Cardiac muscle injury caused by ischemia and reperfusion involves impairment of the electrophysiological, contractile, and energy producing systems of the heart, and mitochondria play a key role in cell survival and recovery of function. Several sites of reversible or irreversible mitochondrial dysfunction have been noted in the post-ischemic heart, yet little is know about how the overall protein profile is altered and whether such changes are detrimental or self-protective. Myocardial preconditioning (PC), the most powerful means of protecting the heart against cell death after severe ischemia, clearly illustrates this dichotomy. With ischemia-induced PC, a series of short sublethal ischemic events trigger a powerful cellular defense response that ultimately limits infarct size. PC involves two components: an early window (classical) with duration of minutes to 2-3 hours and a second window (late or delayed) that develops at about 12 hours and last days. The two windows differ in duration and the robustness of protection and the underlying cellular mechanism(s) responsible for each. Since both the protection and injury associated with ischemia involve major changes at the level of the mitochondria, the focus of this project will be to characterize how ischemia, or other preconditioning stimuli, affect the mitochondrial proteome. Taking this objective to the next level, we will assess how the changes in the mitochondrial protein profile influence the control of oxidative phosphorylation (OxPhos), using both computational methods and experiments on intact muscles from post-ischemic hearts. Our underlying hypothesis is that PC alters the mitochondrial subproteome in such a manner that the detrimental changes elicited with a severe and lethal insult are mitigated with respect to naive myocardium and that in-depth temporal analysis will reveal the underlying mechanism. In other words, comparison of mitochondrial proteomic alterations in naive and PC myocardium will reveal the underlying proteins that are central to the cell's struggle for survival. By carrying out a systematic in-depth proteomic investigation of the mitochondrial subproteome, including careful quantification and characterization of individual proteins and protein complexes, coupled with a plan to assess the functional effects of such changes, we will be able to understand how manipulating OxPhos can result in cardioprotection, with the eventual goal of inducing these changes in high risk patients to reduce the myocardial damage associated with acute MI.
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