Dynamic regulation of metabolism and respiration by endogenously produced nitric oxide protects against oxidative stress

One of the many biological functions of nitric oxide is the ability to protect cells from oxidative stress. To investigate the potential contribution of low steady state levels of nitric oxide generated by endothelial nitric oxide synthase (eNOS) and the mechanisms of protection against H2O2, spontaneously transformed human ECV304 cells, which normally do not express eNOS, were stably transfected with a green fluorescent-tagged eNOS cDNA. The eNOS-transfected cells were found to be resistant to injury and delayed death following a 2-h exposure to H2O2 (50-150 muM). Inhibition of nitric oxide synthesis abolished the protective effect against H2O2 exposure. The ability of nitric oxide to protect cells depended on the presence of respiring mitochondria as ECV304+eNOS cells with diminished mitochondria respiration (rho (-)) are injured to the same extent as nontransfected ECV304 cells and recovery of mitochondrial respiration restores the ability of nitric oxide to protect against H2O2-induced death. Nitric oxide also found to have a profound effect in cell metabolism, because ECV304+eNOS cells had lower steady state levels of ATP and higher utilization of glucose via the glycolytic pathway than ECV304 cells. However, the protective effect of nitric oxide against H2O2 exposure is not reproduced in ECV304 cells after treatment with azide and oligomycin suggesting that the dynamic regulation of respiration by nitric oxide represent a critical and unrecognized primary line of defense against oxidative stress.