfibrozil significantly relaxed all induced forms of contraction in the rat tail artery, acutely at the higher test levels and after a delay of a few hours at the lower test levels. The highest test level of gemfibrozil (400 microM) also completely abolished spontaneously-occurring contractile activity of the isolated uterus and duodenum and markedly suppressed it in the bladder. This is the first evidence that a fibrate drug can directly relax smooth muscle contractions, either induced by various contractile agents or

  5. Mechanotransduction in colonic smooth muscle cells.

    Science.gov (United States)

    Young, S H; Ennes, H S; Mayer, E A

    1997-11-15

    We evaluated mechanisms which mediate alterations in intracellular biochemical events in response to transient mechanical stimulation of colonic smooth muscle cells. Cultured myocytes from the circular muscle layer of the rabbit distal colon responded to brief focal mechanical deformation of the plasma membrane with a transient increase in intracellular calcium concentration ([Ca2+]i) with peak of 422.7 +/- 43.8 nm above an average resting [Ca2+]i of 104.8 +/- 10.9 nM (n = 57) followed by both rapid and prolonged recovery phases. The peak [Ca2+]i increase was reduced by 50% in the absence of extracellular Ca2+, while the prolonged [Ca2+]i recovery was either abolished or reduced to less than or = 15% of control values. In contrast, no significant effect of gadolinium chloride (100 microM) or lanthanum chloride (25 microM) on either peak transient or prolonged [Ca2+]i recovery was observed. Pretreatment of cells with thapsigargin (1 microM) resulted in a 25% reduction of the mechanically induced peak [Ca2+]i response, while the phospholipase C inhibitor U-73122 had no effect on the [Ca2+]i transient peak. [Ca2+]i transients were abolished when cells previously treated with thapsigargin were mechanically stimulated in Ca2+-free solution, or when Ca2+ stores were depleted by thapsigargin in Ca2+-free solution. Pretreatment with the microfilament disrupting drug cytochalasin D (10 microM) or microinjection of myocytes with an intracellular saline resulted in complete inhibition of the transient. The effect of cytochalasin D was reversible and did not prevent the [Ca2+]i increases in response to thapsigargin. These results suggest a communication, which may be mediated by direct mechanical link via actin filaments, between the plasma membrane and an internal Ca2+ store.

  6. Endoscopic management of gastrointestinal smooth muscle tumor

    Institute of Scientific and Technical Information of China (English)

    Xiao-Dong Zhou; Nong-Hua Lv; Hong-Xia Chen; Chong-Wen Wang; Xuan Zhu; Ping Xu; You-Xiang Chen

    2007-01-01

    AIM: To systematically evaluate the efficacy and safety of endoscopic resection of gastrointestinal smooth muscle tumors (SMTs, including leiomyoma and leiomyosarcoma) and to review our preliminary experiences on endoscopic diagnosis of gastrointestinal SMTs.METHODS: A total of 69 patients with gastrointestinal SMT underwent routine endoscopy in our department.Endoscopic ultrasonography (EUS) was also performed in 9 cases of gastrointestinal SMT. The sessile submucosal gastrointestinal SMTs with the base smaller than 2 cm in diameter were resected by "pushing" technique or "grasping and pushing" technique while the pedunculated SMTs were resected by polypectomy. For those SMTs originating from muscularis propria or with the base size ≥ 2 cm, ordinary biopsy technique was performed in tumors with ulcers while the "Digging" technique was performed in those without ulcers.RESULTS: 54 cases of leiomyoma and 15 cases of leiomyosarcoma were identified. In them, 19 cases of submucosal leiomyoma were resected by "pushing"technique and 10 cases were removed by "grasping and pushing" technique. Three cases pedunculated submucosal leiomyoma were resected by polypectomy.No severe complications developed during or after the procedure. No recurrence was observed. The diagnostic accuracy of ordinary and the "Digging" biopsy technique was 90.0% and 94.1%, respectively.CONCLUSION: Endoscopic resection is a safe and effective treatment for leiomyomas with the base size ≤2 cm. The "digging" biopsy technique would be a good option for histologic diagnosis of SMTs.

  7. Smooth muscle phenotypic modulation--a personal experience.

    Science.gov (United States)

    Campbell, Julie H; Campbell, Gordon R

    2012-08-01

    The idea that smooth muscle cells can exist in multiple phenotypic states depending on the functional demands placed upon them has been around for >5 decades. However, much of the literature today refers to only recent articles, giving the impression that it is a new idea. At the same time, the current trend is to delve deeper and deeper into transcriptional regulation of smooth muscle genes, and much of the work describing the change in biology of the cells in the different phenotypic states does not appear to be known. This loss of historical perspective regarding the biology of smooth muscle phenotypic modulation is what the current article has tried to mitigate.

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

    Energy Technology Data Exchange (ETDEWEB)

    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.

  9. Treatment of Muscle-Invasive Bladder Cancer in Older Patients.

    Science.gov (United States)

    Skinner, Eila C

    2016-01-01

    Treatment of muscle-invasive bladder cancer in older patients is challenging. Definitive therapy of localized disease requires either surgery or radiation therapy, ideally combined with systemic chemotherapy. However, current population data suggest that less than half of patients older than age 70 are offered such treatments. We will review tools available to assess the fitness of older patients for surgery, alternatives, and tips for perioperative patient treatment.

  10. Proteomics research on muscle-invasive bladder transitional cell carcinoma

    Directory of Open Access Journals (Sweden)

    Cao Yan

    2011-06-01

    Full Text Available Abstract Background Aimed to facilitate candidate biomarkers selection and improve network-based multi-target therapy, we perform comparative proteomics research on muscle-invasive bladder transitional cell carcinoma. Laser capture microdissection was used to harvest purified muscle-invasive bladder cancer cells and normal urothelial cells from 4 paired samples. Two-dimensional liquid chromatography tandem mass spectrometry was used to identify the proteome expression profile. The differential proteins were further analyzed using bioinformatics tools and compared with the published literature. Results A total of 885/890 proteins commonly appeared in 4 paired samples. 295/337 of the 488/493 proteins that specific expressed in tumor/normal cells own gene ontology (GO cellular component annotation. Compared with the entire list of the international protein index (IPI, there are 42/45 GO terms exhibited as enriched and 9/5 exhibited as depleted, respectively. Several pathways exhibit significantly changes between cancer and normal cells, mainly including spliceosome, endocytosis, oxidative phosphorylation, etc. Finally, descriptive statistics show that the PI Distribution of candidate biomarkers have certain regularity. Conclusions The present study identified the proteome expression profile of muscle-invasive bladder cancer cells and normal urothelial cells, providing information for subcellular pattern research of cancer and offer candidate proteins for biomarker panel and network-based multi-target therapy.

  11. Anisotropic Smoothing Improves DT-MRI-Based Muscle Fiber Tractography.

    Directory of Open Access Journals (Sweden)

    Amanda K W Buck

    Full Text Available To assess the effect of anisotropic smoothing on fiber tracking measures, including pennation angle, fiber tract length, and fiber tract number in the medial gastrocnemius (MG muscle in healthy subjects using diffusion-weighted magnetic resonance imaging (DW-MRI.3T DW-MRI data were used for muscle fiber tractography in the MG of healthy subjects. Anisotropic smoothing was applied at three levels (5%, 10%, 15%, and pennation angle, tract length, fiber tract number, fractional anisotropy, and principal eigenvector orientation were quantified for each smoothing level.Fiber tract length increased with pre-fiber tracking smoothing, and local heterogeneities in fiber direction were reduced. However, pennation angle was not affected by smoothing.Modest anisotropic smoothing (10% improved fiber-tracking results, while preserving structural features.

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

    Directory of Open Access Journals (Sweden)

    Bart Spronck

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

  13. Smooth Muscle Precursor Cells Derived from Human Pluripotent Stem Cells for Treatment of Stress Urinary Incontinence

    Science.gov (United States)

    Wang, Zhe; Li, Yan Hui; Wei, Yi; Green, Morgaine; Wani, Prachi; Zhang, Pengbo; Pera, Renee Reijo; Chen, Bertha

    2016-01-01

    There is great interest in using stem cells (SC) to regenerate a deficient urethral sphincter in patients with urinary incontinence. The smooth muscle component of the sphincter is a significant contributor to sphincter function. However, current translational efforts for sphincter muscle restoration focus only on skeletal muscle regeneration because they rely on adult mesenchymal SC as cell source. These adult SC do not yield sufficient smooth muscle cells (SMCs) for transplantation. We may be able to overcome this limitation by using pluripotent stem cell (PSC) to derive SMCs. Hence, we sought to investigate whether smooth muscle precursor cells (pSMCs) derived from human PSCs can restore urethral function in an animal model generated by surgical urethrolysis and ovariectomy. Rats were divided into four groups: control (no intervention), sham saline (surgery + saline injection), bladder SMC (surgery + human bladder SMC injection), and treatment (surgery + pSMC injection, which includes human embryonic stem cell (hESC) H9-derived pSMC, episomal reprogrammed induced pluripotent stem cells (iPSCs)-derived pSMC, or viral reprogrammed iPSC-derived pSMC). pSMCs (2 × 106 cells/rat) were injected periurethrally 3 weeks postsurgery. Leak point pressure (LPP) and baseline external urethral sphincter electromyography were measured 5 weeks postinjection. Both iPSC-derived pSMC treatment groups showed significantly higher LPP compared to the sham saline group, consistent with restoration of urethral sphincter function. While the difference between the H9-derived pSMC treatment and sham saline group was not significant, it did show a trend toward restoration of the LPP to the level of intact controls. Our data indicate that pSMCs derived from human PSCs (hESC and iPSC) can restore sphincter function. PMID:26785911

  14. Airway smooth muscle cell proliferation is increased in asthma

    NARCIS (Netherlands)

    Johnson, P R; Roth, Michael; Tamm, M; Hughes, J Margaret; Ge, Q; King, G; Burgess, J K; Black, J L

    2001-01-01

    UNLABELLED: Increased airway smooth muscle (ASM) within the bronchial wall of asthmatic patients has been well documented and is likely to be the result of increased muscle proliferation. We have for the first time been able to culture ASM cells from asthmatic patients and to compare their prolifera

  15. UROTENSIN II RECEPTOR IN THE RAT AIRWAY SMOOTH MUSCLE AND ITS EFFECT ON THE RAT AIRWAY SMOOTH MUSCLE CELLS PROLIFERATION

    Institute of Scientific and Technical Information of China (English)

    陈亚红; 赵鸣武; 刘秀华; 姚婉贞; 杨军; 张肇康; 唐朝枢

    2001-01-01

    Objective. To investigate the characteristics of urotensin II (U-II) receptor in the rat airway smooth muscleand the effect and signal transduction pathway of U-II on the proliferation of airway smooth muscle cells.Methods. Using 125-UII binding assay to measure the Bmax and Kd of U-II receptor. Using the 3H-TdRincorporation to deter mine the effect of U-II on the proliferation of airway smooth muscle cells and its signal transduc-tion pathway. Using Fura-2/AM to measure the effect of U-II on the cytosolic free calcium concentration.Results. 1. 125I-UⅡ binding increased with the time and reached saturation at 45min. The Bmax was(ll. 36 +0.37)fmol/mg pr and Kd was (4.46 +0.61)nmol/L. 2. U-II increased 3H-TdR incorporation of theairway smooth muscle cells in a dose-dependent manner. 3. H7, PDg8059 and nicardipine, inhibitors of PKC,MAPK, calcium cha.nnel, respectively, significantly inhibited U-II-stimulated 3H-TdR incorporation of airwaysmooth muscle cells. W7, inhibitor of CaM-PK, had no effect. 4. Cyclosporin A, inhibitor of CaN, inhibited3H-TdRincorporation ofthe airway smooth muscle cells induced by U-Ⅱl in a dose-dependent manner. 5. U-Ⅱlpromot-ed cy-tosolic free calcium concentration increase by 18%.Conclusions. 1. There was U-II receptor in the rat airway smooth muscle. 2. The effect of U-II-stimulated-3H-TdR incorporation of airway smooth muscle cells was mediated by such signal transduction pathway as Ca2 +.PKC, MAPK and Ca.N, etc.``

  16. Loss of LMOD1 impairs smooth muscle cytocontractility and causes megacystis microcolon intestinal hypoperistalsis syndrome in humans and mice

    Science.gov (United States)

    Halim, Danny; Wilson, Michael P.; Oliver, Daniel; Brosens, Erwin; Verheij, Joke B. G. M.; Han, Yu; Nanda, Vivek; Lyu, Qing; Doukas, Michael; Stoop, Hans; Brouwer, Rutger W. W.; van IJcken, Wilfred F. J.; Slivano, Orazio J.; Burns, Alan J.; Christie, Christine K.; de Mesy Bentley, Karen L.; Brooks, Alice S.; Tibboel, Dick; Xu, Suowen; Jin, Zheng Gen; Djuwantono, Tono; Yan, Wei; Alves, Maria M.; Hofstra, Robert M. W.; Miano, Joseph M.

    2017-01-01

    Megacystis microcolon intestinal hypoperistalsis syndrome (MMIHS) is a congenital visceral myopathy characterized by severe dilation of the urinary bladder and defective intestinal motility. The genetic basis of MMIHS has been ascribed to spontaneous and autosomal dominant mutations in actin gamma 2 (ACTG2), a smooth muscle contractile gene. However, evidence suggesting a recessive origin of the disease also exists. Using combined homozygosity mapping and whole exome sequencing, a genetically isolated family was found to carry a premature termination codon in Leiomodin1 (LMOD1), a gene preferentially expressed in vascular and visceral smooth muscle cells. Parents heterozygous for the mutation exhibited no abnormalities, but a child homozygous for the premature termination codon displayed symptoms consistent with MMIHS. We used CRISPR-Cas9 (CRISPR-associated protein) genome editing of Lmod1 to generate a similar premature termination codon. Mice homozygous for the mutation showed loss of LMOD1 protein and pathology consistent with MMIHS, including late gestation expansion of the bladder, hydronephrosis, and rapid demise after parturition. Loss of LMOD1 resulted in a reduction of filamentous actin, elongated cytoskeletal dense bodies, and impaired intestinal smooth muscle contractility. These results define LMOD1 as a disease gene for MMIHS and suggest its role in establishing normal smooth muscle cytoskeletal–contractile coupling. PMID:28292896

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

    OpenAIRE

    Qingqing Wang; Ning Zhao; Simone Kennard; Brenda Lilly

    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 N...

  18. Cellular mechanisms of myogenic activity in gastric smooth muscle.

    Science.gov (United States)

    Suzuki, H

    2000-06-01

    In many regions of the intestine, a thin layer of interstitial cells of Cajal (ICC) lie in the myenteric region, between the circular and longitudinal muscle layers. ICC are connected by gap junctions to surrounding ICC and also with circular and longitudinal smooth muscle cells, forming a large electrical syncytium. Damage of the ICC causes a disorder in the patterns of rhythmic activity. Isolated ICC produce a rhythmic oscillation of the membrane potential. All these observations have led to the suggestion that ICC may be the pacemaker cell responsible for intestinal activity. Gastric smooth muscles generate slow oscillatory membrane potential changes (slow waves) and spike potentials. The activity is considered to be linked to the metabolism in the cell. Three types of cells located in the gastric wall (circular and longitudinal smooth muscle cells and ICC) produce synchronized electrical responses with different shapes. The electrical responses appear to originate in ICC and then spread to the smooth muscle layers, indicating that ICC may also be the pacemaker cells responsible for gastric activity. However, isolated circular smooth muscle tissues spontaneously generate regenerative potentials, suggesting that there are at least two sites for the initiation of spontaneous activity in the stomach. Regenerative potentials persist in the presence of Ca-antagonists and are inhibited by agents which disrupt intracellular Ca(2+) homeostasis. Depolarization of the membrane elicits regenerative potentials after a long delay and the potentials have long refractory periods. This suggests that an unidentified 2nd messenger may be formed during the delay between membrane depolarization and the initiation of a regenerative potential. In gastric muscles of mutant mice which do not express inositol trisphosphate (InsP(3)) receptors, spike potentials but not slow waves are generated, suggesting the possible involvement of InsP(3) in the initiation of spontaneous activity.

  19. Identification of the smooth muscle excitatory receptors for ergot alkaloids.

    Science.gov (United States)

    Innes, I R

    1962-08-01

    In cats under sodium pentobarbitone anaesthesia the first dose of ergotamine (50 mug/kg) invariably caused retraction of the nictitating membrane and a rise of arterial blood pressure. However, the responses to the dose of ergotamine were strikingly reduced when the cats were previously treated with the adrenaline antagonists phenoxybenzamine (5 mg/kg) or ergotamine (50 mug/kg). Further experiments to identify receptors for ergotamine were carried out on three different isolated smooth muscle preparations: rabbit aorta, rat uterus and dog retractor penis.Receptors for adrenaline were selectively protected by high concentrations of adrenaline throughout exposure of the preparation to a blocking concentration of ergotamine or phenoxybenzamine. Protected muscles responded to ergotamine; unprotected muscles did not. Muscles where receptors for acetylcholine, histamine or 5-hydroxytryptamine were protected by high concentrations of these drugs did not respond to ergotamine. Ergometrine, which has no blocking action on adrenaline receptors, behaved in the same way as ergotamine; muscles which were protected by adrenaline against blockade by phenoxybenzamine responded to ergometrine, but unprotected muscles did not. The stimulant actions of adrenaline, ergotamine and ergometrine were also protected against the blocking action of phenoxybenzamine by treating the muscle with a high concentration of ergometrine instead of adrenaline. It is concluded that, in smooth muscle which can be excited by adrenaline, ergotamine and ergometrine act by combining with adrenaline receptors, and that ergotamine may therefore be regarded not only as an adrenaline antagonist but also as a partial agonist since it excites the same receptors.

  20. JunB mediates basal- and TGFβ1-induced smooth muscle cell contractility.

    Directory of Open Access Journals (Sweden)

    Aruna Ramachandran

    Full Text Available Smooth muscle contraction is a dynamic process driven by acto-myosin interactions that are controlled by multiple regulatory proteins. Our studies have shown that members of the AP-1 transcription factor family control discrete behaviors of smooth muscle cells (SMC such as growth, migration and fibrosis. However, the role of AP-1 in regulation of smooth muscle contractility is incompletely understood. In this study we show that the AP-1 family member JunB regulates contractility in visceral SMC by altering actin polymerization and myosin light chain phosphorylation. JunB levels are robustly upregulated downstream of transforming growth factor beta-1 (TGFβ1, a known inducer of SMC contractility. RNAi-mediated silencing of JunB in primary human bladder SMC (pBSMC inhibited cell contractility under both basal and TGFβ1-stimulated conditions, as determined using gel contraction and traction force microscopy assays. JunB knockdown did not alter expression of the contractile proteins α-SMA, calponin or SM22α. However, JunB silencing decreased levels of Rho kinase (ROCK and myosin light chain (MLC20. Moreover, JunB silencing attenuated phosphorylation of the MLC20 regulatory phosphatase subunit MYPT1 and the actin severing protein cofilin. Consistent with these changes, cells in which JunB was knocked down showed a reduction in the F:G actin ratio in response to TGFβ1. Together these findings demonstrate a novel function for JunB in regulating visceral smooth muscle cell contractility through effects on both myosin and the actin cytoskeleton.

  1. Phenotype modulation of airway smooth muscle in asthma

    NARCIS (Netherlands)

    Wright, David B.; Trian, Thomas; Siddiqui, Sana; Pascoe, Chris D.; Johnson, Jill R.; Dekkers, Bart G. J.; Dakshinamurti, Shyamala; Bagchi, Rushita; Burgess, Janette K.; Kanabar, Varsha; Ojo, Oluwaseun O.

    2013-01-01

    The biological responses of airway smooth muscle (ASM) are diverse, in part due to ASM phenotype plasticity. ASM phenotype plasticity refers to the ability of ASM cells to change the degree of a variety of functions, including contractility, proliferation, migration and secretion of inflammatory med

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

    Science.gov (United States)

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

    2004-01-01

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

  3. Congenital smooth muscle hamartoma of the palpebral conjunctiva.

    Science.gov (United States)

    Mora, L Evelyn; Rodríguez-Reyes, Abelardo A; Vera, Ana M; Rubio, Rosa Isela; Mayorquín-Ruiz, Mariana; Salcedo, Guillermo

    2012-01-01

    Smooth muscle hamartoma is defined as a disorganized focus or an overgrowth of mature smooth muscle, generally with low capacity of autonomous growth and benign behavior. The implicated tissues are mature and proliferate in a disorganized fashion. A healthy 5-day-old Mexican boy was referred to the authors' hospital in México city for evaluation of a "cystic" lesion of the right eye that had been noted since birth. The pregnancy and delivery were unremarkable. On physical examination, there was a reddish-pink soft lesion with a tender "cystic" appearance, which was probably emerging from the upper eyelid conjunctiva, which measured 2.7 cm in its widest diameter and transilluminated. Ultrasound imaging revealed an anterior "cystic" lesion with normally formed phakic eye. An excisional biopsy was performed, and the lesion was dissected from the upper tarsal subconjunctival space. Subsequent histologic and immunohistochemical findings were consistent with the diagnosis of congenital smooth muscle hamartoma (CSMH) of the tarsal conjunctiva. The authors' research revealed that only one case of CSMH localized in the conjunctiva (Roper GJ, Smith MS, Lueder GT. Congenital smooth muscle hamartoma of the conjunctival fornix. Am J Ophthalmol. 1999;128:643-4) has been reported to date in the literature. To the best of the authors' knowledge, this current case would be the second case reported of CSMH in this anatomic location. Therefore, the authors' recommendation is to include CSMH in the differential diagnosis of a cystic mass that presents in the fornix and palpebral conjunctiva.

  4. beta-Catenin regulates airway smooth muscle contraction

    NARCIS (Netherlands)

    Jansen, Sepp R.; Van Ziel, Anna M.; Baarsma, Hoeke A.; Gosens, Reinoud

    2010-01-01

    Jansen SR, Van Ziel AM, Baarsma HA, Gosens R. beta-Catenin regulates airway smooth muscle contraction. Am J Physiol Lung Cell Mol Physiol 299: L204-L214, 2010. First published May 14, 2010; doi:10.1152/ajplung.00020.2010.-beta-Catenin is an 88-kDa member of the armadillo family of proteins that is a

  5. Airway smooth muscle and fibroblasts in the pathogenesis of asthma

    NARCIS (Netherlands)

    Johnson, Peter R A; Burgess, Janette K

    2004-01-01

    Asthma is a disease characterized by marked structural changes within the airway wall. These changes include deposition of extracellular matrix proteins and an increase in the numbers of airway smooth muscle cells and subepithelial fibroblasts. Both these cell types possess properties that would ena

  6. Airway smooth muscle - Its relationship to the extracellular matrix

    NARCIS (Netherlands)

    Black, Judith L.; Burgess, Janette K.; Johnson, Peter R.A.

    2003-01-01

    The airway smooth muscle cell has a variety of properties, which confer on it the ability to participate actively in the inflammatory process and the remodeling events, which accompany severe, persistent asthma. Among these properties is its relationship to the extracellular matrix (ECM) with which

  7. Plasticity of cerebrovascular smooth muscle cells after subarachnoid hemorrhage

    DEFF Research Database (Denmark)

    Edvinsson, Lars; Larsen, Stine Schmidt; Maddahi, Aida

    2014-01-01

    , inflammatory reactions, and microthrombosis. Additionally, a large body of evidence indicates that vascular plasticity plays an important role in SAH pathophysiology, and this review aims to summarize our current knowledge on the phenotypic changes of vascular smooth muscle cells of the cerebral vasculature...

  8. Extracellular proteolysis and the migrating vascular smooth muscle cell

    NARCIS (Netherlands)

    Leeuwen, R.T.J. van

    1996-01-01

    Smooth muscle cells (SMC) form the major cell type in the arterial blood vessels. In the undamaged vessel wall they remain in a contractile state characterized by the absence of cell division, a low metabolic activity and a high actin-myosin content. As a reaction to injury of the vessel wall they c

  9. HSP20 phosphorylation and airway smooth muscle relaxation

    Directory of Open Access Journals (Sweden)

    Mariam Ba

    2009-06-01

    Full Text Available Mariam Ba1, Cherie A Singer1, Manoj Tyagi2, Colleen Brophy3, Josh E Baker4, Christine Cremo4, Andrew Halayko5, William T Gerthoffer21Department of Pharmacology, University of Nevada School of Medicine, Reno, NV, USA; 2Department of Biochemistry and Molecular Biology, University of South Alabama, Mobile, AL, USA; 3Harrington Department of Biochemistry, Arizona State University, Tempe, AZ, USA; 4Department of Biochemistry and Molecular Biology, University of Nevada, Reno, NV, USA; 5Departments of Physiology and Internal Medicine, University of Manitoba, Winnipeg, MB, CanadaAbstract: HSP20 (HSPB6 is a small heat shock protein expressed in smooth muscles that is hypothesized to inhibit contraction when phosphorylated by cAMP-dependent protein kinase. To investigate this hypothesis in airway smooth muscle (ASM we showed that HSP20 was constitutively expressed as well as being inducible in cultured hASM cells by treatment with 1 µM isoproterenol or 10 µM salmeterol. In contrast, a mixture of proinflammatory mediators (interleukin-1β, tumor necrosis factor α, and interferon γ inhibited expression of HSP20 by about 50% in 48 hours. To determine whether phosphorylation of HSP20 is sufficient to induce relaxation, canine tracheal smooth muscle was treated with a cell permeant phosphopeptide that mimics the phosphorylation of HSP20. The HSP20 phosphopeptide antagonized carbacholinduced contraction by 60% with no change in myosin light chain phosphorylation. Recombinant full length HSP20 inhibited skeletal actin binding to smooth muscle myosin subfragment 1 (S1, and recombinant cell permeant TAT-HSP20 S16D mutant reduced F-actin filaments in cultured hASM cells. Carbachol stimulation of canine tracheal smooth muscle tissue caused redistribution of HSP20 from large macromolecular complexes (200–500 kDa to smaller complexes (<60 kDa. The results are consistent with HSP20 expression and macromolecular structure being dynamically regulated in airway

  10. Efficacy of transurethral resection of bladder tumor in treatment of elderly muscle-invasive bladder cancer

    Institute of Scientific and Technical Information of China (English)

    Yu Zhou; Min He; Hong-Tao Jia; Yun-Fei Li; Mao-Hua Luo

    2016-01-01

    Objective:To study the clinical efficacy of transurethral resection of bladder tumor (TURBT) in the treatment of elderly muscle-invasive bladder cancer (MIBC), and observe the changes of serum IGF-1, VEGF, BLCA-4, and IL-8 levels before and after treatment.Methods: A total of 56 elderly patients with MIBC who were admitted in our hospital were included in the study and divided into the treatment group (n=30) and the control group (n=26). The patients in the control group were given radical cystectomy, while the patients in the treatment group were given TURBT and bladder irrigation chemotherapy after operation. The surgical complications and survival in the two groups were observed. The serum IGF-1 and VEGF levels, and urine BLCA-4 and IL-8 levels before and after treatment in the two groups were detected.Results:The difference of serum IGF-1 and VEGF levels before operation between the two groups was not statistically significant (P>0.05). The serum IGF-1 and VEGF levels 7 d after operation were significantly reduced when compared with before operation (P<0.01), and the difference between the two groups was statistically significant (P<0.05 orP<0.01). The tumor-free survival rate 2 and 3 years after operation in the observation group were significantly higher than those in the control group (P<0.05).Conclusions: TURBT in the treatment of elderly MIBC has a preferable clinical effect, and can shorten the operation time, with a small trauma and less side effects.

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

    Institute of Scientific and Technical Information of China (English)

    Dale D. TANG

    2008-01-01

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

  12. Novel Simulation Model of Non-Muscle Invasive Bladder Cancer

    DEFF Research Database (Denmark)

    Patel, Sanjay R; Dinh, Tuan; Noah-Vanhoucke, Joyce

    2015-01-01

    Introduction: There have been no randomized controlled trials (RCTs) evaluating the clinical or economic benefit of mitomycin C intravesical therapy vs. radical cystectomy in patients with high-risk non-muscle invasive bladder cancer (NMIBC). We used the Archimedes computational model to simulate...... RCT comparing radical cystectomy versus intravesical mitomycin C (MMC) therapy to evaluate the clinical and economic outcomes for BCG-refractory NMIBC as well demonstrate the utility of computer based models to simulate a clinical trial. Methods: The Archimedes model was developed to generate...... and is more cost-effective when compared to those undergoing MMC. Simulation of clinical trials using computational models similar to the Archimedes model can overcome shortcomings of real-world clinical trials and may prove useful in the face of current medical cost-conscious era....

  13. Menthol inhibiting parasympathetic function of tracheal smooth muscle

    Science.gov (United States)

    Wang, Hsing-Won; Liu, Shao-Cheng; Chao, Pin-Zhir; Lee, Fei-Peng

    2016-01-01

    Menthol is used as a constituent of food and drink, tobacco and cosmetics nowadays. This cold receptor agonist has been used as a nasal inhalation solution in the daily life. The effect of menthol on nasal mucosa in vivo is well known; however, the effect of the drug on tracheal smooth muscle has been rarely explored. Therefore, during administration of the drug for nasal symptoms, it might also affect the trachea via oral intake or inhalation. We used our preparation to test the effectiveness of menthol on isolated rat tracheal smooth muscle. A 5 mm long portion of rat trachea was submersed in 30 ml Krebs solution in a muscle bath at 37ºC. Changes in tracheal contractility in response to the application of a parasympathetic mimetic agent were measured using a transducer connected to a Pentium III computer equipped with polygraph software. The following assessments of menthol 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 the drug on electrically induced tracheal smooth muscle contractions. Results indicated that addition of a parasympathetic mimetic to the incubation medium caused the trachea to contract in a dose-dependent manner. Addition of menthol at doses of 10-5 M or above elicited a relaxation response to 10-6 M methacholine-induced contraction. Menthol could also inhibit electrical field stimulation (EFS) induced spike contraction. However, it alone had a minimal effect on the basal tension of trachea as the concentration increased. We concluded that the degree of drug-induced tracheal contraction or relaxation was dose-dependent. In addition, this study indicated that high concentrations of menthol might actually inhibit parasympathetic function of the trachea. PMID:27994497

  14. Eye features in three Danish patients with multisystemic smooth muscle dysfunction syndrome

    DEFF Research Database (Denmark)

    Moller, Hans Ulrik; Fledelius, Hans C; Milewicz, Dianna M

    2012-01-01

    A de novo mutation of the ACTA2 gene encoding the smooth muscle cell α-actin has been established in patients with multisystemic smooth muscle dysfunction syndrome associated with patent ductus arteriosus and mydriasis present at birth....

  15. Mechanisms mediating cholinergic antral circular smooth muscle contraction in rats

    Institute of Scientific and Technical Information of China (English)

    Helena F Wrzos; Tarun Tandon; Ann Ouyang

    2004-01-01

    AIM: To investigate the pathway (s) mediating rat antral circular smooth muscle contractile responses to the cholinomimetic agent, bethanechol and the subtypes of muscarinic receptors mediating the cholinergic contraction.METHODS: Circular smooth muscle strips from the antrum of Sprague-Dawley rats were mounted in muscle baths in Krebs buffer. Isometric tension was recorded. Cumulative concentration-response curves were obtained for (+)-cisdioxolane (cD), a nonspecific muscarinic agonist, at 10-8-10-4 mol/L, in the presence of tetrodotoxin (TTX, 10-7 mol/L).Results were normalized to cross sectional area. A repeat concentration-response curve was obtained after incubation of the muscle for 90 min with antagonists for M1 (pirenzepine),M2 (methoctramine) and M3 (darifenacin) muscarinic receptor subtypes. The sensitivity to PTX was tested by the ip injection of 100 mg/kg of PTX 5 d before the experiment. The antral circular smooth muscles were removed from PTX-treated and non-treated rats as strips and dispersed smooth muscle cells to identify whether PTX-linked pathway mediated the contractility to bethanechol.RESULTS: A dose-dependent contractile response observed with bethanechol, was not affected by TTX. The pretreatment of rats with pertussis toxin decreased the contraction induced by bethanechol. Lack of calcium as Well as the presence of the L-type calcium channel blocker, nifedipine, also inhibited the cholinergic contraction, with a reduction in response from 2.5±0.4 g/mm2 to 1.2±0.4 g/mm2 (P<0.05). The doseresponse curves were shifted to the right by muscarinic antagonists in the following order of affinity: darifenacin(M3)>methocramine (M2)>pirenzepine (M1).CONCLUSION: The muscarinic receptors-dependent contraction of rat antral circular smooth muscles was linked to the signal transduction pathway(s) involving pertussis-toxin sensitive GTP-binding proteins and to extracellular calcium via L-type voltage gated calcium channels. The presence of the

  16. Structural limits on force production and shortening of smooth muscle.

    Science.gov (United States)

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

    2013-02-01

    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 elements

  17. Contractile proteins of endothelial cells, platelets and smooth muscle.

    Science.gov (United States)

    Becker, C G; Nachman, R L

    1973-04-01

    In experiments described herein it was observed, by direct and indirect immunofluorescence technics, that rabbit antisera to human platelet actomyosin (thrombosthenin) stained mature megakaryocytes, blood platelets, endothelial cells and smooth muscle cells of arteries and veins, endothelial cells of liver sinusoids and certain capillaries, uterine smooth muscle cells, myoepithelial cells, perineurial cells of peripheral nerves and "fibroblastic" cells of granulation tissue. The specificity of immunohistologic staining was confirmed by appropriate absorption and blocking studies and immunodiffusional analysis in agarose gel. It was also observed by immunodiffusional analysis in agarose gel, electrophoresis of actomyosin fragments in polyacrylamide gels, immune inhibition of actomyosin ATPase activity and immune aggregation of platelets that uterine and platelet actomyosin are partially, but not completely, identical.

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

    Science.gov (United States)

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

    2000-02-01

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

  19. Icariin on relaxation effect of corpus cavernosum smooth muscle

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    NO-cGMP pathway in penile corpus cavernosal smooth muscle plays an important role in penile erection. The level of cGMP is regulated by a balance between the rate of synthesis by guanylate cyclase and the rate of hydrolytic breakdown to guanosine 5′monophosphate (GMP) by phos- phodiesterase 5(PDE5). Icariin is isolated from natural drug Epimedii herba, it is shown to have the relaxation effect on corpus cavernosal smooth muscle of rabbit (IC50: 4×10-4 mol/L), and the mechanism of the relaxation effect of Icariin on corpus cavernosum believed to have the inhibiting effect on PDE5 and activation of NO-cGMP pathway to enhancing penile erection.

  20. Modulation of the Cholinergic Mechanisms in the Bronchial Smooth Muscle.

    Science.gov (United States)

    1984-06-01

    Ginsborg and Hirst, 1q72; Sawynok and Jhamandas, 1976), although theopylline has not shown to be a specific adenosine receptor antagonist in all the tissues... theopylline and other cyclic nucletide phosphodiesterase inhibitors. Acta Pharmacol. Toxicol. 45, 336-344. Fredholm, B.B. and P. Hedqvist, 1980...51 mM) evoked release of [3H]-Ach from cholinergic nerves in the bronchial smooth muscle. The effect of theopylline (I mM) on the response to

  1. Action on ileal smooth muscle of synthetic detergents and pardaxin.

    Science.gov (United States)

    Primor, N

    1986-01-01

    Pardaxin (PX), a toxic and repellent substance isolated from the Red Sea flatfish, causes a sharp ball-like profile of drop of saline placed on a hydrophobic film to turn into a flattened one. This effect results with a decrease of the contact angle (theta) from 96 degrees to a maximum of 42 degrees at 10(-4) M of PX. The action of sodium dodecyl sulphate (SDS), a synthetic anionic detergent, benzalkonium chloride (BAC) cationic detergent and pardaxin (PX) a toxic protein with detergent properties, were studied in the ileal guinea-pig longitudinal smooth muscle preparation. SDS (4 X 10(-4) M) and PX (5 X 10(-6) M) diminished the muscle contractile response to field stimulation (0.1 Hz, 1 msec) and to acetylcholine (Ach) and to histamine and elicited a prolonged (4-6 min) TTX-insensitive muscle contraction. The dose dependence of muscle contraction to SDS and PX was found to be sigmoidal and occurred over a narrow range of concentrations. The SDS- but not PX-induced muscle contraction could be reduced by diphenhydramine (H1 antihistamine). BAC (10(-5)-10(-4) M) suppressed the muscle's contractile response to electrical stimulation (0.1 Hz, 1 msec), to Ach, histamine and 5-hydroxytryptamine but did not produce muscle contraction. PX at concentrations higher than 5 X 10(-6) M is a potent detergent and at this concentration shares several pharmacological similarities with SDS.

  2. Smooth muscle FGF/TGFβ cross talk regulates atherosclerosis progression.

    Science.gov (United States)

    Chen, Pei-Yu; Qin, Lingfeng; Li, Guangxin; Tellides, George; Simons, Michael

    2016-07-01

    The conversion of vascular smooth muscle cells (SMCs) from contractile to proliferative phenotype is thought to play an important role in atherosclerosis. However, the contribution of this process to plaque growth has never been fully defined. In this study, we show that activation of SMC TGFβ signaling, achieved by suppression of SMC fibroblast growth factor (FGF) signaling input, induces their conversion to a contractile phenotype and dramatically reduces atherosclerotic plaque size. The FGF/TGFβ signaling cross talk was observed in vitro and in vivo In vitro, inhibition of FGF signaling increased TGFβ activity, thereby promoting smooth muscle differentiation and decreasing proliferation. In vivo, smooth muscle-specific knockout of an FGF receptor adaptor Frs2α led to a profound inhibition of atherosclerotic plaque growth when these animals were crossed on Apoe(-/-) background and subjected to a high-fat diet. In particular, there was a significant reduction in plaque cellularity, increase in fibrous cap area, and decrease in necrotic core size. In agreement with these findings, examination of human coronary arteries with various degrees of atherosclerosis revealed a strong correlation between the activation of FGF signaling, loss of TGFβ activity, and increased disease severity. These results identify SMC FGF/TGFβ signaling cross talk as an important regulator of SMC phenotype switch and document a major contribution of medial SMC proliferation to atherosclerotic plaque growth.

  3. Spontaneously tonic smooth muscle has characteristically higher levels of RhoA/ROK compared with the phasic smooth muscle.

    Science.gov (United States)

    Patel, Chirag A; Rattan, Satish

    2006-11-01

    The internal anal sphincter (IAS) tone is important for the rectoanal continence. The RhoA/Rho kinase (ROK) pathway has been associated with the agonist-induced sustained contraction of the smooth muscle, but its role in the spontaneously tonic smooth muscle is not known. Present studies compared expression of different components of the RhoA/ROK pathway between the IAS (a truly tonic SM), the rectal smooth muscle (RSM) (a mixture of phasic and tonic), and anococcygeus smooth muscle (ASM) (a purely phasic SM) of rat. RT-PCR and Western blot analyses were performed to determine RhoA, ROCK-II, CPI-17, MYPT1, and myosin light-chain 20 (MLC20). Phosphorylated CPI-17 at threonine-38 residue (p(Thr38)-CPI-17), MYPT1 at threonine-696 residue (p(Thr696)-MYPT1), and MLC20 at threonine-18/serine-19 residues (p(Thr18/Ser19)-MLC20) were also determined in the basal state and after pretreatment with the ROK inhibitor Y 27632. In addition, we compared the effect of Y 27632 on the basal isometric tension and ROK activity in the IAS vs. the RSM. Our data show the highest levels of RhoA, ROCK-II, CPI-17, MLC20, and of phospho-MYPT1, -CPI-17, and -MLC20 in the IAS followed by in the RSM and ASM. Conversely, MYPT1 levels were lowest in the IAS and highest in the ASM. In the IAS, Y 27632 caused a concentration-dependent decrease in the basal tone, levels of phospho-MYPT1, -CPI-17, and -MLC20, and ROK activity. We conclude that RhoA/ROK plays a critical role in the basal tone in the IAS by the inhibition of MLC phosphatase via the phosphorylation of MYPT1 and CPI-17.

  4. A fibroblast-associated antigen: Characterization in fibroblasts and immunoreactivity in smooth muscle differentiated stromal cells

    DEFF Research Database (Denmark)

    Rønnov-Jessen, Lone; Celis, Julio E.; van Deurs, Bo

    1992-01-01

    Fibroblasts with smooth muscle differentiation are frequently derived from human breast tissue. Immunofluorescence cytochemistry of a fibroblast-associated antigen recognized by a monoclonal antibody (MAb), 1B10, was analyzed with a view to discriminating smooth muscle differentiated fibroblasts...... from vascular smooth muscle cells. The antigen was detected on the cell surface and in cathepsin D-positive and acridine orange-accumulating vesicular compartments of fibroblasts. Ultrastructurally, the antigen was revealed in coated pits and in endosomal and lysosomal structures. 1B10 recognized three...... immunoreactivity was specific to fibroblasts and smooth muscle differentiated fibroblasts within the context of vascular smooth muscle cells....

  5. Angiogenesis is induced by airway smooth muscle strain.

    Science.gov (United States)

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

    2007-10-01

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

  6. Endoscopic gold fiducial marker placement into the bladder wall to optimize radiotherapy targeting for bladder-preserving management of muscle-invasive bladder cancer: feasibility and initial outcomes.

    Directory of Open Access Journals (Sweden)

    Maurice M Garcia

    Full Text Available BACKGROUND AND PURPOSE: Bladder radiotherapy is a management option for carefully selected patients with muscle-invasive bladder cancer. However, the inability to visualize the tumor site during treatment and normal bladder movement limits targeting accuracy and increases collateral radiation. A means to accurately and reliably target the bladder during radiotherapy is needed. MATERIALS AND METHODS: Eighteen consecutive patients with muscle-invasive bladder cancer (T1-T4 elected bladder-preserving treatment with maximal transurethral resection (TUR, radiation and concurrent chemotherapy. All underwent endoscopic placement of 24-K gold fiducial markers modified with micro-tines (70 [2.9×0.9 mm.]; 19 [2.1×0.7 mm. into healthy submucosa 5-10 mm. from the resection margin, using custom-made coaxial needles. Marker migration was assessed for with intra-op bladder-filling cystogram and measurement of distance between markers. Set-up error and marker retention through completion of radiotherapy was confirmed by on-table portal imaging. RESULTS: Between 1/2007 and 7/2012, a total of 89 markers (3-5 per tumor site were placed into 18 patients of mean age 73.6 years. Two patients elected cystectomy before starting treatment; 16/18 completed chemo-radiotherapy. All (100% markers were visible with all on-table (portal, cone-beam CT, fluoroscopy, plain-film, and CT-scan imaging. In two patients, 1 of 4 markers placed at the tumor site fell-out (voided during the second half of radiotherapy. All other markers (80/82, 98% were present through the end of radio-therapy. No intraoperative (e.g. uncontrolled bleeding, collateral injury or post-operative complications (e.g. stone formation, urinary tract infection, post-TUR hematuria >48 hours occurred. Use of micro-tined fiducial tumor-site markers afforded a 2 to 6-fold reduction in bladder-area targeted with high-dose radiation. DISCUSSION: Placement of the micro-tined fiducial markers into the bladder was

  7. Dihydroartemisinin-Stimulated Hyperplasia of Rat Lung Smooth Muscles

    Directory of Open Access Journals (Sweden)

    Utoh-Nedosa U. Anastasia

    2012-01-01

    Full Text Available Problem statement: Dihydroartemisinin was shown to produce two types of inhibitory effects on the cardiac muscles of rats. It was also shown to stimulate haemopoiesis in the lungs, liver, spleen, intestine and kidney of rats. This study attempted to find out the nature of the effect of oral dihydroartemisinin on the lungs of Wistar albino rats. Approach: The effects of dihydroartemisinin on the tissues of the lungs of wistar albino rats were investigated with five doses of Dihydroartemisinin (DHA administered for 5 days by oral intubation. The five tested doses were 1 mg kg-1, a repeated dose of 1, 2, 60 and 80 mg kg-1 DHA. Results: Histopathological examination of the tissue micrographs of the lungs of the dihydroartemisinin treated rats showed that in comparism with those of the controls, DHA had no adverse effects on the tissues of the lungs of the rats but rather produced a direct stimulatory effect on the smooth muscles of the lungs. This stimulation caused hyperplasia of these tissues which was observable histologically in tissue micrographs of the lungs. These effects of dihydroartemisinin on the tissues of the lungs of Wistar albino rats were dose, repetition and time dependent. Conclusion: These growth hormone-like stimulatory effects of dihydroartemisinin on the smooth muscles of the lungs suggest that DHA enhanced the functioning capacity of the lungs of the DHA-treated rats. These results suggest that dihydroartemisinin has possible respiration enhancement effects.

  8. Mouse bladder wall injection.

    Science.gov (United States)

    Fu, Chi-Ling; Apelo, Charity A; Torres, Baldemar; Thai, Kim H; Hsieh, Michael H

    2011-07-12

    Mouse bladder wall injection is a useful technique to orthotopically study bladder phenomena, including stem cell, smooth muscle, and cancer biology. Before starting injections, the surgical area must be cleaned with soap and water and antiseptic solution. Surgical equipment must be sterilized before use and between each animal. Each mouse is placed under inhaled isoflurane anesthesia (2-5% for induction, 1-3% for maintenance) and its bladder exposed by making a midline abdominal incision with scissors. If the bladder is full, it is partially decompressed by gentle squeezing between two fingers. The cell suspension of interest is intramurally injected into the wall of the bladder dome using a 29 or 30 gauge needle and 1 cc or smaller syringe. The wound is then closed using wound clips and the mouse allowed to recover on a warming pad. Bladder wall injection is a delicate microsurgical technique that can be mastered with practice.

  9. Mechanics of smooth muscle in isolated single microvessels.

    Science.gov (United States)

    Gore, R W; Davis, M J

    1984-01-01

    In vivo studies on frog mesenteric arterioles (4) indicate that segmental differences in the response of microvessels to physical and chemical stimuli can be explained simply in terms of the length-tension characteristics of vascular smooth muscle at different points along the vascular tree. Studies on single, isolated arterioles in vitro were initiated to examine more closely the validity of this explanation for regional response differences. This paper reports some of the results. First-, second-, and third-order arterioles (18-60 micron i.d.) were dissected from hamster cheek pouches. The vessels were cannulated with a modified Burg microperfusion system, and their mechanical properties studied using the methods described by Duling and Gore. Vessels were activated in four stages with K+ and norepinephrine. During activation, transmural pressures were adjusted to minimize vascular smooth-muscle shortening. Active pressure-diameter curves were recorded while adjusting transmural pressure through the range 5 to 400 cm H20 in 5-25 cm steps. Vessel dimensions were measured with a videomicrometer. Passive curves were obtained after equilibration overnight in Ca2+-free medium. The vessels were then fixed and prepared for histologic sectioning, and measurements of vessel-wall composition were made. The Laplace relationship was used to construct length-tension diagrams, and the histologic data were used to normalize the dimensional data to smooth-muscle lengths. Maximum active tension of second-order arterioles (1,170 dynes/cm) was two times previous values reported by Gore et al. This was due presumably to refinements in techniques and dissection procedures. Maximum active stress averaged 3.9 X 10(+6) dynes/cm2 for second-order arterioles. This number is identical to data obtained from hog carotid strips by Dillon et al.

