Abstract 12874: A Novel Mechanism in the Pathogenesis of Heart Failure With Preserved Ejection Fraction Through Hexokinase Mitochondrial Dislocation and Substrate Shuttling Into the O-Glcnacylation Machinery

Abstract

Introduction: Endothelial cell (EC) dysfunction underlies the pathogenesis of heart failure with preserved ejection fraction (HFpEF), however, the mechanism is unclear. ECs are highly dependent on glycolysis; however, it is not known how their metabolic profile is altered in the setting of HFpEF. Hexokinase 1 (HK1) carries the first step in glycolysis and is mostly bound to the mitochondria in ECs. Objective: We sought to elucidate the role of EC glucose metabolism, and specifically HK1, in the development of HFpEF. Results: We first showed that among glycolytic enzymes, HK1 is highly specific to ECs, suggesting the significance of HK1 in EC physiology. Although HK1 mRNA and protein levels are not changed, we noted increased dislocation of HK1 from the mitochondria in ECs from HFpEF mice. To study the role of HK1 dislocation in the development of HFpEF, we generated mice with mitochondrial-binding domain of the endogenous HK1 replaced with Flag tag (ΔE1HK1). ΔE1HK1mice develop impaired cardiac relaxation at 20 weeks of age and HFpEF by 40 weeks, and ECs from these mice displayed impaired angiogenic potential. Since HK1 is involved in glucose metabolism, we then performed metabolomic studies and demonstrated that intermediates in hexosamine biosynthetic pathway (HBP) are significantly decreased in ECs from ΔE1HK1 mice. To assess the mechanism for the decrease in HBP, we assessed protein modification, and noted reduced protein N-glycosylation and increased O-GlcNAcylation in ECs from ΔE1HK1. We also demonstrated that inhibition of the key enzyme in O-GlcNAcylation (OGT) reverses the EC dysfunction noted in ΔE1HK1 mice, indicating that O-GlcNAcylation is responsible for the angiogenic defect in ECs from ΔE1HK1. Finally, to provide a mechanism for altered protein glycosylation with HK1 mitochondrial dislocation, we showed that HK1 associates with N-glycosylation machinery when it is attached to mitochondria, while it binds to OGT when it is dislocated from mitochondria. Conclusion: Our studies demonstrate a role for HK1 mitochondrial binding and protein O-GlcNAcylation in the pathogenesis of HFpEF, and that HK1 cellular localization plays a major role in the fate of HBP intermediates into either N-glycosylation or O-GlcNAcylation machinery in ECs.