Mechanism of Preconditioning and Cardiocyte Apoptosis

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Cardiocyte apoptosis has been identified in many cardiac conditions associated with ischemia-reperfusion (IR) injury. Preconditioning, single or multiple brief periods of IR before a prolonged coronary artery occlusion, reduced myocardial infarct size and apoptosis in IR. The goal of this proposal is to elucidate intracellular signaling pathways by which preconditioning prevents cardiocyte apoptosis. Preconditioning generates oxygen radicals that correlate with cardioprotection in IR. Protein kinase C (PKC) and KATP channels are downstream signals of oxygen radicals. However, preconditioning produces many effects relevant to IR in intact animals and activates a series of signal transduction cascades in cardiocytes. Which effect accounts for endogenous cardioprotection remains unclear. We hypothesize that during preconditioning oxygen radicals open mitochodrial KATP channels through PKCe activation is the primary signal transduction pathway that blocks apoptosis in IR. To test this hypothesis, we will measure oxygen radical production directly in in vitro neonatal rat cardiomyocytes. We will use recombinant adenoviral vectors to overexpress in cardiocytes a wild type PKCe carrying its full length of cDNA (AdCMVPKCe-FL) or a dominant negative PKCe mutant (AdCMVPKCe-DN). The effects of these constructs on cardiocyte function, apoptosis, and infarction will be examined in both in vitro and in vivo rat models of IR. Understanding the role of specific signaling pathways by which preconditioning blocks cardiocyte apoptosis and developing local modulation of the pathways through somatic gene transfer may provide novel therapies for the management of many clinical disorders.