Stretch activates heparin-binding EGF-like growth factor expression in bladder smooth muscle cells

1998 ◽  
Vol 275 (5) ◽  
pp. C1247-C1254 ◽  
Author(s):  
John M. Park ◽  
Joseph G. Borer ◽  
Michael R. Freeman ◽  
Craig A. Peters
Keyword(s):  
Smooth Muscle ◽  
Growth Factor ◽  
Smooth Muscle Cells ◽  
Fold Increase ◽  
Muscle Cells ◽  
Mechanical Stretch ◽  
Mrna Levels ◽  
Ang Ii ◽  
Heparin Binding ◽  
Bladder Smooth Muscle

Cultured rat bladder smooth muscle cells (SMC) were grown on collagen-coated silicone membranes and subjected to continuous cycles of stretch-relaxation. Semiquantitative RT-PCR analysis revealed a time-dependent increase in heparin-binding epidermal growth factor (EGF)-like growth factor (HB-EGF) mRNA levels after stretch, with maximal levels appearing after 4 h. Immunostaining for proHB-EGF revealed higher levels of HB-EGF protein in the stretched than in the nonstretched SMC. The ANG II receptor type 1 antagonist losartan markedly suppressed stretch-activated HB-EGF expression. ANG II levels were 3.3-fold higher in the stretch- than in the non-stretch-conditioned media. Stretch stimulation of bladder SMC that had been transiently transfected with an HB-EGF promoter-luciferase expression construct resulted in an 11-fold increase in reporter activity. Mechanical stretch induced a 4.7-fold increase in tritiated thymidine incorporation rate, and this was reduced by 25% in the presence of losartan. We conclude that mechanical stretch activates HB-EGF gene expression in bladder SMC and that this is mediated in part by autocrine ANG II secretion.

Mechanical stretch regulates cell survival in human bladder smooth muscle cells in vitro

2002 ◽  
Vol 283 (6) ◽  
pp. F1192-F1199 ◽  
Author(s):  
David J. Galvin ◽  
R. William G. Watson ◽  
James I. Gillespie ◽  
Hugh Brady ◽  
John M. Fitzpatrick
Keyword(s):  
Smooth Muscle ◽  
Growth Factor ◽  
Smooth Muscle Cells ◽  
Muscle Cells ◽  
Mechanical Stretch ◽  
Bladder Smooth Muscle ◽  
Human Bladder ◽  
Cell Hypertrophy ◽  
Growth Factor Production

Our understanding of the pathophysiology of the overactive bladder is poor. It has been proposed that localized contractions result in the abnormal stretching of bladder smooth muscle. We hypothesize that stretch regulates the cellular processes that determine tissue size. The purpose of this study was to investigate the effect of stretch on apoptosis, proliferation, cell hypertrophy, and growth factor production in human bladder smooth muscle cells in vitro. Normal human detrusor muscle was obtained from patients undergoing radical cystectomy for invasive bladder cancer, and primary cultures were established. Cells were mechanically stretched on flexible plates at a range of pressures and times. Apoptosis was assessed by propidium iodide incorporation and flow cytometry. Radiolabeled thymidine and amino acid incorporation were used to assess proliferation and cell hypertrophy. ELISA and RT-PCR were used to assess growth factor production. Mechanical stretch inhibits apoptosis in a time- and dose-dependent manner and was associated with increases in the antiapoptotic proteins heat shock protein-70 and cIAP-1. Stretch also increases smooth muscle cell proliferation and hypertrophy, but hypertrophy is the more dominant response. These changes were associated with increases in IGF-1 and basic FGF and a decrease in transforming growth factor-β1. Mechanical stretch regulates apoptosis, proliferation, and cell hypertrophy in human bladder smooth muscle cells.

Obstruction stimulates COX-2 expression in bladder smooth muscle cells via increased mechanical stretch

1999 ◽  
Vol 276 (1) ◽  
pp. F129-F136 ◽  
Author(s):  
John M. Park ◽  
Tianxin Yang ◽  
Lois J. Arend ◽  
Jürgen B. Schnermann ◽  
Craig A. Peters ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Outlet Obstruction ◽  
Muscle Cells ◽  
Mechanical Stretch ◽  
Type I ◽  
Mrna Levels ◽  
Dependent Manner ◽  
Bladder Smooth Muscle ◽  
Cox 2

Studies were performed to investigate the regulatory mechanism of bladder cyclooxygenase-2 (COX-2) expression after outlet obstruction. In situ hybridization of murine bladder tissues using COX-2-specific riboprobes demonstrated that COX-2 expression was induced predominantly in the bladder smooth muscle cells after outlet obstruction. To study the effect of increased mechanical stretch on COX isoform expression, cultured rat bladder smooth muscle cells were grown on silicone elastomer-bottomed plates coated with collagen type I and were subjected to continuous cycles of stretch/relaxation for variable duration. COX-1 mRNA levels did not change with stretch. COX-2 expression increased in a time-dependent manner after stretch, with maximal mRNA and protein levels occurring after 4 h. PGE2 levels increased more than 40-fold in the culture media after stretch, consistent with increased COX activity, and this was reduced to near completion in the presence of a COX-2 inhibitor, NS-398. Exposure to stretch over a 48-h period induced a 4.7 ± 0.6-fold increase in tritiated thymidine incorporation rate. This increase in DNA synthesis was markedly suppressed when the cells were stretched in the presence of NS-398. We conclude that in bladder obstruction COX-2 activation occurs predominantly in the smooth muscle cells in response to mechanical stretch. Our findings also suggest that stretch-activated COX-2 expression may participate in bladder smooth muscle cell proliferation and thereby play a role in pathological bladder wall thickening after obstruction.

