Poster, Podium & Video Sessions
Presentation Authors: Mahendra Kashyap*, Christopher Chermansky, Naoki Yoshimura, Pradeep Tyagi, Pittsburgh, PA
Introduction: MicroRNAs (miRs) are involved in the post-transcriptional regulation of gene expression, and there is growing evidence for their pathological role in overactive bladder (OAB). We recently reported on the perturbed expression of miR-132 in acetic acid induced bladder overactivity (BO). Here, we investigate the functional significance of miR-132 overexpression in bladder and its effect on acetylcholinesterase (AChE) and other target genes in absence of acetic acid exposure.
Methods: Under isoflurane anesthesia, adult female Sprague-Dawley rats were either given bladder wall injection of either 10µg of plasmid vector pLV-[hsa-mir-132] or reporter plasmid encoding luciferase gene complexed with protamine and liposomes in a volume of 80µl. Bladder wall injection was performed at 4 separate sites (20 µL at each anterior, posterior and bilateral) with a 30-gauge needle. 7 days after transfection, transurethral open cystometry under urethane anesthesia ( 1g/kg s.c) was performed and harvested bladder was weighed prior to bladder strip contractility, histology and quantitative real time PCR analysis for AChE, Nerve growth factor (NGF), connexin-43 (Cx-43), monocyte chemoattractant protein (MCP-1) and soluble intracellular adhesion molecule (sICAM-1).
Results: Bladder wall transfection of miR-132 plasmid upregulated the miR-132 expression and evoked BO in absence of acetic acid. The intercontractile interval of 22.28± 1.49 min noted in luciferase transfected group was significantly reduced to 10.5± 0.8 min, while bladder weight was raised from 118.4± 1.12mg to 146.8 ±7.66mg and so was the contractile response of strips to KCl and electric field after transfection of miR-132 plasmid(*p<0.01, n=5). These changes in miR-132 group were associated with a 3 fold downregulation of AChE, 5 fold upregulation of NGF and MCP-1, while a 2 fold upregulation of Cx 43 and sICAM-1.
Conclusions: Observed BO and bladder hypertrophy following exogenous overexpression of miR-132 demonstrate the pathological role of miR-132 in OAB. The miR-132 mediated downregulation of AChE and upregulation of Cx43 explain the enhanced detrusor strip contractility. Upregulation of NGF and chemokines by miR-132 support it as a putative mediator of communication between neural and immune cells of bladder and therefore a suitable OAB drug target.
Source Of Funding: NIH DK088836
University of Pittsburgh
Dr. Mahendra P. Kashyap successfully completed his PhD from Jamia Hamdard University (New Delhi) in association with at Indian Toxicology Research Institute, Lucknow- India. His major research focus was to develop and validate the new and more sensitive human stem cell based in vitro models to understand the mechanisms involved in the pesticide-induced developmental neurotoxicity and therapeutic interventions. Following the isolation and characterization of human umbilical cord blood stem cells, these cells were allowed to differentiate into neuronal cells. At various time points of neuronal maturity; expression and activity of markers involved in neuronal development, injury and repair were studied in presence and absent of toxicants such as Organophosphate pesticide. After completion of PhD, he moved to United State at the University of Pittsburgh School of medicine as a Research Associate in June, 2012. During his postdoctoral training, he has been involved in deciphering the role of different neurotrophins such as NGF and BDNF in the Lower Urinary tract diseases such as Overactive Bladder/Interstitial Cystitis and Prostatitis in normal as well as in Spinal cord injured animals. Presently, his research is focused on the development of antisense/ antagomirs based novel therapeutic interventions in preclinical models of Lower Urinary tract dysfunctions.
Saturday, May 13
9:30 AM – 11:30 AM