Abstract 284: Elevation of Cardiac NAD+ Levels Improves Mitochondrial Function in Mice Lacking Sirtuin 3

Abstract

The mitochondria rely heavily on the ratio between NAD+ and its reduced form NADH to maintain proper function and generate 95% of cardiac cellular energy. Sirtuin 3 (SIRT3) is the major mitochondrial deacetylase and its activity level has been linked to the NAD+/NADH ratio. Numerous studies have shown that deletion of SIRT3 results in hyperacetylation and impairment of mitochondrial enzymes involved in fatty acid metabolism and in the mitochondrial respiratory chain. Because impairments in these processes have shown to be associated with decreases in the NAD+/NADH ratio and contribute to deficits in energy production, we hypothesized that the SIRT3 knockout mice (SIRT3 -/- ) would have a decreased NAD+/NADH ratio and that normalizing the ratio would improve mitochondrial respiratory function and inner membrane potential. In the present study, we observed increases in both NAD+ (1.1-fold, n=3, p<0.05) and NADH (1.5-fold) in cardiac tissue from 14-week old SIRT3 -/- mice compared to wild-type (WT) controls, resulting in a 26±2.2% reduction in the NAD+/NADH ratio. These changes correlated with decreased Complex I ADP stimulated respiration (173±16 vs 118±14 nmolO 2 /min/mg for WT and SIRT3 -/- , respectively, n=3, p<0.05) and suppressed mitochondrial membrane potential. Intraperitoneal injection (I.P.) of Nicotinamide Riboside (NR) increased NAD+ levels in cardiac tissue lysates (WT 1.5-fold and SIRT3 -/- 1.4-fold, n=5, p<0.05) and in mitochondria isolated (WT 1.9-fold and SIRT3 -/- 1.7-fold) from the mice with no significant changes in NADH levels. Therefore, the NR I.P. injections normalized the NAD+/NADH ratio, partially restored the Complex I supported mitochondrial respiration (123±11 vs 157±8 nmolO 2 /min/mg for vehicle and NR treated SIRT3 -/- mice, respectively, n=3, p<0.05), and improved mitochondrial membrane potential in the SIRT3 -/- mice. These results suggest that increasing cardiac NAD+ levels can rescue mitochondrial dysfunction independent of SIRT3 protein deacetylation and warrants further investigation. The next step will be to test whether increasing cardiac NAD+ levels can improve mitochondrial function and reduce injury in SIRT3 -/- mice subjected to chronic heart stress induced by transverse aortic constriction surgery.