Mitochondrial Metabolism in Cardiac Ischemia-Reperfusion Injury

Thesis (Ph.D.)--University of Rochester. School of Medicine & Dentistry. Dept. of Pharmacology and Physiology, 2018.Ischemia-reperfusion (IR) injury is the underlying pathology of acute myocardial infarction. The severity of IR injury is driven by mitochondrial metabolism and modulated by mitochondrial quality control mechanisms. The Krebs’ cycle metabolite succinate is central to IR injury. Succinate accumulates during ischemia and its oxidation at reperfusion drives injury. The mechanism of ischemic succinate accumulation is controversial, and is proposed to involve reversal of mitochondrial complex-II. Here, using stable isotope resolved metabolomics, we demonstrated that complex-II reversal was possible in hypoxic mitochondria, but was not the primary source of succinate in hypoxic cardiomyocytes or ischemic hearts. Rather, in these intact systems succinate primarily originated from canonical Krebs' cycle activity, partly supported by aminotransferase anaplerosis and glycolysis from glycogen. Augmentation of canonical Krebs' cycle activity with dimethyl-α-ketoglutarate both increased ischemic succinate accumulation and drove substrate-level phosphorylation by succinyl-CoA synthetase, improving ischemic energetics. Although ⅔ of ischemic succinate accumulation was extracellular, the remaining ⅓ was metabolized during early reperfusion, wherein acute complex-II inhibition was protective. These results highlight a bifunctional role for succinate: its complex-II independent accumulation being beneficial in ischemia, and its complex-II dependent oxidation being detrimental at reperfusion. IR injury is also modulated by mitochondrial quality control mechanisms, such as autophagy. Mild mitochondrial uncoupling induces mitochondrial autophagy and both have separately been shown to protect against cardiac IR injury. However, a link between mild uncoupling and autophagy within the context of cardiac IR injury or cardioprotection has not been explored experimentally or therapeutically. Previously, we identified cloxyquin via phenotypic screening as a cardioprotective compound. Here, we demonstrated that cloxyquin mildly uncoupled mitochondria and induced autophagy. Cloxyquin-induced cardioprotection was blocked by the autophagy inhibitor chloroquine. Finally, we showed that cloxyquin-induced cardioprotection translated to an in vivo model of cardiac IR injury. Overall, these data suggest that enhanced insights into the molecular events that take place at mitochondria during ischemia may be exploited for therapeutic benefit in cardiac IR injury