Pilot--Akt as a Central Regulator of Bladder Muscle Gene Expression

Children'S Hospital Boston, Boston MA

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Abstract

Urinary tract obstruction (UTO) and the pathophysiologic tissue remodeling associated with it lead to significant morbidity in humans. The molecular mechanisms underlying tissue renewal in the genitourinary tract are largely unknown. However, certain phenotypic changes such as smooth muscle cell (SMC) hyperplasia and hypertrophy, and re-expression of fetal genes, parallel those that occur in cardiac hypertrophy, suggesting that similar signaling mechanisms operate in muscle cells of the heart and bladder exposed to hypertrophic stimuli. Recent studies have identified the phosphoinositide-3-kinase (PI3K)/Akt kinase pathway as a pivotal regulator of signals that lead to cardiac hypertrophy. The role of this pathway in modulating bladder smooth muscle function is undefined. We present preliminary data that stretch- and platelet-derived growth factor-BB (PDGF-BB)-induced DNA synthesis in bladder SMC is dependent on signaling through PI3K, and that DNA synthesis and Akt phosphorylation are inhibited in the presence of pharmacologic inhibitors of PI3K. These findings suggest the hypothesis that similar activation profiles are elicited in SMC exposed to mechanical or growth factor signals. The Specific Aims of the project are to: 1) determine whether signaling throught Akt is necessary and sufficient for bladder SMC growth; 2) identify downstream transcriptional targets of Akt in bladder SMC following mechanical or growth factor stimulation in vitro and in an ex vivo model of bladder overdistension; 3) determine the role of the GATA transcription factors in the response of bladder SMC to stretch and growth factor treatment and their regulation by Akt. We will employ activated and dominant-negative Akt constructs to confirm the involvement of Akt signaling in target gene regulation and growth control in bladder SMC. Members of the GATA family are known to regulate gene expression in an Akt-dependent manner in the heart in response to hypertrophic stimuli, suggesting that they may perform an analogous function in the bladder exposed to overload. These studies will allow us to clarify the role of Akt in regulation of bladder SMC growth and to identify which genes represent transcriptional targets of Akt in response to mechanical and growth factor stimulation.