  10. Multiple congenital familial smooth muscle hamartoma in two siblings.

    Science.gov (United States)

    García-Gavín, Juan; Pérez-Pérez, Lidia; Allegue, Francisco; Pérez-Pedrosa, Alberto; Ortíz-Rey, Jose Antonio; Zulaica, Ander

    2012-05-15

    Smooth muscle hamartoma (SMH) is a cutaneous malformation mainly composed of a disorganized proliferation of normal muscle fibers that arise from arrector pili. It usually presents as a single congenital lesion that frequently involves the back and the lower limbs. Unusual clinical presentations, such as atypical localizations, multiple disseminated lesions, and generalized forms have been rarely described. In 2001, Gualandri et al. reported the presence of multiple SMH in three members of the same family, namely two brothers and their mother. This is, as far as we know, the only familial case reported in the English literature. We herein describe a similar case affecting two siblings who presented with identical congenital lesions in the same location.

  11. [Biomechanics and bio-energetics of smooth muscle contraction. Relation to bronchial hyperreactivity].

    Science.gov (United States)

    Coirault, C; Blanc, F X; Chemla, D; Salmeron, S; Lecarpentier, Y

    2000-06-01

    Mechanical studies of isolated muscle and analysis of molecular actomyosin interactions have improved our understanding of the pathophysiology of airway smooth muscle. Mechanical properties of airway smooth muscle are similar to those of other smooth muscles. Airway smooth muscle exhibits spontaneous intrinsic tone and its maximum shortening velocity (Vmax) is 10-30 fold lower than in striated muscle. Smooth muscle myosin generates step size and elementary force per crossbridge interaction approximately similar to those of skeletal muscle myosin. Special slow cycling crossbridges, termed latch-bridges, have been attributed to myosin light chain dephosphorylation. From a mechanical point of view, it has been shown that airway hyperresponsiveness is characterized by an increased Vmax and an increased shortening capacity, with no significant change in the force-generating capacity.

  12. Biophysical induction of vascular smooth muscle cell podosomes.

    Directory of Open Access Journals (Sweden)

    Na Young Kim

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

  13. MURC deficiency in smooth muscle attenuates pulmonary hypertension

    Science.gov (United States)

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

    2016-01-01

    Emerging evidence suggests that caveolin-1 (Cav1) is associated with pulmonary arterial hypertension. MURC (also called Cavin-4) is a member of the cavin family, which regulates caveolar formation and functions together with caveolins. Here, we show that hypoxia increased Murc mRNA expression in the mouse lung, and that Murc-null mice exhibited attenuation of hypoxia-induced pulmonary hypertension (PH) accompanied by reduced ROCK activity in the lung. Conditional knockout mice lacking Murc in smooth muscle also resist hypoxia-induced PH. MURC regulates the proliferation and migration of pulmonary artery smooth muscle cells (PASMCs) through Rho/ROCK signalling. Cav1 suppresses RhoA activity in PASMCs, which is reversed by MURC. MURC binds to Cav1 and inhibits the association of Cav1 with the active form of Gα13, resulting in the facilitated association of the active form of Gα13 with p115RhoGEF. These results reveal that MURC has a function in the development of PH through modulating Rho/ROCK signalling. PMID:27546070

  14. Smooth muscle archvillin is an ERK scaffolding protein.

    Science.gov (United States)

    Gangopadhyay, Samudra S; Kengni, Edouard; Appel, Sarah; Gallant, Cynthia; Kim, Hak Rim; Leavis, Paul; DeGnore, Jon; Morgan, Kathleen G

    2009-06-26

    ERK influences a number of pathways in all cells, but how ERK activities are segregated between different pathways has not been entirely clear. Using immunoprecipitation and pulldown experiments with domain-specific recombinant fragments, we show that smooth muscle archvillin (SmAV) binds ERK and members of the ERK signaling cascade in a domain-specific, stimulus-dependent, and pathway-specific manner. MEK binds specifically to the first 445 residues of SmAV. B-Raf, an upstream regulator of MEK, constitutively interacts with residues 1-445 and 446-1250. Both ERK and 14-3-3 bind to both fragments, but in a stimulus-specific manner. Phosphorylated ERK is associated only with residues 1-445. An ERK phosphorylation site was determined by mass spectrometry to reside at Ser132. A phospho-antibody raised to this site shows that the site is phosphorylated during alpha-agonist-mediated ERK activation in smooth muscle tissue. Phosphorylation of SmAV by ERK decreases the association of phospho-ERK with SmAV. These results, combined with previous observations, indicate that SmAV serves as a new ERK scaffolding protein and provide a mechanism for regulation of ERK binding, activation, and release from the signaling complex.

  15. Effects of lubiprostone on human uterine smooth muscle cells.

    Science.gov (United States)

    Cuppoletti, John; Malinowska, Danuta H; Chakrabarti, Jayati; Ueno, Ryuji

    2008-06-01

    Lubiprostone, a bicyclic fatty acid derivative and member of a new class of compounds called prostones, locally activates ClC-2 Cl(-) channels without activation of prostaglandin receptors. The present study was specifically designed to test and compare lubiprostone and prostaglandin effects at the cellular level using human uterine smooth muscle cells. Effects on [Ca(2+)](i), membrane potential and [cAMP](i) in human uterine smooth muscle cells were measured. 10 nM lubiprostone significantly decreased [Ca(2+)](i) from 188 to 27 nM, which was unaffected by 100 nM SC-51322, a prostaglandin EP receptor antagonist. In contrast 10nM PGE(2) and PGE(1) both increased [Ca(2+)](i) 3-5-fold which was blocked by SC-51322. Similarly, lubiprostone and prostaglandins had opposite/different effects on membrane potential and [cAMP](i). Lubiprostone caused SC-51322-insensitive membrane hyperpolarization and no effect on [cAMP](i). PGE(2) and PGE(1) both caused SC-51322-sensitive membrane depolarization and increased [cAMP](i). Lubiprostone has fundamentally different cellular effects from prostaglandins that are not mediated by EP receptors.

  16. Microintegrating smooth muscle cells into a biodegradable, elastomeric fiber matrix.

    Science.gov (United States)

    Stankus, John J; Guan, Jianjun; Fujimoto, Kazuro; Wagner, William R

    2006-02-01

    Electrospinning permits fabrication of biodegradable elastomers into matrices that can resemble the scale and mechanical behavior of the native extracellular matrix. However, achieving high-cellular density and infiltration with this technique remains challenging and time consuming. We have overcome this limitation by electrospraying vascular smooth muscle cells (SMCs) concurrently with electrospinning a biodegradable, elastomeric poly(ester urethane)urea (PEUU). Trypan blue staining revealed no significant decrease in cell viability from the fabrication process and electrosprayed SMCs spread and proliferated similar to control unprocessed SMCs. The resulting SMC microintegrated PEUU constructs were cultured under static conditions or transmural perfusion. Higher cell numbers resulted with perfusion culture with 131% and 98% more viable cells versus static culture at days 4 and 7 (pfibers after perfusion culture. SMC microintegrated PEUU was strong, flexible and anisotropic with tensile strengths ranging from 2.0 to 6.5 MPa and breaking strains from 850 to 1,700% dependent on the material axis. The ability to microintegrate smooth muscle or other cell types into a biodegradable elastomer fiber matrix embodies a novel tissue engineering approach that could be applied to fabricate high cell density elastic tissue mimetics, blood vessels or other cardiovascular tissues.

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

    Science.gov (United States)

    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

  18. MicroRNAs dynamically remodel gastrointestinal smooth muscle cells.

    Directory of Open Access Journals (Sweden)

    Chanjae Park

    Full Text Available Smooth muscle cells (SMCs express a unique set of microRNAs (miRNAs which regulate and maintain the differentiation state of SMCs. The goal of this study was to investigate the role of miRNAs during the development of gastrointestinal (GI SMCs in a transgenic animal model. We generated SMC-specific Dicer null animals that express the reporter, green fluorescence protein, in a SMC-specific manner. SMC-specific knockout of Dicer prevented SMC miRNA biogenesis, causing dramatic changes in phenotype, function, and global gene expression in SMCs: the mutant mice developed severe dilation of the intestinal tract associated with the thinning and destruction of the smooth muscle (SM layers; contractile motility in the mutant intestine was dramatically decreased; and SM contractile genes and transcriptional regulators were extensively down-regulated in the mutant SMCs. Profiling and bioinformatic analyses showed that SMC phenotype is regulated by a complex network of positive and negative feedback by SMC miRNAs, serum response factor (SRF, and other transcriptional factors. Taken together, our data suggest that SMC miRNAs are required for the development and survival of SMCs in the GI tract.

  19. Identification of C-Kit-Positive Interstitial Cells in the Dog Lower Urinary Tract and Relationship with Smooth Muscle and Nerves. Hypotheses for a Likely Pacemaker Role.

    Directory of Open Access Journals (Sweden)

    Silvana Arrighi

    2010-01-01

    Full Text Available The aim of this work was to give an evidence of the likely presence of interstitial cells in the canine lower urinary tract and to study their possible interactions with the musculature and the intramural innervation. Cryosections of normal canine bladder and urethra were immunofluorescently labelled with c-kit, a transmembrane, tyrosine kinase growth factor receptor, known to be expressed on the interstitial cells of Cajal (ICCs of the gut. The relationship with antiactin positive smooth muscle cells and PGP9.5-positive intramural innervation was also investigated by confocal microscopy. Anti-c-kit labelling demonstrated a network of elongated and branched c-kit positive cells, which were located in interstitial spaces, oriented in parallel to the smooth muscle bundles that form the bladder muscular layer, irrespective of dog sex. Cells with a similar localization were also PAS- and NADPH-diaphorase-positive. A contact between c-kit immunofluorescent cells and intramural innervation was demonstrated, too. The roles of interstitial cells might include regulation of smooth muscle activity of the bladder detrusor, integrating neuronal signals during urine storage and voiding.

  20. Identification of C-kit-positive interstitial cells in the dog lower urinary tract and relationship with smooth muscle and nerves. Hypotheses for a likely pacemaker role.

    Science.gov (United States)

    Arrighi, Silvana; Bosi, Giampaolo; Groppetti, Debora; Cremonesi, Fausto

    2010-01-01

    The aim of this work was to give an evidence of the likely presence of interstitial cells in the canine lower urinary tract and to study their possible interactions with the musculature and the intramural innervation. Cryosections of normal canine bladder and urethra were immunofluorescently labelled with c-kit, a transmembrane, tyrosine kinase growth factor receptor, known to be expressed on the interstitial cells of Cajal (ICCs) of the gut. The relationship with antiactin positive smooth muscle cells and PGP9.5-positive intramural innervation was also investigated by confocal microscopy. Anti-c-kit labelling demonstrated a network of elongated and branched c-kit positive cells, which were located in interstitial spaces, oriented in parallel to the smooth muscle bundles that form the bladder muscular layer, irrespective of dog sex. Cells with a similar localization were also PAS- and NADPH-diaphorase-positive. A contact between c-kit immunofluorescent cells and intramural innervation was demonstrated, too. The roles of interstitial cells might include regulation of smooth muscle activity of the bladder detrusor, integrating neuronal signals during urine storage and voiding.

  1. Identification of C-Kit-Positive Interstitial Cells in the Dog Lower Urinary Tract and Relationship with Smooth Muscle and Nerves. Hypotheses for a Likely Pacemaker Role.

    Science.gov (United States)

    Arrighi, Silvana; Bosi, Giampaolo; Groppetti, Debora; Cremonesi, Fausto

    2010-01-01

    The aim of this work was to give an evidence of the likely presence of interstitial cells in the canine lower urinary tract and to study their possible interactions with the musculature and the intramural innervation. Cryosections of normal canine bladder and urethra were immunofluorescently labelled with c-kit, a transmembrane, tyrosine kinase growth factor receptor, known to be expressed on the interstitial cells of Cajal (ICCs) of the gut. The relationship with antiactin positive smooth muscle cells and PGP9.5-positive intramural innervation was also investigated by confocal microscopy. Anti-c-kit labelling demonstrated a network of elongated and branched c-kit positive cells, which were located in interstitial spaces, oriented in parallel to the smooth muscle bundles that form the bladder muscular layer, irrespective of dog sex. Cells with a similar localization were also PAS- and NADPH-diaphorase-positive. A contact between c-kit immunofluorescent cells and intramural innervation was demonstrated, too. The roles of interstitial cells might include regulation of smooth muscle activity of the bladder detrusor, integrating neuronal signals during urine storage and voiding. PMID:20706651

  2. Fibronectin matrix polymerization regulates smooth muscle cell phenotype through a Rac1 dependent mechanism.

    Directory of Open Access Journals (Sweden)

    Feng Shi

    Full Text Available Smooth muscle cells are maintained in a differentiated state in the vessel wall, but can be modulated to a synthetic phenotype following injury. Smooth muscle phenotypic modulation is thought to play an important role in the pathology of vascular occlusive diseases. Phenotypically modulated smooth muscle cells exhibit increased proliferative and migratory properties that accompany the downregulation of smooth muscle cell marker proteins. Extracellular matrix proteins, including fibronectin, can regulate the smooth muscle phenotype when used as adhesive substrates. However, cells produce and organize a 3-dimensional fibrillar extracellular matrix, which can affect cell behavior in distinct ways from the protomeric 2-dimensional matrix proteins that are used as adhesive substrates. We previously showed that the deposition/polymerization of fibronectin into the extracellular matrix can regulate the deposition and organization of other extracellular matrix molecules in vitro. Further, our published data show that the presence of a fibronectin polymerization inhibitor results in increased expression of smooth muscle cell differentiation proteins and inhibits vascular remodeling in vivo. In this manuscript, we used an in vitro cell culture system to determine the mechanism by which fibronectin polymerization affects smooth muscle phenotypic modulation. Our data show that fibronectin polymerization decreases the mRNA levels of multiple smooth muscle differentiation genes, and downregulates the levels of smooth muscle α-actin and calponin proteins by a Rac1-dependent mechanism. The expression of smooth muscle genes is transcriptionally regulated by fibronectin polymerization, as evidenced by the increased activity of luciferase reporter constructs in the presence of a fibronectin polymerization inhibitor. Fibronectin polymerization also promotes smooth muscle cell growth, and decreases the levels of actin stress fibers. These data define a Rac1

  3. Smooth Muscle Hgs Deficiency Leads to Impaired Esophageal Motility

    Science.gov (United States)

    Chen, Jicheng; Hou, Ning; Zhang, Chong; Teng, Yan; Cheng, Xuan; Li, Zhenhua; Ren, Jie; Zeng, Jian; Li, Rui; Wang, Wei; Yang, Xiao; Lan, Yu

    2015-01-01

    As a master component of endosomal sorting complex required for transport proteins, hepatocyte growth factor-regulated tyrosine kinase substrate (Hgs) participates multiple cellular behaviors. However, the physiological role of Hgs in smooth muscle cells (SMCs) is by far unknown. Here we explored the in vivo function of Hgs in SMCs by using a conditional gene knockout strategy. Hgs deficiency in SMCs uniquely led to a progressive dilatation of esophagus with a remarkable thinning muscle layer. Of note, the mutant esophagus showed a decreased contractile responsiveness to potassium chloride and acetylcholine stimulation. Furthermore, an increase in the inhibitory neurites along with an intense infiltration of T lymphocytes in the mucosa and muscle layer were observed. Consistently, Hgs deficiency in SMCs resulted in a disturbed expression of a set of genes involved in neurotrophin and inflammation, suggesting that defective SMC might be a novel source for excessive production of cytokines and chemokines which may trigger the neuronal dysplasia and ultimately contribute to the compromised esophageal motility. The data suggest potential implications in the pathogenesis of related diseases such as gastroesophageal reflux disease. PMID:26078721

  4. Troponin T3 expression in skeletal and smooth muscle is required for growth and postnatal survival: characterization of Tnnt3(tm2a(KOMP)Wtsi) mice.

    Science.gov (United States)

    Ju, Yawen; Li, Jie; Xie, Chao; Ritchlin, Christopher T; Xing, Lianping; Hilton, Matthew J; Schwarz, Edward M

    2013-09-01

    The troponin complex, which consists of three regulatory proteins (troponin C, troponin I, and troponin T), is known to regulate muscle contraction in skeletal and cardiac muscle, but its role in smooth muscle remains controversial. Troponin T3 (TnnT3) is a fast skeletal muscle troponin believed to be expressed only in skeletal muscle cells. To determine the in vivo function and tissue-specific expression of Tnnt3, we obtained the heterozygous Tnnt3+/flox/lacZ mice from Knockout Mouse Project (KOMP) Repository. Tnnt3(lacZ/+) mice are smaller than their WT littermates throughout development but do not display any gross phenotypes. Tnnt3(lacZ/lacZ) embryos are smaller than heterozygotes and die shortly after birth. Histology revealed hemorrhagic tissue in Tnnt3(lacZ/lacZ) liver and kidney, which was not present in Tnnt3(lacZ/+) or WT, but no other gross tissue abnormalities. X-gal staining for Tnnt3 promoter-driven lacZ transgene expression revealed positive staining in skeletal muscle and diaphragm and smooth muscle cells located in the aorta, bladder, and bronchus. Collectively, these findings suggest that troponins are expressed in smooth muscle and are required for normal growth and breathing for postnatal survival. Moreover, future studies with this mouse model can explore TnnT3 function in adult muscle function using the conditional-inducible gene deletion approach

  5. Whole-cell recordings of calcium and potassium currents in acutely isolated smooth muscle cells

    Institute of Scientific and Technical Information of China (English)

    Qing Cai; Zhong-Liang Zhu; Xiao-Li Fan

    2006-01-01

    AIM: To record calcium and potassium currents in acutely isolated smooth muscle cells of mesenteric arterial branches in rats.METHODS: Smooth muscle cells were freshly isolated by collagenase digest and mechanical trituration with polished pipettes. Patch clamp technique in whole-cell mode was employed to record calcium and potassium currents.RESULTS: The procedure dissociated smooth muscle cells without impairing the electrophysiological characteristics of the cells. The voltage-gated Ca2+ and potassium currents were successfully recorded using whole-cell patch clamp configuration.CONCLUSION: The method dissociates smooth muscle cells from rat mesenteric arterial branches. Voltage-gated channel currents can be recorded in this preparation.

  6. Hospitalization for transurethral bladder resection reduces quality of life in Danish patients with non-muscle-invasive bladder tumour

    DEFF Research Database (Denmark)

    Mogensen, Karin; Christensen, Karl B.; Vrang, Marie-Louise;

    2016-01-01

    Objective The aim of the study was to evaluate the impact of transurethral resection of bladder tumour (TURBT) on patients' quality of life (QoL) and to validate a tool to quantify problems associated with TURBT in a Danish population. Materials and methods A prospective study was carried out using...... a combination of questionnaires and interviews. The study included 165 consecutive patients undergoing a TURBT owing to non-muscle-invasive bladder cancer (NMIBC) from 1 May 2011 to 30 April 2012. Seven patients were selected for interviews. The Danish translation of the QLQ-NMIBC24 Quality of Life...... Questionnaire for NMIBC, from the European Organisation for Research and Treatment of Cancer (EORTC), was used. The interviews were semi-structured. The reliability of the subscales quantifying QoL as defined by the EORTC was tested by computing Cronbach's coefficient alpha and confirmatory factor analysis...

  7. Membrane Currents in Airway Smooth Muscle: Mechanisms and Therapeutic Implications

    Directory of Open Access Journals (Sweden)

    Luke J Janssen

    1997-01-01

    Full Text Available Electrophysiological and pharmacological techniques were used to characterize the membrane conductance changes underlying spasmogen-evoked depolarization in airway smooth muscle (ASM. Changes included a transient activation of chloride ion channels and prolonged suppression of potassium ion channels; both changes are triggered by release of internally sequestered calcium ion and in turn cause opening of voltage-dependent calcium channels. The resultant influx of calcium ions contributes to contraction as well as to refilling of the internal calcium ion pool. Bronchodilators, on the other hand, act in part through activation of potassium channels, with consequent closure of calcium channels. The tools used to study ion channels in ASM are described, and the investigations of the roles of ion channels in ASM physiology (autacoid-evoked depolarization and hyperpolarization and pathophysiology (airway hyperresponsiveness are summarized. Finally, how the relationship between ion channels and ASM function/dysfunction may relate to the treatment of asthma and related breathing disorders is discussed.

  8. Transforming growth factor-β and smooth muscle differentiation

    Institute of Scientific and Technical Information of China (English)

    2012-01-01

    Transforming growth factor(TGF)-β family members are multifunctional cytokines regulating diverse cel- lular functions such as growth,adhesion,migration, apoptosis,and differentiation.TGF-βs elicit their effects via specific typeⅠand typeⅡserine/threonine kinase receptors and intracellular Smad transcription factors. Knockout mouse models for the different components of the TGF-β signaling pathway have revealed their critical roles in smooth muscle cell(SMC)differentia- tion.Genetic studies in humans have linked mutations in these signaling components to specific cardiovascular disorders such as aorta aneurysm and congenital heart diseases due to SMC defects.In this review,the current understanding of TGF-β function in SMC differentiation is highlighted,and the role of TGF-βsignaling in SMC- related diseases is discussed.

  9. Bronchoprotective effect of simulated deep inspirations in tracheal smooth muscle.

    Science.gov (United States)

    Pascoe, Christopher D; Donovan, Graham M; Bossé, Ynuk; Seow, Chun Y; Paré, Peter D

    2014-12-15

    Deep inspirations (DIs) taken before an inhaled challenge with a spasmogen limit airway responsiveness in nonasthmatic subjects. This phenomenon is called bronchoprotection and is severely impaired in asthmatic subjects. The ability of DIs to prevent a decrease in forced expiratory volume in 1 s (FEV1) was initially attributed to inhibition of airway narrowing. However, DIs taken before methacholine challenge limit airway responsiveness only when a test of lung function requiring a DI is used (FEV1). Therefore, it has been suggested that prior DIs enhance the compliance of the airways or airway smooth muscle (ASM). This would increase the strain the airway wall undergoes during the subsequent DI, which is part of the FEV1 maneuver. To investigate this phenomenon, we used ovine tracheal smooth muscle strips that were subjected to shortening elicited by acetylcholine with or without prior strain mimicking two DIs. The compliance of the shortened strip was then measured in response to a stress mimicking one DI. Our results show that the presence of "DIs" before acetylcholine-induced shortening resulted in 11% greater relengthening in response to the third DI, compared with the prior DIs. This effect, although small, is shown to be potentially important for the reopening of closed airways. The effect of prior DIs was abolished by the adaptation of ASM to either shorter or longer lengths or to a low baseline tone. These results suggest that DIs confer bronchoprotection because they increase the compliance of ASM, which, consequently, promotes greater strain from subsequent DI and fosters the reopening of closed airways.

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

    Science.gov (United States)

    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.

  11. Oxygen mediates vascular smooth muscle relaxation in hypoxia.

    Directory of Open Access Journals (Sweden)

    Jessica Dada

    Full Text Available The activation of soluble guanylate cyclase (sGC by nitric oxide (NO and other ligands has been extensively investigated for many years. In the present study we considered the effect of molecular oxygen (O2 on sGC both as a direct ligand and its affect on other ligands by measuring cyclic guanosine monophosphate (cGMP production, as an index of activity, as well as investigating smooth muscle relaxation under hypoxic conditions. Our isolated enzyme studies confirm the function of sGC is impaired under hypoxic conditions and produces cGMP in the presence of O2, importantly in the absence of NO. We also show that while O2 could partially affect the magnitude of sGC stimulation by NO when the latter was present in excess, activation by the NO independent, haem-dependent sGC stimulator 3-(5'-hydroxymethyl-2'-furyl-1-benzylindazole (YC-1 was unaffected. Our in vitro investigation of smooth muscle relaxation confirmed that O2 alone in the form of a buffer bolus (equilibrated at 95% O2/5% CO2 had the ability to dilate vessels under hypoxic conditions and that this was dependent upon sGC and independent of eNOS. Our studies confirm that O2 can be a direct and important mediator of vasodilation through an increase in cGMP production. In the wider context, these observations are key to understanding the relative roles of O2 versus NO-induced sGC activation.

  12. IP3 receptors regulate vascular smooth muscle contractility and hypertension

    Science.gov (United States)

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

    2016-01-01

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

  13. Circular smooth muscle contributes to esophageal shortening during peristalsis

    Institute of Scientific and Technical Information of China (English)

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

    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 ± 1.64 degrees,84.87± 1.04 degrees and 83.72 ± 1

  14. Bladder cancer: utility of MRI in detection of occult muscle-invasive disease

    Energy Technology Data Exchange (ETDEWEB)

    Rosenkrantz, Andrew B. [Dept. of Radiology, NYU Langone Medical Center, New York (United States)], E-mail: Andrew.rosenkrantz@nyumc.org; Mussi, Thais C. [Dept. of Radiology, NYU Langone Medical Center, New York (United States); Hospital Israelita Albert Einstein, Sao Paulo (Brazil); Melamed, Jonathan [Dept. of Pathology, NYU Langone Medical Center, New York (United States); Taneja, Samir S.; Huang, William C. [Dept. of Urology, Div. of Urologic Oncology, NYU Langone Medical Center, New York (United States)

    2012-07-15

    Background. The presence of muscularis propria invasion by bladder cancer is a key factor in prognosis and treatment decisions, although may be missed by biopsy due to sampling error. MRI has shown potential for detection of muscle invasion but has not specifically been evaluated for this purpose in the setting of bladder cancer patients without evidence of muscle invasion on initial biopsy. Purpose. To evaluate the role of MRI in detection of muscularis propria invasion by bladder cancer following a pathologic diagnosis of non-invasive tumor. Material and Methods. This retrospective study included 23 patients who underwent pelvic MRI following a pathologic diagnosis of bladder cancer without muscularis propria invasion and in whom additional histologic evaluation was performed following MRI. Two radiologists in consensus reviewed T2-weighted images to identify those cases suspicious for muscle invasion on MRI. The radiologists identified whether cases suspicious for invasion demonstrated disruption of the T2-hypointense muscularis layer of the bladder wall, peri-vesical fat stranding, and peri-vesical soft tissue nodularity. Findings were compared with pathologic results obtained after MRI. Results. Suspicion was raised for muscle invasion in eight of 23 cases, four of which exhibited invasion on follow-up pathology. No case without suspicion on MRI exhibited invasion on follow-up pathology. Therefore, sensitivity and specificity were 100% and 79%, respectively. Among individual findings, muscularis disruption on T2WI exhibited sensitivity of 100% and specificity of 79%, peri-vesical fat stranding exhibited sensitivity and specificity of 50% and 84%, and peri-vesical soft tissue nodularity exhibited sensitivity and specificity of 25% and 100%. Conclusion. MRI demonstrated high sensitivity for detection of muscle invasion in cases of bladder cancer without invasion on initial histologic assessment. Muscularis disruption on T2WI appeared to exhibit a better

  15. Pasteur effect in vascular and intestinal smooth muscle.

    Science.gov (United States)

    Pettersson, G; Lundholm, L

    1985-01-01

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

  16. Beta adrenoreceptors in the rabbit bladder detrusor muscle

    Energy Technology Data Exchange (ETDEWEB)

    Anderson, G.F.; Marks, B.H.

    1984-02-01

    This study examines the beta adrenergic receptors of the rabbit detrusor smooth muscle, employing (/sup 125/I)iodocyanopindolol (ICYP) as a ligand for the binding of beta adrenergic receptors. Saturation binding experiments on the isolated membrane fraction yielded a KD for ICYP of 14.7 pM and a maximum binding of 147.6 fmol/mg of protein. Displacement of labeled ICYP by a series of beta adrenergic agents yielded the following KD values for the combined high and low affinity binding sites: I-propranolol, 0.76 nM; ICI 118,551, 1.7 nM; zinterol, 38.0 nM; metoprolol, 3.5 microM; and practolol, 61.4 microM. When these displacement experimental results were compared to KD values from other reported binding studies with ICYP for beta adrenoreceptors, both the order of potency and the KD values indicated primarily beta-2 adrenergic receptor subtypes. Computer program Scatfit analysis of the displacement curves indicated a single slope and affinity constant for all five beta adrenergic agents. Hofstee plots for zinterol, ICI 118,551 and metoprolol, however, were not linear and indicated that minor populations of beta-1 adrenoreceptors were also present as both high and low affinity binding sites could be defined. It is concluded that the primary receptor population is beta-2 and that this tissue is heterogenous with a small population of beta-1 adrenoreceptors representing approximately 13 to 23% of the total beta adrenoreceptor population.

  17. Increased proinflammatory responses from asthmatic human airway smooth muscle cells in response to rhinovirus infection

    NARCIS (Netherlands)

    Oliver, Brian G G; Johnston, Sebastian L; Baraket, Melissa; Burgess, Janette K; King, Nicholas J C; Roth, Michael; Lim, Sam; Black, Judith L

    2006-01-01

    BACKGROUND: Exacerbations of asthma are associated with viral respiratory tract infections, of which rhinoviruses (RV) are the predominant virus type. Airway smooth muscle is important in asthma pathogenesis, however little is known about the potential interaction of RV and human airway smooth muscl

  18. Insulin-Induced Laminin Expression Promotes a Hypercontractile Airway Smooth Muscle Phenotype

    NARCIS (Netherlands)

    Dekkers, Bart G. J.; Schaafsma, Dedmer; Tran, Thai; Zaagsma, Johan; Meurs, Herman

    2009-01-01

    Airway smooth muscle (ASM) plays a key role in the development of airway hyperresponsiveness and remodeling in asthma, which may involve maturation of ASM cells to a hypercontractile phenotype. In vitro studies have indicated that long-term exposure of bovine tracheal smooth muscle (BTSM) to insulin

  19. (Endo)cannabinoid signaling in human bronchial epithelial and smooth muscle cells

    NARCIS (Netherlands)

    Gkoumassi, Effimia

    2007-01-01

    We investigated the pathways used by various (endo)cannabinoids in regulating intracellular calcium homeostasis, adenylyl cyclase and ERK signaling, in bronchial epithelial cells as well as smooth muscle cells. In DDT1 MF2 smooth muscle cells the synthetic cannabinoid CP55,940 increases [Ca2+]i by a

  20. CD40 and OX40 ligand are increased on stimulated asthmatic airway smooth muscle

    NARCIS (Netherlands)

    Burgess, Janette K; Blake, Anita E; Boustany, Sarah; Johnson, Peter R A; Armour, Carol L; Black, Judith L; Hunt, Nicholas H; Hughes, J Margaret

    2005-01-01

    BACKGROUND: Severe, persistent asthma is characterized by airway smooth muscle hyperplasia, inflammatory cell infiltration into the smooth muscle, and increased expression of many cytokines, including IL-4, IL-13, IL-1beta, and TNF-alpha. These cytokines have the potential to alter the expression of

  1. Regulation of GPCR-mediated smooth muscle contraction : implications for asthma and pulmonary hypertension

    NARCIS (Netherlands)

    Wright, D B; Tripathi, S; Sikarwar, A; Santosh, K T; Perez-Zoghbi, J; Ojo, O O; Irechukwu, N; Ward, J P T; Schaafsma, D

    2013-01-01

    Contractile G-protein-coupled receptors (GPCRs) have emerged as key regulators of smooth muscle contraction, both under healthy and diseased conditions. This brief review will discuss some key topics and novel insights regarding GPCR-mediated airway and vascular smooth muscle contraction as discusse

  2. Treatment Options Available for Bacillus Calmette-Guerin Failure in Non-muscle-invasive Bladder Cancer

    NARCIS (Netherlands)

    Yates, D.R.; Brausi, M.A.; Catto, J.W.; Dalbagni, G.; Roupret, M.; Shariat, S.F.; Sylvester, R.J.; Witjes, J.A.; Zlotta, A.R.; Palou-Redorta, J.

    2012-01-01

    CONTEXT: Intravesical bacillus Calmette-Guerin (BCG) is a standard conservative treatment for patients with high-risk non-muscle-invasive bladder cancer (NMIBC). Many patients will experience recurrence or progression following BCG and are termed BCG failures. OBJECTIVE: To summarise the current tre

  3. Evaluation of tissue and urinary survivin expression in non-muscle-invasive bladder cancer

    Directory of Open Access Journals (Sweden)

    S. Sharaf

    2012-12-01

    Conclusion: Urinary survivin is a useful marker for non-invasive detection of non-muscle-invasive bladder cancer recurrence. Its detection is better using ELISA technique than WB and there is no correlation between its expression in tissue and urine.

  4. Treatment and outcome in muscle invasive bladder cancer : a population-based survey

    NARCIS (Netherlands)

    Leliveld, Anna M.; Doornweerd, Benjamin H. J.; Bastiaannet, Esther; Schaapveld, Michael; de Jong, Igle J.

    2010-01-01

    OBJECTIVE: To assess treatments and survival of patients with muscle invasive bladder cancer (MIBC) in the Comprehensive Cancer Center Northern Netherlands (CCCN) region. STUDY DESIGN AND SETTING: Retrospective cohort analysis. Data of 548 patients with MIBC diagnosed between 1997 and 2002 were coll

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

    Science.gov (United States)

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

    2001-11-09

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

  6. Some properties of the smooth muscle of mouse vas deferens.

    Science.gov (United States)

    Holman, M E; Taylor, G S; Tomita, T

    1977-04-01

    1. Contractions of the mouse vas deferens in response to electrical stimulation differ form those recorded form the guinea-pig vas deferens in that they are abolished by tetrodotoxin. 2. Changes in membrane potentials were recorded form the smooth muscle of both preparations in response to stimulation with current pulses applied by an intracellular electrode and by alrge extracellular plate electrodes. 3. Both preparations behaved similarly in response to intracellular stimulation. Electrotonic potentials in response to extracellular current pulses spread in a longitudinal direction in the guinea-pig vas deferens in accordance with the cable-like properties of this preparation. In contrast, no longitudinal spread of eletrotonus was observed in the mouse vas deferens. 4. Responses to nerve stimulation differed in the two preparations. In the guinea-pig, single stimuli caused excitatory junction potentials (e.j.p.s) which gave rise to action potentials. Some cells from the mouse vas deferens showed similar e.j.p.s and action potentials, although the threshold for the initiation of action potentials was lower and more variable. 5. The majority of cells in the mouse vas deferens failed to show action potentials in response to a single stimuli even though the amplitude of e.j.p.s was from 35 to 40 mV. This was probably due to the large resting membrane potentials of these cells, as all-or-nothing action potentials could be evoked if successive e.j.p.s were allowed to sum with each other or if a depolarizing current pulse was applied at the peak of an e.j.p. 6. The nature of the response to nerve stimulation recorded from differnt cells in the mouse vas deferens could be correlated with the amplitude and time course of the response of the same cell to intracellular stimulation. 7. It is concluded that individual smooth muscle cells in both preparations are probably coupled electrically but that there are few, if any, low resistance pathways in the longitudinal direction

  7. Sympathetically evoked Ca2+ signaling in arterial smooth muscle

    Institute of Scientific and Technical Information of China (English)

    Wei-jin ZANG; Joseph ZACHARIA; Christine LAMONT; Withrow Gil WIER

    2006-01-01

    The sympathetic nervous system plays an essential role in the control of total peripheral vascular resistance and blood flow, by controlling the contraction of small arteries. Perivascular sympathetic nerves release ATP, norepinephrine (NE) and neuropeptide Y. This review summarizes our knowledge of the intracellular Ca2+ signals that are activated by ATP and NE, acting respectively on P2X1 and α1 adrenoceptors in arterial smooth muscle. Each neurotransmitter produces a unique type of post-synaptic Ca2+ signal and associated contraction. The neural release of ATP and NE is thought to vary markedly with the pattern of nerve activity, probably reflecting both pre- and post-synaptic mechanisms. Finally, we show that Ca2+ signaling during neurogenic contractions activated by trains of sympathetic nerve fiber action potentials are in fact significantly different from that elicited by simple bath application of exogenous neurotransmitters to isolated arteries (a common experimental technique), and end by identifying important questions remaining in our understanding of sympathetic neurotransmission and the physiological regulation of contraction of small arteries.

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

    Science.gov (United States)

    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.

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

    Science.gov (United States)

    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.

  10. DIAGNOSTIC IMPLICATIONS OF IMMUNOHISTOCHEMICAL MARKERS IN UTERINE SMOOTH MUSCLE TUMORS

    Institute of Scientific and Technical Information of China (English)

    朱雪琼; 石一复; 陈晓端; 吴裕中

    2004-01-01

    Objective: To evaluate the diagnostic implications of immunohistochemical markers in uterine smooth muscle tumors. Methods: Formalin-fixed paraffin-embedded tissue blocks were selected from 17 uterine leiomyosarcomas, 40 uterine unusual leiomyomas and 25 uterine usual leiomyomas. Utilizing immunohistochemical techniques with antigen retrieval, serial sections of each tumor for immunoreactivity with myogenic markers, ovarian steroid receptors, CD44v3, proliferating cell nuclear antigen and mast cells were assessed. Results: Although the myogenic markers and CD44v3 showed less frequent positivity in uterine leiomyosarcomas than those in unusual leiomyomas, they were not reliable markers for differentiating leiomyosarcoma from leiomyoma. Uterine leiomyosarcoma tended to have lower ovarian steroid receptors immunoreactivity rates than leiomyoma. Leiomyoma tended to have a higher quantity of intratumoral mast cells than leiomyosarcoma, while the expression of proliferating cell nuclear antigen was lower in them. Conclusion: Because the estimation of mitotic count was subject to significant variation, the immunohistochemical expression of ovarian steroid receptors, mast cells and proliferating cell nuclear antigen seemed to be helpful for the discrimination of unusual leiomyoma from leiomyosarcoma.

  11. Arterial Myogenic Activation through Smooth Muscle Filamin A

    Directory of Open Access Journals (Sweden)

    Kevin Retailleau

    2016-03-01

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

  12. Epigenetic Control of Smooth Muscle Cell Identity and Lineage Memory.

    Science.gov (United States)

    Gomez, Delphine; Swiatlowska, Pamela; Owens, Gary K

    2015-12-01

    Vascular smooth muscle cells (SMCs), like all cells, acquire a cell-specific epigenetic signature during development that includes acquisition of a unique repertoire of histone and DNA modifications. These changes are postulated to induce an open chromatin state (referred to as euchromatin) on the repertoire of genes that are expressed in differentiated SMC, including SMC-selective marker genes like Acta2 and Myh11, as well as housekeeping genes expressed by most cell types. In contrast, genes that are silenced in differentiated SMC acquire modifications associated with a closed chromatin state (ie, heterochromatin) and transcriptional silencing. Herein, we review mechanisms that regulate epigenetic control of the differentiated state of SMC. In addition, we identify some of the major limitations in the field and future challenges, including development of innovative new tools and approaches, for performing single-cell epigenetic assays and locus-selective editing of the epigenome that will allow direct studies of the functional role of specific epigenetic controls during development, injury repair, and disease, including major cardiovascular diseases, such as atherosclerosis, hypertension, and microvascular disease, associated with diabetes mellitus.

  13. SREBP inhibits VEGF expression in human smooth muscle cells.

    Science.gov (United States)

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

    2006-03-31

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

  14. Histone deacetylase 8 regulates cortactin deacetylation and contraction in smooth muscle tissues.

    Science.gov (United States)

    Li, Jia; Chen, Shu; Cleary, Rachel A; Wang, Ruping; Gannon, Olivia J; Seto, Edward; Tang, Dale D

    2014-08-01

    Histone deacetylases (HDACs) are a family of enzymes that mediate nucleosomal histone deacetylation and gene expression. Some members of the HDAC family have also been implicated in nonhistone protein deacetylation, which modulates cell-cycle control, differentiation, and cell migration. However, the role of HDACs in smooth muscle contraction is largely unknown. Here, HDAC8 was localized both in the cytoplasm and the nucleus of mouse and human smooth muscle cells. Knockdown of HDAC8 by lentivirus-encoding HDAC8 shRNA inhibited force development in response to acetylcholine. Treatment of smooth muscle tissues with HDAC8 inhibitor XXIV (OSU-HDAC-44) induced relaxation of precontracted smooth muscle tissues. In addition, cortactin is an actin-regulatory protein that undergoes deacetylation during migration of NIH 3T3 cells. In this study, acetylcholine stimulation induced cortactin deacetylation in mouse and human smooth muscle tissues, as evidenced by immunoblot analysis using antibody against acetylated lysine. Knockdown of HDAC8 by RNAi or treatment with the inhibitor attenuated cortactin deacetylation and actin polymerization without affecting myosin activation. Furthermore, expression of a charge-neutralizing cortactin mutant inhibited contraction and actin dynamics during contractile activation. These results suggest a novel mechanism for the regulation of smooth muscle contraction. In response to contractile stimulation, HDAC8 may mediate cortactin deacetylation, which subsequently promotes actin filament polymerization and smooth muscle contraction.

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

    Science.gov (United States)

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

    2007-01-01

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

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  18. THE RECURRENCE AFTER ORGAN-SAVING SURGERY OF PATIENTS WITH MUSCLE-INVASIVE BLADDER CANCER

    Directory of Open Access Journals (Sweden)

    T. A. Sveklina

    2014-08-01

    Full Text Available The article represents the study of frequency and nature of the recurrence and survival rate (common, oncology-specific, disease-free after organ-saving surgery of patients with muscle-invasive bladder cancer stages T2b and T3a. Oncology-speсific and disease-free survival rates were much higher if full diagnosis of bladder mucosa, the adjuvant intravesical chemotherapy had been on pre-operative and intra-operative stages than in the absence of these diagnosis and therapy. Recurrentes of bladder cancer which appeared in the absence of diagnosis and combination therapy, statistically reliably occured at another location other than the zone of operation, stage of recurrentes and degree of differentiation of recurrents were less than the original tumor. This information confirms the existence of foci of cancer in situ which have not been identified on the diagnostic stage.

  19. Cigarette Smoke and Estrogen Signaling in Human Airway Smooth Muscle

    Directory of Open Access Journals (Sweden)

    Venkatachalem Sathish

    2015-06-01

    Full Text Available Aims: Cigarette smoke (CS in active smokers and second-hand smoke exposure exacerbate respiratory disorders such as asthma and chronic bronchitis. While women are known to experience a more asthmatic response to CS than emphysema in men, there is limited information on the mechanisms of CS-induced airway dysfunction. We hypothesize that CS interferes with a normal (protective bronchodilatory role of estrogens, thus worsening airway contractility. Methods: We tested effects of cigarette smoke extract (CSE on 17β-estradiol (E2 signaling in enzymatically-dissociated bronchial airway smooth muscle (ASM obtained from lung samples of non-smoking female patients undergoing thoracic surgery. Results: In fura-2 loaded ASM cells, CSE increased intracellular calcium ([Ca2+]i responses to 10µM histamine. Acute exposure to physiological concentrations of E2 decreased [Ca2+]i responses. However, in 24h exposed CSE cells, although expression of estrogen receptors was increased, the effect of E2 on [Ca2+]i was blunted. Acute E2 exposure also decreased store-operated Ca2+ entry and inhibited stromal interaction molecule 1 (STIM1 phosphorylation: effects blunted by CSE. Acute exposure to E2 increased cAMP, but less so in 24h CSE-exposed cells. 24h CSE exposure increased S-nitrosylation of ERα. Furthermore, 24h CSE-exposed bronchial rings showed increased bronchoconstrictor agonist responses that were not reduced as effectively by E2 compared to non-CSE controls. Conclusion: These data suggest that CS induces dysregulation of estrogen signaling in ASM, which could contribute to increased airway contractility in women exposed to CS.

  20. Mechanisms of Cigarette Smoke Effects on Human Airway Smooth Muscle.

    Directory of Open Access Journals (Sweden)

    Mark E Wylam

    Full Text Available Cigarette smoke contributes to or exacerbates airway diseases such as asthma and COPD, where airway hyperresponsiveness and airway smooth muscle (ASM proliferation are key features. While factors such as inflammation contribute to asthma in part by enhancing agonist-induced intracellular Ca(2+ ([Ca(2+]i responses of ASM, the mechanisms by which cigarette smoke affect ASM are still under investigation. In the present study, we tested the hypothesis that cigarette smoke enhances the expression and function of Ca(2+ regulatory proteins leading to increased store operated Ca(2+ entry (SOCE and cell proliferation. Using isolated human ASM (hASM cells, incubated in the presence and absence cigarette smoke extract (CSE we determined ([Ca(2+]i responses and expression of relevant proteins as well as ASM proliferation, reactive oxidant species (ROS and cytokine generation. CSE enhanced [Ca(2+]i responses to agonist and SOCE: effects mediated by increased expression of TRPC3, CD38, STIM1, and/or Orai1, evident by attenuation of CSE effects when siRNAs against these proteins were used, particularly Orai1. CSE also increased hASM ROS generation and cytokine secretion. In addition, we found in the airways of patients with long-term smoking history, TRPC3 and CD38 expression were significantly increased compared to life-long never-smokers, supporting the role of these proteins in smoking effects. Finally, CSE enhanced hASM proliferation, an effect confirmed by upregulation of PCNA and Cyclin E. These results support a critical role for Ca(2+ regulatory proteins and enhanced SOCE to alter airway structure and function in smoking-related airway disease.

  1. Cooling-induced contraction in ovine airways smooth muscle.

    Science.gov (United States)

    Mustafa, S M; Pilcher, C W; Williams, K I

    1999-02-01

    The mechanism of cold-induced bronchoconstriction is poorly understood. This prompted the present study whose aim was to determine the step-wise direct effect of cooling on smooth muscle of isolated ovine airways and analyse the role of calcium in the mechanisms involved. Isolated tracheal strips and bronchial segments were suspended in organ baths containing Krebs' solution for isometric tension recording. Tissue responses during stepwise cooling from 37 to 5 degrees C were examined. Cooling induced a rapid and reproducible contraction proportional to cooling temperature in ovine tracheal and bronchial preparations which was epithelium-independent. On readjustment to 37 degrees C the tone returned rapidly to basal level. Maximum contraction was achieved at a temperature of 5 degrees C for trachea and 15 degrees C for bronchiole. Cooling-induced contractions (CIC) was resistant to tetrodotoxin (1; 10 micrometer), and not affected by the muscarinic antagonist atropine (1 micrometer) or the alpha-adrenergic antagonist phentolamine (1 micrometer), or the histamine H1-antagonist mepyramine (1 micrometer) or indomethacin (1 micrometer). Ca2+ antagonists (nifedipine and verapamil) and Mn2+ raised tracheal but not bronchiolar tone and augmented CIC. Incubation in Ca2+-free, EGTA-containing Krebs' solution for 5 min had no effect on CIC, although it significantly reduced KCl-induced contraction by up to 75%. Cooling inhibited Ca2+ influx measured using 45Ca2+ uptake. Caffeine (100 micrometer) significantly inhibited CIC. The results show that cooling-induced contractions do not appear to involve activation of nerve endings, all surface reception systems or Ca2+ influx. However, CIC is mainly dependent on release of intracellular Ca2+.

