Abstract 929: Nitric Oxide Generated by eNOS Exerts Cardioprotection by Increasing the Mitochondrial Biogenesis and Decreasing Electron Leakage for Superoxide Production by Mitochondrial Succinate Cytochrome c Reductase.
Nitric oxide (NO) produced from endothelial cells has the capacity to modulate mitochondrial functions in regulation of metabolism, respiration, and mitochondrial biogenesis. The succinate ubiquinone cytochrome c reductase (SCR or supercomplex containing Complex II and Complex III) is a crucial segment of mitochondrial electron transport chain. SCR has been identified as an essential part in the mediation of superoxide generation, and an alternative target of NO in mitochondria during oxidative stress. In this work, we have assessed the hypothesis that NO generated from endothelial NO synthase (eNOS or NOS3) exerts a protective effect on myocardial SCR by decreasing its superoxide generation activity (SGA) and increasing its biogenesis. When the myocardial tissue from eNOS knockout [eNOS(−/−)] mice were subjected to electron transfer activity (ETA) analysis, the ETA of the SCR in eNOS(−/−) myocardium was significantly decreased by 29.6±8.5% (n=9, P<0.05). The ETA of individual Complex II and Complex III were decreased by 21.8±5.2% (n=7, P <0.05) and 30.9±6.5% (n=7, P<0.05) respectively. The mRNA expressions of Complex II and Complex III in eNOS(−/−) myocardium were decreased by 48.7% and 33.3% respectively as measured by quantitative real-time PCR. The mRNA expression level of transcription coactivator, PGC-1α (PPAR-γ coactivator), was decreased by 25.6%. The mRNA level of mitochondrial transcription factor A (TFAM) was deceased 15%, indicating a decrease of mitochondrial biogenesis in eNOS(−/−) myocardium. The SCR isolated from the heart mitochondria after repeat exposures to low dosage of NO (1 μM) in the presence of succinate showed a 100% ETA preserved and a 50% decrease of SGA as measured by EPR spin-trapping using 5-diethoxylphosphoryl-5methyl-1-pyroline N-oxide (DEPMPO). Under this condition, formation of nitrosothiol at the 70 kDa FAD-binding subunit of SCR was further detected by using biotin switch method. Furthermore, O2 ·− generation by SCR also induced oxidative inhibition of the ETA of SCR in vitro. In conclusion, NO produced by eNOS protects mitochondrial function in the heart by modulating the network of biogenesis program and decreasing mitochondrial SCR-mediated superoxide production through nitrosothiol formation.