NAD + Redox Imbalance Exacerbates Diabetic Cardiomyopathy via Enhanced Troponin I Phosphorylation, Impaired Energetics and Prolonged Cardiac Relaxation

Abstract

Diabetes is a major risk factor of heart failure. Diabetes and heart failure have been linked to NAD redox imbalance, whose role in the progression of diabetic cardiomyopathy (DCM) is far from established. Streptozotocin (STZ)-induced diabetes in C57BL/6 mice for 16 weeks promoted declines in systolic and diastolic function and lowered cardiac NAD/NADH ratio. We employed cardiac-specific Ndufs4-KO mice (cKO), which exhibit lowered cardiac NAD/NADH ratio with normal baseline function and energetics, to determine the role of NAD redox imbalance in DCM. Plasma metabolomic analysis showed similar diabetic stress in diabetic control and cKO mice. Systolic (fractional shortening) and diastolic (E’/A’ ratio; isovolumic relaxation time, IVRT) dysfunctions were further exacerbated in diabetic cKO mice in both male and female cohorts. Collagen levels and transcript analyses of fibrosis and extracellular matrix-dependent pathways did not show changes in diabetic cKO hearts, suggesting that the exacerbated cardiac dysfunction was likely due to cardiomyocyte dysfunction. We next analyzed protein acetylation-dependent pathways to account for the exacerbated decline of function in diabetic cKO hearts. NAD redox imbalance in diabetic cKO hearts promoted protein acetylation, including SOD2 acetylation (SOD2-K68Ac). This SOD2 acetylation inhibits its antioxidant function and we observed elevated protein oxidation levels in diabetic cKO hearts. To gain further insights how NAD redox imbalance may regulate cardiomyocyte function, phosphorylation levels of myosin binding protein C (MyBPC) and troponin I (TnI) were examined. MyBPC phosphorylation at S282 is suppressed in failing hearts and remained unchanged in diabetic cKO hearts. TnI phosphorylation at S150 (TnI-S150Pi) increases myofilament calcium sensitivity and prolongs calcium dissociation, while TnI-S23/24Pi decreases calcium sensitivity and accelerates calcium dissociation. In diabetic cKO hearts, levels of TnI-S150Pi were elevated while TnI-S23/24Pi remained unchanged. TnI-S150Pi is sensitive to AMPK, the master energy sensor, and we found that ATP levels decreased, and AMP/ATP ratio increased in diabetic cKO hearts. Elevation of cardiac NAD levels by nicotinamide phosphoribosyltransferase (NAMPT) normalized NAD redox balance and alleviated systolic and diastolic dysfunction. Importantly, elevated TnI-S150Pi, exacerbated diastolic dysfunction and prolonged IVRT in diabetic cKO hearts were all restored by NAMPT over-expression. The data collectively suggest that NAD redox imbalance promotes impairments in cellular energetics and diastolic function via enhancing TnI-S150Pi, and is a critical mediator of the progression of DCM.