  2. Ex Vivo Smooth Muscle Pharmacological Effects of a Novel Bradykinin-Related Peptide, and Its Analogue, from Chinese Large Odorous Frog, Odorrana livida Skin Secretions

    Science.gov (United States)

    Xiang, Jie; Wang, Hui; Ma, Chengbang; Zhou, Mei; Wu, Yuxin; Wang, Lei; Guo, Shaodong; Chen, Tianbao; Shaw, Chris

    2016-01-01

    Bradykinin-related peptides (BRPs) are one of the most extensively studied frog secretions-derived peptide families identified from many amphibian species. The diverse primary structures of BRPs have been proven essential for providing valuable information in understanding basic mechanisms associated with drug modification. Here, we isolated, identified and characterized a dodeca-BRP (RAP-L1, T6-BK), with primary structure RAPLPPGFTPFR, from the skin secretions of Chinese large odorous frogs, Odorrana livida. This novel peptide exhibited a dose-dependent contractile property on rat bladder and rat ileum, and increased the contraction frequency on rat uterus ex vivo smooth muscle preparations; it also showed vasorelaxant activity on rat tail artery smooth muscle. In addition, the analogue RAP-L1, T6, L8-BK completely abolished these effects on selected rat smooth muscle tissues, whilst it showed inhibition effect on bradykinin-induced rat tail artery relaxation. By using canonical antagonist for bradykinin B1 or B2 type receptors, we found that RAP-L1, T6-BK -induced relaxation of the arterial smooth muscle was very likely to be modulated by B2 receptors. The analogue RAP-L1, T6, L8-BK further enhanced the bradykinin inhibitory activity only under the condition of co-administration with HOE140 on rat tail artery, suggesting a synergistic inhibition mechanism by which targeting B2 type receptors. PMID:27690099

  3. Effects of neoadjuvant chemotherapy on pathological parameters and survival in patients undergoing radical cystectomy for muscle-invasive bladder cancer

    OpenAIRE

    ÇAĞLAYAN, Alper; Akbulut, Ziya; Atmaca, Ali Fuat; Altinova,Serkan; KILIÇ, Metin; Balbay, Mevlana Derya

    2012-01-01

    Aim: To evaluate the effect of neoadjuvant chemotherapy on tumor pathology and patient survival in patients with muscle-invasive bladder cancer undergoing radical cystectomy. Neoadjuvant chemotherapy is believed to prevent micrometastasis and provide pathological downstaging. Materials and methods: Between June 2004 and March 2009, 74 patients with muscle-invasive bladder cancer were treated with radical cystectomy. Patients fit to receive chemotherapy were administered systemic chemotherapy...

  4. A contribution to improved radiotherapy for muscle invading urinary bladder cancer

    Energy Technology Data Exchange (ETDEWEB)

    Muren, Ludvig PAul

    2002-07-01

    Cystectomy has traditionally been regarded the treatment of choice for muscle invading urinary bladder cancer in most countries. Radiotherapy has been offered patients considered unfit for cystectomy. Since the contraindications of surgery are frequent among bladder cancer patients, a substantial amount of patients with muscle invading bladder cancer (typically 50%) are still managed primarily with radiation. Recently, a tri-modality, organsparing treatment (trans-urethral resection and radio-chemotherapy) has been proposed for bladder cancer, like in the management of a range of other common malignancies. This approach may provide as high control rates as cystectomy yet maintain a higher quality of life for selected patient groups. In both the radical radiotherapy and the combined modality approach, high radiation doses are needed to improve local disease control. Radiation dose escalation requires improved conformation of dose distributions. This PhD programme aimed to develop improved conformal radiotherapy procedures in the management of patients with muscle invading urinary bladder cancer. In the initial phase of this work, computer-controlled movement of the linear accelerator collimator jaws during beam delivery was applied to shape so-called partially wedged beams (PWBs), that were designed specifically to tailor the dose distribution in bladder irradiation closer to the defined bladder target. The dosimetric verification and treatment planning implementation of this beam delivery concept were addressed, and we documented that these dynamic beams were delivered as accurately as standard beams. Particular attention was given to the BMS-96 diode array system, as it was adapted to dynamic beam dosimetry. Next, the potential clinical impact of these beams was analysed. In a retrospectively study of a set of urinary bladder treatment plans, the PWBs were seen to improve the dose homogeneity inside the bladder target as well as to reduce normal tissue (small

  5. Effect of Montelukast on bradykinin-induced contraction of isolated tracheal smooth muscle of guinea pig

    Directory of Open Access Journals (Sweden)

    A Noor

    2011-01-01

    Conclusion: It is concluded that montelukast significantly inhibits, in a dose-dependent manner, the bradykinin-induced contraction of the guinea pig tracheal smooth muscle, and alludes to an interaction between the bradykinin and leukotriene mediators.

  6. End-stage renal disease causes an imbalance between endothelial and smooth muscle progenitor cells

    NARCIS (Netherlands)

    Westerweel, Peter E; Hoefer, Imo E; Blankestijn, Peter J; de Bree, Petra; Groeneveld, Dafna; van Oostrom, Olivia; Braam, Branko; Koomans, Hein A; Verhaar, Marianne C

    2007-01-01

    Patients with end-stage renal disease (ESRD) on hemodialysis have an increased risk of cardiovascular disease (CVD). Circulating endothelial progenitor cells (EPC) contribute to vascular regeneration and repair, thereby protecting against CVD. However, circulating smooth muscle progenitor cells (SPC

  7. Focal adhesion kinase regulates collagen I-induced airway smooth muscle phenotype switching

    NARCIS (Netherlands)

    Dekkers, Bart G J; Spanjer, Anita I R; van der Schuyt, Robert D; Kuik, Willem Jan; Zaagsma, Johan; Meurs, Herman

    2013-01-01

    Increased extracellular matrix (ECM) deposition and airway smooth muscle (ASM) mass are major contributors to airway remodeling in asthma. Recently, we demonstrated that the ECM protein collagen I, which is increased surrounding asthmatic ASM, induces a proliferative, hypocontractile ASM phenotype.

  8. Microfibrillar-associated protein 4 modulates airway smooth muscle cell phenotype in experimental asthma

    DEFF Research Database (Denmark)

    Pilecki, Bartosz; Schlosser, Anders; Wulf-Johansson, Helle

    2015-01-01

    . In the current study we investigated the role of MFAP4 in experimental allergic asthma. METHODS: MFAP4-deficient mice were subjected to alum/ovalbumin and house dust mite induced models of allergic airway disease. In addition, human healthy and asthmatic primary bronchial smooth muscle cell cultures were used...... to evaluate MFAP4-dependent airway smooth muscle responses. RESULTS: MFAP4 deficiency attenuated classical hallmarks of asthma, such as eosinophilic inflammation, eotaxin production, airway remodelling and hyperresponsiveness. In wild-type mice, serum MFAP4 was increased after disease development...... and correlated with local eotaxin levels. MFAP4 was expressed in human bronchial smooth muscle cells and its expression was upregulated in asthmatic cells. Regarding the underlying mechanism, we showed that MFAP4 interacted with integrin αvβ5 and promoted asthmatic bronchial smooth muscle cell proliferation...

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

    DEFF Research Database (Denmark)

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

    2010-01-01

    Recently, overexpression of the genes TMEM16A and TMEM16B has been shown to produce currents qualitatively similar to native Ca(2+)-activated Cl(-) currents (I(ClCa)) in vascular smooth muscle. However, there is no information about this new gene family in vascular smooth muscle, where Cl......(-) channels are a major depolarizing mechanism. Qualitatively similar Cl(-) currents were evoked by a pipette solution containing 500 nM Ca(2+) in smooth muscle cells isolated from BALB/c mouse portal vein, thoracic aorta, and carotid artery. Quantitative PCR using SYBR Green chemistry and primers specific...... for transmembrane protein (TMEM) 16A or the closely related TMEM16B showed TMEM16A expression as follows: portal vein > thoracic aorta > carotid artery > brain. In addition, several alternatively spliced variant transcripts of TMEM16A were detected. In contrast, TMEM16B expression was very low in smooth muscle...

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

    DEFF Research Database (Denmark)

    Chen, Xiaoping; Yang, Dachun; Ma, Shuangtao

    2010-01-01

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

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

    Science.gov (United States)

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

    2014-01-01

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

  12. Smooth muscle pharmacology in the isolated virgin and pregnant rat uterus and cervix.

    Science.gov (United States)

    Darios, Emma S; Seitz, Bridget; Watts, Stephanie W

    2012-06-01

    Uterine smooth muscle function is established, but comparatively little is known about cervical smooth muscle pharmacology. We performed a proof-of-principle experiment that smooth muscle was expressed in the cervix in both virgin and pregnant rats, using the uterus as a comparator. We tested whether all tissues were pharmacologically responsive to contractile and relaxant agonists. Immunohistochemistry revealed the expression of smooth muscle α-actin in all tissues. The isolated tissue bath was used to measure isometric contractility of uterine strips and whole cervices from virgin and pregnant (day 11 ± 2) female Sprague-Dawley rats. We tested classic activators of uterine smooth muscle contraction and relaxation in both uterus and cervix. All tissues contracted to the depolarizing agent potassium chloride, prostaglandin F2α, muscarinic cholinergic agonist carbachol [2-[(aminocarbonxyl)oxy]-N,N,N-trimethylethanaminium chloride], and 5-hydroxytryptamine. Unlike other tissues, the pregnant cervix did not contract to oxytocin, but the oxytocin receptor was present. Both cervix and uterus (virgin and pregnant) had concentration-dependent, near-complete relaxation to the adrenergic agonist norepinephrine and adenylate cyclase activator forskolin [(3R,4aR,5S,6S,6aS,10S,10aR,10bS)-6,10-10b-trihydroxy-3,4a,7,10a-pentamethyl-1-oxo-3-vinyldodecahydro-1H-benzo[f] chroment-5-yl acetate]. The β-adrenergic receptor agonist isoproterenol was less potent in pregnant cervix versus virgin by ∼10-fold. All tissues, particularly the cervix, responded poorly to the nitric-oxide donor sodium nitroprusside, relaxing ∼20% maximally. These findings support the importance of smooth muscle in the cervix, the use of the isolated cervix in pharmacological studies, and a similarity between smooth muscle pharmacology of the nonpregnant uterus and cervix. This work highlights the unappreciated smooth muscle function of the cervix versus uterus and cervical changes in pharmacology during

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

    Directory of Open Access Journals (Sweden)

    Dong-Hai Liu

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

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

    Institute of Scientific and Technical Information of China (English)

    田利民

    2014-01-01

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

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

    Science.gov (United States)

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

    2014-01-01

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

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

    Directory of Open Access Journals (Sweden)

    Thomas R Whitesell

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

  17. Mast cell numbers in airway smooth muscle and PC(20)AMP in asthma and COPD

    NARCIS (Netherlands)

    Liesker, J. J. W.; ten Hacken, N. H. T.; Rutgers, S. R.; Zeinstra-Smith, M.; Postma, D. S.; Timens, W.

    2007-01-01

    Introduction: Most patients with asthma and many patients with COPD show bronchial hyperresponsiveness to adenosine (BHRAMP). BHRAMP may be caused by release of mast cell histamine, which induces smooth muscle contraction. Aim of the study: To evaluate whether mast cell numbers in airway smooth musc

  18. Airway smooth muscle phenotype and function : interactions with current asthma therapies

    NARCIS (Netherlands)

    Halayko, A J; Tran, T; Ji, S Y; Yamasaki, A; Gosens, R

    2006-01-01

    Asthma incidence has climbed markedly in the past two decades despite an increased use of medications that suppress airway inflammation and repress contraction of smooth muscle that encircles the airways. Asthmatics exhibit episodes of airway inflammation that potentiates reversible airway smooth mu

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

    Directory of Open Access Journals (Sweden)

    Amy Y Hsiao

    Full Text Available 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.

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

    Science.gov (United States)

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

    2015-01-01

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

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

    Directory of Open Access Journals (Sweden)

    Chintamani eJoshi

    2012-06-01

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

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

    Directory of Open Access Journals (Sweden)

    Tong Luo

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

  3. Nitric oxide mediates stretch-induced Ca2+ release via activation of phosphatidylinositol 3-kinase-Akt pathway in smooth muscle.

    Directory of Open Access Journals (Sweden)

    Bin Wei

    Full Text Available BACKGROUND: Hollow smooth muscle organs such as the bladder undergo significant changes in wall tension associated with filling and distension, with attendant changes in muscle tone. Our previous study indicated that stretch induces Ca(2+ release occurs in the form of Ca(2+ sparks and Ca(2+ waves in urinary bladder myocytes. While, the mechanism underlying stretch-induced Ca2+ release in smooth muscle is unknown. METHODOLOGY/PRINCIPAL FINDINGS: We examined the transduction mechanism linking cell stretch to Ca(2+ release. The probability and frequency of Ca(2+ sparks induced by stretch were closely related to the extent of cell extension and the time that the stretch was maintained. Experiments in tissues and single myocytes indicated that mechanical stretch significantly increases the production of nitric oxide (NO and the amplitude and duration of muscle contraction. Stretch-induced Ca(2+ sparks and contractility increases were abrogated by the NO inhibitor L-NAME and were also absent in eNOS knockout mice. Furthermore, exposure of eNOS null mice to exogenously generated NO induced Ca(2+ sparks. The soluble guanylyl cyclase inhibitor ODQ did not inhibit SICR, but this process was effectively blocked by the PI3 kinase inhibitors LY494002 and wortmannin; the phosphorylation of Akt and eNOS were up-regulated by 204+/-28.6% and 258+/-36.8% by stretch, respectively. Moreover, stretch significantly increased the eNOS protein expression level. CONCLUSIONS/SIGNIFICANCE: Taking together, these results suggest that stretch-induced Ca2+ release is NO dependent, resulting from the activation of PI3K/Akt pathway in smooth muscle.

  4. Correlation between the distribution of smooth muscle or non muscle myosins and alpha-smooth muscle actin in normal and pathological soft tissues.

    Science.gov (United States)

    Benzonana, G; Skalli, O; Gabbiani, G

    1988-01-01

    The distribution of smooth muscle (SM) and non muscle myosins was compared with that of alpha-SM actin in various normal and pathological tissues and in cultured cells by means of indirect immunofluorescence using a monoclonal antibody specific for alpha-SM actin [anti-alpha sm-1, Skalli et al., 1986b] and two polyclonal antibodies raised against bovine aortic myosin (ABAM) and human platelet myosin (AHPM), respectively. In normal tissues ABAM stained vascular and parenchymal smooth muscle cells (SMC), myoepithelial cells and myoid cells of the testis in a pattern similar to that reported by other authors with antisera raised against non vascular SM myosin. Cells stained with ABAM were always positive for anti-alpha sm-1. In human and experimental atheromatous plaques, most cells were positive for AHPM; a variable proportion was also stained for ABAM plus anti-alpha sm-1. Myofibroblasts from rat granulation tissue, Dupuytren's nodule and stroma from breast carcinoma were constantly positive for AHPM and negative for ABAM; however, myofibroblasts from Dupuytren's nodule and breast carcinoma were anti-alpha sm-1 positive. Early primary cultures of rat aortic SMC were positive for ABAM and anti-alpha sm-1 and became negative for ABAM and positive for AHPM after a few days in culture. They remained positive for AHPM and anti-alpha sm-1 after passages; the staining of AHPM and anti-alpha sm-1 appeared to be colocalized along the same stress fibers. These results may be relevant for the understanding of SMC function and adaptation, and show that in non malignant SMC proliferation, alpha-SM actin represents a more general marker of SM origin than SM myosin.

  5. Shortening induced effects on force (re)development in pig urinary smooth muscle

    NARCIS (Netherlands)

    E. van Asselt (Els); J.J.M. Pel (Johan); R. van Mastrigt (Ron)

    2007-01-01

    textabstractIntroduction: When muscle is allowed to shorten during an active contraction, the maximum force that redevelops after shortening is smaller than the isometric force at the same muscle length without prior shortening. We studied the course of force redevelopment after shortening in smooth

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

    Science.gov (United States)

    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…

  7. Dual ERK and phosphatidylinositol 3-kinase pathways control airway smooth muscle proliferation : differences in asthma

    NARCIS (Netherlands)

    Burgess, Janette K; Lee, Jin Hee; Ge, Qi; Ramsay, Emma E; Poniris, Maree H; Parmentier, Johannes; Roth, Michael; Johnson, Peter R A; Hunt, Nicholas H; Black, Judith L; Ammit, Alaina J

    2008-01-01

    Hyperplasia of airway smooth muscle (ASM) within the bronchial wall of asthmatic patients has been well documented and is likely due to increased muscle proliferation. We have shown that ASM cells obtained from asthmatic patients proliferate faster than those obtained from non-asthmatic patients. In

  8. YFa and analogs: Investigation of opioid receptors in smooth muscle contraction

    Institute of Scientific and Technical Information of China (English)

    Krishan Kumar; Ritika Goyal; Annu Mudgal; Anita Mohan; Santosh Pasha

    2011-01-01

    AIM: To study the pharmacological profile and inhibition of smooth muscle contraction by YFa and its analogs in conjunction with their receptor selectivity. METHODS: The effects of YFa and its analogs (D-Ala2) YFa, Y (D-Ala2) GFMKKKFMRF amide and Des-Phe- YGGFMKKKFMR amide in guinea pig ileum (GPI) and mouse vas deferens (MVD) motility were studied using an isolated tissue organ bath system, and morphine and DynA (1-13) served as controls. Acetylcholine was used for muscle stimulation. The observations were validated by specific antagonist pretreatment experiments using naloxonazine, naltrindole and norbinaltorphimine norBNI. RESULTS: YFa did not demonstrate significant inhibition of GPI muscle contraction as compared with morphine (15% vs 62%, P = 0.0002), but moderate inhibition of MVD muscle contraction, indicating the role of κ opioid receptors in the contraction. A moderate inhibition of GPI muscles by (Des-Phe) YFa revealed the role of anti-opiate receptors in the smooth muscle contraction. (D-Ala-2) YFa showed significant inhibition of smooth muscle contraction, indicating the involvement of mainly d receptors in MVD contraction. These results were supported by specific antagonist pretreatment assays. CONCLUSION: YFa revealed its side-effect-free analgesic properties with regard to arrest of gastrointestinal transit. The study provides evidences for the involvement of κ and anti-opioid receptors in smooth muscle contraction.

  9. Shortening induced effects on force (re)development in pig urinary smooth muscle

    OpenAIRE

    van Asselt, Els; Pel, Johan; van Mastrigt, Ron

    2007-01-01

    textabstractIntroduction: When muscle is allowed to shorten during an active contraction, the maximum force that redevelops after shortening is smaller than the isometric force at the same muscle length without prior shortening. We studied the course of force redevelopment after shortening in smooth muscle to unravel the mechanism responsible for this deactivation. Method: In a first series of measurements the shortening velocity was varied resulting in different shortening amplitudes. In a s...

  10. Time course of isotonic shortening and the underlying contraction mechanism in airway smooth muscle.

    Science.gov (United States)

    Syyong, Harley T; Raqeeb, Abdul; Paré, Peter D; Seow, Chun Y

    2011-09-01

    Although the structure of the contractile unit in smooth muscle is poorly understood, some of the mechanical properties of the muscle suggest that a sliding-filament mechanism, similar to that in striated muscle, is also operative in smooth muscle. To test the applicability of this mechanism to smooth muscle function, we have constructed a mathematical model based on a hypothetical structure of the smooth muscle contractile unit: a side-polar myosin filament sandwiched by actin filaments, each attached to the equivalent of a Z disk. Model prediction of isotonic shortening as a function of time was compared with data from experiments using ovine tracheal smooth muscle. After equilibration and establishment of in situ length, the muscle was stimulated with ACh (100 μM) until force reached a plateau. The muscle was then allowed to shorten isotonically against various loads. From the experimental records, length-force and force-velocity relationships were obtained. Integration of the hyperbolic force-velocity relationship and the linear length-force relationship yielded an exponential function that approximated the time course of isotonic shortening generated by the modeled sliding-filament mechanism. However, to obtain an accurate fit, it was necessary to incorporate a viscoelastic element in series with the sliding-filament mechanism. The results suggest that a large portion of the shortening is due to filament sliding associated with muscle activation and that a small portion is due to continued deformation associated with an element that shows viscoelastic or power-law creep after a step change in force.

  11. Smooth muscle cell-derived carbon monoxide is a regulator of vascular cGMP.

    Science.gov (United States)

    Morita, T; Perrella, M A; Lee, M E; Kourembanas, S

    1995-02-28

    Carbon monoxide (CO) is a product of the enzyme heme oxygenase (HO; EC 1.14.99.3). In vascular smooth muscle cells, exogenously administered CO increases cyclic guanosine 3',5'-monophosphate (cGMP), which is an important regulator of vessel tone. We report here that smooth muscle cells produce CO via HO and that it regulates cGMP levels in these cells. Hypoxia, which has profound effects on vessel tone, significantly increased the transcriptional rate of the HO-1 gene resulting in corresponding increases of its mRNA and HO enzymatic activity. In addition, under the same conditions, rat aortic and pulmonary artery smooth muscle cells accumulated high levels of cGMP following a similar time course to that of HO-1 production. The increased accumulation of cGMP in smooth muscle cells required the enzymatic activity of HO, since it was abolished by a specific HO inhibitor, tin protoporphyrin. In contrast, N omega-nitro-L-arginine, a potent inhibitor of nitric oxide (NO) synthesis, had no effect on cGMP produced by smooth muscle cells, indicating that NO is not responsible for the activation of guanylyl cyclase in this setting. Furthermore, conditioned medium from hypoxic smooth muscle cells stimulated cGMP production in recipient cells and this stimulation was completely inhibited by tin protoporphyrin or hemoglobin, an inhibitor of CO production and a scavenger of CO, respectively. This report shows that HO-1 is expressed by vascular smooth muscle cells and that its product, CO, may regulate vascular tone under physiologic and pathophysiologic (such as hypoxic) conditions.

  12. Pentosan polysulfate decreases prostate smooth muscle proliferation and extracellular matrix turnover.

    Science.gov (United States)

    Elliot, S J; Zorn, B H; McLeod, D G; Moul, J W; Nyberg, L; Striker, L J; Striker, G E

    2003-01-01

    Benign prostatic hyperplasia (BPH) involves proliferation of smooth muscle cells and increased deposition of extracellular matrix (ECM). We recently found that pentosan polysulfate (PPS) has marked effects on growth and ECM of smooth muscle cells derived from vascular tissues. We examined smooth muscle cells cultured from human prostates and the effects of PPS on their growth and ECM production. Fragments of surgical prostatectomy specimens were diced, digested with collagenase (0.01%), and placed in culture medium supplemented with 20% fetal bovine serum. Outgrowths of elongated cells were characterized by light microscopic examination and immunohistochemical techniques by the presence of F-actin, alpha-smooth muscle actin, and myosin, which is a characteristic of smooth muscle cells. Two independent isolates were propagated, and growth curves and ECM production were assessed in the presence and absence of PPS (10 or 100 microg/ml). PPS decreased cell number beginning at day 1 and throughout the incubation period, up to 4 days. The amount of the ECM degradative enzymes, metallo-proteinases MMP-9 and MMP-2, was examined by zymography. PPS did not alter the amount of MMP-2 in the supernatants but MMP-9 was increased 234.4 +/- 17.23-fold over control cells. Tissue inhibitor of MMP (TIMPS), examined by reverse zymography, increased 200% over control. The amount of alpha I type (IV) and alpha I type (I) collagen released in the supernatant, measured by ELISA, significantly decreased in PPS-treated cultures. In conclusion, we found that the administration of PPS decreased proliferation as well as ECM production in prostate smooth muscle. Since smooth muscle proliferation and ECM are involved in the pathophysiology of BPH, PPS may have therapeutic potential.

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

    Science.gov (United States)

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

    2015-02-01

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

  14. Effects and its possible mechanism of Radix Saposhnikoviae on rat colonic smooth muscle in vitro

    Institute of Scientific and Technical Information of China (English)

    Zhenqing Liu; Tao Lü; Ping Hu; Muxin Wei

    2009-01-01

    Objective: To determine the effect of different concentrations of Radix Saposhnikoviae (RS) on the contraction of smooth muscle strips and the Ca2. mobilization of cultured smooth muscle cells of rat colon and its possible mechanism of action. Methods: Strips of rat colon longitudinal muscle were prepared and smooth muscle cells from rat colon were isolated and cultured. In the experiments, in vitro muscle strips were suspended in an organ bath and the contraction of the strips was recorded. In the cell-experiments, intracellular Ca2+ was assessed using fluorescent intensity (FI) of smooth muscle cells loaded with Fluo-4/AM, measured with a laser scanning confocal microscope and related software. Results: In the in vitro experiment, RS (0.02, 0.2, 2, 20 g/L) inhibited contraction of muscle strips in a concentration-dependent manner, and this inhibition was significant for the three higher RS concentrations (P < 0.01) for both Peak (the maximal contraction amplitude) and Area (the area under curves), Similarly, RS inhibited Ach-induced contraction. In these experiments the inhibition of the Peak values in the RS 2 and 20 g/L groups was significant (P < 0.01), as was the inhibition of the Area values in all RS groups (P < 0.05). Naloxone and propranolol did not significantly affect the inhibitory effect of RS on smooth muscle contractility, while phentolamine significantly reduced the inhibitory effect (P < 0.01). In experiments using primary smooth muscle cell cultures in Ca2+-containing buffer, the post-treatment fluorescence of cells in the RS 0.2, 2 and 20 g/L groups differed significantly from pre-treatment values (P < 0.05), and the percent inhibition of fluorescence in the RS 2 g/L and 20 g/L groups was significant (P < 0.01). However, in Ca2+-free buffer, FS had no significant effect on cell fluorescence. Conclusion: RS inhibited both the spontaneous and Ach-stimulated contraction of rat colonic smooth muscle strips. This RS effect appeared to involve a

  15. Dysfunctional interaction of C/EBPα and the glucocorticoid receptor in asthmatic bronchial smooth-muscle cells

    NARCIS (Netherlands)

    Roth, Michael; Johnson, Peter R.A.; Borger, Peter; Bihl, Michel P.; Rüdiger, Jochen J.; King, Gregory G.; Ge, Qi; Hostettler, Katrin; Burgess, Janette K.; Black, Judith L.; Tamm, Michael

    2004-01-01

    BACKGROUND: Increased proliferation of bronchial smooth-muscle cells may lead to increased muscle mass in the airways of patients with asthma. The antiproliferative effect of glucocorticoids in bronchial smooth-muscle cells in subjects without asthma is mediated by a complex of the glucocorticoid re

  16. Systemic therapy in muscle-invasive and metastatic bladder cancer: current trends and future promises.

    Science.gov (United States)

    Aragon-Ching, Jeanny B; Trump, Donald L

    2016-09-01

    Bladder urothelial cancers remain an important urologic cancer with limited treatment options in the locally advanced and metastatic setting. While neoadjuvant chemotherapy for locally advanced muscle-invasive cancers has shown overall survival benefit, clinical uptake in practice have lagged behind. Controversies surrounding adjuvant chemotherapy use are also ongoing. Systemic therapies for metastatic bladder cancer have largely used platinum-based therapies without effective standard second-line therapy options for those who fail, although vinflunine is approved in Europe as a second-line therapy based on a Phase III trial, and most recently, atezolizumab, a checkpoint inhibitor, was approved by the US FDA. Given increasing recognition of mutational signatures expressed in urothelial carcinomas, several promising agents with use of VEGF-targeted therapies, HER2-directed agents and immunotherapies with PD-1/PD-L1 antibodies in various settings are discussed herein.

  17. Characterization of cyclic AMP accumulation in cultured human corpus cavernosum smooth muscle cells.

    Science.gov (United States)

    Palmer, L S; Valcic, M; Melman, A; Giraldi, A; Wagner, G; Christ, G J

    1994-10-01

    Intracavernous pharmacotherapy relies heavily on the use of vasoactive agents which act by increasing intracellular cAMP levels in human corpus cavernosum smooth muscle. Yet little is known about the cAMP generating system in this tissue, and how it may affect observed patient variability. Thus, the goal of these studies was to better characterize the biochemistry of cAMP formation in human corpus cavernosum smooth muscle, and thus provide more insight into the mechanisms of corporal smooth muscle relaxation in vivo. We studied both receptor and nonreceptor mediated increases in cAMP formation in short-term cultures of human corpus cavernosum smooth muscle cells. Both isoproterenol (ISO) and prostaglandin E1 (PGE1) produced concentration-dependent increases in cAMP, but histamine, serotonin and vasoactive intestinal polypeptide did not. Forskolin, a relatively specific activator of adenylate cyclase, was also a potent stimulant of cAMP formation in these cells. Moreover, there was a direct correlation between the degree of forskolin-induced cAMP accumulation in cultured corporal smooth muscle cells and the magnitude of the forskolin-induced relaxation response of precontracted isolated corporal smooth muscle strips. Prostaglandin E1 and ISO concentration response curves (CRCs) were then assayed in the absence and presence of subthreshold forskolin (0.1 microM.). In the presence of forskolin, the calculated maximal PGE1-induced cAMP accumulation (Emax) was significantly greater than that elicited by PGE1 alone, ISO alone, or ISO + forskolin (p protocol was used to demonstrate that both 80:20 and 70:30 FMRs (but not 95:5 or 90:10), were associated with significantly greater cAMP Emax values than that observed for PGE1 alone (p < or = 0.01). These data provide direct evidence that the degree of cAMP formation in cultured corporal smooth muscle cells is strongly correlated with the magnitude of relaxation of isolated corporal smooth muscle strips. In addition, since

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

    Science.gov (United States)

    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.

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

    Science.gov (United States)

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

    2008-09-01

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

  20. Effect of 1,1-dimethylphenyl 1,4-piperazinium on mouse tracheal smooth muscle responsiveness.

    Science.gov (United States)

    Dorion, G; Israël-Assayag, E; Beaulieu, M J; Cormier, Y

    2005-06-01

    Bronchial hyperresponsiveness is one of the main features of asthma. A nicotinic receptor agonist, 1,1-dimethylphenyl 1,4-piperazinium (DMPP), has been shown to have an inhibitory effect on airway response to methacholine in an in vivo model of asthma. The aims of this study were to 1) verify whether nicotinic acetylcholine receptors (nAChR) were present on mouse tracheal smooth muscle, 2) verify whether bronchoprotection observed in mice was due to a direct effect on airway smooth muscle, and 3) compare the effects of nicotinic agonists to that of salbutamol. Alpha3-, alpha4-, and alpha7-nAChR subunits were detected by immunofluorescence on tracheal tissues from normal BALB/c mice. The effect of DMPP on tracheal responsiveness was verified by an isometric method. Tracheas were isolated from normal mice, placed in organ baths, and contracted with a single dose of methacholine. Cumulative doses of DMPP or salbutamol were added to the baths. Results show that mouse tracheal smooth muscle is positive for alpha4- and alpha7-nAChR subunits and that the epithelium is positive for alpha3-, alpha4-, and alpha7-subunits. DMPP induced a greater dose-dependent relaxation of tracheal smooth muscles precontracted with methacholine than with salbutamol. These results suggest that the smooth muscle-relaxing effect of DMPP could have some interest in the treatment of obstructive pulmonary diseases.

  1. Gliosarcoma with prominent smooth muscle component (gliomyosarcoma: A report of 10 cases

    Directory of Open Access Journals (Sweden)

    Manisha Khanna

    2011-01-01

    Full Text Available Background and Aim: Gliosarcoma (GS is an uncommon malignant tumor of the brain, consisting of malignant glial, usually a glioblastoma (GB, as well as sarcomatous component; the latter is usually in the form of fibrosarcoma. We report a series of 10 GSs with prominent smooth muscle component, which is a rare occurrence. Settings and Design: Out of a series of 225 cases of GB admitted in our hospital, 10 were diagnosed as GS with prominent smooth muscle component, gliomyosarcoma (GMS. Materials and Methods: This is an observational study based on the experience with 225 cases of GB, encountered between 1995 and 2008, in our hospital. The tumors showing prominent spindle cell component were stained with reticulin and 20 with strongly positive reticulin stain were diagnosed as GS. They were further studied by immunohistochemical staining for glial fibrillary acidic protein (GFAP, smooth muscle actin (SMA, desmin and factor VIII antigen. Results: Out of 225 cases of GB, 20 were diagnosed as GS. Ten of these showed prominent smooth muscle component and were diagnosed as GMS. They revealed varying degrees of SMA and factor VIII Ag positivity. In the sarcomatous component, SMA and factor VIII positive cells were seen close to the vessel walls as well as away from them. Conclusion: GMS containing prominent smooth muscle component may not be as rare as has been reported in the literature. Both GS and GMS appear to arise from the vessel wall at least in some cases, suggesting their possible vascular origin.

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

    Science.gov (United States)

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

    2016-02-01

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

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

    Science.gov (United States)

    Rattan, Satish; Ali, Mehboob

    2015-04-01

    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.

  4. Rosuvastatin inhibits the smooth muscle cell proliferation by targeting TNFα mediated Rho kinase pathway

    Institute of Scientific and Technical Information of China (English)

    Xiao Sun; Hao Tong; Man Zhang; Xiao-Hang Wang

    2012-01-01

    Objective To investigate whether Tumor Necrosis Factor-alpha (TNFα) is capable of activating Rho kinase pathway which leads to smooth muscle cell proliferation and the intervention function of Rosuvastatin, and clarify the mechanism and intervention manner of anti-atherosclerosis by Rosuvastatin. Methods Wistar neonate rat smooth muscle cells were cultured, and the activity of cell proliferation was determined by methyl thiazolyl tetrazolium (MTT). The expression of Rho kinase genes after the stimulation of TNFα was evaluated by RT-PCR. Western blot method was used to measure the protein expression of proliferating cell nuclear antigen (PCNA) after TNFα stimulation and Rosuvastatin intervention in smooth muscle cell. Results The TNFα stimulation significantly enhanced the expression of Rho kinase and increased the expression of PCNA protein in smooth muscle cells (P < 0.05). These effects were positively correlated with prolonged treatment whereas additional Rosuvastatin administration inhibited the above-mentioned effects (P < 0.05). Conclusions The activation of TNFα mediated Rho kinase signaling pathway can significantly promote smooth muscle cell proliferation, and Rosuvastatin can not only inhibit this pathway but also the induced proliferation.

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

    Science.gov (United States)

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

    2015-12-01

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

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

    Science.gov (United States)

    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

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

    Science.gov (United States)

    Absher, M; Baldor, L

    1991-01-01

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

  8. Relaxation of uterine and aortic smooth muscle by glaucolides D and E from Vernonia liatroides.

    Science.gov (United States)

    Campos, María; Oropeza, Martha; Ponce, Héctor; Fernández, Jaquelina; Jimenez-Estrada, Manuel; Torres, Héctor; Reyes-Chilpa, Ricardo

    2003-01-01

    Vernonia spp. (Asteraceae) are used in herbolaria in Latin America in menstrual and stomach disorders, suggesting smooth muscle relaxing properties of some of their chemical constituents. For pharmacological support for this belief, sesquiterpene lactones glaucolides D and E were assayed on isolated rat smooth muscle. Glaucolide E proved more potent than glaucolide D to relax high KCl- or noradrenaline-induced contractions in aorta and to relax the high KCl-contraction in uterus. Hirsutinolide-type sesquiterpene lactone also was tested but displayed no effect. Relaxation of smooth muscle by structurally related sesquiterpene lactone parthenolide has been attributed mainly to the alpha-methylene gamma-lactone moiety; because glaucolides D and E lack this functional group, their relaxant properties may rely on other alkylating sites such as C10 of the germacra-1(10),4-diene-4-epoxide skeleton.

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

    Science.gov (United States)

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

    1998-12-01

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

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

    Science.gov (United States)

    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.

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

    Directory of Open Access Journals (Sweden)

    Amin Mokhtari-Zaer

    2015-08-01

    Full Text Available 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.

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

    Science.gov (United States)

    Kelynack, Kristen J; Holt, Stephen G

    2016-01-01

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

  13. Smooth muscles and stem cells of embryonic guts express KIT, PDGFRRA, CD34 and many other stem cell antigens: suggestion that GIST arise from smooth muscles and gut stem cells.

    Science.gov (United States)

    Terada, Tadashi

    2013-01-01

    Gastrointestinal stromal tumor (GIST) is believed to original from interstitial cells of (ICC) present in Auerbach's nerve plexus. GIST frequently shows gain-of-function mutations of KIT and PDGFRA. In practical pathology, GIST is diagnosed by positive immunostaining or KIT and/or CD34. The author herein demonstrates that human embryonic gastrointestinal tract smooth muscles (HEGITSM) and human embryonic stem gastrointestinal cells (HEGISC) consistently express KIT, CD34, NCAM, PDGFRA and other stem cell (SC) antigens NSE, synaptophysin, chromogranin, bcl-2, ErbB, and MET throughout the embryonic development of 7-40 gestational week (GW). CK14 was negative. The author examines 42 cases (7-40 GW) of embryonic GI tract (EGI). The HEGISM, HEGIST, and gall bladder smooth muscles (SM) were consistently positive for KIT, CD34, NCAM, PDGFRA, synaptophysin, chromogranin, NSE, bcl-2, ErbB2, and MET in foregut, stomach, GB, midgut, and hindgut throughout the fetal life (7-40 GW). The stem cells (SC) were seen to create the SM, nerves, ICC, and other all structures of GI tract. In adult gastrointestinal walls (n=30), KIT, CD34, PDGFRA, and S100 proteins were expressed in Auerbach's nerve plexus and ICC. The bronchial and vascular SM of embryos did not express these molecules. In GIST, frequent expressions of KIT (100%, 30/30), CD34 (90%, 27/30), and PDGFRA (83%, 25/30) were seen. In general, characteristics of tumors recapitulate their embryonic life. Therefore, it is strongly suggested that GIST may be originated from GI SM and/or GI SC in addition to ICC.

  14. Novel intravesical therapies for non-muscle-invasive bladder cancer refractory to BCG.

    Science.gov (United States)

    Barlow, Lamont J; Seager, Catherine M; Benson, Mitchell C; McKiernan, James M

    2010-01-01

    The definitive treatment for patients with non-muscle-invasive bladder cancer (NMIBC) who fail to respond to intravesical BCG is cystectomy. When a patient is deemed BCG-refractory and cannot or will not undergo cystectomy, alternative intravesical therapy may be the most effective way to minimize recurrence and progression. A number of immunotherapeutic and chemotherapeutic agents have been given intravesically over the years, and several recently and currently investigated novel agents appear to be particularly promising for the management of BCG-refractory NMIBC. The most effective treatments in the future will likely utilize targeted therapies based on the underlying genetic mutations associated with each individual diagnosis of NMIBC.

  15. Neoadjuvant therapy in muscle-invasive bladder cancer: a model for rational accelerated drug development.

    Science.gov (United States)

    Balar, Arjun V; Milowsky, Matthew I

    2015-05-01

    Since the advent of cisplatin-based combination therapy in the management of muscle-invasive and advanced bladder cancer, there has been little progress in improving outcomes for patients. Novel therapies beyond cytotoxic chemotherapy are needed. The neoadjuvant paradigm lends to acquiring ample pretreatment and posttreatment tumor tissue as a standard of care, which enables comprehensive biomarker analyses to better understand mechanisms of both response and resistance, which will aid drug development. This article discusses the evolution of neoadjuvant therapy as standard treatment and the role it may serve toward the development of novel therapies.

  16. Heterozygous de novo and inherited mutations in the smooth muscle actin (ACTG2) gene underlie megacystis-microcolon-intestinal hypoperistalsis syndrome.

    Science.gov (United States)

    Wangler, Michael F; Gonzaga-Jauregui, Claudia; Gambin, Tomasz; Penney, Samantha; Moss, Timothy; Chopra, Atul; Probst, Frank J; Xia, Fan; Yang, Yaping; Werlin, Steven; Eglite, Ieva; Kornejeva, Liene; Bacino, Carlos A; Baldridge, Dustin; Neul, Jeff; Lehman, Efrat Lev; Larson, Austin; Beuten, Joke; Muzny, Donna M; Jhangiani, Shalini; Gibbs, Richard A; Lupski, James R; Beaudet, Arthur

    2014-03-01

    Megacystis-microcolon-intestinal hypoperistalsis syndrome (MMIHS) is a rare disorder of enteric smooth muscle function affecting the intestine and bladder. Patients with this severe phenotype are dependent on total parenteral nutrition and urinary catheterization. The cause of this syndrome has remained a mystery since Berdon's initial description in 1976. No genes have been clearly linked to MMIHS. We used whole-exome sequencing for gene discovery followed by targeted Sanger sequencing in a cohort of patients with MMIHS and intestinal pseudo-obstruction. We identified heterozygous ACTG2 missense variants in 15 unrelated subjects, ten being apparent de novo mutations. Ten unique variants were detected, of which six affected CpG dinucleotides and resulted in missense mutations at arginine residues, perhaps related to biased usage of CpG containing codons within actin genes. We also found some of the same heterozygous mutations that we observed as apparent de novo mutations in MMIHS segregating in families with intestinal pseudo-obstruction, suggesting that ACTG2 is responsible for a spectrum of smooth muscle disease. ACTG2 encodes γ2 enteric actin and is the first gene to be clearly associated with MMIHS, suggesting an important role for contractile proteins in enteric smooth muscle disease.

  17. [Local expression of cytokeratins 8, 17 and 18 in the mesenchyme and smooth muscles in the early stages of human organogenesis].

    Science.gov (United States)

    Bozhok, Iu M; Bannikov, G A; Tavokina, L V; Svitkina, T M; Troianovskiĭ, S M

    1989-01-01

    Expression of cytokeratins 7, 8, 17, 18 in human embryos and fetuses of 6.5-13 weeks was studied using light and electron immunocytochemistry and immunoelectroblotting with the monoclonal antibodies. Cytokeratins 8 and 18 were expressed in 6.5-8 week old embryos not only in epithelium but also in mesenchyme of allantois, urogenital sinus, Wolffian and Mullerian ducts, mesentery, urinary bladder and certain regions of colon, rectum and atrium cordis walls. Furthermore, starting from the 10th week smooth-muscle cells of ring layer in caudal part of rectum bound antibodies against cytokeratin 17 in addition to those against cytokeratins 8 and 18. Corresponding mesenchymal and smooth-muscle cells of adult individuals did not react with either of them. Cytokeratins were still synthesized when mesenchymal cells of embryonic intestine wall were cultivated in vitro. Intermediate filaments of these cells contain cytokeratins 8 and 18, as demonstrated by electron immunocytochemistry and immunoelectroblotting. Thus, the expression of cytokeratins is not restricted to adult and embryonic epithelial tissues but is also characteristic of mesenchyme and smooth muscle differentiation in human embryos and fetuses.

  18. Heterozygous de novo and inherited mutations in the smooth muscle actin (ACTG2 gene underlie megacystis-microcolon-intestinal hypoperistalsis syndrome.

    Directory of Open Access Journals (Sweden)

    Michael F Wangler

    2014-03-01

    Full Text Available Megacystis-microcolon-intestinal hypoperistalsis syndrome (MMIHS is a rare disorder of enteric smooth muscle function affecting the intestine and bladder. Patients with this severe phenotype are dependent on total parenteral nutrition and urinary catheterization. The cause of this syndrome has remained a mystery since Berdon's initial description in 1976. No genes have been clearly linked to MMIHS. We used whole-exome sequencing for gene discovery followed by targeted Sanger sequencing in a cohort of patients with MMIHS and intestinal pseudo-obstruction. We identified heterozygous ACTG2 missense variants in 15 unrelated subjects, ten being apparent de novo mutations. Ten unique variants were detected, of which six affected CpG dinucleotides and resulted in missense mutations at arginine residues, perhaps related to biased usage of CpG containing codons within actin genes. We also found some of the same heterozygous mutations that we observed as apparent de novo mutations in MMIHS segregating in families with intestinal pseudo-obstruction, suggesting that ACTG2 is responsible for a spectrum of smooth muscle disease. ACTG2 encodes γ2 enteric actin and is the first gene to be clearly associated with MMIHS, suggesting an important role for contractile proteins in enteric smooth muscle disease.

  19. A continuum model for excitation-contraction of smooth muscle under finite deformations.

    Science.gov (United States)

    Sharifimajd, Babak; Stålhand, Jonas

    2014-08-21

    The main focus in most of the continuum based muscle models is the mechanics of muscle contraction while other physiological processes governing muscle contraction, e.g., cell membrane excitation and activation, are ignored. These latter processes are essential to initiate contraction and to determine the amount of generated force, and by excluding them, the developed model cannot replicate the true behavior of the muscle in question. The aim of this study is to establish a thermodynamically and physiologically consistent framework which allows us to model smooth muscle contraction by including cell membrane excitability and kinetics of myosin phosphorylation, along with dynamics of smooth muscle contraction. The model accounts for these processes through a set of coupled dissipative constitutive equations derived by applying first principles. To show the performance of the derived model, it is evaluated for two different cases: a chemo-mechanical study of pig taenia coli cells where the excitation process is excluded, and an electro-chemo-mechanical study of rat myometrium. The results show that the model is able to replicate important aspects of the smooth muscle excitation-contraction process.

  20. Value of counting positive PHH3 cells in the diagnosis of uterine smooth muscle tumors.

    Science.gov (United States)

    Pang, Shu-Jie; Li, Cheng-Cheng; Shen, Yan; Liu, Yian-Zhu; Shi, Yi-Quan; Liu, Yi-Xin

    2015-01-01

    The diagnosis of uterine smooth muscle tumors including leiomyosarcomas (LMS), smooth muscle tumors of uncertain malignant potential (STUMP), bizarre (atypical) leiomyoma (BLM), mitotically active leiomyoma (MAL) and leiomyoma (LM) depends on a combination of microscopic features, such as mitoses, cytologic atypia, and coagulative tumor cell necrosis. However, a small number of these tumors still pose difficult diagnostic challenges. The assessment of accurate mitotic figures (MF) is one of the major parameters in the proper classification of uterine smooth muscle tumors. This assessment can be hampered by the presence of increased number of apoptotic bodies or pyknotic nuclei, which frequently mimic mitoses. Phospho-histone H3 (PHH3) is a recently described immunomarker specific for cells undergoing mitoses. In our study, we collected 132 cases of uterine smooth muscle tumors, including 26 LMSs, 16 STUMPs, 30 BLMs, 30 MALs and 30 LMs. We used mitosis specific marker PHH3 to count mitotic indexes (MI) of uterine smooth muscle tumors and compared with the mitotic indexes of hematoxylin and eosin (H&E). There is a positive correlation with the number of mitotic figures in H&E-stained sections and PHH3-stained sections (r=0.944, P0.05). The counting value of PHH3 in LMSs have significantly higher than STUMPs, BLMs, MALs and LMs (Pnumber of mitotic indexes in H&E. To conclude, our results show that counting PHH3 is a useful index in the diagnosis of uterine smooth muscle tumors and it can provide a more accurate index instead of the time-honored mitotic figure counts at a certain ratio.

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

    Science.gov (United States)

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

    2017-02-01

    The role of smooth muscle 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.

  2. Smooth muscle cell-derived carbon monoxide is a regulator of vascular cGMP.

    OpenAIRE

    Morita, T.; Perrella, M A; Lee, M E; Kourembanas, S

    1995-01-01

    Carbon monoxide (CO) is a product of the enzyme heme oxygenase (HO; EC 1.14.99.3). In vascular smooth muscle cells, exogenously administered CO increases cyclic guanosine 3',5'-monophosphate (cGMP), which is an important regulator of vessel tone. We report here that smooth muscle cells produce CO via HO and that it regulates cGMP levels in these cells. Hypoxia, which has profound effects on vessel tone, significantly increased the transcriptional rate of the HO-1 gene resulting in correspondi...

