You have accessJournal of UrologyUrodynamics/Lower Urinary Tract Dysfunction/Female Pelvic Medicine: Basic Research & Pathophysiology II1 Apr 2018MP38-06 RAPAMYCIN REGULATION OF CORE CLOCK GENES IS ASSOCIATED WITH ALTERED DE-OBSTRUCTION SIGNATURES COORDINATE WITH PHYSIOLOGY Annette SCHRODER, Karen Aitken, Jia-Xin Jiang, Aliza Siebenaller, Thenuka THANABALASINGHAM, Martin Sidler, and Darius Bagli Annette SCHRODERAnnette SCHRODER More articles by this author , Karen AitkenKaren Aitken More articles by this author , Jia-Xin JiangJia-Xin Jiang More articles by this author , Aliza SiebenallerAliza Siebenaller More articles by this author , Thenuka THANABALASINGHAMThenuka THANABALASINGHAM More articles by this author , Martin SidlerMartin Sidler More articles by this author , and Darius BagliDarius Bagli More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2018.02.1233AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES Although rapamycin, during stretch, altered matrix, hypoxia and partial outlet obstruction (PBO), improves bladder smooth muscle phenotype and gene expression patterns (Aitken, 2010, Am.J.Path., Schroder, JUrol, 2013, Jiang, PLOSONE, 2015), the effect of rapamycin on gene expression during de-obstruction(dOB) is unknown. We hypothesized that rapamycin-induced changes in gene expression during dOB could uncover new pathways with functional relevance. METHODS Sprague-Dawley female rats underwent PBO by tying a silk suture around the proximal urethra and a 0.9mm steel rod, leaving only the suture in place. PBO animals were randomized to 6 week PBO or 6 week PBO plus de-obstruction (dOB). dOB were randomized to vehicle (saline) or rapamycin for 6 weeks. Shams were performed over 6 and 12 weeks. High-throughput QPCR was performed to identify genes dysregulated during dOB normalized with rapamycin treatment and were tested by students' t-test and Pearson's correlation with non-invasive functional tests which were performed prior to bladder harvest. After identifying significant dOB genes, their gene products were added exogenously to human SMC for 48 hours, fixed and stained immunofluorescently for SMC markers. We used transcription factor binding site programs (DAVID, OPOSSUM) to identify new networks. Circadian/core clock dependency of target genes and SMC markers was queried by applying a circadian modulator, the reverbα agonist, SR9009, to BSMC and testing expression of the dOB genes by QPCR over 1 day and SMC markers by immunofluorescence at 18 hours. RESULTS IGFBP7, BMP2, and SOD3 correlated with functional improvements in rapamycin treated animals. We identified the E-box bound by circadian regulators CLOCK/BMAL or NPAS2/BMAL as a potential common mechanism of transcriptional regulation amongst the three genes. Obstruction and dOB upregulated mRNA expression of CLOCK and NPAS2, above sham levels, while Rapamycin downregulated mRNA of CLOCK, NPAS2, alongside the 3 physiologic genes, p<0.01. BMAL, DEC1/2, and other core clock genes were not significantly altered during rapamycin treatment of dOB vs other groups. Exogenous IGFBP7 increased SMC hypertrophy in vitro. SR9009 had time-specific effects on expression of several core clock genes alongside BMP2 and IGFBP7, p<0.025. CONCLUSIONS Rapamycin improves core clock and dOB target gene expression, suggesting that circadian E-box binding regulators are under transcriptional control by de-obstruction and rapamycin, coordinate with changes in BSMC phenotype. © 2018FiguresReferencesRelatedDetails Volume 199Issue 4SApril 2018Page: e502 Advertisement Copyright & Permissions© 2018MetricsAuthor Information Annette SCHRODER More articles by this author Karen Aitken More articles by this author Jia-Xin Jiang More articles by this author Aliza Siebenaller More articles by this author Thenuka THANABALASINGHAM More articles by this author Martin Sidler More articles by this author Darius Bagli More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...