L-2-Hydroxyglutarate and Metabolic Remodeling in Hypoxia

Project Summary. Cellular hypoxia-reoxygenation or organ ischemia-reperfusion promotes intracellular redox stress, which can lead to cellular dysfunction and injury through the generation of reactive oxygen species (ROS). In prior work, we identified the role of L(S)-2-hydroxyglutarate (L2HG) as a unique metabolic derivative of α-ketoglutarate (2-oxoglutarate, or 2OG) that increases in hypoxia or ischemia. In the ongoing cycle of this award, we have shown that L2HG decreases glycolysis and increases pentose- phosphate pathway (PPP) activity as a unique mechanism for promoting redox protection in endothelial cells and in the myocardium. In this next proposed four-year cycle, we turn our attention to the relationship between L2HG and branched-chain ketoacids (BCKAs), metabolic derivatives of branched- chain amino acids (BCAAs), and their role in redox protection in the heart. Upon examination of the metabolome in hypoxic cardiovascular cells, we observed that BCKAs are significantly increased compared with normoxic cells, and that this increase in the BCKA pool is also an important determinant of redox protection. Abundant preliminary data provided in this application demonstrate that the redox protection afforded by BCKAs is mediated by Hif-1α stabilization, and comes about by a unique mechanism: an increase in BCKAs leads to an increase in BCAAs, which is associated with an increase in 2OG generation from glutamate via reamination of BCKAs. The increase in 2OG in the reductive environment of hypoxia leads to an increase in L2HG, with accompanying redox-protective metabolic reprogramming. Thus, the central hypothesis of this proposal is that an hypoxia-induced accumulation of BCKAs in endothelial cells (ECs) and cardiomyocytes (CM) increases L2HG, which together promote redox protection. To test this hypothesis, we propose three specific aims: 1) we will determine the effects of hypoxia on BCAA and BCKA metabolism in ECs and CMs; 2) we will identify the molecular and metabolic determinants linking BCKA metabolism with L2HG generation, and their consequences for Hif stabilization in ECs and CMs; and 3) we will study the effects of the BCKA-L2HG metabolic axis in hypoxia or ischemia on redox protection and function in ECs, CMs, and the myocardium. The results of these studies should provide useful insight into the complex interplay of metabolic intermediates that govern redox potential and redox protection in the myocardium, and its potential consequences for ischemia-mediated cardiovascular diseases.