  3. Integrin mobilizes intracellular Ca(2+) in renal vascular smooth muscle cells

    DEFF Research Database (Denmark)

    Chan, W L; Holstein-Rathlou, N H; Yip, K P

    2001-01-01

    Peptides with the Arg-Gly-Asp (RGD) motif induce vasoconstriction in rat afferent arterioles by increasing the intracellular Ca(2+) concentration ([Ca(2+)](i)) in vascular smooth muscle cells (VSMC). This finding suggests that occupancy of integrins on the plasma membrane of VSMC might affect...... vascular tone. The purpose of this study was to determine whether occupancy of integrins by exogenous RGD peptides initiates intracellular Ca(2+) signaling in cultured renal VSMC. When smooth muscle cells were exposed to 0.1 mM hexapeptide GRGDSP, [Ca(2+)](i) rapidly increased from 91 +/- 4 to 287 +/- 37 n...

  4. Steroids and antihistamines synergize to inhibit rat's airway smooth muscle contractility.

    Science.gov (United States)

    Liu, Shao-Cheng; Chu, Yueng-Hsiang; Kao, Chuan-Hsiang; Wu, Chi-Chung; Wang, Hsing-Won

    2015-06-01

    Both glucocorticoids and H1-antihistamines were widely used on patients with allergic rhinitis (AR) and obstructive airway diseases. However, their direct effects on airway smooth muscle were not fully explored. In this study, we tested the effectiveness of prednisolone (Kidsolone) and levocetirizine (Xyzal) on isolated rat trachea submersed in Kreb's solution in a muscle bath. Changes in tracheal contractility in response to the application of parasympathetic mimetic agents were measured. The following assessments of the drug were performed: (1) effect on tracheal smooth muscle resting tension; (2) effect on contraction caused by 10(-6) M methacholine; (3) effect of the drug on electrical field stimulation (EFS) induced tracheal smooth muscle contractions. The result revealed sole use of Kidsolone or Xyzal elicited no significant effect or only a little relaxation response on tracheal tension after methacholine treatment. The tension was 90.5 ± 7.5 and 99.5 ± 0.8 % at 10(-4) M for Xyzal and 10(-5) M for Kidsolone, respectively. However, a dramatically spasmolytic effect was observed after co-administration of Kidsolone and Xyzal and the tension dropped to 67.5 ± 13.6 %, with statistical significance (p antihistamines to dramatically relax the trachea smooth muscle within minutes. Therefore, for AR patients with acute asthma attack, combined use of those two drugs is recommended.

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

    Energy Technology Data Exchange (ETDEWEB)

    Salvucci, Fernando Pablo; Schiavone, Jonathan; Craiem, Damian; Barra, Juan Gabriel [Facultad de Ingenieria y Ciencias Exactas y Naturales, Universidad Favaloro Av. Belgrano 1723 - Buenos Aires (Argentina)

    2007-11-15

    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 {eta}. 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 {eta}. 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 {eta} were calculated in control state and during muscle activation. The elastic modulus E did not present significant changes with heart rate. The viscous modulus {eta} decreased 49% with a two-fold acceleration in heart rate from 80 to 160 bpm. However, the product {eta} HR remained stable. The viscous modulus {eta} 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 {eta} when HR increased might be related to smooth muscle relaxation mediated by endothelium activity, which was stimulated by flow increase. We conclude that HR can modulate arterial wall viscoelasticity through endothelium-dependent mechanisms.

  6. Feasibility of organ preservation in muscle-invasive transitional cell carcinoma bladder: A single institutional approach

    Directory of Open Access Journals (Sweden)

    Chhaya Roy

    2015-01-01

    Full Text Available Background: Trimodality treatment initial transurethral resection of the bladder tumor [TURBT] followed by concurrent chemotherapy and radiation and organ preservation have been gradually replacing the radical cystectomy in muscle-invasive transitional cell carcinoma (TCC of bladder. Aims: The aims of this study is to determine the clinical effectiveness, safety and protocol completion rate of trimodality treatment in muscle-invasive TCC of the bladder. Settings and Design: Prospective randomized and open-labeled study. Subjects and Methods: Patients with TCC of bladder, American Joint Committee on Cancer tumor node metastasis (TNM Bladder Cancer Staging (2002 T2-3, N0, M0. Were underwent TURBT followed by three cycles of neoadjuvant chemotherapy with methotrexate, vinblastine, adriamycin, and cisplatin regimen. The patients were then randomized to receive either concurrent cisplatin 75 mg/m 2 in week 1 and 4 (arm-A or no cisplatin (arm-B along with external beam radiation therapy (EBRT 45 Gy, in 25 fractions over 5 weeks. 4 weeks after completion of the initial phase of treatment, all patients were re-evaluated with TURBT. Those with complete remission (CR received additional 15 Gy of EBRT in 8 fractions, while patients with residual disease were recommended for immediate radical cystectomy. All the patients of arm-B received boost dose of 15 Gy of EBRT. Statistical Analysis Used: The major statistical endpoints of this study were the CR rate at 8 weeks post-concurrent chemoradiotherapy (CCRT and only radiotherapy. Statistical significance was accepted at the P < 0.05 (two-sided level. Statistical analysis was performed entirely using the Statistical Package for the Social Sciences for Windows, version 17 (SPSS Inc., Chicago, IL, U.S.A.. Results: 8 weeks after completion of treatment 13/16 (81% patients were in CR in CCRT arm (arm-A compare to 6/15 (40% patients receiving radiation only (arm-B. Conclusions: Patients, after TURBT receiving CCRT

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

    Energy Technology Data Exchange (ETDEWEB)

    Kumar, V.S.; Robinson, H.; O-Neall-Hennessey, E.; Reshetnikova, L.; Brown, J. H.; Szent-Gyorgyi, A. G.; Cohen, C.

    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 result provides additional evidence that phosphorylation of the RLC is unlikely to act as an on-switch in regulation of scallop catch muscle myosin.

  8. Pharmacomechanical coupling in smooth muscle may involve phosphatidylinositol metabolism.

    OpenAIRE

    Baron, C B; Cunningham, M.; Strauss, J F; Coburn, R F

    1984-01-01

    Cholinergic contraction of canine trachealis muscle, a contraction that primarily utilizes membrane potential-independent mechanisms for activating contractile proteins (pharmacomechanical coupling), is associated with a decline in the phosphatidylinositol pool, an increase in the phosphatidic acid and diacylglycerol pools, and an increased incorporation of 32PO4 into phosphatidylinositol. We found that these changes occur during development of the contraction and during maintenance of tensio...

  9. Infiltration of hypertrophic esophageal smooth muscle by mast cells and basophils.

    Science.gov (United States)

    Tung, H N; Schulze-Delrieu, K; Shirazi, S

    1993-01-01

    Partial obstruction leads to chronic distension and muscular hypertrophy of the opossum esophagus. The smooth muscle cells of the circular muscle layer enlarge, become pleomorphic and are surrounded by an amorphous ground substance in the extracellular space. Here we describe the histological and ultrastructural features of a peculiar cellular infiltrate in the hypertrophic smooth muscle. The infiltrate consisted uniquely of mast cells and basophils. In per unit area, the number of mast cells increased from 0.9 +/- 0.1 cells in controls to 3.7 +/- 0.2 in hypertrophic smooth muscle; the corresponding numbers for basophils were 2.5 +/- 0.2 and 7.2 +/- 0.3 cells. Cells were seen primarily in the septal spaces of the circular muscle layer and at the interface of the circular and longitudinal muscle layer. The cytoplasm of basophils is normally packed with round and oval granules. The granules stain metachromatically and with varying intensity on Wright-Giemsa stains. On transmission electronmicroscopy, granules display a membrane and a great diversity in the structure of their luminal contents. In hypertrophic muscle, most granules were discharging their contents into the cytoplasm or extracellular space. The membranes of adjacent empty granules then fused to form a chain of vacuoles. Similar changes occurred also in the mast cells which differed from the basophil by their lack of nuclear lobulation and by the greater homogeneity of their cytoplasmic granules. It is possible that these inflammatory cells are involved in the reconstruction of the smooth muscle and its connective tissue which occur during esophageal distension and hypertrophy.

  10. Treatment results of radiation therapy for muscle-invasive bladder cancer

    Energy Technology Data Exchange (ETDEWEB)

    Langsenlehner, Tanja; Doeller, Carmen; Stranzl-Lawatsch, Heidi; Kapp, Karin S. [Univ. Clinic of Therapeutic Radiology and Oncology, Medical Univ. of Graz (Austria); Quehenberger, Franz [Inst. for Medical Informatics, Statistics and Documentation, Medical Univ. of Graz (Austria); Langsenlehner, Uwe [Internal Outpatient Dept., Steiermaerkische GKK, Graz (Austria); Pummer, Karl [Dept. of Urology, Medical Univ. of Graz (Austria)

    2010-04-15

    Purpose: To assess local control and survival rates in patients with muscle-invasive bladder cancer treated with external-beam radiotherapy and to investigate prognostic factors. Patients and methods: Between 1997 and 2007, 75 patients (male, n = 58; female, n = 17, median age, 74.2 years) with localized transitional cell carcinoma of the bladder (T2, n = 34; T3, n = 32; T4, n = 9) not suitable for radical surgery due to advanced age, comorbidity or inoperability underwent external-beam radiotherapy without simultaneous chemotherapy at the University Clinic of Therapeutic Radiology and Oncology, Medical University of Graz, Austria. A conformal four-field technique was used in all patients to treat the tumor and regional lymph nodes with single daily fractions of 1.8-2 Gy to a total dose of 50-50.4 Gy, followed by a cone-down to encompass the empty bladder which was boosted to 70-70.4 Gy. All patients had undergone transurethral tumor resection prior to radiotherapy which was macroscopically incomplete in 62 patients. Results: Complete response was achieved in 65% of patients. Actuarial 3-year local control and metastases-free survival rates were 52.5% and 63.7%, 3-year local recurrence-free survival rate in complete responders was 71%. In univariate analysis, hydronephrosis, lymph vessel invasion, and macroscopic residual tumor were significantly predictive of disease progression. Hydronephrosis and lymph vessel invasion were also associated with a higher risk of local recurrence. The actuarial 3-year progression-free and overall survival rates were 40.1% and 56.9%, respectively. Conclusion: Radiotherapy is an effective treatment option in terms of local control and survival even in elderly patients with locally advanced bladder cancer not suitable for cystectomy. (orig.)

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

    Science.gov (United States)

    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. Different muscarinic receptor subtypes modulate proliferation of primary human detrusor smooth muscle cells via Akt/PI3K and map kinases.

    Science.gov (United States)

    Arrighi, Nicola; Bodei, Serena; Zani, Danilo; Michel, Martin C; Simeone, Claudio; Cosciani Cunico, Sergio; Spano, Pierfranco; Sigala, Sandra

    2013-08-01

    While acetylcholine (ACh) and muscarinic receptors in the bladder are mainly known for their role in the regulation of smooth muscle contractility, in other tissues they are involved in tissue remodelling and promote cell growth and proliferation. In the present study we have used primary cultures of human detrusor smooth muscle cells (HDSMCs), in order to investigate the role of muscarinic receptors in HDSMC proliferation. Samples were obtained as discarded tissue from men >65 years undergoing radical cystectomy for bladder cancer and cut in pieces that were either immediately frozen or placed in culture medium for the cell culture establishment. HDSMCs were isolated from samples, propagated and maintained in culture. [(3)H]-QNB radioligand binding on biopsies revealed the presence of muscarinic receptors, with a Kd of 0.10±0.02nM and a Bmax of 72.8±0.1fmol/mg protein. The relative expression of muscarinic receptor subtypes, based on Q-RT-PCR, was similar in biopsies and HDSMC with a rank order of M2≥M3>M1>M4>M5. The cholinergic agonist carbachol (CCh, 1-100μM) concentration-dependently increased [(3)H]-thymidine incorporation (up to 46±4%). This was concentration-dependently inhibited by the general muscarinic receptor antagonist atropine and by subtype-preferring antagonists with an order of potency of darifenacin >4-DAMP>AF-DX 116. The CCh-induced cell proliferation was blocked by selective PI-3 kinase and ERK activation inhibitors, strongly suggesting that these intracellular pathways mediate, at least in part, the muscarinic receptor-mediated cell proliferation. This work shows that M2 and M3 receptors can mediate not only HDSM contraction but also proliferation; they may also contribute bladder remodelling including detrusor hypertrophy.

  13. Time-dependent effects of castration on the bladder function and histological changes in the bladder and blood vessels.

    Science.gov (United States)

    Magari, Tomohiro; Shibata, Yasuhiro; Arai, Seiji; Kashiwagi, Bunzo; Suzuki, Keiji; Suzuki, Kazuhiro

    2014-01-01

    We examined the effect of androgens on bladder blood flow (BBF), bladder function and histological changes in castrated male rats. Male Wistar rats were classified into unoperated group (control group), groups castrated at the age of 8 weeks (group 8wPC) and groups castrated at the age of 4 weeks (group 4wPC). Each rat was used at the age of 20 weeks. BBF was measured using fluorescent microspheres. Bladder cystometry was performed without anesthesia or restraint; the bladder was first irrigated with saline and then with 0.25% acetic acid (AA) solution. Maximum voiding pressure and voiding interval were measured. The bladder and iliac artery were histologically examined for differences in smooth muscle and quantity of collagen fiber to analyze the effect of castration on the smooth muscle content. No differences were noted in BBF following castration. The voiding intervals for all groups were shortened (P control group (Pcontrol group (Pblood vessels.

  14. Extracellular matrix proteins modulate asthmatic airway smooth muscle cell proliferation via an autocrine mechanism

    NARCIS (Netherlands)

    Johnson, Peter R A; Burgess, Janette K; Underwood, P Anne; Au, Wendy; Poniris, Maree H; Tamm, Michael; Ge, Qi; Roth, Michael; Black, Judith L

    2004-01-01

    BACKGROUND: Airway remodeling is a key feature of persistent asthma and includes alterations in the extracellular matrix protein profile around the airway smooth muscle (ASM) and hyperplasia of the ASM. We have previously shown that nonasthmatic ASM cells in culture produce a range of extracellular

  15. Molecular and functional characterization of Kv7 K+ channel in murine gastrointestinal smooth muscles

    DEFF Research Database (Denmark)

    Jepps, Thomas Andrew; Greenwood, Iain A; Moffatt, James D

    2009-01-01

    Members of the K(v)7 voltage-gated K(+) channel family are important determinants of cardiac and neuronal membrane excitability. Recently, we and others have shown that K(v)7 channels are also crucial regulators of smooth muscle activity. The aim of the present study was to assess the K(v)7 expre...

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

    DEFF Research Database (Denmark)

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

    2006-01-01

    Objective: The vascular extracellular matrix (ECM) can affect smooth muscle cell (SMC) adhesion, migration and proliferation-events that are important during the atherosclerotic process. Fibulin-2 is a member of the ECM protein family of fibulins and has been found to cross-link versican/hyaluron...

  17. Mounier-Kuhn syndrome: a case of tracheal smooth muscle remodeling.

    Science.gov (United States)

    Cook, Daniel P; Adam, Ryan J; Abou Alaiwa, Mahmoud H; Eberlein, Michael; Klesney-Tait, Julia A; Parekh, Kalpaj R; Meyerholz, David K; Stoltz, David A

    2017-02-01

    Mounier-Kuhn syndrome is a rare clinical disorder characterized by tracheobronchial dilation and recurrent lower respiratory tract infections. While the etiology of the disease remains unknown, histopathological analysis of Mounier-Kuhn airways demonstrates that the disease is, in part, characterized by cellular changes in airway smooth muscle.

  18. Croton sonderianus essential oil samples distinctly affect rat airway smooth muscle.

    Science.gov (United States)

    Pinho-da-Silva, L; Mendes-Maia, P V; do Nascimento Garcia, T M; Cruz, J S; de Morais, S M; Coelho-de-Souza, A N; Lahlou, S; Leal-Cardoso, J H

    2010-08-01

    Plants of the genus Croton have been used extensively in the northeast of Brazil for treating various clinical conditions. Previous studies have demonstrated that the essential oil of some specimens of Croton sp. have a relaxing effect on tracheal smooth muscle. Our study aimed to characterize the effects of Croton sonderianus essential oil samples, collected at 1:00 pm (EO-13) and 9:00 pm (EO-21), on rat tracheal smooth muscle. The two samples were submitted to gas chromatography (GC) and mass spectrometry (MS) analysis to identify their components. Rat tracheal smooth muscle strips were used to assess the biological activity. The major constituents of EO-21 were: spathulenol (18.32%), beta-caryophyllene (14.58%) and caryophyllene oxide (8.54%) and the major constituents of EO-13 were bicyclogermacrene (16.29%), beta-phellandrene (15.42%) and beta-caryophyllene (13.82%). These samples showed an antispasmodic effect on tracheal smooth muscle strips pre-contracted with high K+ concentration (80 mM) or with acetylcholine. EO-21 increased baseline tonus while EO-13 provoked a decrease. These results demonstrated that EO-13 and EO-21 have different chemical composition and showed myorelaxant activity. In conclusion, EO-13 and EO-21 may have potential therapeutic use in the treatment of bronchospasm.

  19. Allergic sensitization enhances the contribution of Rho-kinase to airway smooth muscle contraction

    NARCIS (Netherlands)

    Schaafsma, D.; Gosens, Reinout; Bos, I.S.T.; Meurs, Herman; Zaagsma, Hans; Nelemans, Herman

    2004-01-01

    1 Repeated allergen challenge has been shown to increase the role of Rho-kinase in airway smooth muscle (ASM) contraction. We considered the possibility that active allergic sensitization by itself, that is, without subsequent allergen exposure, could be sufficient to enhance Rho-kinase-mediated ASM

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

    NARCIS (Netherlands)

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

    2001-01-01

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

  1. Effect of pharmacologically induced smooth muscle activation on permeability in murine colitis

    Directory of Open Access Journals (Sweden)

    Freek J. Zijlstra

    2003-01-01

    Full Text Available Background: Both intestinal permeability and contractility are altered in inflammatory bowel disease. Little is known about their mutual relation. Therefore, an in vitro organ bath technique was developed to investigate the simultaneous effects of inflammation on permeability and smooth muscle contractility in different segments of the colon.

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

    Science.gov (United States)

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

    2016-08-10

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

  3. Glucocorticosteroids and beta(2)-Adrenoceptor Agonists Synergize to Inhibit Airway Smooth Muscle Remodeling

    NARCIS (Netherlands)

    Dekkers, Bart G. J.; Pehlic, Adnan; Mariani, Raissa; Bos, I. Sophie T.; Meurs, Herman; Zaagsma, Johan

    2012-01-01

    Airway remodeling, including increased airway smooth muscle (ASM) mass and contractility, contributes to increased airway narrowing in asthma. Increased ASM mass may be caused by exposure to mitogens, including platelet-derived growth factor (PDGF) and collagen type I, which induce a proliferative,

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

    Science.gov (United States)

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

    1989-02-01

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

  5. Airway smooth muscle dynamics : a common pathway of airway obstruction in asthma

    NARCIS (Netherlands)

    An, S S; Bai, T R; Bates, J H T; Black, J L; Brown, R H; Brusasco, V; Chitano, P; Deng, L; Dowell, M; Eidelman, D H; Fabry, B; Fairbank, N J; Ford, L E; Fredberg, J J; Gerthoffer, W T; Gilbert, S H; Gosens, R; Gunst, S J; Halayko, A J; Ingram, R H; Irvin, C G; James, A L; Janssen, L J; King, G G; Knight, D A; Lauzon, A M; Lakser, O J; Ludwig, M S; Lutchen, K R; Maksym, G N; Martin, J G; Mauad, T; McParland, B E; Mijailovich, S M; Mitchell, H W; Mitchell, R W; Mitzner, W; Murphy, T M; Paré, P D; Pellegrino, R; Sanderson, M J; Schellenberg, R R; Seow, C Y; Silveira, P S P; Smith, P G; Solway, J; Stephens, N L; Sterk, P J; Stewart, A G; Tang, D D; Tepper, R S; Tran, T; Wang, L

    2007-01-01

    Excessive airway obstruction is the cause of symptoms and abnormal lung function in asthma. As airway smooth muscle (ASM) is the effecter controlling airway calibre, it is suspected that dysfunction of ASM contributes to the pathophysiology of asthma. However, the precise role of ASM in the series o

  6. Pulmonary surfactant in the airway physiology: a direct relaxing effect on the smooth muscle.

    Science.gov (United States)

    Calkovska, A; Uhliarova, B; Joskova, M; Franova, S; Kolomaznik, M; Calkovsky, V; Smolarova, S

    2015-04-01

    Beside alveoli, surface active material plays an important role in the airway physiology. In the upper airways it primarily serves in local defense. Lower airway surfactant stabilizes peripheral airways, provides the transport and defense, has barrier and anti-edematous functions, and possesses direct relaxant effect on the smooth muscle. We tested in vitro the effect of two surfactant preparations Curosurf® and Alveofact® on the precontracted smooth muscle of intra- and extra-pulmonary airways. Relaxation was more pronounced for lung tissue strip containing bronchial smooth muscle as the primary site of surfactant effect. The study does not confirm the participation of ATP-dependent potassium channels and cAMP-regulated epithelial chloride channels known as CFTR chloride channels, or nitric oxide involvement in contractile response of smooth muscle to surfactant.By controlling wall thickness and airway diameter, pulmonary surfactant is an important component of airway physiology. Thus, surfactant dysfunction may be included in pathophysiology of asthma, COPD, or other diseases with bronchial obstruction.

  7. CD40 and OX40 ligand are differentially regulated on asthmatic airway smooth muscle

    NARCIS (Netherlands)

    Krimmer, D I; Loseli, M; Hughes, J M; Oliver, B G G; Moir, L M; Hunt, N H; Black, J L; Burgess, J K

    2009-01-01

    BACKGROUND: CD40 and OX40 Ligand (OX40L) are cell-surface molecules expressed on airway smooth muscle (ASM) that can enhance inflammatory cell activation and survival. The aim of this study was to examine the effect of tumour necrosis factor-alpha (TNF-alpha) and interferon-gamma (IFN-gamma) on ASM

  8. Comparison of gel contraction mediated by airway smooth muscle cells from patients with and without asthma

    NARCIS (Netherlands)

    Matsumoto, Hisako; Moir, Lyn M; Oliver, Brian G G; Burgess, Janette K; Roth, Michael; Black, Judith L; McParland, Brent E

    2007-01-01

    BACKGROUND: Exaggerated bronchial constriction is the most significant and life threatening response of patients with asthma to inhaled stimuli. However, few studies have investigated the contractility of airway smooth muscle (ASM) from these patients. The purpose of this study was to establish a me

  9. Connective tissue growth factor induces extracellular matrix in asthmatic airway smooth muscle

    NARCIS (Netherlands)

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

    2006-01-01

    Transforming growth factor (TGF)-beta and connective tissue growth factor may be implicated in extracellular matrix protein deposition in asthma. We have recently reported that TGF-beta increased connective tissue growth factor expression in airway smooth muscle cells isolated from patients with ast

  10. The effect of asthma therapeutics on signalling and transcriptional regulation of airway smooth muscle function

    NARCIS (Netherlands)

    Ammit, Alaina J; Burgess, Janette K; Hirst, Stuart J; Hughes, J Margaret; Kaur, Manminder; Lau, Justine Y; Zuyderduyn, Suzanne

    2009-01-01

    SCOPE OF THE REVIEW: Our knowledge of the multifunctional nature of airway smooth muscle (ASM) has expanded rapidly in the last decade, but the underlying molecular mechanisms and how current therapies for obstructive airway diseases, such as asthma and chronic obstructive pulmonary disease (COPD),

  11. Regulation of actin dynamics by wnt-5a : Implications for human airway smooth muscle contraction

    NARCIS (Netherlands)

    Koopmans, Tim; Kumawat, Kuldeep; Menzen, Mark; Halayko, Andrew; Gosens, Reinoud

    2016-01-01

    An important pathophysiological feature of asthma is airway hyperresponsiveness (AHR), characterized by exaggerated bronchoconstriction in which the airway smooth muscle (ASM) is fundamentally involved. How the ASM in asthmatics differs from that in non-asthmatics is a current focus for research. We

  12. A mechanism for arteriolar remodeling based on maintenance of smooth muscle cell activation

    DEFF Research Database (Denmark)

    Jacobsen, Jens Christian Brings; Mulvany, Michael John; Holstein-Rathlou, N.-H.

    2008-01-01

    Structural adaptation in arterioles is part of normal vascular physiology but is also seen in disease states such as hypertension. Smooth muscle cell (SMC) activation has been shown to be central to microvascular remodeling. We hypothesize that, in a remodeling process driven by SMC activation...

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

    DEFF Research Database (Denmark)

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

    2016-01-01

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

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

    DEFF Research Database (Denmark)

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

    2007-01-01

    in the transformation of human vascular smooth muscle cells (HVSMC) to osteoblast-like cells. In this study, we evaluated the effect of BMP-2, BMP-7 and transforming growth factor beta (TGF-beta1) on the secretion and mRNA expression of OPG and its ligands receptor activator of nuclear factor-kappabeta ligand (RANKL...

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

    Energy Technology Data Exchange (ETDEWEB)

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

    2015-08-15

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

  16. High glucose enhance expression of matrix metalloproteinase—2 in smooth muscle cells

    Institute of Scientific and Technical Information of China (English)

    HAOFeng; YUJin-De

    2003-01-01

    AIM:To investigate the effects of high glucose on expression of matrix metalloproteinase-2(MMP-2) in rat aortic smooth muscle cells and the influence of matrix remodeling on atherogenesis in diabetic patients. METHODS: The smooth muscle cells were cultured from the thoracic aorta of Sprague-Dawley (SD) rat. MMP-2 mRNA was determined by reverse transcriptase-polymerase chain reaction(RT-PCR),MMP-2 protein was measured by Western blotting, and MMP-2 activity in conditioned medium was observed by zymography. RESULTS:In comparison with the control, there was no difference in the expression of MMP-2 when glucose concentration was 1g/L,whereas MMP-2 activity in smooth muscle cells was significantly increased by the glucose 5 g/L(P<0.01). CONCLUSION:High glucose enhanced the expression and activity of MMP-2 in smooth muscle cells, which may provide an explanation for the phenomenon that diabetes patients are prone to have atherosclerotic lesions.

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

    DEFF Research Database (Denmark)

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

    2012-01-01

    Micro-anatomical structures in tissues have potential physiological effects. In arteries and arterioles smooth muscle cells and endothelial cells are separated by the internal elastic lamina, but the two cell layers often make contact through micro protrusions called myoendothelial junctions. Cross...

  18. Functional characterization of serum- and growth factor-induced phenotypic changes in intact bovine tracheal smooth muscle

    NARCIS (Netherlands)

    Gosens, R; Meurs, H; Bromhaar, MMG; McKay, S; Nelemans, SA; Zaagsma, J

    2002-01-01

    1 The present study aims to investigate whether phenotypic changes, reported to occur in cultured isolated airway smooth muscle (ASM) cells, are of relevance to intact ASM. Moreover, we aimed to gain insight into the signalling pathways involved. 2 Culturing of bovine tracheal smooth muscle (BTSM) s

  19. Effect of fenoterol-induced constitutive beta(2)-adrenoceptor activity on contractile receptor function in airway smooth muscle

    NARCIS (Netherlands)

    de Vries, B; Roffel, AF; Zaagsma, J; Meurs, H

    2001-01-01

    In the present study, we investigated the effect of fenoterol-induced constitutive beta (2)-adrenoceptor activity on muscarinic receptor agonist- and histamine-induced bovine tracheal smooth muscle contractions. Bovine tracheal smooth muscle strips were incubated with 10 muM fenoterol or vehicle for

  20. Effects of fenoterol on beta-adrenoceptor and muscarinic M-2 receptor function in bovine tracheal smooth muscle

    NARCIS (Netherlands)

    De Vries, B; Roffel, AF; Kooistra, JM; Meurs, H; Zaagsma, J

    2001-01-01

    Prolonged (18 h) incubation of isolated bovine tracheal smooth muscle with the beta (2)-adrenoceptor agonist fenoterol (10 muM) induced desensitization of isoprenaline-induced adenylyl cyclase activity in bovine tracheal smooth muscle membranes, characterized by a 25% decrease in maximal effect (E-m

  1. [Effect of adrenaline on the proliferation of the tunica media smooth muscle cells of rat aorta in culture].

    Science.gov (United States)

    Blaes, N; Bourdillon, M C; Crouzet, B; Suplisson, A; Boissel, J P

    1980-03-24

    The proliferation of Rat medial aortic smooth muscle cells in secondary cultures is increased with adrenalin. The maximal effect is obtained after 3 days and the increase is dose-dependent. Thus adrenalin might be one of the factors responsible for the proliferation of smooth muscle cells that could play a key role in the formation of the atherosclerotic plaque in vivo.

  2. File list: ALL.CDV.20.AllAg.Coronary_artery_smooth_muscle [Chip-atlas[Archive

    Lifescience Database Archive (English)

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  3. File list: Oth.CDV.50.AllAg.Coronary_artery_smooth_muscle [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Oth.CDV.50.AllAg.Coronary_artery_smooth_muscle hg19 TFs and others Cardiovascular Coronary arte...osciencedbc.jp/kyushu-u/hg19/assembled/Oth.CDV.50.AllAg.Coronary_artery_smooth_muscle.bed ...

  4. File list: ALL.CDV.05.AllAg.Coronary_artery_smooth_muscle [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available ALL.CDV.05.AllAg.Coronary_artery_smooth_muscle hg19 All antigens Cardiovascular Coronary arte...ttp://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/ALL.CDV.05.AllAg.Coronary_artery_smooth_muscle.bed ...

  5. File list: Oth.CDV.20.AllAg.Coronary_artery_smooth_muscle [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Oth.CDV.20.AllAg.Coronary_artery_smooth_muscle hg19 TFs and others Cardiovascular Coronary arte...osciencedbc.jp/kyushu-u/hg19/assembled/Oth.CDV.20.AllAg.Coronary_artery_smooth_muscle.bed ...

  6. File list: ALL.CDV.50.AllAg.Coronary_artery_smooth_muscle [Chip-atlas[Archive

    Lifescience Database Archive (English)

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  7. File list: ALL.CDV.10.AllAg.Coronary_artery_smooth_muscle [Chip-atlas[Archive

    Lifescience Database Archive (English)

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  8. File list: Oth.CDV.10.AllAg.Coronary_artery_smooth_muscle [Chip-atlas[Archive

    Lifescience Database Archive (English)

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  9. File list: Oth.CDV.05.AllAg.Coronary_artery_smooth_muscle [Chip-atlas[Archive

    Lifescience Database Archive (English)

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  10. PAT1 (SLC36A1) shows nuclear localization and affect growth of smooth muscle cells from rats

    DEFF Research Database (Denmark)

    Jensen, Anne; Figueiredo-Larsen, Evan Manuel; Holm, René

    2014-01-01

    the localization and function of PAT1 in smooth muscle cells (SMCs). The PAT1 protein was found in smooth muscles from rat intestine and in the embryonic rat aorta cell line A7r5. Immunolocalization and cellular fractionation studies revealed that the majority of the PAT1 protein located within the cell nucleus...

  11. Rapid effects of phytoestrogens on human colonic smooth muscle are mediated by oestrogen receptor beta.

    LENUS (Irish Health Repository)

    Hogan, A M

    2012-02-01

    Epidemiological studies have correlated consumption of dietary phytoestrogens with beneficial effects on colon, breast and prostate cancers. Genomic and non-genomic mechanisms are responsible for anti-carcinogenic effects but, until now, the effect on human colon was assumed to be passive and remote. No direct effect on human colonic smooth muscle has previously been described. Institutional research board approval was granted. Histologically normal colon was obtained from the proximal resection margin of colorectal carcinoma specimens. Circular smooth muscle strips were microdissected and suspended under 1g of tension in organ baths containing oxygenated Krebs solution at 37 degrees C. After an equilibration period, tissues were exposed to diarylpropionitrile (DPN) (ER beta agonist) and 1,3,5-tris(4-hydroxyphenyl)-4-propyl-1H-pyrazole (PPT) (ER alpha agonist) or to the synthetic phytoestrogen compounds genistein (n=8), daidzein (n=8), fisetin (n=8) and quercetin (n=8) in the presence or absence of fulvestrant (oestrogen receptor antagonist). Mechanism of action was investigated by inhibition of downstream pathways. The cholinergic agonist carbachol was used to induce contractile activity. Tension was recorded isometrically. Phytoestrogens inhibit carbachol-induced colonic contractility. In keeping with a non-genomic, rapid onset direct action, the effect was within minutes, reversible and similar to previously described actions of 17 beta oestradiol. No effect was seen in the presence of fulvestrant indicating receptor modulation. While the DPN exerted inhibitory effects, PPT did not. The effect appears to be reliant on a p38\\/mitogen activated protein kinase mediated induction of nitric oxide production in colonic smooth muscle. The present data set provides the first description of a direct effect of genistein, daidzein, fisetin and quercetin on human colonic smooth muscle. The presence of ER in colonic smooth muscle has been functionally proven and the beta

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

    Science.gov (United States)

    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.

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

    Science.gov (United States)

    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.

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

    Energy Technology Data Exchange (ETDEWEB)

    Stepp, M.A.

    1986-01-01

    Aortic smooth muscles cells have been implicated in the etiology of lesions which occur in atherosclerosis and hypertension. Both diseases involve proliferation of smooth muscle cells and accumulation of excessive amounts of extracellular matrix proteins, including collagen type I and type III produced by the smooth muscle cells. To better understand the sites of regulation of collagen biosynthesis and to correlate these with the growth rate of the cells, cultured bovine aortic smooth muscle cells were studied as a function of the number of days (3 to 14) in second passage. Cells grew rapidly up to day 6 when confluence was reached. The total incorporation of (/sup 3/H)-proline into proteins was highest at day 3 and decreased to a constant level after the cultures reached confluence. In contrast, collagen protein production was lowest before confluence and continued to increase over the entire time course of the experiments. cDNA clones for the ..cap alpha..1 and ..cap alpha..2 chains of type I and the ..cap alpha..1 chain of type III collagen were used to quantitate the steady state level of collagen mRNAs. RNA was tested in a cell-free translation system. Changes in the translational activity of collagen mRNAs parallelled the observed increases in collagen protein production. Thus, at later time points, collagen mRNAs are more active in directing synthesis of preprocollagens, even though less collagen mRNA is present. The conclusion is that the site of regulation of the expression of collagen genes is a function of the growth rate of cultured smooth muscle cells.

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

    Science.gov (United States)

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

    2015-02-01

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

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

    Institute of Scientific and Technical Information of China (English)

    Iain L O BUXTON; Nathanael HEYMAN; Yi-ying WU; Scott BARNETT; Craig ULRICH

    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.

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

    Science.gov (United States)

    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.

  18. Diverse expression of delayed rectifier K+ channel subtype transcripts in several types of smooth muscles of the rat.

    Science.gov (United States)

    Ohya, S; Tanaka, M; Watanabe, M; Maizumi, Y

    2000-06-01

    Diverse expression of voltage dependent K+ (Kv) channels was examined in smooth muscles (SMs); carotid artery (CA), mesenteric artery (MA), urinary bladder (UB), and vas deferens (VD) of the rat, using RT-PCR based analyses. Among eight Kv channel subtypes examined (Kv 1.1, Kv 1.2, Kv 1.5, Kv 1.6, Kv 2.1, Kv 2.2, Kv 3.1, and Kv 3.2), expression of three delayed rectifier Kv (KD) channel (Kv 1.2, Kv 1.5, and Kv 2.1) transcripts was observed in these SMs. To determine precisely the expression levels of the transcripts encoding K(D) subtypes, those of three K(D) subtypes (Kv 1.2, Kv 1.5, and Kv 2.1) were determined by competitive PCR. In vascular SM tissues, CA and MA, Kv 1.2 and Kv 1.5 transcripts were expressed at relatively high levels, whereas in visceral SM tissues, UB and VD, Kv 2.1 transcripts were expressed at the relatively high levels. These results suggest that the diverse expression of K(D) subtypes is, at least in part, responsible for differences in electrical excitability and also for the variation of the electrophysiological and pharmacological phenotypes as tonic and phasic SMs.

  19. Bladder contractility is modulated by Kv7 channels in pig detrusor

    DEFF Research Database (Denmark)

    Svalø, Julie; Bille, Michala; Parameswaran Theepakaran, Neeraja;

    2013-01-01

    Kv7 channels are involved in smooth muscle relaxation, and accordingly we believe that they constitute potential targets for the treatment of overactive bladder syndrome. We have therefore used myography to examine the function of Kv7 channels in detrusor, i.e. pig bladder, with a view...

  20. Chromosomal imbalance in the progression of high-risk non-muscle invasive bladder cancer

    Directory of Open Access Journals (Sweden)

    Ørntoft Torben

    2009-05-01

    Full Text Available Abstract Background Non-muscle invasive bladder neoplasms with invasion of the lamina propria (stage T1 or high grade of dysplasia are at "high risk" of progression to life-threatening cancer. However, the individual course is difficult to predict. Chromosomal instability (CI is associated with high tumor stage and grade, and possibly with the risk of progression. Methods To investigate the relationship between CI and subsequent disease progression, we performed a case-control-study of 125 patients with "high-risk" non-muscle invasive bladder neoplasms, 67 with later disease progression, and 58 with no progression. Selection criteria were conservative (non-radical resections and full prospective clinical follow-up (> 5 years. We investigated primary lesions in 59, and recurrent lesions in 66 cases. We used Affymetrix GeneChip® Mapping 10 K and 50 K SNP microarrays to evaluate genome wide chromosomal imbalance (loss-of-heterozygosity and DNA copy number changes in 48 representative tumors. DNA copy number changes of 15 key instability regions were further investigated using QPCR in 101 tumors (including 25 tumors also analysed on 50 K SNP microarrays. Results Chromosomal instability did not predict any higher risk of subsequent progression. Stage T1 and high-grade tumors had generally more unstable genomes than tumors of lower stage and grade (mostly non-primary tumors following a "high-risk" tumor. However, about 25% of the "high-risk" tumors had very few alterations. This was independent of subsequent progression. Recurrent lesions represent underlying field disease. A separate analysis of these lesions did neither reflect any difference in the risk of progression. Of specific chromosomal alterations, a possible association between loss of chromosome 8p11 and the risk of progression was found. However, the predictive value was limited by the heterogeneity of the changes. Conclusion Chromosomal instability (CI was associated with "high risk

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

    Science.gov (United States)

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

    2016-01-15

    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.

  2. Signal transduction of bombesin-induced circular smooth muscle cell contraction in cat esophagus

    Institute of Scientific and Technical Information of China (English)

    Sung-Uk Park; Chang-Yell Shin; Jung-Su Ryu; Hyen-O La; Sun-Young Park; Hyun-Ju Song; Young-Sil Min; Dong-Seok Kim; Uy-Dong Sohn

    2006-01-01

    AIM: To investigate the mechanism of bombesin-induced circular smooth muscle cell contraction in cat esophagus.METHODS: Specific G protein or phospholipase C involved in cat esophagus contraction was identified,muscle cells were permeabilized with saponin. After permeabilization of muscle cells, the Gi3 antibody inhibited bombesin-induced smooth muscle cell contraction.RESULTS: Incubation of permeabilized circular muscle cells with PLC-β3 antibody could inhibit bombesin-induced contraction. H-7, chelerythrine (PKC inhibitor)and genistein (protein tyrosine kinase inhibitor) inhibited bombesin-induced contraction, but DAG kinase inhibitor,R59949, could not inhibit it. To examine which mitogenactivated protein kinase (MAPK) was involved in bombesin-induced contraction, the specific MAPK inhibitors (MEK inhibitor, PD98059 and p38 MAPK inhibitor, SB202190)were used. Preincubation of PD98059 blocked the contraction induced by bombesin in a concentration-dependent manner. However, SB202190 had no effects on contraction.CONCLUSION: Bombesin-induced circular muscle cell contraction in cat esophagus is madiated via a PKC or a PTK-dependent pathway or p44/p42 MAPK pathway.

  3. A novel bioreactor to simulate urinary bladder mechanical properties and compliance for bladder functional tissue engineering

    Institute of Scientific and Technical Information of China (English)

    WEI Xin; LI Dao-bing; XU Feng; WANG Yan; ZHU Yu-chun; LI Hong; WANG Kun-jie

    2011-01-01

    Background Bioreactors are pivotal tools for generating mechanical stimulation in functional tissue engineering study.This study aimed to create a bioreactor that can simulate urinary bladder mechanical properties, and to investigate the effects of a mechanically stimulated culture on urothelial cells and bladder smooth muscle cells.Methods We designed a bioreactor to simulate the mechanical properties of bladder. A pressure-record system was used to evaluate the mechanical properties of the bioreactor by measuring the pressure in culture chambers. To test the biocompatibility of the bioreactor, viabilities of urothelial cells and smooth muscle cells cultured in the bioreactor under static and mechanically changed conditions were measured after 7-day culture. To evaluate the effect of mechanical stimulations on the vital cells, urethral cells and smooth muscle cells were cultured in the simulated mechanical conditions. After that, the viability and the distribution pattern of the cells were observed and compared with cells cultured in non-mechanical stimulated condition.Results The bioreactor system successfully generated waveforms similar to the intended programmed model while maintaining a cell-seeded elastic membrane between the chambers. There were no differences between viabilities of urothelial cells ((91.90±1.22)% vs. (93.14±1.78)%, P >0.05) and bladder smooth muscle cells ((93.41±1.49)% vs.(92.61±1.34)%, P >0.05). The viability of cells and tissue structure observation after cultured in simulated condition showed that mechanical stimulation was the only factor affected cells in the bioreactor and improved the arrangement of cells on silastic membrane.Conclusions This bioreactor can effectively simulate the physiological and mechanical properties of the bladder.Mechanical stimulation is the only factor that affected the viability of cells cultured in the bioreactor. The bioreactor can change the growth behavior of urothelial cells and bladder smooth

  4. Signal pathways involved in emodin-induced contraction of smooth muscle cells from rat colon

    Institute of Scientific and Technical Information of China (English)

    Tao Ma; Qing-Hui Qi; Jian Xu; Zuo-Liang Dong; Wen-Xiu Yang

    2004-01-01

    AIM: To investigate the effects induced by emodin on single smooth muscle cells from rat colon in vitro, and to determine the signal pathways involved.METHODS: Cells were isolated from the muscle layers of Wistar rat colon by enzymatic digestion. Cell length was measured by computerized image micrometry. Intracellular Ca2+ ([Ca2+]i) signals were studied using the fluorescent Ca2+ indicator fluo-3 and confocal microscopy. PKCα distribution at rest state or after stimulation was measured with immunofluorescence confocal microscopy.RESULTS: (1) Emodin dose-dependently caused colonic smooth muscle cells contraction; (2) emodin induced an increase in intracellular Ca2+ concentration; (3) the contractile responses induced by emodin were respectively inhibited by preincubation of the cells with ML-7 (an inhibitorof MLCK)and calphostin C (an inhibitor of PKC); (4) Incubation of cells with emodin caused translocation of PKCα from cytosolic area to the membrane.CONCLUSION: Emodin has a direct contractile effect on colonic smooth muscle cell. This signal cascade induced by emodin is initiated by increased [Ca2+]i and PKCα translocation,which in turn lead to the activation of MLCK and the suppression of MLCP. Both of them contribute to the emodininduced contraction.

  5. Laminin on urinary bladder matrix promotes proliferation of bone marrow mesenchymal stem cells and differentiation into smooth muscle cells%UBM材料上的Laminin蛋白促进负载骨髓间充质干细胞增殖和平滑肌分化

    Institute of Scientific and Technical Information of China (English)

    魏桂枝; 刘晶霞; 徐萍萍; 倪兵; 刘禄斌; 梁志清

    2015-01-01

    目的 探讨猪膀胱细胞外基质(urinary bladder matrix,UBM)上的Laminin蛋白对大鼠骨髓间充质干细胞(bone marrow mesenchymal stem cells,BMSCs)增殖和向平滑肌分化的影响.方法 采用机械分离法、酶消化法及冷冻干燥法等方法对新鲜猪膀胱进行脱细胞处理,采用蛋白与支架共孵育,制备出UBM+Laminin及UBM+Anti-Laminin复合支架.采用贴壁法培养BMSCs,流式细胞仪鉴定BMSCs.采用P3代BMSCs,分别接种于UBM+Laminin、UBM及UBM+Anti-Laminin上.采用CCK-8检测、HE染色观察及免疫组化鉴定等方法检测Laminin蛋白对BMSCs增殖、迁移和分化的影响.结果 流式细胞仪鉴定BMSCs CD29阳性表达率为98%,CD31阳性表达率为4.56%,CD34阳性率为3.30%.3种材料均能支持BMSCs生长,且随共孵育时间的延长,D(450)值逐渐升高,其中UBM+Laminin复合材料组细胞D(450)最高(P<0.01).HE染色结果显示UBM+ Laminin组细胞接种第3天形成黏附层,并向深部迁移,第14天可达材料致密层;UBM组第7天仅少许细胞向深层迁移,第14天未达基底层;UBM+Anti-Laminin组仅在支架表层生长.免疫组化检测显示UBM+Laminin组、UBM组、UBM+ Anti-Laminin组细胞α-SMA表达分别呈阳性、弱阳性和阴性.结论 UBM材料上Laminin蛋白能促进负载BMSCs增殖、迁移、并有诱导其向平滑肌方向分化的作用.

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

    Science.gov (United States)

    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

  7. Radiotherapy concurrent with weekly gemcitabine after transurethral tumor resection in muscle ınvasive bladder cancer

    Directory of Open Access Journals (Sweden)

    Umut Demirci

    2015-01-01

    Full Text Available Objective: In this report, we determined the efficacy and the toxicity of low dose weekly gemcitabine with radiotherapy, in medically unfit or refused surgery muscle-invasive bladder cancer (BC patients. Materials and Methods: From 2008 to 2012, 15 patients were included into the retrospective analysis. Weekly gemcitabine was administered at a rate of 50 mg/m 2 with a median dose of 63 Gy radiotherapy. Results: The median age was 69 (range, 55-86. Median follow-up was 15 months (range, 5-53 months. A complete response was achieved in 12 patients (80%. Median progression free survival and overall survival were 15 months (range, 7-23 months and 18 months (range not calculated, respectively. Local recurrence was found in 3 patients (20% and distant recurrence was found in 5 patients (33.3% for the entire group. While salvage surgery was performed on 1 patient, salvage chemotherapy was delivered for 4 patients. Treatment was well tolerated, there was no treatment interruption or instances of toxic death. A serious toxicity (grade 3 cystitis was seen in only 1 patient. Conclusions: Multimodality treatment of muscle invasive BC proved a feasible and effective treatment option. Gemcitabine based chemoradiation is an active treatment option with a low toxicity profile for patients with muscle invasive BC, who are not suitable medically or refused to surgery.

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

    Science.gov (United States)

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

    2006-01-01

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

  9. Increased Expression of RhoA in Epithelium and Smooth Muscle of Obese Mouse Models: Implications for Isoprenoid Control of Airway Smooth Muscle and Fibroblasts

    Directory of Open Access Journals (Sweden)

    Kristie R. Ross

    2013-01-01

    Full Text Available The simultaneous rise in the prevalence of asthma and obesity has prompted epidemiologic studies that establish obesity as a risk factor for asthma. The alterations in cell signaling that explain this link are not well understood and warrant investigation so that therapies that target this asthma phenotype can be developed. We identified a significant increase in expression of the small GTPase RhoA in nasal epithelial cells and tracheal smooth muscle cells from leptin-deficient (ob/ob mice compared to their wild-type counterparts. Since RhoA function is dependent on isoprenoid modification, we sought to determine the role of isoprenoid-mediated signaling in regulating the viability and proliferation of human airway smooth muscle cells (ASM and normal human lung fibroblasts (NHLF. Inhibiting isoprenoid signaling with mevastatin significantly decreased the viability of ASM and NHLF. This inhibition was reversed by geranylgeranyl pyrophosphate (GGPP, but not farnesyl pyrophosphate (FPP, suggesting specificity to the Rho GTPases. Conversely, increasing isoprenoid synthesis significantly increased ASM proliferation and RhoA protein expression. RhoA expression is inherently increased in airway tissue from ob/ob mice, and obesity-entrained alterations in this pathway may make it a novel therapeutic target for treating airway disease in the obese population.

