Heart disease is the major cause of death in chronic renal failure (CRF) patients. Increases in ROS (reactive oxygen species), stress and circulating inhibitors of Na/K-ATPase have been well documented in CRF patients treated with hemodialysis. Moreover, Na/K-ATPase is an important signal transduction element in cardiac myocytes. Based on our prior work, we believe that interaction between ROS and Na/K-ATPase activates multiple signaling pathways that are important for regulation of cell growth and gene expression in cardiac myocytes. Further, interaction between ROS and other circulating pump inhibitor can cause a significant inhibition of the enzyme through both transcriptional and post-translational mechanisms. Such inhibition of the enzyme will impair the ability of cardiac myocytes to extrude Na+, thus Ca2+ through Na+/Ca2+ exchanger. This certainly represents an important risk factor for development of diastolic dysfunction of the heart in CRF patients. Clearly, it is important to study how ROS interact with Na/K-ATPase and the roles of such interaction in regulation of cardiac growth, gene expression and cardiac contractile function.We, therefore, proposed the following three specific aims to address these issues.
Specific Aim I will test the hypotheses that Na/K-ATPase serves as a receptor for ROS and that inhibition of Na/K-ATPase by ROS recruits and activates Src, resulting in assembly of a signaling complex and subsequent activation of the Ras/MAPK cascade.
Specific Aim 2 will dissect pathways by which ROS post-translationally regulate Na/K-ATPase.
Specific Aim 3 will test the hypothesis that activation of Ras/MAPKs and inhibition of Na/K-ATPase regulate intracellular Ca2+([Ca2+]i) and contractility in response to increased ROS stress, and profile ROS-induced changes in gene expression and protein structures in cardiac myocytes. We proposed to use a combination of proteomics, adenovirus-mediated gene expression, cDNA expression array, representation difference analysis, confocal fluorescence microscopy, and other molecular biology techniques to critically test our working hypotheses. We expect that these basic investigations will contribute to our understanding of the biology of Na/K-ATPase, uremic cardiomyopathy and provide new information for developing novel therapies addressing the serious and common problem of heart diseases in CRF patients. Ca2+ and contractility in response to increased ROS stress, and profile ROS-induced changes in gene expression and protein structures in cardiac myocytes. We proposed to use a combination of proteomics, adenovirus-mediated gene expression, cDNA expression array, representation difference analysis, confocal fluorescence microscopy, and other molecular biology techniques to critically test our working hypotheses. We expect that these basic investigations will contribute to our understanding of the biology of Na/K-ATPase, uremic cardiomyopathy and provide new information for developing novel therapies addressing the serious and common problem of heart diseases in CRF patients.
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