Project Summary Cardiovascular diseases are currently the major cause of morbidity and mortality in the world. This is due to the inability of the adult mammalian heart to replace damaged tissue following injury. Identifying novel approaches towards regenerating heart tissue following injury has significant therapeutic potential for heart failure patients. Until recently, complete heart regeneration following injury has been observed only in lower vertebrates. However, the ability of neonatal mice to regenerate their hearts following injury for a brief window after birth indicates that uncovering the evolutionarily conserved mechanisms of cardiac regeneration has great potential to treat human heart failure. The transition from embryonic/neonatal states to an adult state is accompanied with a metabolic switch for energy utilization from glycolysis to oxidative phosphorylation. This metabolic switch leads to a significant increase in reactive oxygen species (ROS) production from the mitochondria, which causes cardiomyocyte DNA damage and results in cardiomyocyte cell cycle exit and loss of the endogenous cardiac regeneration potential. What regulates this metabolic switch, and whether individual metabolites can regulate ROS production and cardiac tissue regeneration remains unknown. Increased ROS production in ischemic tissues has been demonstrated to occur as a result of the accumulation of the mitochondrial metabolite succinate, and inhibition of succinate dehydrogenase (SDH) blocks succinate accumulation and is cardioprotective against redox insult during ischemia/reperfusion (IR) injury. Thus, we hypothesized that changes in oxygen levels following birth might trigger succinate accumulation and ROS production, which contributes to cardiomyocyte cell-cycle exit in the postnatal heart. Our preliminary results demonstrate that injection of succinate in the neonatal mouse heart results in inhibition of neonatal cardiomyocyte proliferation and regeneration. Conversely, inhibition of SDH by malonate treatment after birth extends the window of cardiomyocyte proliferation and regeneration in juvenile mice. Administration of Atpenin A5, a potent inhibitor of SDH, induces a regenerative response in juvenile mice similar to malonate, demonstrating a central role for SDH inhibition in promoting cardiomyocyte proliferation and regeneration following injury. Remarkably, malonate treatment of adult mice following MI stimulates cardiomyocyte proliferation, revascularization, and results in complete restoration of cardiac structure and function following infarction. More importantly, malonate treatment at 1-week post-MI following the establishment of infarction and reduction of cardiac function results in myocardial regeneration and restoration of cardiac function over time. Our overarching hypothesis is that malonate metabolically reprograms the adult mammalian heart to a regenerative state via SDH inhibition. Our goal in this proposal is to dissect the cellu...