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

    Energy Technology Data Exchange (ETDEWEB)

    Josephson, Matthew P.; Sikkink, Laura A. [Department of Biochemistry and Molecular Biology, Mayo Clinic, Rochester, MN 55905 (United States); Penheiter, Alan R. [Molecular Medicine Program, Mayo Clinic, Rochester, MN 55905 (United States); Burghardt, Thomas P., E-mail: burghardt@mayo.edu [Department of Biochemistry and Molecular Biology, Mayo Clinic, Rochester, MN 55905 (United States); Department of Physiology and Biomedical Engineering, Mayo Clinic, Rochester, MN 55905 (United States); Ajtai, Katalin [Department of Biochemistry and Molecular Biology, Mayo Clinic, Rochester, MN 55905 (United States)

    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 chain 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 significant

  11. Role of Ryanodine Receptor Subtypes in Initiation and Formation of Calcium Sparks in Arterial Smooth Muscle: Comparison with Striated Muscle

    Directory of Open Access Journals (Sweden)

    Maik Gollasch

    2009-01-01

    Full Text Available Calcium sparks represent local, rapid, and transient calcium release events from a cluster of ryanodine receptors (RyRs in the sarcoplasmic reticulum. In arterial smooth muscle cells (SMCs, calcium sparks activate calcium-dependent potassium channels causing decrease in the global intracellular [Ca2+] and oppose vasoconstriction. This is in contrast to cardiac and skeletal muscle, where spatial and temporal summation of calcium sparks leads to global increases in intracellular [Ca2+] and myocyte contraction. We summarize the present data on local RyR calcium signaling in arterial SMCs in comparison to striated muscle and muscle-specific differences in coupling between L-type calcium channels and RyRs. Accordingly, arterial SMC Cav1.2 L-type channels regulate intracellular calcium stores content, which in turn modulates calcium efflux though RyRs. Downregulation of RyR2 up to a certain degree is compensated by increased SR calcium content to normalize calcium sparks. This indirect coupling between Cav1.2 and RyR in arterial SMCs is opposite to striated muscle, where triggering of calcium sparks is controlled by rapid and direct cross-talk between Cav1.1/Cav1.2 L-type channels and RyRs. We discuss the role of RyR isoforms in initiation and formation of calcium sparks in SMCs and their possible molecular binding partners and regulators, which differ compared to striated muscle.

  12. Effect of oxidative stress on Rho kinase II and smooth muscle contraction in rat stomach.

    Science.gov (United States)

    Al-Shboul, Othman; Mustafa, Ayman

    2015-06-01

    Recent studies have shown that both Rho kinase signaling and oxidative stress are involved in the pathogenesis of a number of human diseases, such as diabetes mellitus, hypertension, and atherosclerosis. However, very little is known about the effect of oxidative stress on the gastrointestinal (GI) smooth muscle Rho kinase pathway. The aim of the current study was to investigate the effect of oxidative stress on Rho kinase II and muscle contraction in rat stomach. The peroxynitrite donor 3-morpholinosydnonimine (SIN-1), hydrogen peroxide (H2O2), and peroxynitrite were used to induce oxidative stress. Rho kinase II expression and ACh-induced activity were measured in control and oxidant-treated cells via specifically designed enzyme-linked immunosorbent assay (ELISA) and activity assay kits, respectively. Single smooth muscle cell contraction was measured via scanning micrometry in the presence or absence of the Rho kinase blocker, Y-27632 dihydrochloride. All oxidant agents significantly increased ACh-induced Rho kinase II activity without affecting its expression level. Most important, oxidative stress induced by all three agents augmented ACh-stimulated muscle cell contraction, which was significantly inhibited by Y-27632. In conclusion, oxidative stress activates Rho kinase II and enhances contraction in rat gastric muscle, suggesting an important role in GI motility disorders associated with oxidative stress.

  13. Differences in the primary culture, purification and biological characteristics between endothelial cells and smooth muscle cells from rat aorta

    Institute of Scientific and Technical Information of China (English)

    Shaobo Hu; Zifang Song; Qichang Zheng; Jun Nie

    2009-01-01

    Objective: To investigate the differences of primary culture, purification and biological characteristics between endothelial cells and smooth muscle cells from rat aorta. Methods: Endothelial cells were obtained using the vascular ring adherence, collagenase digestion method and an improved vascular ring adherence method, while smooth muscle cells were separated from tissue sections of rat aorta. Clones of endothelial cells were selected by limiting dilution assay. Both cell types were identified using specific cell immunofluorescent markers,and phase contrast microscopy was used to observe the morphological disparity between endothelial cells and smooth muscle cells at the single cell and colony level. Cell proliferation was determined by the cell counting kit-8. Differences between endothelial cells and smooth muscle cells were evaluated in trypsin digestion 6me, attachment time and recovery after cryopreservation. Results: Endothelial cells were obtained by all three methods. The improved vascular ring method provided the most reproducible results. Cells were in good condition, and of high purity. Smooth muscle cells were cultured successfully by the tissue fragment culture method. Clonal expansion of singleendothelial cells was attained. The two cell types expressed their respective specific markers, and the rate of proliferation of smooth muscle cells exceeded that of endothelial cells. Endothelial cells were more sensitive to trypsin digestion than smooth muscle cells. In addition, they had a shorter adherence time and better recovery following cryopreservation than smooth muscle cells. Conclusion: The improved vascular ring method was optimal for yielding endothelial cells. Limiting dilution is a novel and valid method for purifying primary endothelial cells from rat aorta. Primary rat endothelial cell and vascular smooth muscle cell cultures exhibited different morphological characteristics, proliferation rate, adherence time, susceptibility to trypsin

  14. Deranged smooth muscle α-actin as a biomarker of intestinal pseudo-obstruction: a controlled multinational case series

    Science.gov (United States)

    Knowles, C H; Silk, D B A; Darzi, A; Veress, B; Feakins, R; Raimundo, A H; Crompton, T; Browning, E C; Lindberg, G; Martin, J E

    2004-01-01

    Background and aims: Chronic idiopathic intestinal pseudo-obstruction (CIIP) is a severe motility disorder associated with significant morbidity. Several histopathological (neuropathic and myopathic) phenotypes have been described but only a single adult with jejunal smooth (circular) muscle α-actin deficiency. We present a prospective multinational case series investigating smooth muscle α-actin deficiency as a biomarker of this disease. Methods: A total of 115 fully clinically and physiologically (including prolonged (24 hour) ambulatory jejunal manometry) characterised CIIP patients from three European centres were studied. Immunohistochemical localisation of actins and other cytoskeletal proteins were performed on laparoscopic full thickness jejunal biopsies and compared with adult controls. Distribution of α-actin was also characterised in other gut regions and in the developing human alimentary tract. Results: Twenty eight of 115 (24%) CIIP patient biopsies had absent (n = 22) or partial (n = 6) jejunal smooth muscle α-actin immunostaining in the circular muscle layer. In contrast, smooth muscle α-actin staining was preserved in the longitudinal muscle and in adult jejunal controls (n = 20). Comparative study of other adult alimentary tract regions and fetal small intestine, suggested significant spatial and temporal variations in smooth muscle α-actin expression. Conclusions: The ability to modulate α-smooth muscle actin expression, evident in development, is maintained in adult life and may be influenced by disease, rendering it a valuable biomarker even in the absence of other structural abnormalities. PMID:15479676

  15. Smooth Muscle Tumor Originating in the Pleura: A Case Report and Updated Literature Review

    Science.gov (United States)

    Zarubin, Vadim; Zarineh, Alireza

    2016-01-01

    Smooth muscle tumors (SMTs) of the pleura are exceptionally rare. At present and to the best of these authors' knowledge, there are only 17 cases reported in the literature. We describe a case of a 51-year-old woman who complained of left sided pleuritic chest pain. Further, computed tomography (CT) revealed a left sided localized pleural-based mass involving the 9th rib. She underwent an interventional radiology guided percutaneous core biopsy of the lesion, which disclosed a “Smooth Muscle Tumor of Undetermined Malignant Potential (SMT-UMP).” A video-assisted thoracoscopic surgery (VATS) was performed for diagnosis and treatment purposes. Resections of the pleural-based mass and 9th rib were performed. SMT-UMP was the definitive diagnosis. PMID:27747117

  16. Smooth Muscle Tumor Originating in the Pleura: A Case Report and Updated Literature Review

    Directory of Open Access Journals (Sweden)

    Santiago Fabián Moscoso Martínez

    2016-01-01

    Full Text Available Smooth muscle tumors (SMTs of the pleura are exceptionally rare. At present and to the best of these authors’ knowledge, there are only 17 cases reported in the literature. We describe a case of a 51-year-old woman who complained of left sided pleuritic chest pain. Further, computed tomography (CT revealed a left sided localized pleural-based mass involving the 9th rib. She underwent an interventional radiology guided percutaneous core biopsy of the lesion, which disclosed a “Smooth Muscle Tumor of Undetermined Malignant Potential (SMT-UMP.” A video-assisted thoracoscopic surgery (VATS was performed for diagnosis and treatment purposes. Resections of the pleural-based mass and 9th rib were performed. SMT-UMP was the definitive diagnosis.

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

    Science.gov (United States)

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

    2012-02-01

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

  18. Endometrial stromal sarcoma with smooth muscle and glandular differentiation of the feline uterus.

    Science.gov (United States)

    Sato, T; Maeda, H; Suzuki, A; Shibuya, H; Sakata, A; Shirai, W

    2007-05-01

    The intra-abdominal tumor developing in the uterus and lung of a domestic Shorthair cat was examined histopathologically and immunohistochemically. The tumor showed a proliferation of both endometrial stromal and smooth muscle cells accompanied by prominent vasculature. There were well-differentiated endometrial glands, and tubuli made up a monolayer of eosinophilic cuboidal epithelium. Immunohistochemically, the spindle-shaped cells and half of the stromal-like cells reacted to caldesmon and desmin antibodies. The neoplastic epithelium expressed AE1/AE3 cytokeratin. Feline endometrial stromal tumor has, to the best of our knowledge, not been reported previously and has smooth muscle and glandular components that are a unique variant to the human counterpart.

  19. Prediction of peak forces for a shortening smooth muscle tissue subjected to vibration.

    Science.gov (United States)

    Pidaparti, Ramana M; Dhanaraj, Nandhini; Meiss, Richard A

    2008-01-01

    The objective of the present study is to investigate the peak forces for a tracheal smooth muscle tissue subjected to an applied longitudinal vibration following isotonic shortening. A non-linear finite element analysis was carried out to simulate the vibratory response under experimental conditions that corresponds to forced length oscillations at 33 Hz for 1 second. The stiffness change and hysteresis estimated from the experimental data was used in the analysis. The finite element results of peak forces are compared to the experimental data obtained. The comparison of results indicate that the approach and the vibratory response obtained may be useful for describing the cross-bridge de-attachments within the cells as well as connective tissue connections characteristic of tracheal smooth muscle tissue.

  20. A function for filamentous alpha-smooth muscle actin: Retardation of motility in human breast fibroblasts

    DEFF Research Database (Denmark)

    Rønnov-Jessen, Lone; Petersen, Ole William

    1996-01-01

    Actins are known to comprise six mammalian isoforms of which beta- and gamma-nonmuscle actins are present in all cells, whereas alpha-smooth muscle (alpha-sm) actin is normally restricted to cells of the smooth muscle lineages. alpha-Sm actin has been found also to be expressed transiently...... reactions. Here, we show that the presence of alpha-sm actin is a signal for retardation of migratory behavior in fibroblasts. Comparison in a migration assay of fibroblast cell strains with and without alpha-sm actin revealed migratory restraint in alpha-sm actin-positive fibroblasts. Electroporation...... in certain nonmuscle cells, in particular fibroblasts, which are referred to as myofibroblasts. The functional significance of alpha-sm actin in fibroblasts is unknown. However, myofibroblasts appear to play a prominent role in stromal reaction in breast cancer, at the site of wound repair, and in fibrotic...

  1. siRNA-mediated knockdown of endogenously expressed bestrophin in smooth muscles

    DEFF Research Database (Denmark)

    Larsen, Per; Matchkov, Vladimir; Nilsson, Holger;

     We have recently characterized in smooth muscle cells a unique cGMP-dependent Ca2+-activated Cl- current (ICl(cGMP-Ca)) that co-exists with a "classical" Ca2+-activated Cl- current. We hypothesized that bestrophin-4 (a product of the VMD2-like 3 gene) could be responsible for the ICl(cGMP-Ca) ba...... is responsible for the ICl(cGMP-Ca) in smooth muscle cells. This study presents a novel efficient technique for specific downregulation of gene expression in blood vessels, much needed in studies of vascular function....... was controlled by qPCR and Western blot. The downregulation of bestrophin-4 expression (by 88% in mRNA) with siRNA was a associated with significant reduction (by 83%) of the ICl(cGMP-Ca) while the "classical" Ca2+-activated Cl- current was not affected. Our studies provide evidence that bestrophin-4...

  2. The effects of wild-type and mutant SOD1 on smooth muscle contraction

    Directory of Open Access Journals (Sweden)

    Nikolić-Kokić Aleksandra

    2015-01-01

    Full Text Available In this work we compared the mutated liver copper zinc-containing superoxide dismutase (SOD1 protein G93A of the transgenic rat model of familial amyotrophic lateral sclerosis (FALS, to wild-type (WT rat SOD1. We examined their enzymatic activities and effects on isometric contractions of uteri of healthy virgin rats. G93A SOD1 showed a slightly higher activity than WT SOD1 and, in contrast to WT SOD1, G93A SOD1 did not induce smooth muscle relaxation. This result indicates that effects on smooth muscles are not related to SOD1 enzyme activity and suggest that heterodimers of G93A SOD1 form an ion-conducting pore that diminishes the relaxatory effects of SOD1. We propose that this type of pathogenic feedback affects neurons in FALS. [Projekat Ministarstva nauke Republike Srbije, br. 173014 i br. 175083

  3. Atrial natriuretic factor inhibits mitogen-induced growth in aortic smooth muscle cells.

    Science.gov (United States)

    Baldini, P M; De Vito, P; Fraziano, M; Mattioli, P; Luly, P; Di Nardo, P

    2002-10-01

    Atrial natriuretic factor (ANF) is a polypeptide able to affect cardiovascular homeostasis exhibiting diuretic, natriuretic, and vasorelaxant activities. ANF shows antimitogenic effects in different cell types acting through R(2) receptor. Excessive proliferation of smooth muscle cells is a common phenomenon in diseases such as atherosclerosis, but the role of growth factors in the mechanism which modulate this process has yet to be clarified. The potential antimitogenic role of ANF on the cell growth induced by growth factors appears very intriguing. Aim of the present study was to investigate the possible involvement of ANF on rat aortic smooth muscle (RASM) cells proliferation induced by known mitogens and the mechanism involved. Our data show that ANF, at physiological concentration range, inhibits RASM cell proliferation induced by known mitogens such as PDGF and insulin, and the effect seems to be elicited through the modulation of phosphatidic acid (PA) production and MAP kinases involvement.

  4. Increased fibroblast telomerase expression precedes myofibroblast α-smooth muscle actin expression in idiopathic pulmonary fibrosis

    Directory of Open Access Journals (Sweden)

    Daniel Reis Waisberg

    2012-09-01

    Full Text Available OBJECTIVE: This study sought to identify the relationship between fibroblast telomerase expression, myofibroblasts, and telomerase-mediated regulatory signals in idiopathic pulmonary fibrosis. METHODS: Thirty-four surgical lung biopsies, which had been obtained from patients with idiopathic pulmonary fibrosis and histologically classified as usual interstitial pneumonia, were examined. Immunohistochemistry was used to evaluate fibroblast telomerase expression, myofibroblast α-smooth muscle actin expression and the tissue expression of inter leu kin-4, transforming growth factor-β, and basic fibroblast growth factor. The point-counting technique was used to quantify the expression of these markers in unaffected, collapsed, mural fibrosis, and honeycombing areas. The results were correlated to patient survival. RESULTS: Fibroblast telomerase expression and basic fibroblast growth factor tissue expression were higher in collapsed areas, whereas myofibroblast expression and interleukine-4 tissue expression were higher in areas of mural fibrosis. Transforming growth factor-β expression was higher in collapsed, mural fibrosis and honeycombing areas in comparison to unaffected areas. Positive correlations were found between basic fibroblast growth factor tissue expression and fibroblast telomerase expression and between interleukin-4 tissue expression and myofibroblast α-smooth muscle actin expression. Negative correlations were observed between interleukin-4 expression and basic fibroblast growth factor tissue expression in areas of mural fibrosis. Myofibroblast α-smooth muscle actin expression and interleukin-4 tissue expression in areas of mural fibrosis were negatively associated with patient survival. CONCLUSION: Fibroblast telomerase expression is higher in areas of early remodeling in lung tissues demonstrating typical interstitial pneumonia, whereas myofibroblast α-smooth muscle actin expression predominates in areas of late remodeling

  5. Diabetic conditions promote binding of monocytes to vascular smooth muscle cells and their subsequent differentiation

    OpenAIRE

    Meng, Li; Park, Jehyun; Cai, Qiangjun; Lanting, Linda; Reddy, Marpadga A; Natarajan, Rama

    2009-01-01

    Diabetes is associated with significantly accelerated rates of atherosclerosis, key features of which include the presence of excessive macrophage-derived foam cells in the subendothelial space. We examined the hypothesis that enhanced monocyte-vascular smooth muscle cell (VSMC) interactions leading to subendothelial monocyte retention and differentiation to macrophages under diabetic conditions may be underlying mechanisms. Human aortic VSMCs (HVSMCs) treated with diabetic stimuli high gluco...

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

    Energy Technology Data Exchange (ETDEWEB)

    Joo, Hyung Joon; Seo, Ha-Rim [Department of Cardiology, Cardiovascular Center, College of Medicine, Korea University, Seoul (Korea, Republic of); Jeong, Hyo Eun [Department of Mechanical Engineering, Korea University, Seoul (Korea, Republic of); Choi, Seung-Cheol; Park, Jae Hyung; Yu, Cheol Woong; Hong, Soon Jun [Department of Cardiology, Cardiovascular Center, College of Medicine, Korea University, Seoul (Korea, Republic of); Chung, Seok [Department of Mechanical Engineering, Korea University, Seoul (Korea, Republic of); Lim, Do-Sun, E-mail: dslmd@kumc.or.kr [Department of Cardiology, Cardiovascular Center, College of Medicine, Korea University, Seoul (Korea, Republic of)

    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 endothelial 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.

  7. Transforming growth factor-beta as a differentiating factor for cultured smooth muscle cells.

    Science.gov (United States)

    Gawaziuk, J P; X; Sheikh, F; Cheng, Z-Q; Cattini, P A; Stephens, N L

    2007-10-01

    The aim of the present study was to determine whether the development of supercontractile smooth muscle cells, contributing to the nonspecific hyperreactivity of airways in asthmatic patients, is due to transforming growth factor (TGF)-beta. In cultured smooth muscle cells starved by removal of 10% foetal bovine serum for 7 days, growth arrest was seen; 30% became elongated and demonstrated super contractility. Study of conditioned medium suggested that the differentiating factor was TGF-beta. Sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) was carried out on conditioned medium from the arrested cells. Two protein bands were identified as matrix metalloproteinase (MMP)-2 and TGF-beta1. To determine second messenger signalling by SMAD2, Western blotting and confocal microscopy were employed. Conditioned medium from arrested cultures showed the presence of MMP-2 and TGF-beta1, as revealed by SDS-PAGE; 68- and 25-kDa bands were seen. Differentiation was confirmed by upregulation of marker proteins, smooth muscle type myosin heavy chain and myosin light chain kinase. Confirmation was obtained by downregulating these proteins with decorin treatment, which reduces the levels of active TGF-beta and an adenoviral dominant-negative vector coding for a mutated type II TGF-beta-receptor. Activation of second messenger signalling was demonstrated immunocytochemically by the presence of phosphorylated SMAD2 and SMAD4. Transforming growth factor-beta is likely to be the differentiating factor responsible for the development of these supercontractile smooth muscle cells. The development of such cells in vivo after cessation of an asthmatic attack could contribute to the nonspecific hyperreactivity of airways seen in patients.

  8. Characterisation of K+ Channels in Human Fetoplacental Vascular Smooth Muscle Cells

    OpenAIRE

    Brereton, Melissa F.; Mark Wareing; Rebecca L Jones; 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...

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

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    Tharciano Luiz Teixeira Braga da Silva

    2015-01-01

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

  10. TRPC3 Regulates Release of Brain-Derived Neurotrophic Factor From Human Airway Smooth Muscle

    OpenAIRE

    Vohra, Pawan K.; Thompson, Michael A.; Sathish, Venkatachalem; Kiel, Alexander; Jerde, Calvin; Pabelick, Christina M.; Singh, Brij B.; Prakash, Y. S.

    2013-01-01

    Exogenous brain-derived neurotrophic factor (BDNF) enhances Ca2+ signaling and cell proliferation in human airway smooth muscle (ASM), especially with inflammation. Human ASM also expresses BDNF, raising the potential for autocrine/paracrine effects. The mechanisms by which ASM BDNF secretion occurs are not known. Transient receptor potential channels (TRPCs) regulate a variety of intracellular processes including store-operated Ca2+ entry (SOCE; including in ASM) and secretion of factors suc...

  11. The role of urine markers, white light cystoscopy and fluorescence cystoscopy in recurrence, progression and follow-up of non-muscle invasive bladder cancer

    NARCIS (Netherlands)

    Karaoglu, I.; Heijden, A.G. van der; Witjes, J.A.

    2014-01-01

    Non-muscle invasive bladder cancer (NMIBC) accounts for approximately 70 % of all bladder cancer cases and represents a heterogeneous pathological entity, characterized by a variable natural history and oncological outcome. The combination of cystoscopy and urine cytology is considered the gold stan

  12. Increased proinflammatory responses from asthmatic human airway smooth muscle cells in response to rhinovirus infection

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    King Nicholas JC

    2006-05-01

    Full Text Available Abstract Background Exacerbations of asthma are associated with viral respiratory tract infections, of which rhinoviruses (RV are the predominant virus type. Airway smooth muscle is important in asthma pathogenesis, however little is known about the potential interaction of RV and human airway smooth muscle cells (HASM. We hypothesised that rhinovirus induction of inflammatory cytokine release from airway smooth muscle is augmented and differentially regulated in asthmatic compared to normal HASM cells. Methods HASM cells, isolated from either asthmatic or non-asthmatic subjects, were infected with rhinovirus. Cytokine production was assayed by ELISA, ICAM-1 cell surface expression was assessed by FACS, and the transcription regulation of IL-6 was measured by luciferase activity. Results RV-induced IL-6 release was significantly greater in HASM cells derived from asthmatic subjects compared to non-asthmatic subjects. This response was RV specific, as 5% serum- induced IL-6 release was not different in the two cell types. Whilst serum stimulated IL-8 production in cells from both subject groups, RV induced IL-8 production in only asthmatic derived HASM cells. The transcriptional induction of IL-6 was differentially regulated via C/EBP in the asthmatic and NF-κB + AP-1 in the non-asthmatic HASM cells. Conclusion This study demonstrates augmentation and differential transcriptional regulation of RV specific innate immune response in HASM cells derived from asthmatic and non-asthmatics, and may give valuable insight into the mechanisms of RV-induced asthma exacerbations.

  13. Von Willebrand factor inhibits mature smooth muscle gene expression through impairment of Notch signaling.

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    He Meng

    Full Text Available Von Willebrand factor (vWF, a hemostatic protein normally synthesized and stored by endothelial cells and platelets, has been localized beyond the endothelium in vascular disease states. Previous studies have implicated potential non-hemostatic functions of vWF, but signaling mechanisms underlying its effects are currently undefined. We present evidence that vWF breaches the endothelium and is expressed in a transmural distribution pattern in cerebral small vessel disease (SVD. To determine the potential molecular consequences of vWF permeation into the vessel wall, we also tested whether vWF impairs Notch regulation of key smooth muscle marker genes. In a co-culture system using Notch ligand expressing cells to stimulate Notch in A7R5 cells, vWF strongly inhibited both the Notch pathway and the activation of mature smooth muscle gene promoters. Similar repressive effects were observed in primary human cerebral vascular smooth muscle cells. Expression of the intracellular domain of NOTCH3 allowed cells to bypass the inhibitory effects of vWF. Moreover, vWF forms molecular complexes with all four mammalian Notch ectodomains, suggesting a novel function of vWF as an extracellular inhibitor of Notch signaling. In sum, these studies demonstrate vWF in the vessel wall as a common feature of cerebral SVD; furthermore, we provide a plausible mechanism by which non-hemostatic vWF may play a novel role in the promotion of vascular disease.

  14. Non-selective cation channels mediate chloroquine-induced relaxation in precontracted mouse airway smooth muscle.

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    Ting Zhang

    Full Text Available Bitter tastants can induce relaxation in precontracted airway smooth muscle by activating big-conductance potassium channels (BKs or by inactivating voltage-dependent L-type Ca2+ channels (VDLCCs. In this study, a new pathway for bitter tastant-induced relaxation was defined and investigated. We found nifedipine-insensitive and bitter tastant chloroquine-sensitive relaxation in epithelium-denuded mouse tracheal rings (TRs precontracted with acetylcholine (ACH. In the presence of nifedipine (10 µM, ACH induced cytosolic Ca2+ elevation and cell shortening in single airway smooth muscle cells (ASMCs, and these changes were inhibited by chloroquine. In TRs, ACH triggered a transient contraction under Ca2+-free conditions, and, following a restoration of Ca2+, a strong contraction occurred, which was inhibited by chloroquine. Moreover, the ACH-activated whole-cell and single channel currents of non-selective cation channels (NSCCs were blocked by chloroquine. Pyrazole 3 (Pyr3, an inhibitor of transient receptor potential C3 (TRPC3 channels, partially inhibited ACH-induced contraction, intracellular Ca2+ elevation, and NSCC currents. These results demonstrate that NSCCs play a role in bitter tastant-induced relaxation in precontracted airway smooth muscle.

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

    Science.gov (United States)

    Kaneda, Tae; Tsuruoka, Shuichi; Fujimura, Akio

    2010-12-15

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

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

    Science.gov (United States)

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

    1979-01-01

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

  17. Magnesium used in bioabsorbable stents controls smooth muscle cell proliferation and stimulates endothelial cells in vitro.

    Science.gov (United States)

    Sternberg, Katrin; Gratz, Matthias; Koeck, Kathleen; Mostertz, Joerg; Begunk, Robert; Loebler, Marian; Semmling, Beatrice; Seidlitz, Anne; Hildebrandt, Petra; Homuth, Georg; Grabow, Niels; Tuemmler, Conny; Weitschies, Werner; Schmitz, Klaus-Peter; Kroemer, Heyo K

    2012-01-01

    Magnesium-based bioabsorbable cardiovascular stents have been developed to overcome limitations of permanent metallic stents, such as late stent thrombosis. During stent degradation, endothelial and smooth muscle cells will be exposed to locally high magnesium concentrations with yet unknown physiological consequences. Here, we investigated the effects of elevated magnesium concentrations on human coronary artery endothelial and smooth muscle cell (HCAEC, HCASMC) growth and gene expression. In the course of 24 h after incubation with magnesium chloride solutions (1 or 10 mM) intracellular magnesium level in HCASMC raised from 0.55 ± 0.25 mM (1 mM) to 1.38 ± 0.95 mM (10 mM), while no increase was detected in HCAEC. Accordingly, a DNA microarray-based study identified 69 magnesium regulated transcripts in HCAEC, but 2172 magnesium regulated transcripts in HCASMC. Notably, a significant regulation of various growth factors and extracellular matrix components was observed. In contrast, viability and proliferation of HCAEC were increased at concentrations of up to 25 mM magnesium chloride, while in HCASMC viability and proliferation appeared to be unaffected. Taken together, our data indicate that magnesium halts smooth muscle cell proliferation and stimulates endothelial cell proliferation, which might translate into a beneficial effect in the setting of stent associated vascular injury.

  18. Biomathematical pattern of EMG signal propagation in smooth muscle of the non-pregnant porcine uterus

    Science.gov (United States)

    Domino, Malgorzata; Pawlinski, Bartosz; Gajewski, Zdzislaw

    2017-01-01

    Uterine contractions are generated by myometrial smooth muscle cells (SMCs) that comprise most of the myometrial layer of the uterine wall. Aberrant uterine motility (i.e., hypo- or hyper-contractility or asynchronous contractions) has been implicated in the pathogenesis of infertility due to the failure of implantation, endometriosis and abnormal estrous cycles. The mechanism whereby the non-pregnant uterus initiates spontaneous contractions remains poorly understood. The aim of the present study was to employ linear synchronization measures for analyzing the pattern of EMG signal propagation (direction and speed) in smooth muscles of the non-pregnant porcine uterus in vivo using telemetry recording system. It has been revealed that the EMG signal conduction in the uterine wall of the non-pregnant sow does not occur at random but it rather exhibits specific directions and speed. All detectable EMG signals moved along the uterine horn in both cervico-tubal and tubo-cervical directions. The signal migration speed could be divided into the three main types or categories: i. slow basic migration rhythm (SBMR); ii. rapid basic migration rhythm (RBMR); and iii. rapid accessory migration rhythm (RAMR). In conclusion, the EMG signal propagation in smooth muscles of the porcine uterus in vivo can be assessed using a linear synchronization model. Physiological pattern of the uterine contractile activity determined in this study provides a basis for future investigations of normal and pathologicall myogenic function of the uterus. PMID:28282410

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

    Science.gov (United States)

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

    2003-08-01

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

  20. Smooth muscle myosin inhibition: a novel therapeutic approach for pulmonary hypertension.

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    David Ho

    Full Text Available OBJECTIVE: Pulmonary hypertension remains a major clinical problem despite current therapies. In this study, we examine for the first time a novel pharmacological target, smooth muscle myosin, and determine if the smooth muscle myosin inhibitor, CK-2019165 (CK-165 ameliorates pulmonary hypertension. MATERIALS AND METHODS: Six domestic female pigs were surgically instrumented to measure pulmonary blood flow and systemic and pulmonary vascular dynamics. Pulmonary hypertension was induced by hypoxia, or infusion of the thromboxane analog (U-46619, 0.1 µg/kg/min, i.v.. In rats, chronic pulmonary hypertension was induced by monocrotaline. RESULTS: CK-165 (4 mg/kg, i.v. reduced pulmonary vascular resistance by 22±3 and 28±6% from baseline in hypoxia and thromboxane pig models, respectively (p<0.01 and 0.01, while mean arterial pressure also fell and heart rate rose slightly. When CK-165 was delivered via inhalation in the hypoxia model, pulmonary vascular resistance fell by 17±6% (p<0.05 while mean arterial pressure and heart rate were unchanged. In the monocrotaline model of chronic pulmonary hypertension, inhaled CK-165 resulted in a similar (18.0±3.8% reduction in right ventricular systolic pressure as compared with sildenafil (20.3±4.5%. CONCLUSION: Inhibition of smooth muscle myosin may be a novel therapeutic target for treatment of pulmonary hypertension.

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

    Science.gov (United States)

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

    2015-11-10

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

  2. Management of gastrointestinal smooth muscle tumors: Seven years experience of a teaching hospital in Saudi Arabia

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    Al Salamah Saleh

    2000-01-01

    Full Text Available Background: Gastrointestinal (GI smooth muscle tumors are a group of intramural GI tumors formerly known as leiomyoma and leiomyosarcoma. This retrospective study was made to review our experience in surgical management of GI smooth muscle tumors. Methods: The clinical records for patients with GI smooth muscle tumors during 1993-1999 were reviewed. Results: Ten tumors were located in stomach. Abdominal pain was the main presenting symptom followed by GI bleeding. The preoperative diagnosis was made by enhanced computerized tomography (CT on eleven patients out of thirteen, while upper GI endoscopy with biopsy identified six tumors out of thirteen patients had the test. Tumors were treated by conservative excision in four and radical excision in fourteen. Histology was leiomyoma in six patients (with mitotic figures < 4 per 10 high power field and leiomyosarcoma was found in twelve patients (with mitotic figures> 4 per 10 high power field. Median follow-up was 4 years. There was one recurrence for leiomyoma (size: 6cm. All patients with leiomyosarcoma presented later with metastasis and died. Conclusion: Size and mitotic activity of tumors are the main determinant factors. Enhanced CT remains the proper diagnostic tool.

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

    Institute of Scientific and Technical Information of China (English)

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

    2016-01-01

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

  4. Relaxant effect of an estrone derivate EA303 on isolated colonic smooth muscle of rabbits

    Institute of Scientific and Technical Information of China (English)

    ZHU Lan; FU Shou-ting; CHEN Guo-liang; ZHOU Qian-qian

    2008-01-01

    Objective To study the relaxant effect mechanism of an estrone derivate EA303 on isolate colonic smooth muscle of rabbits. Methods Preparations of the isolated colonic smooth muscle of rabbits were prepared. The effect of EA303 on potassium channel, β receptor and prostaglandin were studied by observing the difference of relaxant dose-effect curves of EA303 on preparations pre-contracted with BaCl3, High K+ solution and Acetylcholine chloride (ACh) in the absence or presence incubation with glibenclamide (10 μM), propranolol (0.1 μM) and Indometacin (10 μM). Results The relaxant effect of EA303 on contraction caused by BaCl2 and High K+ solution were weakened by glibenclamide inhibiting the opening of K+ channel while the relaxant effect of that on contraction caused by ACh was strengthened, after adding propranolol inhabiting β receptor, EA303 attenuated the relaxant action on contraction caused by BaCl2. EA303 had some relaxant impact on contraction induced by High K+ solution after adding indometacin inhabiting the synthesis the prostaglandin (PG). Conclusions The relaxant effect of EA303 on isolated colonic smooth muscle of rabbits may be related with PG synthesis enzyme, potassium channel and β receptor.

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

    Science.gov (United States)

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

    2014-09-01

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

  6. Experimental modulation of the plasmalemmal microfluidity. Studies on endothelial and aortic smooth muscle cells.

    Science.gov (United States)

    Badea, M; Jinga, V; Hörer, O

    1984-01-01

    The microfluidity of cell membranes has been modified experimentally in endothelial cells and smooth muscle cells of bovine or monkey aorta cultured in vitro. Microfluidity was estimated by fluorescence depolarization measurements of diphenyl-hexatriene (DPH)-labelled cells. In both types of cells investigated, the arachidonic acid at concentration of 90 microM induced an increase in the microfluidity by 26-53% whereas the cholesterol at the same concentration produced a decrease in the microfluidity by 16-25%. The oleic acid in the range of 30 to 90 microM increased the monkey smooth muscle cell membranes microfluidity by 21-33% but did not change the microfluidity of endothelial and bovine aortic smooth muscle cells. The stearic acid did not influence the microfluidity of either type of cells under investigation. Cortisol at 90 microM changed the microfluidity of the bovine aortic endothelial cells plasmalemma depending on the incubation time. Possible factors of error in the physical measurements due to the extracellular localization of DPH have been identified.

  7. Stromal proteome expression profile and muscle-invasive bladder cancer research

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    Niu Haitao

    2012-08-01

    Full Text Available Abstract Background To globally characterize the cancer stroma expression profile of muscle-invasive transitional cell carcinoma and to discuss the cancer biology as well as biomarker discovery from stroma. Laser capture micro dissection was used to harvest purified muscle-invasive bladder cancer stromal cells and normal urothelial stromal cells from 4 paired samples. Two-dimensional liquid chromatography tandem mass spectrometry was used to identify the proteome expression profile. The differential proteins were further analyzed using bioinformatics tools and compared with the published literature. Results We identified 868/872 commonly expressed proteins and 978 differential proteins from 4 paired cancer and normal stromal samples using laser capture micro dissection coupled with two-dimensional liquid chromatography tandem mass spectrometry. 487/491 proteins uniquely expressed in cancer/normal stroma. Differential proteins were compared with the entire list of the international protein index (IPI, and there were 42/42 gene ontology (GO terms exhibited as enriched and 8/5 exhibited as depleted in cellular Component, respectively. Significantly altered pathways between cancer/normal stroma mainly include metabolic pathways, ribosome, focal adhesion, etc. Finally, descriptive statistics show that the stromal proteins with extremes of PI and MW have the same probability to be a biomarker. Conclusions Based on our results, stromal cells are essential component of the cancer, biomarker discovery and network based multi target therapy should consider neoplastic cells itself and corresponding stroma as whole one.

  8. THE ROLE OF GASOTRANSMITTERS IN REGULATING OF THE FUNCTIONS OF SMOOTH MUSCLES: THE POSSIBLE EFFECTOR SYSTEMS

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    I. V. Kovalev

    2014-01-01

    Full Text Available Influence of gasotransmitters carbon monoxide (CO and hydrogen sulfide (H2S on the electrical and contractile activities of smooth muscle cells (SMCs of the guinea pig ureter and rat aorta were studied by methods of double sucrose bridge and mechanography. It has been shown that CO causes a dose-dependent decrease of the contractile response of SMCs of the ureter and rat aorta and also reduces the amplitude and duration of the action potential plateau. Against the background of the action of biologically active substances, agonists α1-adrenergetic and H1-histaminergetic receptors (phenylephrine and histamine, respectively, these effects of CO donor (CORM II were amplified. The inhibitory effect of CO on the parameters of the contractile and electrical activities of smooth muscles is attenuated by blocking potassium channels of plasma membrane with tetraethylammonium (TEA or inhibition of soluble guanylate cyclase (ODQ [1H-[1,2,4]-oxadiazolo[4,3-a]quinoxalin-l-one]. Thus, the effects of carbon monoxide on the electrical and contractile activities of SMCs are associated with an increase potassium conductivity of the membrane or the activation of soluble guanylate cyclase.In experiments with a donor of hydrogen sulfide (NaHS, it was shown, that it has an activating effect on the electrical and contractile activities of smooth muscles of the guinea pig ureter, which is caused by the action of potassium conductivity of the membrane. Activating effect of H2S on the contractile properties of SMCs of the guinea pig ureter decreased by blocking ATP-dependent channels with glibenclamide. Analysis of the effect of H2S on sodium and calcium conductance of the membrane smooth muscles of the ureter using modified sodium-free and TEA- containing Krebs solution showed that the contribution of potassium conductance is mainly sold at high concentrations (100 and 1000 μmol donor NaHS. Probably, that the impact of low concentrations of NaHS (10 μmol on the

  9. Muscle-invasive bladder cancer in a young adult: a case report and a review of the literature.

    Science.gov (United States)

    Nabbout, Philippe; Eldefrawy, Ahmed; Engles, C Dirk; Culkin, Daniel J; Slobodov, Gennady

    2013-01-01

    The peak incidence of bladder cancer (BC) is in the sixth decade of life. Muscle-invasive bladder cancer (MIBC) in young adults is extremely rare. We report a case of MIBC in a 28-year-old smoking male patient. The patient presented with hematuria and flank pain for which he underwent a computerized tomography (CT) scan of the abdomen and pelvis with and without contrast. The CT scan showed a 6 cm mass on the left side of the trigone extending to the left urteric orifice and left hydronephrosis, but no lymphadenopathy was noted. The patient then underwent a left nephrostomy tube placement followed by trans-urethral resection of bladder tumor (TURBT). The tumor involved both ureteric orifices and extended to the prostatic urethra. Complete resection was not feasible. Pathology showed high-grade T1 urothelial carcinoma. CT scan of the chest showed no distant lung metastasis. The patient then elected to undergo radical cystectomy with ileal conduit urinary diversion. Final pathology revealed T2a N0 urothelial carcinoma of the bladder. Our aim is to present our experience and review the literature for the natural history and oncological and quality of life outcomes of urothelial carcinoma of the bladder in young patients.

  10. Calcium ion requirement for acetylcholine-stimulated breakdown of triphosphoinositide in rabbit iris smooth muscle.

    Science.gov (United States)

    Akhtar, R A; Abdel-Latif, A A

    1978-03-01

    Previous studies from this laboratory have established that addition of acetylcholine (ACh) or norepinephrine to 32P-labeled rabbit iris smooth muscle increases significantly the breakdown of triphosphoinositide (TPI) and that these stimulatory effects are blocked by atropine and phentolamine, respectively. The present studies were undertaken in order to show the effect of Ca++ on the ACh-stimulated breakdown of TPI ("TPI effect") in this tissue. Paired iris smooth muscles were prelabeled with 32Pi for 30 minutes at 37 degrees C in Ca++-free iso-osmotic salt medium. The prelabeled irises were then washed and incubated for 10 minutes in nonradioactive Ca++-free medium which contained 10 mM 2-deoxyglucose under various conditions. The phospholipids were isolated by means of two-dimensional thin-layer chromatography and their radioactivities were determined. In the absence of Ca++, 50 micrometer ACh increased TPI breakdown and phosphatidic acid (PA) labeling by 16 and 38%, respectively. In the absence of ACh, 0.75 micrometer Ca++ increased TPI breakdown and PA labeling by 11 and 20%, respectively. When both ACh and Ca++ were added, the increase in TPI breakdown and PA labeling rose to 32 and 74%, respectively. The labeling of phosphatidylinositol was found to be insensitive to the presence of Ca++. Ca++ was determined in the iris smooth muscle and it was found to contain 3.13 mumol of Ca++ per g of tissue. This was reduced by 80% after the muscle was washed and incubated in a medium which contained 0.25 micrometer ethyleneglycol bis (beta-aminoethyl ether)-N,N'-tetraacetic acid (EGTA). The TPI effect was abolished by 0.25 micrometer EGTA and restored when excess Ca++ (1.25 micrometer) was added. Concentrations of Ca++ as low as 50 micrometer provoked a TPI effect. Sr++ (2 micrometer), but not Ba++ or Mn++, was found to substitute partially for Ca++. Ionophore A-23187 (20 micrometer) was found to increase the breakdown of TPI and labeling of PA by 11 and 24

  11. Development of transplant vasculopathy in aortic allografts correlates with neointimal smooth muscle cell proliferative capacity and fibrocyte frequency

    NARCIS (Netherlands)

    Onuta, Geanina; van Ark, Joris; Rienstra, Heleen; Boer, Mark Walther; Klatter, Flip A.; Bruggeman, Cathrien A.; Zeebregts, Clark J.; Rozing, Jan; Hillebrands, Jan-Luuk

    2010-01-01

    Objective: Transplant vasculopathy consists of neointima formation in graft vasculature resulting from vascular smooth muscle cell recruitment and proliferation. Variation in the severity of vasculopathy has been demonstrated. Genetic predisposition is suggested as a putative cause of this variation

  12. p42/p44 MAP kinase activation is localized to caveolae-free membrane domains in airway smooth muscle

    NARCIS (Netherlands)

    Gosens, Reinoud; Dueck, Gordon; Gerthoffer, William T; Unruh, Helmut; Zaagsma, Johan; Meurs, Herman; Halayko, Andrew J

    2007-01-01

    Caveolae are abundant plasma membrane invaginations in airway smooth muscle that may function as preorganized signalosomes by sequestering and regulating proteins that control cell proliferation, including receptor tyrosine kinases (RTKs) and their signaling effectors. We previously demonstrated, ho

  13. The effect of trinitrobenzene sulfonic acid on gut-derived smooth muscle cell arachidonic acid metabolism: role of endogenous prostanoids

    Directory of Open Access Journals (Sweden)

    W. E. Longo

    1997-01-01

    Full Text Available The contribution of smooth muscle cells as a potential source of eicosanoid production during inflammatory states remains to be elucidated. We investigated the effect of trinitrobenzene sulfonic acid (TNB, a known pro-inflammatory agent, on jejunal smooth muscle cell eicosanoid production. Human gut-derived smooth muscle cells (HISM were incubated with TNB for 1 hour. Additionally, some cells were preincubated with either dimethylthiourea, or indomethacin for 1 hour before exposure to identical concentrations of TNB. Incubation with TNB led to significant increases in PGE2 and 6-keto PGF-1α release, but not leukotriene B4 release; responses which were both inhibited by dimethylthiourea and indomethacin treatment. Our results suggest that gutderived smooth muscle cells may represent an important source of proinflammatory prostanoids but not leukotrienes during inflammatory states of the intestine. The inhibition of prostanoid activity by thiourea may be mediated by suppression of cyclooxygenase activity in this cell line.

  14. HB-EGF-Promoted Airway Smooth Muscle Cells and Their Progenitor Migration Contribute to Airway Smooth Muscle Remodeling in Asthmatic Mouse.

    Science.gov (United States)

    Wang, Qing; Li, Hequan; Yao, Yinan; Lu, Guohua; Wang, Yuehong; Xia, Dajing; Zhou, Jianying

    2016-03-01

    The airway smooth muscle (ASM) cells' proliferation, migration, and their progenitor's migration are currently regarded as causative factors for ASM remodeling in asthma. Heparin-binding epidermal growth factor (HB-EGF), a potent mitogen and chemotactic factor, could promote ASM cell proliferation through MAPK pathways. In this study, we obtained primary ASM cells and their progenitors from C57BL/6 mice and went on to explore the role of HB-EGF in these cells migration and the underlying mechanisms. We found that recombinant HB-EGF (rHB-EGF) intratracheal instillation accelerated ASM layer thickening in an OVA-induced asthmatic mouse. Modified Boyden chamber assay revealed that rHB-EGF facilitate ASM cell migration in a dose-dependent manner and ASM cells from asthmatic mice had a greater migration ability than that from normal counterparts. rHB-EGF could stimulate the phosphorylation of ERK1/2 and p38 in ASM cells but further migration assay showed that only epidermal growth factor receptor inhibitor (AG1478) or p38 inhibitor (SB203580), but not ERK1/2 inhibitor (PD98059), could inhibit rHB-EGF-mediated ASM cells migration. Actin cytoskeleton experiments exhibited that rHB-EGF could cause actin stress fibers disassembly and focal adhesions formation of ASM cells through the activation of p38. Finally, airway instillation of rHB-EGF promoted the recruitment of bone marrow-derived smooth muscle progenitor cells, which were transferred via caudal vein, migrating into the airway from the circulation. These observations demonstrated that ASM remodeling in asthma might have resulted from HB-EGF-mediated ASM cells and their progenitor cells migration, via p38 MAPK-dependent actin cytoskeleton remodeling.

  15. 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.

    Directory of Open Access Journals (Sweden)

    Wulin Tian

    Full Text Available 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.

  16. 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

    Institute of Scientific and Technical Information of China (English)

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

    2011-01-01

    @@ In this study,we investigated the effects of a combination of Ginkgo biloba extracts (GBE) and phosphodiesterase type 5 (IRDE-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 x 10-5 mol I-1 norepinephrine,GBE (0.01-1 mg ml-1) and mirodenafil (0.01-100 nmoll-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%a18.35% at 0.01 mg ml一'to 52.28%±11.42% at 1 mg ml-1).After pre-treatment with 0.03 mg ml-1of GBE,the relaxant effects of mirodenafil were increased at all concentrations.After tetraethylammonium (TEA) (1 mmoll-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 mmoll-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.