Essential role for extracellular Ca2+ in JNK activation by mechanical stretch in bladder smooth muscle cells

2001 ◽  
Vol 281 (4) ◽  
pp. C1165-C1172 ◽  
Author(s):  
Nobuhiro Kushida ◽  
Yukihito Kabuyama ◽  
Osamu Yamaguchi ◽  
Yoshimi Homma
Keyword(s):  
Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Fold Increase ◽  
Muscle Cells ◽  
Mechanical Stretch ◽  
Mitogen Activated Protein Kinase ◽  
Bladder Smooth Muscle ◽  
Voltage Dependent ◽  
Mitogen Activated Protein ◽  
Jnk Activation

Mechanical stretch has been implicated in phenotypic changes as an adaptive response to stretch stress physically loaded in bladder smooth muscle cells (BSMCs). To investigate stretch-induced signaling, we examined the mitogen-activated protein kinase (MAPK) family using rat primary BSMCs. When BSMCs were subjected to sustained mechanical stretch using collagen-coated silicon membranes, activation of c-Jun NH2-terminal kinase (JNK) was most relevant among three subsets of MAPK family members: the activity was elevated from 5 min after stretch and peaked at 10 min with an 11-fold increase. Activation of p38 was weak compared with that of JNK, and ERK was not activated at all. JNK activation by mechanical stretch was totally dependent on extracellular Ca2+ and inhibited by Gd3+, a blocker of stretch-activated (SA) ion channels. Nifedipine and verapamil, inhibitors for voltage-dependent Ca2+ channels, had no effect on this JNK activation. Moreover, none of the inhibitors pertussis toxin, genistein, wortmannin, or calphostin C affected stretch-induced JNK activation, indicating that G protein-coupled and tyrosine kinase receptors are unlikely to be involved in this JNK activation. On the other hand, W-7, a calmodulin inhibitor, and cyclosporin A, a calcineurin inhibitor, prevented JNK activation by stretch. These results suggest a novel pathway for stretch-induced activation of JNK in BSMCs: mechanical stretch evokes Ca2+ influx via Gd3+-sensitive SA Ca2+ channels, resulting in JNK activation under regulation in part by calmodulin and calcineurin.

Abstract 361: Cyclophilin A Mediates Angiotensin II--Stimulated NADPH Oxidase Activation in Vascular Smooth Muscle Cells

2012 ◽  
Vol 32 (suppl_1) ◽  
Author(s):  
Nwe Nwe Soe ◽  
Mark Sowden ◽  
Patrizia Nigro ◽  
Bradford C Berk
Keyword(s):  
Smooth Muscle ◽  
Plasma Membrane ◽  
Nadph Oxidase ◽  
Smooth Muscle Cells ◽  
Fold Increase ◽  
Muscle Cells ◽  
Dependent Manner ◽  
Ang Ii ◽  
Membrane Translocation ◽  
Ppiase Activity

Objective: Cyclophilin A (CyPA) is a ubiquitously expressed cytosolic protein that possesses PPIase activity and scaffold function. CyPA regulates Angiotensin II (Ang II) induced reactive oxygen species (ROS) production in vascular smooth muscle cells. However, the mechanism of this CyPA regulation remains unclear. We hypothesized that CyPA regulates plasma membrane translocation of NADPH oxidase cytosolic subunit, p47phox, which is required for NADPH oxidase structural organization and activity. Methods and results: Immunofluorescence studies in rat aortic smooth muscle cells revealed that CyPA translocated from the cytosol to the plasma membrane in response to Ang II in a time dependent manner with a peak at 10min (46.4±5.4 fold increase). Mouse Aortic Smooth Muscle Cells (MASM) were isolated from mice lacking CyPA (CyPA-/-) and wild type controls (WT), treated with Ang II (100nM) and immunofluorescence analysis was performed. Ang II induced p47phox plasma membrane translocation at 10min in WT mice. However, p47 phox translocation was significantly inhibited in CyPA -/- MASM. CyPA and p47phox colocalized at the plasma membrane in response to Ang II. Further analysis using subcellular fractionation studies confirmed that Ang II induced p47phox plasma membrane translocation was inhibited in CyPA -/- MASM compared to WT (1.2±2.7 vs 4.3±3.4 fold increase). Coimmunoprecipitation analyses confirmed that Ang II increased CyPA association with p47phox in a time dependent manner (2.5±3.4 fold increase at 10min). Finally, pretreatment with the PPIase activity inhibitor, cyclosporine A (1uM), could not inhibit CyPA association with p47phox and CyPA mediated p47phox translocation to the plasma membrane. Conclusion: These data suggest that Ang II promotes an association between CyPA and p47phox that enhances plasma membrane translocation of p47phox. This is proposed to increase the NADPH oxidase activity thereby increasing cellular ROS production. This process is independent of the PPIase activity of CyPA. Therefore, inhibition of the CyPA and p47phox association could be a future therapeutic target for Ang II induced ROS regulated cardiovascular diseases such as atherosclerosis and abdominal aortic aneurysm formation.

