Abstract 13303: Folic Acid Reverses Nitric Oxide Synthase Uncoupling and Prevents Insulin Resistance-Induced Cardiac Dysfunction: Role Of Ca2+/calmodulin Activated Kinase II (CaMKII)
It has been shown that nitric oxide synthase (NOS) becomes uncoupled in diabetes and switches from producing nitric oxide to superoxide (O2-) enroute to cardiac dysfunction. However, the downstream effector(s) from NOS uncoupling in diabetic cardiomyopathy remain elusive. In this study we examined the impact of folic acid supplement (15mg/kg/d for 4 wks), which reverses NOS uncoupling, on cardiac dysfunction cause by sucrose diet-induced insulin resistance. Insulin resistance was confirmed by glucose tolerance test while whole heart echocardiographic function and isolated cardiomyocyte contractile function were assessed. Tetracaine inhibitable rise in cytosolic Ca2+ was used to measure ryanodine receptor (RYR) leak. O2- production was monitored using DHE staining in the absence or presence of the NOS inhibitor L-NAME to examine NOS uncoupling and oxidative/phosphorylation state of CaMKII was evaluated by western blot. Our data revealed increased NOS dependent O2- production following sucrose diet feeding, (-2.4±5.7 vs. 30.8±7.1) which was reversed with folic acid administration (p<0.05, n=3). Echocardiographic function displayed decreased fractional shortening, (50.4±1.7% vs 38.2±1.4%, n≥7, p<0.05) while cardiomyocytes showed decrease peak shortening, maximal velocity of shortening and relengthening following sucrose feeding, the effects of which were restored by folic acid (n= 3, p<0.05). Insulin resistance decreased intracellular Ca2+ rise and prolonged intracellular Ca2+ clearance, which was nullified by folic acid (n=3, p<0.05). Analysis of CaMKII revealed that insulin resistance promoted phosphorylation (T286) (0.101±0.007 vs. 0.336±0.027) but not oxidation (M281-282), of CaMKII which was reversed by folic acid (n=4-5, p<0.05). Associated with CaMKII activation, an increased resting Ca2+ leak from RYR (1.37±0.45 vs. 3.65±0.64), and spontaneous Ca2+ transients (0.007±0.003 per sec. vs. 0.087±0.018 per sec.) were observed in insulin resistance, both of which were alleviated by folic acid (n=3, p<0.05). In summary, these data suggest that insulin resistance induced CaMKII phosphorylation and RYR impairment leading to cardiac dysfunction can be reversed by elimination of NOS uncoupling with folic acid supplementation.
- © 2012 by American Heart Association, Inc.