  17. Integrin αVβ5 Mediated TGF-β Activation by Airway Smooth Muscle Cells in Asthma

    OpenAIRE

    Tatler, Amanda L; John, Alison E.; Jolly, Lisa; Habgood, Anthony; Porte, Jo; Brightling, Chris; Knox, Alan J; Pang, Linhua; Sheppard, Dean; Huang, Xiaozhu; Jenkins, Gisli

    2011-01-01

    Severe asthma is associated with airway remodelling, characterised by structural changes including increased smooth muscle mass and matrix deposition in the airway, leading to deteriorating lung function. Transforming growth factor-β (TGF-β) is a pleiotropic cytokine leading to increased synthesis of matrix molecules by human airway smooth muscle cells (HASMs) and is implicated in asthmatic airway remodelling. TGF-β is synthesised as a latent complex, sequestered in the extracellular matrix, ...

  18. Effect of pinaverium bromide on stress-induced colonic smooth muscle contractility disorder in rats

    Institute of Scientific and Technical Information of China (English)

    Yun Dai; Jian-Xiang Liu; Jun-Xia Li; Yun-Feng Xu

    2003-01-01

    AIM: To investigate the effect of pinaverium bromide, a Ltype calcium channel blocker with selectivity for the gastrointestinal tract on contractile activity of colonic circular smooth muscle in normal or cold-restraint stressed rats and its possible mechanism.METHODS: Cold-restraint stress was conducted on rats to increase fecal pellets output. Each isolated colonic circular muscle strip was suspended in a tissue chamber containing warm oxygenated Tyrode-Ringer solution. The contractile response to ACh or KCl was measured isometrically on inkwriting recorder. Incubated muscle in different concentrations of pinaverium and the effects of pinaverium were investigated on ACh or KCl-induced contraction. Colon smooth muscle cells were cultured from rats and [Ca2+]i was measured in cell suspension using the Ca2+ fluorescent dye fura-2/AlMl.RESULTS: During stress, rats fecal pellet output increased 61% (P<0.01). Stimulated with ACh or KCl, the muscle contractility was higher in stress than that in control. Pinaverium inhibited the increment of [Ca2+]i and the muscle contraction in response to ACh or KCl in a dose dependent manner. A significant inhibition of pinaverium to ACh or KCl induced [Ca2+]i increment was observed at 10-6 mol/L. The IC50 values for inhibition of ACh induced contraction for the stress and control group were 1.66×10-6 mol/L and 0.91×10-6mol/L, respectively. The ICs0 values for inhibition of KCl induced contraction for the stress and control group were 8.13×10-7 mol/L and 3.80×10-7 mol/L, respectively.CONCLUSION: Increase in [Ca2+]i of smooth muscle cells is directly related to the generation of contraction force in colon. L-type Ca2+ channels represent the main route of Ca2+ entry.Pinaverium inhibits the calcium influx through L-type channels;decreases the contractile response to many kinds of agonists and regulates the stress-induced colon hypermotility.

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

    Science.gov (United States)

    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.

  20. Novel role for the transient potential receptor melastatin 4 channel in guinea pig detrusor smooth muscle physiology.

    Science.gov (United States)

    Smith, Amy C; Hristov, Kiril L; Cheng, Qiuping; Xin, Wenkuan; Parajuli, Shankar P; Earley, Scott; Malysz, John; Petkov, Georgi V

    2013-03-01

    Members of the transient receptor potential (TRP) channel superfamily, including the Ca(2+)-activated monovalent cation-selective TRP melastatin 4 (TRPM4) channel, have been recently identified in the urinary bladder. However, their expression and function at the level of detrusor smooth muscle (DSM) remain largely unexplored. In this study, for the first time we investigated the role of TRPM4 channels in guinea pig DSM excitation-contraction coupling using a multidisciplinary approach encompassing protein detection, electrophysiology, live-cell Ca(2+) imaging, DSM contractility, and 9-phenanthrol, a recently characterized selective inhibitor of the TRPM4 channel. Western blot and immunocytochemistry experiments demonstrated the expression of the TRPM4 channel in whole DSM tissue and freshly isolated DSM cells with specific localization on the plasma membrane. Perforated whole cell patch-clamp recordings and real-time Ca(2+) imaging experiments with fura 2-AM, both using freshly isolated DSM cells, revealed that 9-phenanthrol (30 μM) significantly reduced the cation current and decreased intracellular Ca(2+) levels. 9-Phenanthrol (0.1-30 μM) significantly inhibited spontaneous, 0.1 μM carbachol-induced, 20 mM KCl-induced, and nerve-evoked contractions in guinea pig DSM-isolated strips with IC50 values of 1-7 μM and 70-80% maximum inhibition. 9-Phenanthrol also reduced nerve-evoked contraction amplitude induced by continuous repetitive electrical field stimulation of 10-Hz frequency and shifted the frequency-response curve (0.5-50 Hz) relative to the control. Collectively, our data demonstrate the novel finding that TRPM4 channels are expressed in guinea pig DSM and reveal their critical role in the regulation of guinea pig DSM excitation-contraction coupling.

  1. BK channel activation by NS11021 decreases excitability and contractility of urinary bladder smooth muscle

    DEFF Research Database (Denmark)

    Layne, Jeffrey J; Nausch, Bernhard; Olesen, Søren-Peter

    2009-01-01

    (o)) and whole cell BK channel currents. The frequency of spontaneous action potentials in UBSM strips was reduced by NS11021 from a control value of 20.9 + or - 5.9 to 10.9 + or - 3.7 per minute. NS11021 also reduced the force of UBSM spontaneous phasic contractions by approximately 50%, and this force...... the frequency of spontaneous action potentials....

  2. On the terminology for describing the length-force relationship and its changes in airway smooth muscle.

    Science.gov (United States)

    Bai, Tony R; Bates, Jason H T; Brusasco, Vito; Camoretti-Mercado, Blanca; Chitano, Pasquale; Deng, Lin Hong; Dowell, Maria; Fabry, Ben; Ford, Lincoln E; Fredberg, Jeffrey J; Gerthoffer, William T; Gilbert, Susan H; Gunst, Susan J; Hai, Chi-Ming; Halayko, Andrew J; Hirst, Stuart J; James, Alan L; Janssen, Luke J; Jones, Keith A; King, Greg G; Lakser, Oren J; Lambert, Rodney K; Lauzon, Anne-Marie; Lutchen, Kenneth R; Maksym, Geoffrey N; Meiss, Richard A; Mijailovich, Srboljub M; Mitchell, Howard W; Mitchell, Richard W; Mitzner, Wayne; Murphy, Thomas M; Paré, Peter D; Schellenberg, R Robert; Seow, Chun Y; Sieck, Gary C; Smith, Paul G; Smolensky, Alex V; Solway, Julian; Stephens, Newman L; Stewart, Alastair G; Tang, Dale D; Wang, Lu

    2004-12-01

    The observation that the length-force relationship in airway smooth muscle can be shifted along the length axis by accommodating the muscle at different lengths has stimulated great interest. In light of the recent understanding of the dynamic nature of length-force relationship, many of our concepts regarding smooth muscle mechanical properties, including the notion that the muscle possesses a unique optimal length that correlates to maximal force generation, are likely to be incorrect. To facilitate accurate and efficient communication among scientists interested in the function of airway smooth muscle, a revised and collectively accepted nomenclature describing the adaptive and dynamic nature of the length-force relationship will be invaluable. Setting aside the issue of underlying mechanism, the purpose of this article is to define terminology that will aid investigators in describing observed phenomena. In particular, we recommend that the term "optimal length" (or any other term implying a unique length that correlates with maximal force generation) for airway smooth muscle be avoided. Instead, the in situ length or an arbitrary but clearly defined reference length should be used. We propose the usage of "length adaptation" to describe the phenomenon whereby the length-force curve of a muscle shifts along the length axis due to accommodation of the muscle at different lengths. We also discuss frequently used terms that do not have commonly accepted definitions that should be used cautiously.

  3. Effect of Nateglinide and Glibenclamide on Endothelial Cells and Smooth Muscle Cells from Human Coronary Arteries

    Directory of Open Access Journals (Sweden)

    Seeger H

    2004-01-01

    Full Text Available In the present work the effect of nateglinide and glibenclamide, two different substances used for therapy of diabetes mellitus type 2, were investigated on the synthesis of markers of endothelial function and on the proliferation of smooth muscle cells in vitro. As cell models endothelial and smooth muscle cells from human coronary arteries were used. Both substances were tested at concentrations of 0.1, 1 and 10 mmol/l. As markers of endothelial function prostacyclin, endothelin and plasminogen-activator-inhibitor-1 (PAI-1 were tested. Nateglinide and glibenclamide were similarly able to inhibit endothelial endothelin and PAI-1 synthesis, but only at the highest concentration tested. Endothelial prostacyclin synthesis and proliferation of smooth muscle cells were not significantly changed by both substances. These results indicate that both nateglinide and glibenclamide may have potential in reducing negative long-term effects of diabetes such as atherogenesis. Kurzfassung: Effekt von Nateglinid und Glibenclamid auf Endothel- und Muskelzellen humaner Koronararterien. In der vorliegenden Arbeit wurde die Wirkung von Nateglinid und Glibenclamid, zweier unterschiedlicher Substanzen zur Behandlung des Diabetes mellitus Typ 2, auf die Synthese von Markern der Endothelfunktion und auf die Proliferation glatter Muskelzellen untersucht. Als Zellmodell dienten Endothelzellen und glatte Muskelzellen menschlicher Koronararterien. Beide Substanzen wurden in den Konzentrationen 0,1, 1 und 10 mmol/l getestet. Als Marker der Endothelfunktion dienten Prostazyklin, Endothelin und Plasminogen-Aktivator-Inhibitor-1 (PAI-1. Sowohl Nateglinid als auch Glibenclamid konnten die endotheliale Endothelin- und PAI-1-Produktion in ähnlichem Ausmaß senken, allerdings nur in der höchsten Konzentration. Die Prostazyklinsynthese und die Muskelzellproliferation wurden nicht signifikant beeinflußt. Diese Ergebnisse deuten daraufhin, daß sowohl Nateglinid als auch

  4. Actions of genistein on contractile response of smooth muscle isolated from guinea pig gallbladder

    Institute of Scientific and Technical Information of China (English)

    Ya-Li Luo; Ya-Li Wang; Neng-Lian Li; Tian-Zhen Zheng; Li Zhang; Ya-Li She; Shu-Ming Hu

    2009-01-01

    BACKGROUND: Defective contractile motility of the gallbladder is an important factor for gallstone formation. Estrogen might increase the risk of gallstones and cholecystitis, and estradiol inhibits the contractile activity of isolated strips of guinea pig gallbladder. The potential risks associated with hormone replacement therapy (HRT) include symptomatic gallstones. Phytoestrogen have been used to treat menopause syndromes by replacing traditional estrogen. This experiment aimed to determine the effects of the phytoestrogen genistein on the contractile response of smooth muscle strips isolated from guinea pig gallbladder and its possible mechanism of action. METHODS: Guinea pigs were sacriifced to remove the whole gallbladder. Two or three smooth muscle strips were cut longitudinally. Each strip was suspended in a tissue chamber containing Krebs solution. After 2 hours of equilibration, contractile response indexes were recorded. Different concentrations of genistein were added to the chamber and the contractile responses were measured. Each antagonist was added 2 minutes before genistein to study possible mechanisms. The effect of genistein on calcium-dependent contraction curves and biphasic contraction in calcium-free Krebs solution were measured. RESULTS: Genistein decreased the resting tension dose-dependently, and reduced the mean contractile amplitude and frequency in gallbladder strips. Ranitidine partly inhibited the effect of genistein, but methylene blue, Nω-nitro-L-arginine, and propranolol hydrochloride did not inlfuence this action. Genistein had no signiifcant effects on calcium-dependent contraction. Genistein reduced the ifrst contraction induced by acetylcholine chloride, but did not affect the second contraction caused by CaCl2. CONCLUSIONS: Genistein relaxed smooth muscle isolated from the gallbladder of guinea pigs and this might contribute to the formation of gallstones. The inhibitory action might be related to H2 receptors and

  5. The impact of extracellular and intracellular Ca2+ on ethanol-induced smooth muscle contraction

    Institute of Scientific and Technical Information of China (English)

    Naciye YAKTUBAY DONDAS; Mahir KAPLAN; Derya KAYA; Ergin SiNGiRiK

    2009-01-01

    Aim:To evaluate the impact of extracellular and intracellular Ca~(2+) on contractions induced by ethanol in smooth muscle.Methods: Longitudinal smooth muscle strips were prepared from the gastric fundi of mice. The contractions of smooth muscle strips were recorded with an isometric force displacement transducer.Results: Ethanol (164 mmol/L) produced reproducible contractions in isolated gastric fundal strips of mice. Although lidocaine (50 and 100 μmol/L), a local anesthetic agent, and hexamethonium (100 and 500 μmol/L), a ganglionic blocking agent, failed to affect these contractions, verapamil (1-50 μmol/L) and nifedipine (1-50 μmol/L), selective blockers of L-type Ca~(2+) channels, significantly inhibited the contractile responses of ethanol. Using a Ca~(2+)-free medium nearly eliminated these contractions in the same tissue. Ryanodine (1-50 μmol/L) and ruthenium red (10-100 μmol/L), selective blockers of intracellular Ca~(2+) channels/ryanodine receptors; cyclopiazonic acid (CPA; 1-10 μmol/L), a selective inhibitor of sarcoplasmic reticulum (SR) Ca~(2+)-ATPase; and caffeine (0.5-5 mmol/L), a depleting agent of intracellular Ca~(2+) stores, significantly inhibited the contractile responses induced by ethanol. In addition, the com-bination of caffeine (5 mmol/L) plus CPA (10 μmol/L), and ryanodine (10 μmol/L) plus CPA (10 μmol/L), caused further inhibition of contractions in response to ethanol. This inhibition was significantly different from those associated with caffeine, ryanodine or CPA. Furthermore the combination of caffeine (5 mmol/L), ryanodine (10 μmol/L) and CPA(10 μmol/L) eliminated the contractions induced by ethanol in isolated gastric fundal strips of mice.Conclusion: Both extracellular and intracellular Ca~(2+) may have important roles in regulating contractions induced by ethanol in the mouse gastric fundus.

  6. A key role for STIM1 in store operated calcium channel activation in airway smooth muscle

    Directory of Open Access Journals (Sweden)

    Peel Samantha E

    2006-09-01

    Full Text Available Abstract Background Control of cytosolic calcium plays a key role in airway myocyte function. Changes in intracellular Ca2+ stores can modulate contractile responses, modulate proliferation and regulate synthetic activity. Influx of Ca2+ in non excitable smooth muscle is believed to be predominantly through store operated channels (SOC or receptor operated channels (ROC. Whereas agonists can activate both SOC and ROC in a range of smooth muscle types, the specific trigger for SOC activation is depletion of the sarcoplasmic reticulum Ca2+ stores. The mechanism underlying SOC activation following depletion of intracellular Ca2+ stores in smooth muscle has not been identified. Methods To investigate the roles of the STIM homologues in SOC activation in airway myocytes, specific siRNA sequences were utilised to target and selectively suppress both STIM1 and STIM2. Quantitative real time PCR was employed to assess the efficiency and the specificity of the siRNA mediated knockdown of mRNA. Activation of SOC was investigated by both whole cell patch clamp electrophysiology and a fluorescence based calcium assay. Results Transfection of 20 nM siRNA specific for STIM1 or 2 resulted in robust decreases (>70% of the relevant mRNA. siRNA targeted at STIM1 resulted in a reduction of SOC associated Ca2+ influx in response to store depletion by cyclopiazonic acid (60% or histamine but not bradykinin. siRNA to STIM2 had no effect on these responses. In addition STIM1 suppression resulted in a more or less complete abrogation of SOC associated inward currents assessed by whole cell patch clamp. Conclusion Here we show that STIM1 acts as a key signal for SOC activation following intracellular Ca2+ store depletion or following agonist stimulation with histamine in human airway myocytes. These are the first data demonstrating a role for STIM1 in a physiologically relevant, non-transformed endogenous expression cell model.

  7. Role of M1 receptor in regulation of gastric fundus smooth muscle contraction

    Directory of Open Access Journals (Sweden)

    Marta Gajdus

    2011-09-01

    Full Text Available Background:The subject of this study is determination of the influence of drugs on gastric fundus smooth muscle contraction induced by activation of muscarinic receptors M1. Experiments tested interactions between a receptor agonist, carbachol and muscarinic receptor antagonists, atropine and pirenzepine.Material/Methods:Testing was conducted on tissues isolated from rat’s stomach. Male Wistar rats with weight between 220 g and 360 g were anesthetized by intraperitoneal injection of urethane (120 mg/kg. The stomach was dissected, and later the gastric fundus was isolated. Tissue was placed in a dish for insulated organs with 20 ml in capacity, filled with Krebs fluid. Results contained in the study are average values ± SE. In order to determine statistical significance, the principles of receptor theory were used (Kenakin modification.Results:According to tests, carbachol, in concentrations ranging between 10–8 M to 10–4 M, in a dosage-dependent way induces gastric fundus smooth muscle contraction. Presented results indicate that carbachol meets the conditions posed to full agonists. On the other hand, atropine, a non-selective muscarinic receptor antagonist, causes a concentration-dependent shift of concentration-effect curve (for carbachol to the right, maintaining maximum reaction. According to analysis of the curve determined, we can deduce that atropine meets the conditions posed to competitive antagonists. The use of pirenzepine, a competitive receptor agonist M1, causes shift of concentration-effect curve (for carbachol to the right, maintaining maximum reaction.Conclusions:From the testing conducted on the preparation of the gastric fundus we can deduce that atropine causes shift of concentration-effect curves for carbachol to the right. A similar effect is released by pirenzepine, selectively blocking muscarinic receptors of M1 type. The results indicate that in the preparation of the gastric fundus smooth muscle, M1 type

  8. Tropomyosin variants describe distinct functional subcellular domains in differentiated vascular smooth muscle cells.

    Science.gov (United States)

    Gallant, Cynthia; Appel, Sarah; Graceffa, Philip; Leavis, Paul; Lin, Jim Jung-Ching; Gunning, Peter W; Schevzov, Galina; Chaponnier, Christine; DeGnore, Jon; Lehman, William; Morgan, Kathleen G

    2011-06-01

    Tropomyosin (Tm) is known to be an important gatekeeper of actin function. Tm isoforms are encoded by four genes, and each gene produces several variants by alternative splicing, which have been proposed to play roles in motility, proliferation, and apoptosis. Smooth muscle studies have focused on gizzard smooth muscle, where a heterodimer of Tm from the α-gene (Tmsm-α) and from the β-gene (Tmsm-β) is associated with contractile filaments. In this study we examined Tm in differentiated mammalian vascular smooth muscle (dVSM). Liquid chromatography-tandem mass spectrometry (LC MS/MS) analysis and Western blot screening with variant-specific antibodies revealed that at least five different Tm proteins are expressed in this tissue: Tm6 (Tmsm-α) and Tm2 from the α-gene, Tm1 (Tmsm-β) from the β-gene, Tm5NM1 from the γ-gene, and Tm4 from the δ-gene. Tm6 is by far most abundant in dVSM followed by Tm1, Tm2, Tm5NM1, and Tm4. Coimmunoprecipitation and coimmunofluorescence studies demonstrate that Tm1 and Tm6 coassociate with different actin isoforms and display different intracellular localizations. Using an antibody specific for cytoplasmic γ-actin, we report here the presence of a γ-actin cortical cytoskeleton in dVSM cells. Tm1 colocalizes with cortical cytoplasmic γ-actin and coprecipitates with γ-actin. Tm6, on the other hand, is located on contractile bundles. These data indicate that Tm1 and Tm6 do not form a classical heterodimer in dVSM but rather describe different functional cellular compartments.

  9. Leptin augments coronary vasoconstriction and smooth muscle proliferation via a Rho-kinase-dependent pathway.

    Science.gov (United States)

    Noblet, Jillian N; Goodwill, Adam G; Sassoon, Daniel J; Kiel, Alexander M; Tune, Johnathan D

    2016-05-01

    Leptin has been implicated as a key upstream mediator of pathways associated with coronary vascular dysfunction and disease. The purpose of this investigation was to test the hypothesis that leptin modifies the coronary artery proteome and promotes increases in coronary smooth muscle contraction and proliferation via influences on Rho kinase signaling. Global proteomic assessment of coronary arteries from lean swine cultured with obese concentrations of leptin (30 ng/mL) for 3 days revealed significant alterations in the coronary artery proteome (68 proteins) and identified an association between leptin treatment and calcium signaling/contraction (four proteins) and cellular growth and proliferation (35 proteins). Isometric tension studies demonstrated that both acute (30 min) and chronic (3 days, serum-free media) exposure to obese concentrations of leptin potentiated depolarization-induced contraction of coronary arteries. Inhibition of Rho kinase significantly reduced leptin-mediated increases in coronary artery contractions. The effects of leptin on the functional expression of Rho kinase were time-dependent, as acute treatment increased Rho kinase activity while chronic (3 day) exposure was associated with increases in Rho kinase protein abundance. Proliferation assays following chronic leptin administration (8 day, serum-containing media) demonstrated that leptin augmented coronary vascular smooth muscle proliferation and increased Rho kinase activity. Inhibition of Rho kinase significantly reduced these effects of leptin. Taken together, these findings demonstrate that leptin promotes increases in coronary vasoconstriction and smooth muscle proliferation and indicate that these phenotypic effects are associated with alterations in the coronary artery proteome and dynamic effects on the Rho kinase pathway.

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

    Science.gov (United States)

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

    2009-08-01

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

  11. Expression of Potassium Channels in Uterine Smooth Muscle Cells from Patients with Adenomyosis

    Institute of Scientific and Technical Information of China (English)

    Jing-Hua Shi; Li Jin; Jin-Hua Leng; Jing-He Lang

    2016-01-01

    Background:Adenomyosis (AM) has impaired contraction.This study aimed to explore the expression of potassium channels related to contraction in myometrial smooth muscle cells (MSMCs) of AM.Methods:Uterine tissue samples from 22 patients (cases) with histologically confirmed AM and 12 (controls) with cervical intraepithelial neoplasia were collected for both immunohistochemistry and real-time polymerase chain reaction to detect the expression of large conductance calcium-and voltage-sensitive K+ channel (BKCa)-α/β subunits,voltage-gated potassium channel (Kv) 4.2,and Kv4.3.Student's t-test was used to compare the expression.Results:The BKCa-α/β subunits,Kv4.2,and Kv4.3 were located in smooth muscle cells,glandular epithelium,and stromal cells.However,BKCa-β subunit expression in endometrial glands of the controls was weak,and Kv4.3 was almost undetectable in the controls.The expression of BKCa-α messenger RNA (mRNA) (0.62 ± 0.19-fold decrease,P < 0.05) and Kv4.3 mRNA (0.67 ± 0.20-fold decrease,P < 0.05) decreased significantly in the M SMCs of the control group compared with the AM group.However,there were no significant differences in BKCa-β subunit mRNA or Kv4.2 mRNA.Conclusions:The BKCa-α mRNA and the Kv4.3 mRNA are expressed significantly higher in AM than those in the control group,that might cause the abnormal uterus smooth muscle contractility,change the microcirculation of uterus to accumulate the inflammatory factors,impair the endometrium further,and aggravate the pain.

  12. Expression of Potassium Channels in Uterine Smooth Muscle Cells from Patients with Adenomyosis

    Directory of Open Access Journals (Sweden)

    Jing-Hua Shi

    2016-01-01

    Full Text Available Background: Adenomyosis (AM has impaired contraction. This study aimed to explore the expression of potassium channels related to contraction in myometrial smooth muscle cells (MSMCs of AM. Methods: Uterine tissue samples from 22 patients (cases with histologically confirmed AM and 12 (controls with cervical intraepithelial neoplasia were collected for both immunohistochemistry and real-time polymerase chain reaction to detect the expression of large conductance calcium- and voltage-sensitive K + channel (BKCa-α/β subunits, voltage-gated potassium channel (Kv 4.2, and Kv4.3. Student′s t-test was used to compare the expression. Results: The BKCa-α/β subunits, Kv4.2, and Kv4.3 were located in smooth muscle cells, glandular epithelium, and stromal cells. However, BKCa-β subunit expression in endometrial glands of the controls was weak, and Kv4.3 was almost undetectable in the controls. The expression of BKCa-α messenger RNA (mRNA (0.62 ± 0.19-fold decrease, P < 0.05 and Kv4.3 mRNA (0.67 ± 0.20-fold decrease, P < 0.05 decreased significantly in the MSMCs of the control group compared with the AM group. However, there were no significant differences in BKCa-β subunit mRNA or Kv4.2 mRNA. Conclusions: The BKCa-α mRNA and the Kv4.3 mRNA are expressed significantly higher in AM than those in the control group, that might cause the abnormal uterus smooth muscle contractility, change the microcirculation of uterus to accumulate the inflammatory factors, impair the endometrium further, and aggravate the pain.

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

    Science.gov (United States)

    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.

  14. Effects of pseudophosphorylation mutants on the structural dynamics of smooth muscle myosin regulatory light chain

    Science.gov (United States)

    Espinoza-Fonseca, L. Michel; Colson, Brett A.; Thomas, David D.

    2014-01-01

    We have performed 50 independent molecular dynamics (MD) simulations to determine the effect of pseudophosphorylation mutants on the structural dynamics of smooth muscle myosin (SMM) regulatory light chain (RLC). We previously showed that the N-terminal phosphorylation domain of RLC simultaneously populates two structural states in equilibrium, closed and open, and that phosphorylation at S19 induces a modest shift toward the open state, which is sufficient to activate smooth muscle. However, it remains unknown why pseudophosphorylation mutants poorly mimic phosphorylation-induced activation of SMM. We performed MD simulations of unphosphorylated, phosphorylated, and three pseudophosphorylatedRLC mutants: S19E, T18D/S19D and T18E/S19E. We found that the S19E mutation does not shift the equilibrium toward the open state, indicating that simple charge replacement at position S19 does not mimic the activating effect of phosphorylation, providing a structural explanation for previously published functional data. In contrast, mutants T18D/S19D and T18E/S19E shift the equilibrium toward the open structure and partially activate in vitro motility, further supporting the model that an increase in the mol fraction of the open state is coupled to SMM motility. Structural analyses of the doubly-charged pseudophosphorylation mutants suggest that alterations in an interdomain salt bridge between residues R4 and D100 results in impaired signal transmission from RLC to the catalytic domain of SMM, which explains the low ATPase activity of these mutants. Our results demonstrate that phosphorylation produces a unique structural balance in the RLC. These observations have important implications for our understanding of the structural aspects of activation and force potentiation in smooth and striated muscle. PMID:25091814

  15. Differential effects of fatty acids on glycolysis and glycogen metabolism in vascular smooth muscle.

    Science.gov (United States)

    Barron, J T; Kopp, S J; Tow, J P; Parrillo, J E

    1991-07-10

    The effects of fatty acids of different chain lengths on aerobic glycolysis, lactic acid production, glycogen metabolism and contractile function of vascular smooth muscle were investigated. Porcine carotid artery segments were treated with 50 microM iodoacetate and perchloric acid tissue extracts were then analyzed by 31P-NMR spectroscopy to observe the accumulation of phosphorylated glycolytic intermediates so that the activity of the Embden-Myerhof pathway could be tracked under various experimental paradigms. Aerobic glycolysis and lactate production in resting arteries were almost completely inhibited with 0.5 mM octanoate, partially inhibited with 0.5 mM acetate and unaffected by 0.5 mM palmitate. Inhibition of glycolysis by octanoate was not attributable to inhibition of glucose uptake or glucose phosphorylation. Basal glycogen synthesis was unchanged with palmitate and acetate, but was inhibited by 52% with octanoate incubation. The characteristic glycogenolysis which occurs upon isometric contraction with 80 mM KCl in the absence of fatty acid in the medium was not demonstrable in the presence of any of the fatty acids tested. Glycogen sparing was also demonstrable in norepinephrine contractions with octanoate and acetate, but not with palmitate. Additionally, norepinephrine-stimulated isometric contraction was associated with enhanced synthesis of glycogen amounting to 6-times the basal rate in medium containing octanoate. Contractile responses to norepinephrine were attenuated by 20% in media containing fatty acids. Thus, fatty acids significantly alter metabolism and contractility of vascular smooth muscle. Fatty acids of different chain lengths affect smooth muscle differentially; the pattern of substrate utilization during contraction depends on the contractile agonist and the fatty acid present in the medium.

  16. Diversity of K+ channels in circular smooth muscle of opossum lower esophageal sphincter.

    Science.gov (United States)

    Zhang, Y; Paterson, W G

    2001-07-01

    We previously demonstrated that a balance of K+ and Ca2+-activated Cl- channel activity maintained the basal tone of circular smooth muscle of opossum lower esophageal sphincter (LES). In the current studies, the contribution of major K+ channels to the LES basal tone was investigated in circular smooth muscle of opossum LES in vitro. K+ channel activity was recorded in dispersed single cells at room temperature using patch-clamp recordings. Whole-cell patch-clamp recordings displayed an outward current beginning to activate at -60 mV by step test pulses lasting 400 ms (-120 mV to +100 mV) with increments of 20 mV from holding potential of -80 mV ([K+]I = 150 mM, [K+]o = 2.5 mM). However, no inward rectification was observed. The outward current peaked within 50 ms and showed little or no inactivation. It was significantly decreased by bath application of nifedipine, tetraethylammonium (TEA), 4-aminopyridine (4-AP), and iberiotoxin (IBTN). Further combination of TEA with 4-AP, nifedipine with 4-AP, and IBTN with TEA, or vice versa, blocked more than 90% of the outward current. Ca2+-sensitive single channels were recorded at asymetrical K+ gradients in cell-attached patch-clamp configurations (100.8+/-3.2 pS, n = 8). Open probability of the single channels recorded in inside-out patch-clamp configurations were greatly decreased by bath application of IBTN (100 nM) (Vh = -14.4+/-4.8 mV in control vs. 27.3+/-0.1 mV, n = 3, P < 0.05). These data suggest that large conductance Ca2+-activated K+ and delayed rectifier K+ channels contribute to the membrane potential, and thereby regulate the basal tone of opossum LES circular smooth muscle.

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

    Science.gov (United States)

    Liu, C Y; Sturek, M

    1996-06-01

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

  18. Cholinergic facilitation of neurotransmission to the smooth muscle of the guinea-pig prostate gland.

    Science.gov (United States)

    Lau, W A; Pennefather, J N; Mitchelson, F J

    2000-07-01

    1. Functional experiments have been conducted to assess the effects of acetylcholine and carbachol, and the receptors on which they act to facilitate neurotransmission to the stromal smooth muscle of the prostate gland of the guinea-pig. 2. Acetylcholine and carbachol (0.1 microM - 0.1 mM) enhanced contractions evoked by trains of electrical field stimulation (20 pulses of 0.5 ms at 10 Hz every 50 s with a dial setting of 60 V) of nerve terminals within the guinea-pig isolated prostate. In these concentrations they had negligible effects on prostatic smooth muscle tone. 3. The facilitatory effects of acetylcholine, but not those of carbachol, were further enhanced in the presence of physostigmine (10 microM). 3. The facilitatory effects of carbachol were unaffected by the neuropeptide Y Y(1) receptor antagonist BIBP 3226 ((R)-N(2)-(diphenylacetyl)-N-[(4-hydroxyphenyl)methyl]-arginina mide) (0.3 microM, n=3) or suramin (100 microM, n=5). Prazosin (0.1 microM, n=5) and guanethidine (10 microM, n=5) alone and in combination (n=4), reduced responses to field stimulation and produced rightward shifts of the log concentration-response curves to carbachol. 4. The rank orders of potency of subtype-preferring muscarinic receptor antagonists in inhibiting the facilitatory actions of acetylcholine and carbachol were: pirenzepine > HHSiD (hexahydrosiladifenidol) > pF-HHSiD (para-fluoro-hexahydrosiladifenidol)>/= 5 himbacine, and pirenzepine > HHSiD > himbacine>/= 5 pF-HHSiD, respectively. These profiles suggest that muscarinic receptors of the M(1)-subtype mediate the facilitatory effects of acetylcholine and carbachol on neurotransmission to the smooth muscle of the guinea-pig prostate.

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

    Science.gov (United States)

    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.

  20. Regulation of Guinea Pig Detrusor Smooth Muscle Excitability by 17β-Estradiol: The Role of the Large Conductance Voltage- and Ca2+-Activated K+ Channels.

    Science.gov (United States)

    Provence, Aaron; Hristov, Kiril L; Parajuli, Shankar P; Petkov, Georgi V

    2015-01-01

    Estrogen replacement therapies have been suggested to be beneficial in alleviating symptoms of overactive bladder. However, the precise regulatory mechanisms of estrogen in urinary bladder smooth muscle (UBSM) at the cellular level remain unknown. Large conductance voltage- and Ca2+-activated K+ (BK) channels, which are key regulators of UBSM function, are suggested to be non-genomic targets of estrogens. This study provides an electrophysiological investigation into the role of UBSM BK channels as direct targets for 17β-estradiol, the principle estrogen in human circulation. Single BK channel recordings on inside-out excised membrane patches and perforated whole cell patch-clamp were applied in combination with the BK channel selective inhibitor paxilline to elucidate the mechanism of regulation of BK channel activity by 17β-estradiol in freshly-isolated guinea pig UBSM cells. 17β-Estradiol (100 nM) significantly increased the amplitude of depolarization-induced whole cell steady-state BK currents and the frequency of spontaneous transient BK currents in freshly-isolated UBSM cells. The increase in whole cell BK currents by 17β-estradiol was eliminated upon blocking BK channels with paxilline. 17β-Estradiol (100 nM) significantly increased (~3-fold) the single BK channel open probability, indicating direct 17β-estradiol-BK channel interactions. 17β-Estradiol (100 nM) caused a significant hyperpolarization of the membrane potential of UBSM cells, and this hyperpolarization was reversed by blocking the BK channels with paxilline. 17β-Estradiol (100 nM) had no effects on L-type voltage-gated Ca2+ channel currents recorded under perforated patch-clamp conditions. This study reveals a new regulatory mechanism in the urinary bladder whereby BK channels are directly activated by 17β-estradiol to reduce UBSM cell excitability.

  1. Device for Investigation of Mechanical Tension of Isolated Smooth Muscle Vessels and Airway Segments of Animals

    Science.gov (United States)

    Aleinik, A.; Karpovich, N.; Turgunova, N.; Nosarev, A.

    2016-11-01

    For the purpose of testing and the search for new drug compounds, designed to heal many human diseases, it is necessary to investigate the deformation of experimental tissue samples under influence of these drugs. For this task a precision force sensor for measuring the mechanical tension, produced by isolated ring segments of blood vessels and airways was created. The hardware and software systems for the study of changes in contractile responses of the airway smooth muscles and blood vessels of experimental animals was developed.

  2. The pharmacological properties of K+ currents from rabbit isolated aortic smooth muscle cells.

    OpenAIRE

    Halliday, F. C.; Aaronson, P. I.; Evans, A. M.; Gurney, A M

    1995-01-01

    1. Using the whole-cell patch-clamp technique, the effects of several K+ channel blocking drugs on K+ current recorded from rabbit isolated aortic smooth muscle cells were investigated. 2. Upon depolarization from -80 mV, outward K+ current composed of several distinct components were observed: a transient, 4-aminopyridine (4-AP)-sensitive component (I1) and a sustained component (Isus), comprising a 4-AP-sensitive delayed rectifier current (IK(V)), and a noisy current which was sensitive to ...

  3. Pharmacological role of atorvastatin in myocardium and smooth muscle progenitor cells

    Directory of Open Access Journals (Sweden)

    Suresh Kanna

    2016-06-01

    Full Text Available Atorvastatin is a synthetic 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA reductase inhibitor with a great potency in the reduction of lipids and it has been well documented in both primary and secondary prevention studies. It exhibits pleiotropic properties in both in-vitro and in vivo conditions. Conversely, atorvastatin remain under-utilized in several situations. The main objective of this review is to focuses the pharmacological benefits, pleiotropic properties of the atorvastatin related to smooth muscle proliferation and myocardium. [Int J Basic Clin Pharmacol 2016; 5(3.000: 605-608

  4. Signaling and regulation of G protein-coupled receptors in airway smooth muscle

    Directory of Open Access Journals (Sweden)

    Penn Raymond B

    2003-03-01

    Full Text Available Abstract Signaling through G protein-coupled receptors (GPCRs mediates numerous airway smooth muscle (ASM functions including contraction, growth, and "synthetic" functions that orchestrate airway inflammation and promote remodeling of airway architecture. In this review we provide a comprehensive overview of the GPCRs that have been identified in ASM cells, and discuss the extent to which signaling via these GPCRs has been characterized and linked to distinct ASM functions. In addition, we examine the role of GPCR signaling and its regulation in asthma and asthma treatment, and suggest an integrative model whereby an imbalance of GPCR-derived signals in ASM cells contributes to the asthmatic state.

  5. Effects of pinacidil on proliferation of cultured rabbit airway smooth muscle cells induced by endothelin-1

    Institute of Scientific and Technical Information of China (English)

    WANG Hong; XIE Wei-ping; QI Xu; ZHANG Xi-long

    2005-01-01

    @@ It has been found that the potassium channel dysfunction of the membrane of airway smooth muscle cells (ASMCs) is closely associated with proliferation of ASMCs.1 Preliminary research has demonstrated that pinacidil, an ATP sensitive potassium channel (KATP) opener, could play a remarkable role in the prevention and treatment of antigen induced bronchial asthma in guinea pigs.2 This study was designed to investigate further the role and molecular mechanism of the proliferation of ASMCs: a chief pathological change of the nonacute phase of bronchial asthmatic episodes.

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

    Energy Technology Data Exchange (ETDEWEB)

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

    1987-10-01

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

  7. Roles of NHE-1 in the proliferation and apoptosis of pulmonary artery smooth muscle cells in rats

    Institute of Scientific and Technical Information of China (English)

    姚伟; 钱桂生; 杨晓静

    2002-01-01

    Objective To evaluate the roles of Na+/H+ exchanger-1 (NHE-1)in the proliferation and apoptosis of pulmonary artery smooth muscle cells in rats. Methods Twenty Wistar rats were randomized into control group and 3-week hypoxic group. Intracellular pH (pHi) of the smooth muscle was determined with fluorescence measurement of the pH-sensitive dye BCECF-AM, and the expression of NHE-1 mRNA was detected by reverse transcription polymerase chain reaction (RT-PCR). Primary culture of pulmonary artery smooth muscle cells in vitro was performed. In situ cell death detection kit (TUNEL) was used for studying the effect of specific NHE-1 inhibitor-dimethyl amiloride (DMA) on the apoptosis of muscle cells which had intracellular acidification. Results pHi value and NHE-1 mRNA expression of pulmonary artery smooth muscle cells were significantly higher in the hypoxic group than in the control group (P<0.01, P<0.001). DMA elevated the apoptotic ratio remarkably. The effect was enhanced when DMA concentration increased and the time prolonged. Conclusions With the function of adjusting pHi, NHE-1 may play an important role in the proliferation and apoptosis of pulmonary artery smooth muscle cells.

  8. Focal adhesion kinase antisense oligodeoxynucleotides inhibit human pulmonary artery smooth muscle cells proliferation and promote human pulmonary artery smooth muscle cells apoptosis

    Institute of Scientific and Technical Information of China (English)

    LIN Chun-long; ZHANG Zhen-xiang; XU Yong-jian; NI Wang; CHEN Shi-xin

    2005-01-01

    Background Pulmonary artery smooth muscle cell (PASMC) proliferation plays an important role in pulmonary vessel structural remodelling. At present, the mechanisms related to proliferation of PASMCs are not clear. Focal adhesion kinase (FAK) is a widely expressed nonreceptor protein tyrosine kinase. Recent research indicates that FAK is implicated in signalling pathways which regulate cytoskeletal organization, adhesion, migration, survival and proliferation of cells. Furthermore, there are no reports about the role of FAK in human pulmonary artery smooth muscle cells (HPASMCs). We investigated whether FAK takes part in the intracellular signalling pathway involved in HPASMCs proliferation and apoptosis, by using antisense oligodeoxynucleotides (ODNs) to selectively suppress the expression of FAK protein.Methods Cultured HPASMCs stimulated by fibronectin (40 μg/ml) were passively transfected with ODNs, sense FAK, mismatch sense and antisense-FAK respectively. Expression of FAK, Jun NH2-terminal kinase (JNK), cyclin-dependent kinase 2 (CDK 2) and caspase-3 proteins were detected by immunoprecipitation and Western blots. Cell cycle and cell apoptosis were analysed by flow cytometry. In addition, cytoplasmic FAK expression was detected by immunocytochemical staining.Results When compared with mismatch sense group, the protein expressions of FAK, JNK and CDK 2 in HPASMCs decreased in antisense-FAK ODNs group and increased in sense-FAK ODNs group significantly. Caspase-3 expression upregulated in HPASMCs when treated with antisense ODNs and downregulated when treated with sense ODNs. When compared with mismatch sense ODNs group, the proportion of cells at G1 phase decreased significantly in sense ODNs group, while the proportion of cells at S phase increased significantly. In contrast, compared with mismatch sense ODNs group, the proportion of cells at G1 phase was increased significantly in antisense-FAK ODNs group. The level of cell apoptosis in antisense-FAK group

  9. Multiple skeletal muscle metastases in a case of transitional cell carcinoma of bladder detected by F-18 FDG PET/CT

    Energy Technology Data Exchange (ETDEWEB)

    Kashyap, Raghava; Mittal, Bhagwant Rai; Chakraborty, Dhritiman; Bhattacharya, Anish; Singh, Baljinder [Institute of Medical Education and Research, Chandigarh (India)

    2010-12-15

    We present a case of poorly differentiated muscle invasive transitional cell carcinoma in a 64-year-old male diagnosed with FDG-avid mass in the urinary bladder wall and multiple skeletal muscles visualised on F-18 FDG PET/CT

  10. Uhrf2 is important for DNA damage response in vascular smooth muscle cells.

    Science.gov (United States)

    Luo, Tao; Cui, Shijun; Bian, Chunjing; Yu, Xiaochun

    2013-11-08

    Emerging evidence shows that Uhrf1 plays an important role in DNA damage response for maintaining genomic stability. Interestingly, Uhrf1 has a paralog Uhrf2 in mammals. Uhrf1 and Uhrf2 share similar domain architectures. However, the role of Uhrf2 in DNA damage response has not been studied yet. During the analysis of the expression level of Uhrf2 in different tissues, we found that Uhrf2 is highly expressed in aorta and aortic vascular smooth muscle cells. Thus, we studied the role of Uhrf2 in DNA damage response in aortic vascular smooth muscle cells. Using laser microirradiation, we found that like Uhrf1, Uhrf2 was recruited to the sites of DNA damage. We dissected the functional domains of Uhrf2 and found that the TTD, PHD and SRA domains are important for the relocation of Uhrf2 to the sites of DNA damage. Moreover, depletion of Uhrf2 suppressed DNA damage-induced H2AX phosphorylation and DNA damage repair. Taken together, our results demonstrate the function of Uhrf2 in DNA damage response.

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

    Science.gov (United States)

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

    2016-03-01

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

  12. American ginseng inhibits vascular smooth muscle cell proliferation via suppressing Jak/Stat pathway

    Science.gov (United States)

    Wu, Qi; Wang, Wenjuan; Li, Siying; Nagarkatti, Prakash; Nagarkatti, Mitzi; Windust, Anthony; Wang, Xing Li; Tang, Dongqi; Cui, Taixing

    2014-01-01

    Ethnopharmcological relevance Ginseng, a folk medicine which has been used for thousands of years in Asia, has been promoted for the treatment or prevention of health problems including cardiovascular disease. However, the molecular mechanism of ginseng-induced cardiovascular protection is unclear. Thus, we investigated signaling mechanism by which American ginseng inhibits vascular smooth muscle cell (VSMC) proliferation, a key feature of diverse vascular disease. Materials and methods A standardized crude extract of American ginseng was supplied by the National Research Council of Canada, Institute for National Measurement Standards. Rat aortic smooth muscle cells (RASMCs) were exposed to fetal bovine serum (FBS), platelet derived growth factor (PDGF), insulin, or angiotensin II (Ang II) to induce proliferation that was examined by measuring DNA synthesis and cell numbers. Western blot was applied to determine the activations of Jak, Stat, Akt, and ERK. Results American ginseng inhibited RASMC proliferation induced by FBS, PDGF, insulin or Ang II. American ginseng slightly increased both basal and FBS-, PDGF- or Ang II-induced activities of Akt and ERK in RASMCs; however, it dramatically inhibited the activation of Jak2 and Stat3. Conclusion These results demonstrate that American ginseng inhibits VSMC proliferation through suppressing the Jak/Stat pathway. PMID:23041701

  13. Possible Mechanisms for Functional Antagonistic Effect of Ferula assafoetida on Muscarinic Receptors in Tracheal Smooth Muscle

    Science.gov (United States)

    Kiyanmehr, Majid; Boskabady, Mohammad Hossein; Khazdair, Mohammad Reza; Hashemzehi, Milad

    2016-01-01

    Background The contribution of histamine (H1) receptors inhibitory and/or β-adrenoceptors stimulatory mechanisms in the relaxant property of Ferula assa-foetida. (F. asafoetida) was examined in the present study. Methods We evaluated the effect of three concentrations of F. asafoetida extract (2.5, 5, and 10 mg/mL), a muscarinic receptors antagonist, and saline on methacholine concentration-response curve in tracheal smooth muscles incubated with β-adrenergic and histamine (H1) (group 1), and only β-adrenergic (group 2) receptors antagonists. Results EC50 values in the presence of atropine, extract (5 and 10 mg/mL) and maximum responses to methacholine due to the 10 mg/mL extract in both groups and 5 mg/mL extract in group 1 were higher than saline (P < 0.0001, P = 0.0477, and P = 0.0008 in group 1 and P < 0.0001, P = 0.0438, and P = 0.0107 in group 2 for atropine, 5 and 10 mg/mL extract, respectively). Values of concentration ratio minus one (CR-1), in the presence of extracts were lower than atropine in both groups (P = 0.0339 for high extract concentration in group 1 and P < 0.0001 for other extract concentrations in both groups). Conclusion Histamine (H1) receptor blockade affects muscarinic receptors inhibitory property of F. asafoetida in tracheal smooth muscle PMID:27540324

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

    Science.gov (United States)

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

    1988-01-01

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

  15. Relationship of adrenomedullin expression and microvessel density and prognosis in smooth muscle tumor of uterus

    Institute of Scientific and Technical Information of China (English)

    JIANG Yuan; TIAN Xuehong; YUAN Jie; JIN Yuemei; TAN Yusong

    2007-01-01

    The aim of this paper was to investigate the relationship between the expression of adrenomedullin(ADM)and microvessel density(MVD)and prognosis in smooth muscle tumor of uterus.The expression of ADM was detected using immunohistochemical staining in specimens from 15 normal controls,28 eases of uterine leiomyoma(LE)and 19 eases of uterine leiomyosarcoma(LES).The MVD was assayed by immunostainting with CD34.There was a positive correlation between the ADM expression and MVD in LE and LES respectively(rs=0.823,P<0.01;rs=0.793,P<0.01).The expression of ADM in LE was statistically lower than that in LES(P<0.05).There was a positive correlation between the ADM expression and mitotic figures in LES(P<0.05):the more mitotic figures,the higher levels of the ADM expression and poor prognosis.The ADM is an important angiogenic factor in smooth muscle tumor of uterus.The ADM can be used as an accessory marker in estimating the malignant potency of LE and judging the prognosis of LES,and as a novel molecular target of anti-angiogenic and anticarcinogenic strategies.