PDGF-BB-induced DNA synthesis is delayed by angiotensin II in vascular smooth muscle cells

1998 ◽  
Vol 274 (5) ◽  
pp. H1742-H1748 ◽  
Author(s):  
Gunilla Dahlfors ◽  
Yun Chen ◽  
Maria Wasteson ◽  
Hans J. Arnqvist
Keyword(s):  
Smooth Muscle ◽  
Growth Factor ◽  
Smooth Muscle Cells ◽  
Dna Synthesis ◽  
Receptor Antagonist ◽  
Muscle Cells ◽  
Inhibitory Effect ◽  
Ang Ii ◽  
Β Receptor ◽  
Tgf Β1

The interaction of ANG II with platelet-derived growth factor (PDGF)-BB-induced DNA synthesis was studied in cultured rat aortic smooth muscle cells. PDGF-BB-induced DNA synthesis was delayed (∼6–8 h) by ANG II as shown by a time-course experiment. Losartan, an AT1-receptor antagonist, blocked the transient inhibitory effect of ANG II, whereas the AT2-receptor antagonist PD-123319 had no effect. Autocrine- or paracrine-acting transforming growth factor-β1 (TGF-β1), believed to be a mediator of ANG II-induced inhibitory effects, was not responsible for the delay of PDGF-BB-induced DNA synthesis, because a potent TGF-β1 neutralizing antibody could not reverse this effect of ANG II, nor was the delay of the PDGF-BB effect caused by inhibition of PDGF-β-receptor phosphorylation as shown by Western blot analysis of immunoprecipitated PDGF-β receptor. In conclusion, our results show that ANG II can exert a transient inhibitory effect on PDGF-BB-induced proliferation via the AT1 receptor.

Cyclic stretch activates p38 SAPK2-, ErbB2-, and AT1-dependent signaling in bladder smooth muscle cells

2000 ◽  
Vol 279 (4) ◽  
pp. C1155-C1167 ◽  
Author(s):  
Hiep T. Nguyen ◽  
Rosalyn M. Adam ◽  
Samuel H. Bride ◽  
John M. Park ◽  
Craig A. Peters ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Growth Factor ◽  
Protein Kinase ◽  
Smooth Muscle Cells ◽  
Dna Synthesis ◽  
Mapk Pathway ◽  
Muscle Cells ◽  
Cyclic Stretch ◽  
Bladder Smooth Muscle ◽  
Erk Mapk

Cyclic mechanical stretch of bladder smooth muscle cells (SMC) increases rates of DNA synthesis and stimulates transcription of the gene for heparin-binding epidermal growth factor-like growth factor (HB-EGF), an ErbB1/EGF receptor ligand that has been linked to hypertrophic bladder growth. In this study we sought to clarify the signaling pathways responsible for mechanotransduction of the stretch stimulus. HB-EGF mRNA levels, DNA synthesis, and AP-1/Ets DNA binding activities were induced by repetitive stretch of primary culture rat bladder SMC. Inhibitors of the p38 SAPK2 pathway, the angiotensin receptor type 1 (AT1), and the ErbB2 tyrosine kinase reduced each of these activities, while an inhibitor of the extracellular signal-regulated kinase mitogen-activated protein kinase (Erk-MAPK) pathway had no effect. Stretch rapidly activated stress-activated protein kinase 2 (p38 SAPK2) and Jun NH2-terminal kinase (JNK)/SAPK pathways but not the Erk-MAPK pathway and induced ErbB2 but not ErbB1 phosphorylation. Angiotensin II (ANG II) a bladder SMC mitogen previously linked to the stretch response, did not activate ErbB2, and ErbB2 activation occurred in response to stretch in the presence of an ANG receptor inhibitor, indicating that activation of the AT1-mediated pathway and the ErbB2-dependent pathway occurs by independent mechanisms. p38 SAPK2 and JNK/SAPK signaling also appeared to be independent of the ErbB2 and AT1 pathways. These findings indicate that stretch-stimulated DNA synthesis and gene expression in normal bladder SMC occur via multiple independent receptor systems (e.g., AT1 and ErbB2) and at least one MAPK pathway (p38 SAPK2). Further, we show that the Erk-MAPK pathway, which in most systems is linked to receptor-dependent cell growth responses, is not involved in progression to DNA synthesis or in the response of the HB-EGF gene to mechanical forces.

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