1949 TEMPORAL GENE EXPRESSION PROFILING OF PROTEINS INVOLVED IN CA2+-SENSITIZATION FOLLOWING PARTIAL BLADDER OUTLET OBSTRUCTION: MECHANISM FOR UPREGULATION OF RHOA AND ROKβ GENE EXPRESSION

Abstract

You have accessJournal of UrologyUrodynamics/Incontinence/Female Urology: Non-Neurogenic Voiding Dysfunction1 Apr 20111949 TEMPORAL GENE EXPRESSION PROFILING OF PROTEINS INVOLVED IN CA2+-SENSITIZATION FOLLOWING PARTIAL BLADDER OUTLET OBSTRUCTION: MECHANISM FOR UPREGULATION OF RHOA AND ROKβ GENE EXPRESSION Ettickan Boopathi, Stephen Zderic, Alan Wein, and Samuel Chacko Ettickan BoopathiEttickan Boopathi Glenolden, PA More articles by this author , Stephen ZdericStephen Zderic Philadelphia, PA More articles by this author , Alan WeinAlan Wein Philadelphia, PA More articles by this author , and Samuel ChackoSamuel Chacko Glenolden, PA More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2011.02.2153AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES Partial Bladder Outlet Obstruction (PBOO)-induced remodeling of the detrusor smooth muscle (DSM) is associated with modulation of cell signaling that regulates DSM contraction. The RhoA, Rho-activated kinase (ROK) and CPI-17 have been linked to Ca2+-sensitization, which maintains smooth muscle tone by keeping the myosin regulatory light chain (LC20) in the phosphorylated state by lowering myosin phosphatase activity. In this study, we show the temporal gene expression of proteins involved in the Ca2+-sensitization pathway at both mRNA and protein levels. Furthermore, using cultured murine and human bladder primary smooth muscle cells, we show that the bladder wall stretch, but not hypoxia, is a major factor responsible for the modulation of proteins involved in the Ca2+-sensitization of DSM contraction in PBOO-induced DSM remodeling. METHODS Male C57Bl/6 mice were surgically obstructed and kept for 1, 3, 7 and 14 days. Sham-operated mice served as a control. The bladders were excised, urothelium scarped off with a scalpel, and the serosa was removed. DSM obtained from PBOO and sham control animals were analyzed by PCR and western blotting. For stretch and hypoxia experiments, primary smooth muscle cells from murine and human bladders were seeded on type I collagen-coated silicone sheeting membrane at a density of 1 × 105/well in M199/10%FBS and grown to 80% confluence. Cells were rendered quiescent by incubation in medium containing 1% FBS for 48 hours and then subjected to cyclic stretch or 1 % hypoxia independently for 24 and 48 hours. RESULTS Expression of RhoA, ROKβ and CPI-17 increased in 3, 7 and 14 day- obstructed groups compared to sham-operated control murine bladders both at mRNA and protein levels. There was a significant increase of RhoA and ROKβ expression in response to stretch in cultured murine and human primary DSM cells compared to un-stretched cells. Contrary to this, the expression of RhoA and ROKβ in these cells decreased following 24 and 48 hrs after hypoxia. CONCLUSIONS These results confirm that RhoA, ROKβ and CPI-17 gene expression are upregulated in short- and long-term obstructed murine bladder smooth muscle. Our data is consistent with mechanical stretch of the bladder wall smooth muscle being the major factor in turning on the expression of proteins involved in the Ca2+-sensitization pathway, rather than hypoxia in PBOO. © 2011 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 185Issue 4SApril 2011Page: e779-e780 Advertisement Copyright & Permissions© 2011 by American Urological Association Education and Research, Inc.MetricsAuthor Information Ettickan Boopathi Glenolden, PA More articles by this author Stephen Zderic Philadelphia, PA More articles by this author Alan Wein Philadelphia, PA More articles by this author Samuel Chacko Glenolden, PA More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...