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

    Science.gov (United States)

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

    2016-03-01

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

  17. Propylthiouracil, independent of its antithyroid effect, promotes vascular smooth muscle cells differentiation via PTEN induction.

    Science.gov (United States)

    Chen, Wei-Jan; Pang, Jong-Hwei S; Lin, Kwang-Huei; Lee, Dany-Young; Hsu, Lung-An; Kuo, Chi-Tai

    2010-01-01

    Propylthiouracil (PTU), independent of its antithyroid effect, is recently found to have an antiatherosclerotic effect. The aim of this study is to determine the impact of PTU on phenotypic modulation of vascular smooth muscle cells (VSMCs), as phenotypic modulation may contribute to the growth of atherosclerotic lesions and neointimal formation after arterial injury. Propylthiouracil reduced neointimal formation in balloon-injured rat carotid arteries. In vitro, PTU may convert VSMCs from a serum-induced dedifferentiation state to a differentiated state, as indicated by a spindle-shaped morphology and an increase in the expression of SMC differentiation marker contractile proteins, including calponin and smooth muscle (SM)-myosin heavy chain (SM-MHC). Transient transfection studies in VSMCs demonstrated that PTU induced the activity of SMC marker genes (calponin and SM-MHC) promoters, indicating that PTU up-regulates these genes expression predominantly at the transcriptional level. Furthermore, PTU enhanced the expression of PTEN and inhibition of PTEN by siRNA knockdown blocked PTU-induced activation of contractile proteins expression and promoter activity. In the rat carotid injury model, PTU reversed the down-regulation of contractile proteins and up-regulated PTEN in the neointima induced by balloon injury. Propylthiouracil promotes VSMC differentiation, at lest in part, via induction of the PTEN-mediated pathway. These findings suggest a possible mechanism by which PTU may contribute to its beneficial effects on atherogenesis and neointimal formation after arterial injury.

  18. Airway smooth muscle dynamics: a common pathway of airway obstruction in asthma.

    Science.gov (United States)

    An, S S; Bai, T R; Bates, J H T; Black, J L; Brown, R H; Brusasco, V; Chitano, P; Deng, L; Dowell, M; Eidelman, D H; Fabry, B; Fairbank, N J; Ford, L E; Fredberg, J J; Gerthoffer, W T; Gilbert, S H; Gosens, R; Gunst, S J; Halayko, A J; Ingram, R H; Irvin, C G; James, A L; Janssen, L J; King, G G; Knight, D A; Lauzon, A M; Lakser, O J; Ludwig, M S; Lutchen, K R; Maksym, G N; Martin, J G; Mauad, T; McParland, B E; Mijailovich, S M; Mitchell, H W; Mitchell, R W; Mitzner, W; Murphy, T M; Paré, P D; Pellegrino, R; Sanderson, M J; Schellenberg, R R; Seow, C Y; Silveira, P S P; Smith, P G; Solway, J; Stephens, N L; Sterk, P J; Stewart, A G; Tang, D D; Tepper, R S; Tran, T; Wang, L

    2007-05-01

    Excessive airway obstruction is the cause of symptoms and abnormal lung function in asthma. As airway smooth muscle (ASM) is the effecter controlling airway calibre, it is suspected that dysfunction of ASM contributes to the pathophysiology of asthma. However, the precise role of ASM in the series of events leading to asthmatic symptoms is not clear. It is not certain whether, in asthma, there is a change in the intrinsic properties of ASM, a change in the structure and mechanical properties of the noncontractile components of the airway wall, or a change in the interdependence of the airway wall with the surrounding lung parenchyma. All these potential changes could result from acute or chronic airway inflammation and associated tissue repair and remodelling. Anti-inflammatory therapy, however, does not "cure" asthma, and airway hyperresponsiveness can persist in asthmatics, even in the absence of airway inflammation. This is perhaps because the therapy does not directly address a fundamental abnormality of asthma, that of exaggerated airway narrowing due to excessive shortening of ASM. In the present study, a central role for airway smooth muscle in the pathogenesis of airway hyperresponsiveness in asthma is explored.

  19. beta. -Adrenoceptors in human tracheal smooth muscle: characteristics of binding and relaxation

    Energy Technology Data Exchange (ETDEWEB)

    van Koppen, C.J.; Hermanussen, M.W.; Verrijp, K.N.; Rodrigues de Miranda, J.F.; Beld, A.J.; Lammers, J.W.J.; van Ginneken, C.A.M.

    1987-06-29

    Specific binding of (/sup 125/I)-(-)-cyanopindolol to human tracheal smooth muscle membranes was saturable, stereo-selective and of high affinity (K/sub d/ = 5.3 +/- 0.9 pmol/l and R/sub T/ = 78 +/- 7 fmol/g tissue). The ..beta../sub 1/-selective antagonists atenolol and LK 203-030 inhibited specific (/sup 125/I)-(-)-cyanopindolol binding according to a one binding site model with low affinity in nearly all subjects, pointing to a homogeneous BETA/sub 2/-adrenoceptor population. In one subject using LK 203-030 a small ..beta../sub 1/-adrenoceptor subpopulation could be demonstrated. The beta-mimetics isoprenaline, fenoterol, salbutamol and terbutaline recognized high and low affinity agonist binding sites. Isoprenaline's pK/sub H/- and pK/sub L/-values for the high and low affinity sites were 8.0 +/- 0.2 and 5.9 +/- 0.3 respectively. In functional experiments isoprenaline relaxed tracheal smooth muscle strips having intrinsic tone with a pD/sub 2/-value of 6.63 +/- 0.19. 32 references, 4 figures, 2 tables.

  20. Relaxant Effects of Matrine on Aortic Smooth Muscles of Guinea Pigs

    Institute of Scientific and Technical Information of China (English)

    JIE ZHENG; PING ZHENG; XU ZHOU; LIN YAN; RU ZHOU; XUE-YAN FU; GUI-DONG DAI

    2009-01-01

    Objectives To determine whether matrine, a kind of traditional Chinese medicinal alkaloid, can relax the aortic smooth muscles isolated from guinea pigs and to investigate the mechanism of its relaxant effects. Methods Phenylephrine or potassium chloride concentration-dependent relaxation response of aortic smooth muscles to matrine was studied in the precontracted guinea pigs. Results Matrine (lx104 mol/L-3.3x10-3mol/L) relaxed the endothelium-denuded aortic rings pre-contracted sub-maximally with phenylephrine, in a concentration-dependent manner, and its pre-incubation (3.3x10-3mol/L) produced a significant rightward shift in the phenylephrine dose-response curve, but had no effects on the potassium chloride-induced contraction. The anti-contractile effect of matrine was not reduced by the highly selective ATP-dependent K+ channel blocker glibenclamide (10-5mol/L), either by the non-selective K+channel blocker tetraethylammonium (103mol/L), or by theβ-antagonist propranolol (105 mol/L). In either "normal" or "Ca2+-free'' bathing medium, the phenylephrine-induced contraction was attenuated by matrine (3.3x103 mol/L), indicating that the vasorelaxation was due to inhibition of intracellular and extracellular Ca2+ mobilization. Conclusion Matrine inhibits phenylephrine-induced contractions by inhibiting activation of a-adrenoceptor and interfering with the release of intracellular Ca2+ and the influx of extracellular Ca2+.

  1. Airway Responsiveness: Role of Inflammation, Epithelium Damage and Smooth Muscle Tension

    Directory of Open Access Journals (Sweden)

    K. I. Gourgoulianis

    1999-01-01

    Full Text Available The purpose of this study was the effect of epithelium damage on mechanical responses of airway smooth muscles under different resting tension. We performed acetylcholine (ACh (10-5M-induced contraction on tracheal strips from 30 rabbits in five groups (0.5, 1, 1.5, 2 and 2.5 g before and after epithelium removal. At low resting tension (0.5-1.5g, the epithelium removal decreased the ACh-induced contractions. At 2g resting tension, the epithelium removal increased the ACh-induced contractions of airways with intact epithelium about 20%. At 2.5 g resting tension, the elevation of contraction is about 25% (p<0.01. Consequently, after epithelium loss, the resting tension determines the airway smooth muscles responsiveness. In asthma, mediators such as ACh act on already contracted inflammatory airways, which results in additional increase of contraction. In contrast, low resting tension, a condition that simulates normal tidal breathing, protects from bronchoconstriction even when the epithelium is damaged.

  2. May Sonic Hedgehog proteins be markers for malignancy in uterine smooth muscle tumors?

    Science.gov (United States)

    Garcia, Natalia; Bozzini, Nilo; Baiocchi, Glauco; da Cunha, Isabela Werneck; Maciel, Gustavo Arantes; Soares Junior, José Maria; Soares, Fernando Augusto; Baracat, Edmund Chada; Carvalho, Katia Candido

    2016-04-01

    Several studies have demonstrated that the Sonic Hedgehog signaling pathway (SHH) plays an important role in tumorigenesis and cellular differentiation. We analyzed the protein expression of SHH pathway components and evaluated whether their profile could be useful for the diagnosis, prognosis, or prediction of the risk of malignancy for uterine smooth muscle tumors (USMTs). A total of 176 samples (20 myometrium, 119 variants of leiomyoma, and 37 leiomyosarcoma) were evaluated for the protein expression of the SHH signaling components, HHIP1 (SHH inhibitor), and BMP4 (SHH target) by immunohistochemistry. Western blot analysis was performed to verify the specificity of the antibodies. We grouped leiomyoma samples into conventional leiomyomas and unusual leiomyomas that comprise atypical, cellular, mitotically active leiomyomas and uterine smooth muscle tumors of uncertain malignant potential. Immunohistochemical analysis showed that SMO, SUFU, GLI1, GLI3, and BMP4 expression gradually increased depending on to the histologic tissue type. The protein expression of SMO, SUFU, and GLI1 was increased in unusual leiomyoma and leiomyosarcoma samples compared to normal myometrium. The inhibitor HHIP1 showed higher expression in myometrium, whereas only negative or basal expression of SMO, SUFU, GLI1, and GLI3 was detected in these samples. Strong expression of SHH was associated with poorer overall survival. Our data suggest that the expression of SHH proteins can be useful for evaluating the potential risk of malignancy for USMTs. Moreover, GLI1 and SMO may serve as future therapeutic targets for women with USMTs.

  3. [Response mechanisms of the airway smooth muscle tissue in experimental bronchial spasm].

    Science.gov (United States)

    Zashikhin, A L; Agafonov, Iu V; Barmina, A O

    2009-01-01

    This investigation was aimed at the complex evaluation of the reactivity mechanisms of bronchial smooth muscle tissue (SMT) in experimental bronchial spasm. Morphometric, cytospectrophotometric and electron microscopical analysis demonstrated the presence of three types of smooth muscle cells (SMC) within the bronchial SMT (small, medium, large), that differed in their linear and metabolic parameters. The findings of this study indicate that under the conditions of experimental bronchial spasm development, the ratios of SMC in bronchial SMT are changed with the increase in proportion of small SMC and the elimination of large SMC. In the dynamics of experimental bronchial spasm development, the activation of cytoplasmic synthesis as well as of DNA synthesis was detected mainly in group of small SMC. The reactive-dystrophic changes were marked at the subcellular level, that were most often identified in large SMC resulting in their elimination from population in the dynamics of an experiment. The data obtained suggest that one of the important mechanisms of airway SMT adaptation to the bronchial spasm development is a dynamic reorganization of SMC population.

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

    Science.gov (United States)

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

    2004-03-01

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

  5. Phytoncide, Nanochemicals from Chamaecyparis obtusa, Inhibits Proliferation and Migration of Vascular Smooth Muscle Cells.

    Science.gov (United States)

    Lim, Leejin; Jang, Young-Su; Yun, Je-Jung; Song, Heesang

    2015-01-01

    Phytoncide, nanochemicals extracted from Chamaecyparis obtusa (C. obtusa), is reported to possess many pharmacological activities including immunological stimulating, anti-cancer, antioxidant, and antiinflammatory activities. However, the effect of phytoncide in vascuar diseases, especially on the behavior of vascular smooth muscle cells, has not yet been clearly elucidated. Therefore, in the present study, we investigated the effects of 15 kinds of phytoncide by various extraction conditions from C. obtusa on the proliferation and migration in rat aortic smooth muscle cells (RAoSMCs). First of all, we determined the concentration of each extracts not having cytotoxicity by MTT assay. We observed that the proliferation rate measured using BrdU assay was significantly reduced by supercritical fluid, steam distillation, Me-OH, and hexane extraction fraction in order with higher extent, respectively. Moreover, the treatment of above nanofractions inhibit the migration of RAoSMCs by 40%, 60%, and 30%, respectively, both in 2-D wound healing assay and 3-D boyden chamber assay. Immunoblot revealed that the phosphorylated levels of Akt and ERK were significantly reduced in nanofractions treated RAoSMCs. Taken together, these data suggest that phytoncide extracted from C. obtusa inhibits proliferation and migration in RAoSMCs via the modulation of phosphorylated levels of Akt and ERK. Therefore, phytoncide nanomolecules might be a potential therapeutic approach to prevent or treat atheroscrelosis and restenosis.

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

    Science.gov (United States)

    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.

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

    Institute of Scientific and Technical Information of China (English)

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

    2006-01-01

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

  8. Induction of interleukin-8 production by angiotensin Ⅱ in rat vascular smooth muscle cells

    Institute of Scientific and Technical Information of China (English)

    Zhi Wang; Lili Zhang; Baogui Sun; Qiuyan Dai

    2009-01-01

    Objective:Interleukin-8(IL-8) represents the prototypical chemokine that is made by a wide variety of cell types.Previously studies have suggested that angiotensin Ⅱ(Ang Ⅱ) is involved in atherogenesis through induction ofproinflammatory cytokines such as interleukin-6 or monocyte chemoattractant protein-1 (MCP-1) in vascular smooth muscle cells(VSMCs),while the role orang Ⅱ on IL-8 expression in VSMCs is poorly studied.Methods:In this study,VSMCs were isolated from the thoracic aorta of Sprague-Dawley rats.The expression of smooth muscle α-actin was confirmed by an immunohistochemical method.Semi-quantitative RT-PCR and enzyme-linked immunosorbent assay (ELISA) analyses were conducted to detect IL-8 expression.Results:In the present study we found that Ang Ⅱ significantly increased the expression of IL-8 both at the mRNA and protein levels in rat VSMCs in a dose- and time-dependent manner.Conclusion:These findings suggested that Ang Ⅱ may participate in atherosclerosis through induction of inflammatory mediator in VSMCs.

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

    Science.gov (United States)

    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 (Pmuscle cells is likely due to transient membrane disruption on initiation of flow.

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

    Energy Technology Data Exchange (ETDEWEB)

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

    2013-11-22

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

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

    Science.gov (United States)

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

    2012-01-01

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

  12. Increased activity of chondroitin sulfate-synthesizing enzymes during proliferation of arterial smooth muscle cells

    Energy Technology Data Exchange (ETDEWEB)

    Hollmann, J.; Thiel, J.; Schmidt, A.; Buddecke, E.

    1986-12-01

    Cultured arterial smooth muscle cells incorporate (/sup 35/S)sulfate into the extracellular chondroitin sulfate/dermatan sulfate containing proteoglycans at a higher rate in the phase of logarithmic growth than do non-dividing cells. The cell growth-dependent decrease in /sup 35/S incorporation with increasing cell density is accompanied by a decrease in the activity of chondroitin sulfate-synthesizing enzymes. The specific activity of xylosyl transferase, N-acetylgalactosaminyl transferase I and chondroitin sulfotransferase declines as the cells proceed from low to high densities. The corresponding correlation coefficients are 0.86, 0.91 and 0.89. The ratio of C-60H/C-40H sulfation of chondroitin shows a cell proliferation-dependent decrease indicating an inverse correlation of chondroitin 6-sulfotransferase and chondroitin 4-sulfotransferase activity. The observed changes in the expression of enzyme activities are thought to have some implications in the pathogenesis of arteriosclerosis, the initial stages of which are characterized by proliferation of arterial smooth muscle cells.

  13. The relaxant effect of Ferula assafoetida on smooth muscles and the possible mechanisms

    Directory of Open Access Journals (Sweden)

    Khazdair Mohammad Reza

    2015-04-01

    Full Text Available Asafoetida (Ferula asafoetida an oleo-gum-resin belongs to the Apiaceae family which obtained from the living underground rhizome or tap roots of the plant. F. assa-foetida is used in traditional medicine for the treatment of variety of disorders. Asafoetida is used as a culinary spice and in folk medicine has been used to treat several diseases, including intestinal parasites, weak digestion, gastrointestinal disorders, asthma and influenza. A wide range of chemical compounds including sugars, sesquiterpene coumarins and polysulfides have been isolated from this plant. This oleo-gum-resin is known to possess antifungal, anti-diabetic, anti-inflammatory, anti-mutagenic and antiviral activities. Several studies investigated the effects of F. asafoetida gum extract on the contractile responses induced by acetylcholine, methacholin, histamine and KCl on different smooth muscles. The present review summarizes the information regarding the relaxant effect of asafetida and its extracts on different smooth muscles and the possible mechanisms of this effect.

  14. Effects of Artesunate on Tracheal Smooth Muscle from the Guinea—pig

    Institute of Scientific and Technical Information of China (English)

    Mao-ShengYang; Jian-ChuXiao

    1997-01-01

    Artesunae is a derivative of qinghaosu,with a sesquiterpene structure.The specific action and the clinical uses of artesunate are on the preliminary stage,on the one hand ,artesunate has specific action of both antiinflammation and antivirus,and also has protective effect on the pulmonary alveolar macrophages,whuich may be advantageous to the treatment of the airway non-specific inflammation of asthma,.On the other hand,qinghaousu has the activities to relax vascular smooth muscle and to cause hypotension.The expectorant action,the antitussive action and the antuiasthmatic action of qinghaosu were preported.Artesunate may also have antiasthmatic activity,because the antimalarial potency of artesunate is stronger than that of qinghaosu,and Artesunate can block Ca2+ influx by inhibiting calcium-dependent chloride current.The main aims of this paper are to investigate the site,the mode,and the mechanism of artesunate action on isolated tracheal smooth muscle from the guinea-pig.

  15. Co-cultivation of human aortic smooth muscle cells with epicardial adipocytes affects their proliferation rate.

    Science.gov (United States)

    Ždychová, J; Čejková, S; Králová Lesná, I; Králová, A; Malušková, J; Janoušek, L; Kazdová, L

    2014-01-01

    The abnormal proliferation of vascular smooth muscle cells (VSMC) is thought to play a role in the pathogenesis of atherosclerosis. Adipocytes produce several bioactive paracrine substances that can affect the growth and migration of VSMCs. Our study focuses on the direct effect of the bioactive substances in conditioned media (CM) that was obtained by incubation with primary adipocyte-derived cell lines, including cell lines derived from both preadipocytes and from more mature cells, on the proliferation rate of human aortic smooth muscle cells (HAoSMCs). We used a Luminex assay to measure the adipokine content of the CM and showed that there was a higher concentration of monocyte chemoattractant protein-1 in renal preadipocyte-CM compared with the HAoSMC control (p<0.5). The addition of both renal preadipocyte- and epicardial adipocyte- CM resulted in the elevated production of vascular endothelial growth factor compared with the control HASoSMC CM (p<0.001). The adiponectin content in renal adipocyte-CM was increased compared to all the remaining adipocyte-CM (p<0.01). Moreover, the results showed a higher proliferation rate of HAoSMCs after co-culture with epicardial adipocyte-CM compared to the HAoSMC control (p<0.05). These results suggest that bioactive substances produced by adipocytes have a stimulatory effect on the proliferation of VSMCs.

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

    Institute of Scientific and Technical Information of China (English)

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

    2015-01-01

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

  17. Concurrent chemoradiotherapy improves survival outcome in muscle-invasive bladder cancer

    Energy Technology Data Exchange (ETDEWEB)

    Byun, Sang Jun; Kim, Jin Hee; Oh, Young Kee; Kim, Byung Hoon [Dongsan Medical Center, Keimyung University School of Medicine, Daegu (Korea, Republic of)

    2015-12-15

    To evaluate survival rates and prognostic factors related to treatment outcomes after bladder preserving therapy including transurethral resection of bladder tumor, radiotherapy (RT) with or without concurrent chemotherapy in bladder cancer with a curative intent. We retrospectively studied 50 bladder cancer patients treated with bladder-preserving therapy at Keimyung University Dongsan Medical Center from January 1999 to December 2010. Age ranged from 46 to 89 years (median, 71.5 years). Bladder cancer was the American Joint Committee on Cancer (AJCC) stage II, III, and IV in 9, 27, and 14 patients, respectively. Thirty patients were treated with concurrent chemoradiotherapy (CCRT) and 20 patients with RT alone. Nine patients received chemotherapy prior to CCRT or RT alone. Radiation was delivered with a four-field box technique (median, 63 Gy; range, 48.6 to 70.2 Gy). The follow-up periods ranged from 2 to 169 months (median, 34 months). Thirty patients (60%) showed complete response and 13 (26%) a partial response. All patients could have their own bladder preserved. Five-year overall survival (OS) rate was 37.2%, and the 5-year disease-free survival (DFS) rate was 30.2%. In multivariate analysis, tumor grade and CCRT were statistically significant in OS. Tumor grade was a significant prognostic factor related to OS. CCRT is also considered to improve survival outcomes. Further multi-institutional studies are needed to elucidate the impact of RT in bladder cancer.

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

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Wenrui; Kong, Hui; Zeng, Xiaoning; Wang, Jingjing; Wang, Zailiang; Yan, Xiaopei; Wang, Yanli; Xie, Weiping, E-mail: wpxie@njmu.edu.cn; Wang, Hong, E-mail: hongwang@njmu.edu.cn

    2015-08-15

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

  19. Smooth-muscle-like cells derived from human embryonic stem cells support and augment cord-like structures in vitro.

    Science.gov (United States)

    Vo, Elaine; Hanjaya-Putra, Donny; Zha, Yuanting; Kusuma, Sravanti; Gerecht, Sharon

    2010-06-01

    Engineering vascularized tissue is crucial for its successful implantation, survival, and integration with the host tissue. Vascular smooth muscle cells (v-SMCs) provide physical support to the vasculature and aid in maintaining endothelial viability. In this study, we show an efficient derivation of v-SMCs from human embryonic stem cells (hESCs), and demonstrate their functionality and ability to support the vasculature in vitro. Human ESCs were differentiated in monolayers and supplemented with platelet-derived growth factor-BB (PDGF-BB) and transforming growth factor-beta 1 (TGF-beta1). Human ESC-derived smooth-muscle-like cells (SMLCs) were found to highly express specific smooth muscle cell (SMC) markers--including alpha-smooth muscle actin, calponin, SM22, and smooth muscle myosin heavy chain--to produce and secrete fibronectin and collagen, and to contract in response to carbachol. In vitro tubulogenesis assays revealed that these hESC-derived SMLCs interacted with human endothelial progenitor cell (EPCs) to form longer and thicker cord-like structures in vitro. We have demonstrated a simple protocol for the efficient derivation of highly purified SMLCs from hESCs. These in vitro functional SMLCs interacted with EPCs to support and augment capillary-like structures (CLSs), demonstrating the potential of hESCs as a cell source for therapeutic vascular tissue engineering.

  20. The effects of cannabidiol on the antigen-induced contraction of airways smooth muscle in the guinea-pig.

    Science.gov (United States)

    Dudášová, A; Keir, S D; Parsons, M E; Molleman, A; Page, C P

    2013-06-01

    (-)-Δ(9)-Tetrahydrocannabinol has been demonstrated to have beneficial effects in the airways, but its psychoactive effects preclude its therapeutic use for the treatment of airways diseases. In the present study we have investigated the effects of (-)-cannabidiol, a non-psychoactive component of cannabis for its actions on bronchial smooth muscle in vitro and in vivo. Guinea-pig bronchial smooth muscle contractions induced by exogenously applied spasmogens were measured isometrically. In addition, contractile responses of bronchial smooth muscle from ovalbumin-sensitized guinea-pigs were investigated in the absence or presence of (-)-cannabidiol. Furthermore, the effect of (-)-cannabidiol against ovalbumin-induced airway obstruction was investigated in vivo in ovalbumin-sensitized guinea-pigs. (-)-Cannabidiol did not influence the bronchial smooth muscle contraction induced by carbachol, histamine or neurokinin A. In contrast, (-)-cannabidiol inhibited anandamide- and virodhamine-induced responses of isolated bronchi. A fatty acid amide hydrolase inhibitor, phenylmethanesulfonyl fluoride reversed the inhibitory effect of (-)-cannabidiol on anandamide-induced contractions. In addition, (-)-cannabidiol inhibited the contractile response of bronchi obtained from allergic guinea-pigs induced by ovalbumin. In vivo, (-)-cannabidiol reduced ovalbumin-induced airway obstruction. In conclusion, our results suggest that cannabidiol can influence antigen-induced airway smooth muscle tone suggesting that this molecule may have beneficial effects in the treatment of obstructive airway disorders.

  1. Inhibition of PKCalpha and rhoA translocation in differentiated smooth muscle by a caveolin scaffolding domain peptide.

    Science.gov (United States)

    Taggart, M J; Leavis, P; Feron, O; Morgan, K G

    2000-07-10

    Receptor-coupled contraction of smooth muscle involves recruitment to the plasma membrane of downstream effector molecules PKCalpha and rhoA but the mechanism of this signal integration is unclear. Caveolins, the principal structural proteins of caveolar plasma membrane invaginations, have been implicated in the organization and regulation of many signal transducing molecules. Thus, using laser scanning confocal immunofluorescent microscopy, we tested the hypothesis that caveolin is involved in smooth muscle signaling by investigating caveolin isoform expression and localization, together with the effect of a peptide inhibitor of caveolin function, in intact differentiated smooth muscle cells. All three main caveolin isoforms were identified in uterine, stomach, and ileal smooth muscles and assumed a predominantly plasma membranous localization in myometrial cells. Cytoplasmic introduction of a peptide corresponding to the caveolin-1 scaffolding domain-an essential region for caveolin interaction with signaling molecules--significantly inhibited agonist-induced translocation of both PKCalpha and rhoA. Translocation was unimpaired by a scrambled peptide and was unaltered in sham-treated cells. The membranous localization of caveolins, and direct inhibition of receptor-coupled PKCalpha and rhoA translocation by the caveolin-1 scaffolding domain, supports the concept that caveolins can regulate the integration of extracellular contractile stimuli and downstream intracellular effectors in smooth muscle.

  2. Protective effect of high-dose montelukast on salbutamol-induced homologous desensitisation in airway smooth muscle.

    Science.gov (United States)

    Fogli, Stefano; Stefanelli, Fabio; Martelli, Alma; Daniele, Simona; Testai, Lara; Calderone, Vincenzo; Trincavelli, Maria Letizia; Martini, Claudia; Breschi, Maria Cristina

    2013-12-01

    Montelukast (MK) is a potent cysteinyl-leukotriene receptor antagonist that causes dose-related improvements in chronic asthma. We sought to determine whether MK was able to prevent salbutamol-induced tolerance in airway smooth muscle. Homologous β2-adrenoceptor desensitisation models were established in guinea-pigs and in human bronchial smooth muscle cells (BSMC) by chronic salbutamol administration. Characterisation tools included measurement of the response of tracheal smooth muscle tissues to salbutamol, analysis of gene expression and receptor trafficking, evaluation of intracellular cAMP levels and phosphodiesterase (PDE) activity in human bronchial smooth muscle cells. Salbutamol-induced β2-adrenoceptor desensitisation was characterised by β2-agonist hyporesponsiveness (-30%, p salbutamol. Prolonged salbutamol treatment significantly decreased cAMP synthesis, induced a complete removal of the β2-adrenoceptor from plasma membrane with a parallel increase in the cytosol and increased PDE4D5 gene transcription and PDE activity in human bronchial smooth muscle cells. In homologously desensitised BSMC, MK 30 μM for 24 h was able to prevent salbutamol subsensitivity and such an effect was associated with inhibition of salbutamol-induced PDE4 activity and restoration of membrane β2-adrenoceptor expression and function. These findings suggest the presence of a favourable interaction between MK and β2-adrenoceptor agonists that might improve the therapeutic index of bronchodilators in patients with chronic respiratory diseases.

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

    Science.gov (United States)

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

    2004-02-20

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

  4. [Transurethral en bloc resection of non-muscle invasive bladder cancer. What is the state of the art?].

    Science.gov (United States)

    Kramer, M W; Wolters, M; Abdelkawi, I F; Merseburger, A S; Nagele, U; Gross, A; Bach, T; Kuczyk, M A; Herrmann, T R W

    2012-06-01

    Bladder cancer of the urothelium is the second most common malignancy among urological tumors. In view of a worldwide aging population and the fact that increased incidence rates are associated with higher age, new socioeconomic challenges will appear. Even nowadays the treatment of bladder cancer bears the highest lifetime treatment costs per patient among all forms of cancer. In conjunction with higher comorbidity rates among older patients urologists are facing new challenges in the treatment and care of patients with bladder cancer. The standard treatment for non-muscle invasive bladder cancer (NMIBC) is monopolar transurethral resection using resection loops (TURB). Based on experience in the surgical treatment of benign prostatic hyperplasia, different concepts of en bloc resection of bladder tumors using alternative energy resources (e.g. holmium laser, thulium laser and the water-jet HybridKnife) have been developed. Goals of new treatment modalities are reduction of perioperative and postoperative comorbidities, better pathological work-up of the specimens and increased recurrence-free survival. Postulated advantages using laser devices are a more precise cutting line as well as better hemostasis. The evidential value of this review is limited due to the lack of randomized, prospective studies. However, there is a tendency towards a limitation of perioperative and postoperative morbidities as well as higher chance of well-preserved tissues for better pathohistological evaluation using en bloc resection methods. More studies with long-term follow-up periods and better randomization are needed to clarify whether en bloc strategies provide better long-term oncological survival.

  5. Outcomes of BCG Induction in High-Risk Non-Muscle-Invasive Bladder Cancer Patients (NMIBC): A Retrospective Cohort Study

    Science.gov (United States)

    Ghauri, Rashid; Ahmed, Monis J; Shah, Muhammad F; Nasir, Irfan ul Islam; Siddiqui, Jasim; Ahmed, Irfan; Mir, Khurram

    2017-01-01

    Non-muscle-invasive bladder cancer (NMIBC) is categorized into high-risk and low-risk groups. Although, bacillus Calmette-Guerin (BCG) is the recommended adjuvant therapy of high-risk bladder tumor, optimal schedule (induction versus maintenance) of this therapy is a subject of debate. The objective was to evaluate outcomes of induction BCG in high-risk NMIBC patients at Shaukat Khanum Memorial Cancer Hospital & Research Centre, Pakistan and retrospective cohort study conducted in the department of urology, Shaukat Khanum Memorial Cancer Hospital & Research Centre, Pakistan. Three-year disease-free survival and progression-free survival was the main outcome measure. Data of 68 high-risk (Ta and T1 with G3 or high-grade subtype) bladder cancer patients who underwent transurethral resection followed by six-weekly intravesical BCG instillation was included in the study. Recurrence was described as biopsy-proven bladder cancer; whereas the presence of muscle invasion was considered as progression. Disease-free survival and progression-free survival were defined as time intervals elapsed between the starting date of BCG instillation and recurrence or progression, respectively. Kaplan-Meier curve was employed to estimate the three-year study end-points. Disease-free survival at three years was observed to be 66.2% and progression-free survival at 86.8%. The use of induction BCG alone for high-risk patients of NMIBC is a viable option both in terms of effective disease-free and progression-free survival rates. PMID:28168135

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

    Directory of Open Access Journals (Sweden)

    Heiying Jin

    Full Text Available 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.

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

    Science.gov (United States)

    Liu, Wenrui; Kong, Hui; Zeng, Xiaoning; Wang, Jingjing; Wang, Zailiang; Yan, Xiaopei; Wang, Yanli; Xie, Weiping; Wang, Hong

    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 (KATP) channels have been identified in ASMCs. Mount evidence has suggested that KATP channel openers can reduce airway hyperresponsiveness and alleviate airway remodeling. Opening K(+) channels triggers K(+) efflux, which leading to membrane hyperpolarization, preventing Ca(2+)entry through closing voltage-operated Ca(2+) channels. Intracellular Ca(2+) is the most important regulator of muscle contraction, cell proliferation and migration. K(+) efflux decreases Ca(2+) influx, which consequently influences ASMCs proliferation and migration. As a KATP 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(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 KATP channel antagonist. These findings provide a strong evidence to support that Ipt antagonize the proliferating and migrating effects of PDGF-BB on

  8. Effects of menthol on circular smooth muscle of human colon: analysis of the mechanism of action.

    Science.gov (United States)

    Amato, Antonella; Liotta, Rosa; Mulè, Flavia

    2014-10-05

    Menthol is the major constituent of peppermint oil, an herbal preparation commonly used to treat nausea, spasms during colonoscopy and irritable bowel disease. The mechanism responsible for its spasmolytic action remains unclear. The aims of this study were to investigate the effects induced by menthol on the human distal colon mechanical activity in vitro and to analyze the mechanism of action. The spontaneous or evoked-contractions of the circular smooth muscle were recorded using vertical organ bath. Menthol (0.1 mM-30 mM) reduced, in a concentration-dependent manner, the amplitude of the spontaneous contractions without affecting the frequency and the resting basal tone. The inhibitory effect was not affected by 5-benzyloxytryptamine (1 μM), a transient receptor potential-melastatin8 channel antagonist, or tetrodotoxin (1 μM), a neural blocker, or 1H-[1,2,4] oxadiazolo [4,3-a]quinoxalin-1-one (10 µM), inhibitor of nitric oxide (NO)-sensitive soluble guanylyl cyclase, or tetraethylammonium (10 mM), a blocker of potassium (K+)-channels. On the contrary, nifedipine (3 nM), a voltage-activated L-type Ca2+ channel blocker, significantly reduced the inhibitory menthol actions. Menthol also reduced in a concentration-dependent manner the contractile responses caused by exogenous application of Ca2+ (75-375 μM) in a Ca2+-free solution, or induced by potassium chloride (KCl; 40 mM). Moreover menthol (1-3 mM) strongly reduced the electrical field stimulation (EFS)-evoked atropine-sensitive contractions and the carbachol-contractile responses. The present results suggest that menthol induces spasmolytic effects in human colon circular muscle inhibiting directly the gastrointestinal smooth muscle contractility, through the block of Ca2+ influx through sarcolemma L-type Ca2+ channels.

  9. Smooth muscle cells can express cytokeratins of "simple" epithelium. Immunocytochemical and biochemical studies in vitro and in vivo.

    Science.gov (United States)

    Gown, A M; Boyd, H C; Chang, Y; Ferguson, M; Reichler, B; Tippens, D

    1988-08-01

    Cytokeratins are a set of 19 proteins that together constitute the class of intermediate filament protein expressed by epithelial cells and tumors. Using a panel of 9 different monoclonal anti-cytokeratin antibodies, the authors have performed immunocytochemistry on methanol-fixed, frozen sections and methacarn-fixed, paraffin-embedded tissue of human myometrial specimens. Anomalous cytokeratin expression (ACE) by smooth muscle cells was found in all specimens. Immunoblots of this tissue confirmed the presence of cytokeratin 19, and possibly 8. In addition, immunocytochemical studies demonstrated ACE in human fetal tissues within the intestinal muscularis and the heart, especially in the region of the aortic outflow tract, and in 8 of 19 cases of leiomyosarcoma from adults. Indirect immunofluorescence studies were also performed on cells explanted from myometrial tissue; the overwhelming majority of cells derived from these cultures were smooth muscle cells as verified by expression of muscle actins, and a subpopulation of these cells was found to be cytokeratin-positive. ACE was confirmed in vitro by double labeling experiments demonstrating simultaneous expression of muscle actins and cytokeratins within the same cell. The significance of this smooth muscle cell ACE is unknown, but it may be a phenotypic marker of smooth muscle in a proliferative state. ACE could be a source of confusion in the immunocytochemical analysis of poorly differentiated malignancies if a complete panel of antibodies is not employed.

  10. Are we following the guidelines on non-muscle invasive bladder cancer?

    Directory of Open Access Journals (Sweden)

    Leonardo Oliveira Reis

    2016-02-01

    Full Text Available ABSTRACT Objectives To evaluate the clinical practice of non-muscle invasive bladder cancer (NMIBC treatment in Brazil in relation to international guidelines: Sociedade Brasileira de Urologia (SBU, European Association of Urology (EAU and American Urological Association (AUA. Materials and Methods Cross-sectional study using questionnaires about urological practice on treatment of NMIBC during the 32nd Brazilian Congress of Urology. A total of 650 question forms were answered. Results There were 73% of complete answers (total of 476 question forms. In total, 246 urologists (51.68% lived in the southeast region and 310 (65.13% treat 1 to 3 cases of NMIBC per month. Low risk cancer: Only 35 urologists (7.5% apply the single intravesical dose of immediate chemotherapy with Mitomicin C recommended by the above guidelines. Adjuvant therapy with BCG 2 to 4 weeks after TUR is used by 167 participants (35.1% and 271 urologists (56.9% use only TUR. High risk tumors: 397 urologists (83.4% use adjuvant therapy, 375 (78.8% use BCG 2 to 4 weeks after TUR, of which 306 (64.3% referred the use for at least one year. Intravesical chemotherapy with Mitomicin C (a controversial recommendation was used by 22 urologists (4.6%. BCG dose raised a lot of discrepancies. Induction doses of 40, 80 and 120mg were referred by 105 (22%, 193 (40.4% and 54 (11.3% respectively. Maintenance doses of 40, 80 and 120mg were referred by 190 (48.7%, 144 (37.0% and 32 (8.2% urologists, respectively. Schemes of administration were also varied and the one cited by SWOG protocol was the most used: 142 (29.8%. Conclusion SBU, EAU and AUA guidelines are partially respected by Brazilian urologists, particularly in low risk tumors. In high risk tumors, concordance rates are comparable to international data. Further studies are necessary to fully understand the reasons of such disagreement.

  11. Are we following the guidelines on non-muscle invasive bladder cancer?

    Science.gov (United States)

    Reis, Leonardo Oliveira; Moro, Juliano Cesar; Ribeiro, Luis Fernando Bastos; Voris, Brunno Raphael Iamashita; Sadi, Marcos Vinicius

    2016-01-01

    ABSTRACT Objectives To evaluate the clinical practice of non-muscle invasive bladder cancer (NMIBC) treatment in Brazil in relation to international guidelines: Sociedade Brasileira de Urologia (SBU), European Association of Urology (EAU) and American Urological Association (AUA). Materials and Methods Cross-sectional study using questionnaires about urological practice on treatment of NMIBC during the 32nd Brazilian Congress of Urology. A total of 650 question forms were answered. Results There were 73% of complete answers (total of 476 question forms). In total, 246 urologists (51.68%) lived in the southeast region and 310 (65.13%) treat 1 to 3 cases of NMIBC per month. Low risk cancer: Only 35 urologists (7.5%) apply the single intravesical dose of immediate chemotherapy with Mitomicin C recommended by the above guidelines. Adjuvant therapy with BCG 2 to 4 weeks after TUR is used by 167 participants (35.1%) and 271 urologists (56.9%) use only TUR. High risk tumors: 397 urologists (83.4%) use adjuvant therapy, 375 (78.8%) use BCG 2 to 4 weeks after TUR, of which 306 (64.3%) referred the use for at least one year. Intravesical chemotherapy with Mitomicin C (a controversial recommendation) was used by 22 urologists (4.6%). BCG dose raised a lot of discrepancies. Induction doses of 40, 80 and 120mg were referred by 105 (22%), 193 (40.4%) and 54 (11.3%) respectively. Maintenance doses of 40, 80 and 120mg were referred by 190 (48.7%), 144 (37.0%) and 32 (8.2%) urologists, respectively. Schemes of administration were also varied and the one cited by SWOG protocol was the most used: 142 (29.8%). Conclusion SBU, EAU and AUA guidelines are partially respected by Brazilian urologists, particularly in low risk tumors. In high risk tumors, concordance rates are comparable to international data. Further studies are necessary to fully understand the reasons of such disagreement. PMID:27136464

  12. Could an increase in airway smooth muscle shortening velocity cause airway hyperresponsiveness?

    Science.gov (United States)

    Bullimore, Sharon R; Siddiqui, Sana; Donovan, Graham M; Martin, James G; Sneyd, James; Bates, Jason H T; Lauzon, Anne-Marie

    2011-01-01

    Airway hyperresponsiveness (AHR) is a characteristic feature of asthma. It has been proposed that an increase in the shortening velocity of airway smooth muscle (ASM) could contribute to AHR. To address this possibility, we tested whether an increase in the isotonic shortening velocity of ASM is associated with an increase in the rate and total amount of shortening when ASM is subjected to an oscillating load, as occurs during breathing. Experiments were performed in vitro using 27 rat tracheal ASM strips supramaximally stimulated with methacholine. Isotonic velocity at 20% isometric force (Fiso) was measured, and then the load on the muscle was varied sinusoidally (0.33 ± 0.25 Fiso, 1.2 Hz) for 20 min, while muscle length was measured. A large amplitude oscillation was applied every 4 min to simulate a deep breath. We found that: 1) ASM strips with a higher isotonic velocity shortened more quickly during the force oscillations, both initially (P shortening during the force oscillation protocol (P shortening with increased isotonic velocity could be explained by a change in either the cycling rate of phosphorylated cross bridges or the rate of myosin light chain phosphorylation. We conclude that, if asthma involves an increase in ASM velocity, this could be an important factor in the associated AHR.

  13. Accumulating evidence for increased velocity of airway smooth muscle shortening in asthmatic airway hyperresponsiveness.

    Science.gov (United States)

    Ijpma, Gijs; Matusovsky, Oleg; Lauzon, Anne-Marie

    2012-01-01

    It remains unclear whether airway smooth muscle (ASM) mechanics is altered in asthma. While efforts have originally focussed on contractile force, some evidence points to an increased velocity of shortening. A greater rate of airway renarrowing after a deep inspiration has been reported in asthmatics compared to controls, which could result from a shortening velocity increase. In addition, we have recently shown in rats that increased shortening velocity correlates with increased muscle shortening, without increasing muscle force. Nonetheless, establishing whether or not asthmatic ASM shortens faster than that of normal subjects remains problematic. Endobronchial biopsies provide excellent tissue samples because the patients are well characterized, but the size of the samples allows only cell level experiments. Whole human lungs from transplant programs suffer primarily from poor patient characterization, leading to high variability. ASM from several animal models of asthma has shown increased shortening velocity, but it is unclear whether this is representative of human asthma. Several candidates have been suggested as responsible for increased shortening velocity in asthma, such as alterations in contractile protein expression or changes in the contractile apparatus structure. There is no doubt that more remains to be learned about the role of shortening velocity in asthma.

  14. Transgenic overexpression of pregnancy-associated plasma protein-A in murine arterial smooth muscle accelerates atherosclerotic lesion development

    DEFF Research Database (Denmark)

    Conover, Cheryl A; Mason, Megan A; Bale, Laurie K

    2010-01-01

    of atherosclerotic lesions, we generated transgenic mice that express human PAPP-A in arterial smooth muscle. Four founder lines were characterized for transgenic human PAPP-A mRNA and protein expression, IGFBP-4 protease activity, and tissue specificity. In study I, apolipoprotein E knockout (ApoE KO) mice, a well...... from ApoE KO/Tg compared with ApoE KO mice (P smooth muscle of double ApoE KO/PAPP-A KO mice resulted in a 2.5-fold increase in lesion area (P = 0.002), without an effect...... on lesion number. PAPP-A transgene expression was associated with a significant increase in an IGF-responsive gene (P smooth muscle accelerates lesion progression in a mouse model...

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

    DEFF Research Database (Denmark)

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

    2007-01-01

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

  16. Differences in time to peak carbachol-induced contractions between circular and longitudinal smooth muscles of mouse ileum.

    Science.gov (United States)

    Azuma, Yasu-Taka; Samezawa, Nanako; Nishiyama, Kazuhiro; Nakajima, Hidemitsu; Takeuchi, Tadayoshi

    2016-01-01

    The muscular layer in the GI tract consists of an inner circular muscular layer and an outer longitudinal muscular layer. Acetylcholine (ACh) is the representative neurotransmitter that causes contractions in the gastrointestinal tracts of most animal species. There are many reports of muscarinic receptor-mediated contraction of longitudinal muscles, but few studies discuss circular muscles. The present study detailed the contractile response in the circular smooth muscles of the mouse ileum. We used small muscle strips (0.2 mm × 1 mm) and large muscle strips (4 × 4 mm) isolated from the circular and longitudinal muscle layers of the mouse ileum to compare contraction responses in circular and longitudinal smooth muscles. The time to peak contractile responses to carbamylcholine (CCh) were later in the small muscle strips (0.2 × 1 mm) of circular muscle (5.7 min) than longitudinal muscles (0.4 min). The time to peak contractile responses to CCh in the large muscle strips (4 × 4 mm) were also later in the circular muscle (3.1 min) than the longitudinal muscle (1.4 min). Furthermore, a muscarinic M2 receptor antagonist and gap junction inhibitor significantly delayed the time to peak contraction of the large muscle strips (4 × 4 mm) from the circular muscular layer. Our findings indicate that muscarinic M2 receptors in the circular muscular layer of mouse ileum exert a previously undocumented function in gut motility via the regulation of gap junctions.

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

    Science.gov (United States)

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

    2007-11-01

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

  18. Differentiation of Murine Bone Marrow-Derived Smooth Muscle Progenitor Cells Is Regulated by PDGF-BB and Collagen.

    Directory of Open Access Journals (Sweden)

    Clifford Lin

    Full Text Available Smooth muscle cells (SMCs are key regulators of vascular disease and circulating smooth muscle progenitor cells may play important roles in vascular repair or remodelling. We developed enhanced protocols to derive smooth muscle progenitors from murine bone marrow and tested whether factors that are increased in atherosclerotic plaques, namely platelet-derived growth factor-BB (PDGF-BB and monomeric collagen, can influence the smooth muscle specific differentiation, proliferation, and survival of mouse bone marrow-derived progenitor cells. During a 21 day period of culture, bone marrow cells underwent a marked increase in expression of the SMC markers α-SMA (1.93 ± 0.15 vs. 0.0008 ± 0.0003 (ng/ng GAPDH at 0 d, SM22-α (1.50 ± 0.27 vs. 0.005 ± 0.001 (ng/ng GAPDH at 0 d and SM-MHC (0.017 ± 0.004 vs. 0.001 ± 0.001 (ng/ng GAPDH at 0 d. Bromodeoxyuridine (BrdU incorporation experiments showed that in early culture, the smooth muscle progenitor subpopulation could be identified by high proliferative rates prior to the expression of smooth muscle specific markers. Culture of fresh bone marrow or smooth muscle progenitor cells with PDGF-BB suppressed the expression of α-SMA and SM22-α, in a rapidly reversible manner requiring PDGF receptor kinase activity. Progenitors cultured on polymerized collagen gels demonstrated expression of SMC markers, rates of proliferation and apoptosis similar to that of cells on tissue culture plastic; in contrast, cells grown on monomeric collagen gels displayed lower SMC marker expression, lower growth rates (319 ± 36 vs. 635 ± 97 cells/mm2, and increased apoptosis (5.3 ± 1.6% vs. 1.0 ± 0.5% (Annexin 5 staining. Our data shows that the differentiation and survival of smooth muscle progenitors are critically affected by PDGF-BB and as well as the substrate collagen structure.

  19. Modelling the elastin, collagen and smooth muscle contribution to the duodenal mechanical behaviour in patients with systemic sclerosis

    DEFF Research Database (Denmark)

    Gao, F.; Liao, Donghua; Drewes, Asbjørn Mohr;

    2009-01-01

    wall mechanics, structure and function is important. The aim was to establish a model for differentiating the biomechanical remodelling of elastin, collagen and smooth muscle in the duodenum in SS patients. A duodenal distension protocol was used in six patients and five healthy controls. A theoretical...... model for evaluating the mechanical contributions of elastin, collagen and smooth muscle tone was established. The tension-strain curves computed from pressure and cross-sectional area data were analysed. The elastic modulus of elastin, the relationship between the collagen recruitment, collagen density...

  20. A cyclic GMP-dependent calcium-activated chloride current in smooth-muscle cells from rat mesenteric resistance arteries

    DEFF Research Database (Denmark)

    Matchkov, Vladimir; Aalkjær, Christian; Nilsson, Holger

    2004-01-01

    -PET-cGMP or with a peptide inhibitor of PKG, or with the nonhydrolysable ATP analogue AMP-PNP. Under biionic conditions, the anion permeability sequence of the channel was SCN- > Br- > I- > Cl- > acetate > F- >> aspartate, but the conductance sequence was I- > Br- > Cl- > acetate > F- > aspartate = SCN-. The current had...... conditions of high calcium in the patch-pipette solution, a current similar to the latter could be identified also in the mesenteric artery smooth-muscle cells. We conclude that smooth-muscle cells from rat mesenteric resistance arteries have a novel cGMP-dependent calcium-activated chloride current, which...

  1. Inhibition of rac1 reduces PDGF-induced reactive oxygen species and proliferation in vascular smooth muscle cells.

    OpenAIRE

    2001-01-01

    In vascular smooth muscle cells, reactive oxygen species (ROS) were known to mediate platelet-derived growth factor (PDGF)-induced cell proliferation and NADH/NADPH oxidase is the major source of ROS. NADH/NADPH oxidase is controlled by rac1 in non-phagocytic cells. In this study, we examined whether the inhibition of rac1 by adenoviral-mediated gene transfer of a dominant negative rac1 gene product (Ad.N17rac1) could reduce the proliferation of rat aortic vascular smooth muscle cells (RASMC)...

  2. Ouabain sensitive Na+/K+-pump regulates other membrane transporters in the microdomain of smooth muscle cells

    DEFF Research Database (Denmark)

    Matchkov, Vladimir; Nilsson, Holger; Aalkjær, Christian

    Ouabain, a specific inhibitor of the Na+/K+-pump, has previously been shown to interfere with intercellular communication. We have demonstrated a mechanism of this action of ouabain (1). We have showed that gap junctions between vascular smooth muscle cells (SMCs) are regulated through......+/K+-pump-containing microdomain is functionally linked to KATP channels via the local ion homeostasis and that this interaction can be bidirectional (1;2). [Ca2+]i in individual SMCs was imaged simultaneously with isometric force in rat mesenteric small arteries. Paired cultured rat aortic smooth muscle cells (A7r5) were used...

  3. Oncologic Outcomes after Anterior Exenteration for Muscle Invasive Bladder Cancer in Women

    DEFF Research Database (Denmark)

    Gregg, Justin R; Emeruwa, Curran; Wong, Johnson;

    2016-01-01

    PURPOSE: We investigated oncologic and urinary outcomes after anterior exenteration for urothelial cell carcinoma in females, identifying tumor characteristics associated with female pelvic organ involvement. We hypothesized that a lack of trigonal or bladder floor tumor, intraoperative palpable ...

  4. Activation of protein kinase C accelerates contraction kinetics of airway smooth muscle.

    Science.gov (United States)

    Peiper, U; Knipp, S C; Thies, B; Henke, R

    1996-01-01

    Contraction kinetics of isolated rat tracheal smooth muscle were studied by analysing the increase of force subsequent to force-inhibiting passive length changes lasting 1 s (100 Hz, sinus, 5% of muscle length). Compared with carbachol activation, phorboldibutyrate (PDBu)-induced stimulation of protein kinase C (PKC) demonstrated no significant difference in the extent of force development in the polarized preparation [mean peak force 9.16 +/- 0.37 mN (carbachol) vs. 9.12 +/- 0.37 mN (PDBu)]. However, the time constant calculated for the slow component of post-vibration force recovery was 6.40 +/- 0.29 s after addition of PDBu vs. 22.39 +/- 1.40 s during carbachol activation, indicating a significant phorbol ester-induced acceleration of the cross-bridge cycling rate. In the K-depolarized preparation, treatment with 26.4 microM indolactam (IL) to activate PKC produced muscle relaxation (9.94 +/- 0.16 mN measured 0-30 min after the onset of depolarization vs. 4.13 +/- 0.05 mN measured during 30-60 min of IL treatment). Again, even in the presence of high sarcoplasmic Ca2+ resulting from tonic depolarization, PKC activation was associated with a distinct diminution of the time constant (25.99 +/- 0.79 s during the first 30 min of depolarization vs. 10.32 +/- 0.21 s during 30-60 min of IL treatment). In contrast, addition of 0.035 microM verapamil, 1.5 microM isoproterenol, and 32 microM dibutyryl-cAMP to the bathing medium induced relaxation without affecting the rate of post-vibration force recovery. The results suggest that the calcium-dependent signal cascade (agonist receptor/inositol trisphosphate/ Ca(2+)-calmodulin/myosin light chain kinase) hardly affects the regulation of contraction kinetics in the tonically activated intact smooth muscle preparation. PKC stimulation, however, accelerates actin/myosin interaction kinetics, possibly by inhibition of phosphatase(s).

  5. Relaxation of rabbit cavernous smooth muscle to 17β-estradiol: a non-genomic, NO-independent mechanism

    Institute of Scientific and Technical Information of China (English)

    Sae-ChulKim; Kyung-KunSeo; Soon-ChulMyung; MooYeolLee

    2004-01-01

    Aim: To investigate whether estrogen was involved in relaxation of rabbit cavernous smooth muscle.Methods: Relaxation response of the rabbit cavernous smooth muscles to 17β-estradiol (0.3, 3, 30 and 300 nmol/L) were observed in vitro. The response of the muscle strips to estrogen after incubation with either actinomycin D (10μmol/L) or L-NAME (10μmol/L) were also evaluated. Inside-out mode of patch clamp in a single smooth muscle cell was applied to investigate the Maxi-K channel activities. Results: Estrogen caused a dose-dependent relaxation of the strips precontracted with norepinephrine. The maximal response was noted about 10 minutes after treatment. Theestrogen-induced relaxation was prevented by neither actinomycin D nor L-NAME, suggesting that the response was not mediated by gene transcription or nitric oxide (NO). Application of 17β-estradiol increased the Maxi-K channelactivities. Conclusion: 17β-estradiol may be involved in relaxation of rabbit cavernous smooth muscles via a nongenomic and NO independent mechanism. 17β-estradiol stimulates Maxi-K channel of the rabbit cavernous myocyte.

  6. Photobiomodulation of vascular endothelial and smooth muscle cells in vitro with red laser light

    Science.gov (United States)

    Kipshidze, Nicholas; Keelan, Michael H., Jr.; Horn, Joseph B.; Nikolaychik, Victor

    1996-12-01

    Numerous reports suggest that low power red laser light (LPRLL) is capable of affecting cellular processes in the absence of significant thermal effect. The objective of the present study was to determine the effect of LPRLL on viability, growth, and attachment characteristics of rabbit and human aortic endothelial cells (EC) and smooth muscle cells (SMC) in vitro. All cell cultures were irradiated with single dose LPRLL using a He-Ne continuous wave laser with different energy densities. Assessment of effect on cell viability, growth, and attachment was performed utilizing Alamar Blue assay. Based upon our experiments, we conclude that: 1) stimulation and/or inhibition of cell growth and death can be obtained with LPRLL by varying the energy level, 2) LPRLL increases EC attachment, and 3) EC are more sensitive to photobiomodulation with LPRLL than SMC. These data may have significant importance leading to the establishment of new methods for phototherapy of atherosclerosis and restenosis.

  7. Direct effect of croton oil on intestinal epithelial cells and colonic smooth muscle cells

    Institute of Scientific and Technical Information of China (English)

    Xin Wang; Mei Lan; Han-Ping Wu; Yong-Quan Shi; Ju Lu; Jie Ding; Kai-Cun Wu; Jian-Ping Jin; Dai-Ming Fan

    2002-01-01

    AIM: To investigate the direct effect of croton oil (CO) onhuman intestinal epithelial cells (HIEC) and guinea pigcolonic smooth muscle cells in vitro.METHODS: Growth curves of HIEC were drawn by MTTcolorimetry. The dynamics of cell proliferation was analyzedwith flow cytometry, and morphological changes wereobserved under light and electron microscopy after long-term (6 weeks) treatment with CO. Expression of cyclo-oxygenase2 (COX-2) mRNA was detected by dot blot inHIEC treated with CO. Genes related to CO were screenedby DD-PCR, and the direct effect of CO on the contractilityof isolated guinea pig colonic smooth muscle cells wasobservedRESULTS: High concentration (20- 40 mg @ L 1) Coinhibited cell growth significantly (1, 3, 5, 7d OD sequence:(20 mg@L 1) 0.040± 0.003, 0.081 ± 0.012, 0.147± 0.022,0.024± 0.016; (40 mg@ L-1) 0.033 ± 0.044, 0.056 ± 0.012,0.104 ± 0.010, 0. 189 ± 0.006; OD eontrol 0.031 ± 0.008, 0.096± 0.012, 0.173 ± 0.009, 0.300 ± 0.016, P < 0.01), whichappeared to be related directly to the dosage. Comparedwith the control, the fraction number of cells in G1 phasedecreased from 0.60 to 0.58, while that in S phase increasedfrom 0.30 to 0.34, and DNA index also increased after 6weeks of treatment with CO (the dosage was increasedgradually from 4 to 40 rg@ L-1 ). Light microscopicobservation revealed that cells had karyomegaly, lessplasma and karyoplasm lopsidedness. Electron microscopyalso showed an increase in cell proliferation and in thequantity of abnormal nuclei with pathologic mitosis.Expression of COX-2 mRNA decreased significantly in HIECtreated with CO. Thirteen differential cDNA fragments werecloned from HIEC treated with CO, one of which was 100percent homologous with human mitochondrial cytochromeC oxidase subunit Ⅱ. The length of isolated guinea pigcolonic smooth muscle cells was significantly shortenedafter treatment with CO ( P < 0.05).CONCLUSION: At a high CO concentration ( > 20 mg@ L 1 ),cell growth and

  8. [3H]ouabain binding to cultured rat vascular smooth muscle cells.

    Science.gov (United States)

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

    1984-05-01

    The number of Na+ pump units (Bmax) and the equilibrium dissociation constant (Kd) for ouabain as well as parameters of K+ binding to the Na+ pump were examined in in vitro-grown vascular smooth muscle cells ( VSMC ) derived from Sprague-Dawley rats. The technique to measure these variables utilizes analyses of [3H]ouabain displacement from its VSMC receptors by nonlabeled ouabain and K+. The mean values for Bmax and Kd in the cultured VSMCs were 1.95 X 10(5) receptor sites per single VSMC and 2.68 X 10(-6) M, respectively. The equilibrium dissociation constant for K+ (Ki) was 0.92 mM. K+ binding to the cultured VSMCs demonstrated positive cooperativity with a Hill coefficient (n) of 1.78.

  9. Mutations in smooth muscle alpha-actin (ACTA2) lead to thoracic aortic aneurysms and dissections.

    Science.gov (United States)

    Guo, Dong-Chuan; Pannu, Hariyadarshi; Tran-Fadulu, Van; Papke, Christina L; Yu, Robert K; Avidan, Nili; Bourgeois, Scott; Estrera, Anthony L; Safi, Hazim J; Sparks, Elizabeth; Amor, David; Ades, Lesley; McConnell, Vivienne; Willoughby, Colin E; Abuelo, Dianne; Willing, Marcia; Lewis, Richard A; Kim, Dong H; Scherer, Steve; Tung, Poyee P; Ahn, Chul; Buja, L Maximilian; Raman, C S; Shete, Sanjay S; Milewicz, Dianna M

    2007-12-01

    The major function of vascular smooth muscle cells (SMCs) is contraction to regulate blood pressure and flow. SMC contractile force requires cyclic interactions between SMC alpha-actin (encoded by ACTA2) and the beta-myosin heavy chain (encoded by MYH11). Here we show that missense mutations in ACTA2 are responsible for 14% of inherited ascending thoracic aortic aneurysms and dissections (TAAD). Structural analyses and immunofluorescence of actin filaments in SMCs derived from individuals heterozygous for ACTA2 mutations illustrate that these mutations interfere with actin filament assembly and are predicted to decrease SMC contraction. Aortic tissues from affected individuals showed aortic medial degeneration, focal areas of medial SMC hyperplasia and disarray, and stenotic arteries in the vasa vasorum due to medial SMC proliferation. These data, along with the previously reported MYH11 mutations causing familial TAAD, indicate the importance of SMC contraction in maintaining the structural integrity of the ascending aorta.

  10. Oxidative modification of high density lipoprotein induced by cultured human arterial smooth muscle cells

    Institute of Scientific and Technical Information of China (English)

    江渝; 刘红; 彭家和; 叶治家; 何凤田; 董燕麟; 刘秉文

    2003-01-01

    Objective: To observe the oxidative modification of high density lipoprotein (HDL) induced by cultured human arterial smooth muscle cells (SMCs). Methods: HDL cocultured with SMCs at 37℃ in 48 h was subjected, and native HDL (N-HDL) served as control. Oxidative modification of HDL was identified by using agarose gel electrophoresis. Absorbances of conjugated diene (CD) and lipid hydroperoxide (LOOH) were measured with ultraviolet spectrophotometry at 234 and 560 nm respectively, and fluorescence intensity of thiobarbuturic acid reaction substance (TBARS) with fluorescence spectrophotometry at 550 nm emission wavelength with excitation at 515 nm. Results: In comparison with N-HDL, the electrophoretic mobility of SMCs-cocultured HDL was increased, and the contents of CD, LOOH and TBARS HDL were very significantly higher than those of the control HDL (P<0.01). Conclusion: Oxidative modification of HDL can be induced by human arterial SMCs.

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

    Science.gov (United States)

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

    2000-08-11

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

  12. Genetic analysis of blood vessel formation role of endothelial versus smooth muscle cells.

    Science.gov (United States)

    Carmeliet, P; Collen, D

    1997-11-01

    Formation of new blood vessels is vital during embryogenesis, essential for reproduction and wound healing during adulthood, and required to rescue tissue during ischemia. Neovascularization may, however, also contribute to the pathogenesis of several disorders, including tumorigenesis, diabetic vasculopathy, and chronic inflammation. Initially, blood vessels form as endothelium-lined channels by in situ differentiation of endothelial cells. Subsequently, they sprout and remodel into a highly organized and interconnected vascular network. During further maturation of the blood vessels, a sheet of primitive vascular smooth muscle cells surrounds the endothelium-lined channels, which controls endothelial cell function and provides structural support. Recent molecular analyses have identified candidate molecules that affect these processes. Their in vivo role has been further established by targeted gene manipulation in transgenic mice. This review highlights recent developments in the genetic analysis of blood vessel formation, as deduced from analysis of gene-inactivated mice. (Trends Cardiovasc Med 1997;7:271-281). © 1997, Elsevier Science Inc.

  13. Effects of drug combinations on smooth muscle cell proliferation: an isobolographic analysis.

    Science.gov (United States)

    Parry, Tom J; Thyagarajan, Rathna; Argentieri, Dennis; Falotico, Robert; Siekierka, John; Tallarida, Ronald J

    2006-02-17

    Although sirolimus is a potent inhibitor of vascular smooth muscle cell (VSMC) proliferation and is effective at preventing restenosis in the majority of clinical revascularization procedures employing sirolimus-eluting stents, some VSMC may escape the antiproliferative effects of sirolimus. The present study examines the effects of combining sirolimus with other known cell cycle-specific antiproliferative agents (cladribine, topotecan or etoposide) on cultured coronary artery VSMC proliferation and utilizes a novel isobolographic approach to determine whether sirolimus/antiproliferative agent combinations produce subadditive, additive or supraadditive potentiation of antiproliferative activity. All agents were found to inhibit coronary artery VSMC proliferation in a dose-dependent manner. Cladribine was found to potentiate the antiproliferative activity of sirolimus in either an additive or supraadditive manner, depending upon the cladribine concentration. Topotecan potentiated the sirolimus antiproliferative activity by simple additivity while etoposide yielded subadditive potentiation. The present results demonstrate the utility of isobolographic analysis for identifying and optimizing antiproliferative drug combinations.

  14. Effects of piperitenone oxide on the intestinal smooth muscle of the guinea pig

    Directory of Open Access Journals (Sweden)

    P.J.C. Sousa

    1997-06-01

    Full Text Available We investigated the effects of piperitenone oxide (PO, a major constituent of the essential oil of Mentha x villosa, on the guinea pig ileum. PO (30 to 740 µg/ml relaxed basal tonus without significantly altering the resting membrane potential. In addition, PO relaxed preparations precontracted with either 60 mM K+ or 5 mM tetraethylammonium in a concentration-dependent manner. At concentrations from 0.1 to 10 µg/ml PO potentiated acetylcholine-induced contractions, while higher concentrations (>30 µg/ml blocked this response. These higher PO concentrations also inhibited contractions induced by 60 mM K+. PO also blocked the components of acetylcholine contraction which are not sensitive to nifedipine or to solutions with nominal zero Ca2+ and EGTA. These results show that PO is a relaxant of intestinal smooth muscle and suggest that this activity may be mediated at least in part by an intracellular effect

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

    Science.gov (United States)

    Sinha, Sanjay; Iyer, Dharini; Granata, Alessandra

    2014-06-01

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

  16. Epstein-Barr virus-associated smooth muscle tumour presenting as a parasagittal brain tumour.

    Science.gov (United States)

    Ibebuike, K E; Pather, S; Emereole, O; Ndolo, P; Kajee, A; Gopal, R; Naidoo, S

    2012-11-01

    Dural-based brain tumours, apart from meningiomas, are rare. Epstein-Barr virus (EBV)-associated smooth muscle tumor (SMT) is a documented but rare disease that occurs in immunocompromized patients. These tumours may be located at unusual sites including the brain. We present a 37-year-old patient, positive for the human immunodeficiency virus (HIV), who was admitted after generalized tonic-clonic seizures. MRI and CT scan revealed a dural-based brain tumour, intraoperatively thought to be a meningioma, but with an eventual histological diagnosis of EBV-SMT. Clinically the patient was well postoperatively with a Glasgow coma scale score of 15/15 and no focal neurologic deficit. This case confirms the association between EBV and SMT in patients with HIV/acquired immunodeficiency syndrome (AIDS). It also highlights the need to include EBV-SMT in the differential diagnosis of intracranial mass lesions in patients with HIV/AIDS.

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

    Energy Technology Data Exchange (ETDEWEB)

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

    2015-09-04

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

  18. EFFICACY OF DIFFERENT RESECTIONS ON NON-MUSCLE-INVASIVE BLADDER CANCER AND ANALYSIS OF THE OPTIMAL SURGICAL METHOD.

    Science.gov (United States)

    Chen, G F; Shi, T P; Wang, B J; Wang, X Y; Zang, Q

    2015-01-01

    This study aimed to analyze the clinical efficacy of different resections in treating non-muscle-invasive bladder cancer (NMIBC), including partial cystectomy, transurethral resection of bladder tumor (TURBT) and holmium laser resection of bladder tumor. Two hundred and sixteen patients were recruited with NMIBC who were available for follow-up visits in hospital, including 62 cases treated with partial cystectomy, 90 cases treated with TURBT and 64 cases with holmium laser resection. Analysis was made on the cases with tumor relapse in the two years, on operation time, blood loss, time for indwelling urinary catheter, hospital stay and complications after operation. Results were compared to the clinical efficacy of these operation patterns. It was found that the two-year relapse rate for TURBT group, partial cystectomy group and Holmium laser resection group was 41%, 31%, and 33% respectively, and the difference had no statistical significance (p>0.05). Both the TURBT group and holmium laser resection group had shorter operation time, hospital stay and time for indwelling urinary catheter as well as much less blood loss when compared with the partial cystectomy group; the difference had statistical significance (pspasm. Therefore, this study presumes that holmium laser resection and TURBT are much safer and quicker for recovery and obviously superior to the partial cystectomy.

  19. Time-dependent effects of castration on the bladder function and histological changes in the bladder and blood vessels

    Institute of Scientific and Technical Information of China (English)

    Tomohiro Magari; Yasuhiro Shibata; Seiji Arai; Bunzo Kashiwagi; Keiji Suzuki; Kazuhiro Suzuki

    2014-01-01

    We examined the effect of androgens on bladder blood lfow (BBF), bladder function and histological changes in castrated male rats. Male Wistar rats were classiifed into unoperated group (control group), groups castrated at the age of 8 weeks (group 8wPC) and groups castrated at the age of 4 weeks (group 4wPC). Each rat was used at the age of 20 weeks. BBF was measured using lfuorescent microspheres. Bladder cystometry was performed without anesthesia or restraint;the bladder was ifrst irrigated with saline and then with 0.25%acetic acid (AA) solution. Maximum voiding pressure and voiding interval were measured. The bladder and iliac artery were histologically examined for differences in smooth muscle and quantity of collagen ifber to analyze the effect of castration on the smooth muscle content. No differences were noted in BBF following castration. The voiding intervals for all groups were shortened (P<0.001) following AA irrigation. No signiifcant difference was noted in the maximum voiding pressure. Histological changes were observed in bladder and iliac artery. Smooth muscle/collagen ratio at the bladder was lower in groups 8wPC and 4wPC compared to the control group (P<0.01), while that at the iliac artery was decreased in group 4wPC compared to the control group (P<0.001). In conclusion, our ifndings indicate that castration does not alter BBF, but leads to histological changes in the bladder as well as its associated blood vessels.

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

    Science.gov (United States)

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

    2012-10-15

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

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

    Energy Technology Data Exchange (ETDEWEB)

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

    2012-10-19

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

  2. [Venom of Latrodectus mactans from Chile (Araneae, Theridiidae): effect on smooth muscle].

    Science.gov (United States)

    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.

  3. Endothelial and smooth muscle cells from abdominal aortic aneurysm have increased oxidative stress and telomere attrition.

    Directory of Open Access Journals (Sweden)

    Giuseppe Cafueri

    Full Text Available BACKGROUND: Abdominal aortic aneurysm (AAA is a complex multi-factorial disease with life-threatening complications. AAA is typically asymptomatic and its rupture is associated with high mortality rate. Both environmental and genetic risk factors are involved in AAA pathogenesis. Aim of this study was to investigate telomere length (TL and oxidative DNA damage in paired blood lymphocytes, aortic endothelial cells (EC, vascular smooth muscle cells (VSMC, and epidermal cells from patients with AAA in comparison with matched controls. METHODS: TL was assessed using a modification of quantitative (Q-FISH in combination with immunofluorescence for CD31 or α-smooth muscle actin to detect EC and VSMC, respectively. Oxidative DNA damage was investigated by immunofluorescence staining for 7, 8-dihydro-8-oxo-2'-deoxyguanosine (8-oxo-dG. RESULTS AND CONCLUSIONS: Telomeres were found to be significantly shortened in EC, VSMC, keratinocytes and blood lymphocytes from AAA patients compared to matched controls. 8-oxo-dG immunoreactivity, indicative of oxidative DNA damage, was detected at higher levels in all of the above cell types from AAA patients compared to matched controls. Increased DNA double strand breaks were detected in AAA patients vs controls by nuclear staining for γ-H2AX histone. There was statistically significant inverse correlation between TL and accumulation of oxidative DNA damage in blood lymphocytes from AAA patients. This study shows for the first time that EC and VSMC from AAA have shortened telomeres and oxidative DNA damage. Similar findings were obtained with circulating lymphocytes and keratinocytes, indicating the systemic nature of the disease. Potential translational implications of these findings are discussed.

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

    Science.gov (United States)

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

    2013-01-01

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

  5. Evidence that CFTR is expressed in rat tracheal smooth muscle cells and contributes to bronchodilation

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    Mettey Yvette

    2006-08-01

    Full Text Available Abstract Background The airway functions are profoundly affected in many diseases including asthma, chronic obstructive pulmonary disease (COPD and cystic fibrosis (CF. CF the most common lethal autosomal recessive genetic disease is caused by mutations of the CFTR gene, which normally encodes a multifunctional and integral membrane protein, the CF transmembrane conductance regulator (CFTR expressed in airway epithelial cells. Methods To demonstrate that CFTR is also expressed in tracheal smooth muscle cells (TSMC, we used iodide efflux assay to analyse the chloride transports in organ culture of rat TSMC, immunofluorescence study to localize CFTR proteins and isometric contraction measurement on isolated tracheal rings to observe the implication of CFTR in the bronchodilation. Results We characterized three different pathways stimulated by the cAMP agonist forskolin and the isoflavone agent genistein, by the calcium ionophore A23187 and by hypo-osmotic challenge. The pharmacology of the cAMP-dependent iodide efflux was investigated in detail. We demonstrated in rat TSMC that it is remarkably similar to that of the epithelial CFTR, both for activation (using three benzo [c]quinolizinium derivatives and for inhibition (glibenclamide, DPC and CFTRinh-172. Using rat tracheal rings, we observed that the activation of CFTR by benzoquinolizinium derivatives in TSMC leads to CFTRinh-172-sensitive bronchodilation after constriction with carbachol. An immunolocalisation study confirmed expression of CFTR in tracheal myocytes. Conclusion Altogether, these observations revealed that CFTR in the airways of rat is expressed not only in the epithelial cells but also in tracheal smooth muscle cells leading to the hypothesis that this ionic channel could contribute to bronchodilation.

  6. Modulating activity of M1 receptor to the reaction of ileal smooth muscle

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    Izabela Glaza

    2011-08-01

    Full Text Available Background:The subject of the study was determination of the effect of drugs on ileal smooth muscle contraction induced by activation of M1 type muscarinic receptors. Drugs that have an effect on muscarinic receptors are divided to agonists, with close ties to the receptor and high internal activity and antagonists, with no internal activity. Conducted experiments tested interactions between a broad-spectrum agonist of muscarinic receptors, carbachol and a selective muscarinic receptor antagonist of M1 type, pirenzepine.Material/Methods:Testing was conducted on tissues isolated from rat’s intestine. Male Wistar rats with weight between 220 g and 360 g were anesthetized by intraperitoneal injection of urethane (120 mg/kg. Concentration-effect curves were determined with the use of cumulated concentration method, in accordance with the van Rossum method (1963 in Kenakin modification (2006.Results:The purpose of the study was determination of concentration-effect curves for carbachol. This curve was compared with the curve of receptor occupation depending on concentration of this drug. Based on concentration-effect curves, the average value of EC50 was calculated for carbachol, amounting to 2.44×10–6 [M/l].Conclusions:The results confirmed that atropine is effective in stopping contractions caused by carbachol, meeting the conditions of competitive antagonists. Atropine caused the shift of curves for carbachol to the right. Pirenzepine, selectively blocking muscarinic receptors of M1 type gave similar results. It was proved that in the preparation of gastric fundus smooth muscle, M1 type receptors occur not only presynaptically, but also postsynaptically.

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

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

    2016-04-01

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

  8. Airway hyperresponsiveness; smooth muscle as the principal actor [version 1; referees: 2 approved

    Directory of Open Access Journals (Sweden)

    Anne-Marie Lauzon

    2016-03-01

    Full Text Available Airway hyperresponsiveness (AHR is a defining characteristic of asthma that refers to the capacity of the airways to undergo exaggerated narrowing in response to stimuli that do not result in comparable degrees of airway narrowing in healthy subjects. Airway smooth muscle (ASM contraction mediates airway narrowing, but it remains uncertain as to whether the smooth muscle is intrinsically altered in asthmatic subjects or is responding abnormally as a result of the milieu in which it sits. ASM in the trachea or major bronchi does not differ in its contractile characteristics in asthmatics, but the more pertinent peripheral airways await complete exploration. The mass of ASM is increased in many but not all asthmatics and therefore cannot be a unifying hypothesis for AHR, although when increased in mass it may contribute to AHR. The inability of a deep breath to reverse or prevent bronchial narrowing in asthma may reflect an intrinsic difference in the mechanisms that lead to softening of contracted ASM when subjected to stretch. Cytokines such as interleukin-13 and tumor necrosis factor-α promote a more contractile ASM phenotype. The composition and increased stiffness of the matrix in which ASM is embedded promotes a more proliferative and pro-inflammatory ASM phenotype, but the expected dedifferentiation and loss of contractility have not been shown. Airway epithelium may drive ASM proliferation and/or molecular remodeling in ways that may lead to AHR. In conclusion, AHR is likely multifactorial in origin, reflecting the plasticity of ASM properties in the inflammatory environment of the asthmatic airway.

  9. Role of dystrophin in airway smooth muscle phenotype, contraction and lung function.

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    Pawan Sharma

    Full Text Available Dystrophin links the transmembrane dystrophin-glycoprotein complex to the actin cytoskeleton. We have shown that dystrophin-glycoprotein complex subunits are markers for airway smooth muscle phenotype maturation and together with caveolin-1, play an important role in calcium homeostasis. We tested if dystrophin affects phenotype maturation, tracheal contraction and lung physiology. We used dystrophin deficient Golden Retriever dogs (GRMD and mdx mice vs healthy control animals in our approach. We found significant reduction of contractile protein markers: smooth muscle myosin heavy chain (smMHC and calponin and reduced Ca2+ response to contractile agonist in dystrophin deficient cells. Immunocytochemistry revealed reduced stress fibers and number of smMHC positive cells in dystrophin-deficient cells, when compared to control. Immunoblot analysis of Akt1, GSK3β and mTOR phosphorylation further revealed that downstream PI3K signaling, which is essential for phenotype maturation, was suppressed in dystrophin deficient cell cultures. Tracheal rings from mdx mice showed significant reduction in the isometric contraction to methacholine (MCh when compared to genetic control BL10ScSnJ mice (wild-type. In vivo lung function studies using a small animal ventilator revealed a significant reduction in peak airway resistance induced by maximum concentrations of inhaled MCh in mdx mice, while there was no change in other lung function parameters. These data show that the lack of dystrophin is associated with a concomitant suppression of ASM cell phenotype maturation in vitro, ASM contraction ex vivo and lung function in vivo, indicating that a linkage between the DGC and the actin cytoskeleton via dystrophin is a determinant of the phenotype and functional properties of ASM.

  10. Effect of dexamethasone on voltage-gated Na+ channel in cultured human bronchial smooth muscle cells.

    Science.gov (United States)

    Nakajima, Toshiaki; Jo, Taisuke; Meguro, Kentaro; Oonuma, Hitoshi; Ma, Ji; Kubota, Nami; Imuta, Hiroyuki; Takano, Haruhito; Iida, Haruko; Nagase, Takahide; Nagata, Taiji

    2008-06-06

    Voltage-gated Na(+) channel (I(Na)) encoded by SCN9A mRNA is expressed in cultured human bronchial smooth muscle cells. We investigated the effects of dexamethasone on I(Na), by using whole-cell voltage clamp techniques, reverse transcriptase/polymerase chain reaction (RT-PCR), and quantitative real-time RT-PCR. Acute application of dexamethasone (10(-6) M) did not affect I(Na). However, the percentage of the cells with I(Na) was significantly less in cells pretreated with dexamethasone for 48 h, and the current-density of I(Na) adjusted by cell capacitance in cells with I(Na) was also decreased in cells treated with dexamethasone. RT-PCR analysis showed that alpha and beta subunits mRNA of I(Na) mainly consisted of SCN9A and SCN1beta, respectively. Treatment with dexamethasone for 24-48 h inhibited the expression of SCN9A mRNA. The inhibitory effect of dexamethasone was concentration-dependent, and was observed at a concentration higher than 0.1 nM. The effect of dexamethasone on SCN9A mRNA was not blocked by spironolactone, but inhibited by mifepristone. The inhibitory effects of dexamethasone on SCN9A mRNA could not be explained by the changes of the stabilization of mRNA measured by using actinomycin D. These results suggest that dexamethasone inhibited I(Na) encoded by SCN9A mRNA in cultured human bronchial smooth muscle cells by inhibiting the transcription via the glucocorticoid receptor.

  11. Opioid receptor antagonists increase [Ca2+]i in rat arterial smooth muscle cells in hemorrhagic shock

    Institute of Scientific and Technical Information of China (English)

    Li KAI; Zhong-feng WANG; Yu-liang SHI; Liang-ming LIU; De-yao HU

    2004-01-01

    AIM: To examine the effects of opioid receptor antagonists and norepinephrine on intracellular free Ca2+ concentration ([Ca2+]i) in mesenteric arterial (MA) smooth muscle cells (SMC) isolated from normal and hemorrhagic shocked rats in the vascular hyporesponse stage. METHODS: The rat model of hemorrhagic shock was made by withdrawing blood to decrease the artery mean blood pressure to 3.73-4.26 kPa and keeping at the level for 3 h.[Ca2+]i of vascular smooth muscle cells (VSMC) were detected by the laser scan confocal microscopy. RESULTS:In the hyporesponse VMSC of rats in hemorrhagic shock, selective δ-, κ-, and μ-opioid receptor antagonists (naltrindole, nor-binaltorphimine, and β-funaltrexamine, 100 nmol/L) as well as norepinephrine 5 μmol/L significantly increased [Ca2+]i by 47 %±13 %, 37 %±14 %, 33 %±10 %, and 54 %±17 %, respectively, although their effects were lower than those in the normal rat cells (the increased values were 148 %±54 %, 130 %±44 %, 63 %±17 %and 110 %±38 %, respectively); and the norepinephrine-induced increase in [Ca2+]i was further augmented by three δ-, κ-, and μ-opioid receptor antagonists (50 nmol/L, respectively) application (from 52 %± 16 % to 99 %±29 %,146 %±54 % and 137 %±47 %, respectively). CONCLUSION: The disorder of [Ca2+]i regulation induced by hemorrhagic shock was mediated by opioid receptor and α-adrenoceptor, which may be partly responsible for the vascular hyporesponse, and the opioid receptor antagonists improved the response of resistance arteries to vascular stimulants in decompensatory stage of hemorrhagic shock.

  12. Oxidized low density lipoprotein (LDL) affects hyaluronan synthesis in human aortic smooth muscle cells.

    Science.gov (United States)

    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.

  13. Colon distention induces persistent visceral hypersensitivity by mechanotranscription of pain mediators in colonic smooth muscle cells.

    Science.gov (United States)

    Lin, You-Min; Fu, Yu; Wu, Chester C; Xu, Guang-Yin; Huang, Li-Yen; Shi, Xuan-Zheng

    2015-03-01

    Abdominal pain and distention are major complaints in irritable bowel syndrome. Abdominal distention is mainly attributed to intraluminal retention of gas or solid contents, which may cause mechanical stress to the gut wall. Visceral hypersensitivity (VHS) may account for abdominal pain. We sought to determine whether tonic colon distention causes persistent VHS and if so whether mechanical stress-induced expression (mechanotranscription) of pain mediators in colonic smooth muscle cells (SMCs) plays a role in VHS. Human colonic SMCs were isolated and stretched in vitro to investigate whether mechanical stress upregulates expression of the pain mediator cyclooxygenase-2 (COX-2). Rat colon was distended with a 5-cm-long balloon, and gene expression of COX-2, visceromotor response (VMR), and sensory neuron excitability were determined. Static stretch of colonic SMCs induced marked expression of COX-2 mRNA and protein in a force- and time-dependent manner. Subnoxious tonic distention of the distal colon at ∼30-40 mmHg for 20 or 40 min induced COX-2 expression and PGE2 production in colonic smooth muscle, but not in the mucosa layer. Lumen distention also increased VMR in a force- and time-dependent manner. The increase of VMR persisted for at least 3 days. Patch-clamp experiments showed that the excitability of colon projecting sensory neurons in the dorsal root ganglia was markedly augmented, 24 h after lumen distention. Administration of COX-2 inhibitor NS-398 partially but significantly attenuated distention-induced VHS. In conclusion, tonic lumen distention upregulates expression of COX-2 in colonic SMC, and COX-2 contributes to persistent VHS.

  14. Exploring Arterial Smooth Muscle Kv7 Potassium Channel Function using Patch Clamp Electrophysiology and Pressure Myography

    Science.gov (United States)

    Brueggemann, Lioubov I.; Mani, Bharath K.; Haick, Jennifer; Byron, Kenneth L.

    2012-01-01

    Contraction or relaxation of smooth muscle cells within the walls of resistance arteries determines the artery diameter and thereby controls flow of blood through the vessel and contributes to systemic blood pressure. The contraction process is regulated primarily by cytosolic calcium concentration ([Ca2+]cyt), which is in turn controlled by a variety of ion transporters and channels. Ion channels are common intermediates in signal transduction pathways activated by vasoactive hormones to effect vasoconstriction or vasodilation. And ion channels are often targeted by therapeutic agents either intentionally (e.g. calcium channel blockers used to induce vasodilation and lower blood pressure) or unintentionally (e.g. to induce unwanted cardiovascular side effects). Kv7 (KCNQ) voltage-activated potassium channels have recently been implicated as important physiological and therapeutic targets for regulation of smooth muscle contraction. To elucidate the specific roles of Kv7 channels in both physiological signal transduction and in the actions of therapeutic agents, we need to study how their activity is modulated at the cellular level as well as evaluate their contribution in the context of the intact artery. The rat mesenteric arteries provide a useful model system. The arteries can be easily dissected, cleaned of connective tissue, and used to prepare isolated arterial myocytes for patch clamp electrophysiology, or cannulated and pressurized for measurements of vasoconstrictor/vasodilator responses under relatively physiological conditions. Here we describe the methods used for both types of measurements and provide some examples of how the experimental design can be integrated to provide a clearer understanding of the roles of these ion channels in the regulation of vascular tone. PMID:23007713

  15. TIMP-1 Induces α-Smooth Muscle Actin in Fibroblasts to Promote Urethral Scar Formation

    Directory of Open Access Journals (Sweden)

    Yinglong Sa

    2015-04-01

    Full Text Available Background/Aims: Tissue inhibitor of metalloproteinases-1 (TIMP-1 has been reported to upregulate in urethral scar. However, the underlying molecular mechanisms remain undefined. Methods: Here, we studied levels of TIMP-1 and α-smooth muscle actin (α-SMA in the fibroblasts isolated from urethral scar tissues, compared to the fibroblasts isolated from normal urethra. Then we either overexpressed TIMP-1, or inhibited TIMP-1 by lentiviruses carrying a transgene or a short hairpin small interfering RNA for TIMP-1 in human fibroblasts. We examined the effects of modulation of TIMP-1 on α-SMA, and on epithelial-mesenchymal transition (EMT-related genes. We also studied the underlying mechanisms. Results: We detected significantly higher levels of TIMP-1 and α-smooth muscle actin (α-SMA in the fibroblasts isolated from urethral scar tissues, compared to the fibroblasts isolated from normal urethra. Moreover, the levels of TIMP-1 and α-SMA strongly correlated. Moreover, we found that TIMP-1 significantly increased levels of α-SMA, transforming growth factor β 1 (TGFβ1, Collagen I and some other key factors related to an enhanced EMT, suggesting that TIMP-1 may induce transformation of fibroblasts into myofibroblasts to promote tissue EMT to enhance the formation of urethral scar. Moreover, increases in TIMP-1 also induced an increase in fibroblast cell growth and cell invasion, in an ERK/MAPK-signaling-dependent manner. Conclusion: Our study thus highlights a pivotal role of TIMP-1 in urethral scar formation.

  16. Stromal hedgehog signaling maintains smooth muscle and hampers micro-invasive prostate cancer

    Science.gov (United States)

    Yang, Zhaohui; Peng, Yu-Ching; Gopalan, Anuradha; Gao, Dong; Chen, Yu

    2017-01-01

    ABSTRACT It is widely appreciated that reactive stroma or carcinoma-associated fibroblasts can influence epithelial tumor progression. In prostate cancer (PCa), the second most common male malignancy worldwide, the amount of reactive stroma is variable and has predictive value for tumor recurrence. By analyzing human PCa protein and RNA expression databases, we found smooth muscle cells (SMCs) are decreased in advanced tumors, whereas fibroblasts are maintained. In three mouse models of PCa, PB-MYC, ERG/PTEN and TRAMP, we found the composition of the stroma is distinct. SMCs are greatly depleted in advanced PB-MYC tumors and locally reduced in ERG/PTEN prostates, whereas in TRAMP tumors the SMC layers are increased. In addition, interductal fibroblast-like cells expand in PB-MYC and ERG/PTEN tumors, whereas in TRAMP PCa they expand little and stromal cells invade into intraductal adenomas. Fate mapping of SMCs showed that in PB-MYC tumors the cells are depleted, whereas they expand in TRAMP tumors and interestingly contribute to the stromal cells in intraductal adenomas. Hedgehog (HH) ligands secreted by epithelial cells are known to regulate prostate mesenchyme expansion differentially during development and regeneration. Any possible role of HH signaling in stromal cells during PCa progression is poorly understood. We found that HH signaling is high in SMCs and fibroblasts near tumor cells in all models, and epithelial Shh expression is decreased whereas Ihh and Dhh are increased. In human primary PCa, expression of IHH is the highest of the three HH genes, and elevated HH signaling correlates with high stromal gene expression. Moreover, increasing HH signaling in the stroma of PB-MYC PCa resulted in more intact SMC layers and decreased tumor progression (micro-invasive carcinoma). Thus, we propose HH signaling restrains tumor progression by maintaining the smooth muscle and preventing invasion by tumor cells. Our studies highlight the importance of understanding

  17. Effects of hispidulin, a flavone isolated from Inula viscosa, on isolated guinea-pig smooth muscle.

    Science.gov (United States)

    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.

  18. Regulation of CCL5 expression in smooth muscle cells following arterial injury.

    Directory of Open Access Journals (Sweden)

    Huan Liu

    Full Text Available Chemokines play a crucial role in inflammation and in the pathophysiology of atherosclerosis by recruiting inflammatory immune cells to the endothelium. Chemokine CCL5 has been shown to be involved in atherosclerosis progression. However, little is known about how CCL5 is regulated in vascular smooth muscle cells. In this study we report that CCL5 mRNA expression was induced and peaked in aorta at day 7 and then declined after balloon artery injury, whereas IP-10 and MCP-1 mRNA expression were induced and peaked at day 3 and then rapidly declined.The expression of CCL5 receptors (CCR1, 3 & 5 were also rapidly induced and then declined except CCR5 which expression was still relatively high at day 14 after balloon injury. In rat smooth muscle cells (SMCs, similar as in aorta CCL5 mRNA expression was induced and kept increasing after LPS plus IFN-gamma stimulation, whereas IP-10 mRNA expression was rapidly induced and then declined. Our data further indicate that induction of CCL5 expression in SMCs was mediated by IRF-1 via binding to the IRF-1 response element in CCL5 promoter. Moreover, p38 MAPK was involved in suppression of CCL5 and IP-10 expression in SMCs through common upstream molecule MKK3. The downstream molecule MK2 was required for p38-mediated CCL5 but not IP-10 inhibition. Our findings indicate that CCL5 induction in aorta and SMCs is mediated by IRF-1 while activation of p38 MAPK signaling inhibits CCL5 and IP-10 expression. Methods targeting MK2 expression could be used to selectively regulate CCL5 but not IP-10 expression in SMCs.

  19. Montelukast prevents microparticle-induced inflammatory and functional alterations in human bronchial smooth muscle cells.

    Science.gov (United States)

    Fogli, Stefano; Stefanelli, Fabio; Neri, Tommaso; Bardelli, Claudio; Amoruso, Angela; Brunelleschi, Sandra; Celi, Alessandro; Breschi, Maria Cristina

    2013-10-01

    Microparticles (MPs) are membrane fragments that may play a role in the pathogenesis of chronic respiratory diseases. We aimed to investigate whether human monocytes/macrophage-derived MPs could induce a pro-inflammatory phenotype in human bronchial smooth muscle cells (BSMC) and the effect of montelukast in this setting. Experimental methods included isolation of human monocytes/macrophages and generation of monocyte-derived MPs, RT-PCR analysis of gene expression, immunoenzymatic determination of pro-inflammatory factor release, bioluminescent assay of intracellular cAMP levels and electromobility shift assay analysis of NF-κB nuclear translocation. Stimulation of human BSMC with monocyte-derived MPs induced a pro-inflammatory switch in human BSMC by inducing gene expression (COX-2 and IL-8), protein release in the supernatant (PGE2 and IL-8), and heterologous β2-adrenoceptor desensitization. The latter effect was most likely related to autocrine PGE2 since pre-treatment with COX inhibitors restored the ability of salbutamol to induce cAMP synthesis in desensitized cells. Challenge with MPs induced nuclear translocation of NF-κB and selective NF-κB inhibition decreased MP-induced cytokine release in the supernatant. Montelukast treatment prevented IL-8 release and heterologous β2-adrenoceptor desensitization in human BSMC exposed to monocyte-derived MPs by blocking NF-κB nuclear translocation. These findings provide evidence on the role of human monocyte-derived MPs in the airway smooth muscle phenotype switch as a novel potential mechanism in the progression of chronic respiratory diseases and on the protective effects by montelukast in this setting.

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

    Energy Technology Data Exchange (ETDEWEB)

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

    2013-02-01

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

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

    Energy Technology Data Exchange (ETDEWEB)

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

    2014-11-07

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

  2. Analysis of the Sonic Hedgehog signaling pathway in normal and abnormal bladder development.

    Science.gov (United States)

    DeSouza, Kristin R; Saha, Monalee; Carpenter, Ashley R; Scott, Melissa; McHugh, Kirk M

    2013-01-01

    In this study, we examined the expression of Sonic Hedgehog, Patched, Gli1, Gli2, Gli3 and Myocardin in the developing bladders of male and female normal and megabladder (mgb-/-) mutant mice at embryonic days 12 through 16 by in situ hybridization. This analysis indicated that each member of the Sonic Hedgehog signaling pathway as well as Myocardin displayed distinct temporal and spatial patterns of expression during normal bladder development. In contrast, mgb-/- bladders showed both temporal and spatial changes in the expression of Patched, Gli1 and Gli3 as well as a complete lack of Myocardin expression. These changes occurred primarily in the outer mesenchyme of developing mgb-/- bladders consistent with the development of an amuscular bladder phenotype in these animals. These results provide the first comprehensive analysis of the Sonic Hedgehog signaling pathway during normal bladder development and provide strong evidence that this key signaling cascade is critical in establishing radial patterning in the developing bladder. In addition, the lack of detrusor smooth muscle development observed in mgb-/- mice is associated with bladder-specific temporospatial changes in Sonic Hedgehog signaling coupled with a lack of Myocardin expression that appears to result in altered patterning of the outer mesenchyme and poor initiation and differentiation of smooth muscle