Oxidative stress has been shown to convert endothelial nitric oxide synthase (eNOS) from an NO-producing enzyme to an enzyme that generates superoxide, a process termed NOS uncoupling. This uncoupling of eNOS converts it to function as an NADPH oxidase with superoxide and hydrogen peroxide generation. eNOS uncoupling has been associated with many pathophysiologic conditions, such as heart failure, ischemia/reperfusion injury, hypertension, atherosclerosis, and diabetes. The mechanisms implicated in the uncoupling of eNOS include oxidation of the critical NOS cofactor tetrahydrobiopterin, depletion of l-arginine, and accumulation of methylarginines. All of these prior mechanisms of eNOS-derived reactive oxygen species formation occur primarily at the heme of the oxygenase domain and are blocked by heme blockers or the NOS inhibitor N-nitro-larginine methylester. Recently, we have identified another unique mechanism of redox regulation of eNOS through S-glutathionylation that was shown to be important in cell signaling and vascular disease. Herein, we briefly review the mechanisms of eNOS uncoupling as well as their interrelationships and the evidence for their importance in disease. Antioxid. Redox Signal. 14, 1769-1775.
Oxidative stress has been shown to be of critical importance in the pathogenesis of a wide variety of disease processes including heart attack, stroke, diabetes, hypertension, cancer, excitotoxic brain injury, neurodegenerative disease, and the broad spectrum of inflammatory disease (35). Oxidative stress occurs in cells and tissues because of overproduction of superoxide (O 2 -) and its secondary oxidants that are formed from several sources, including NADPH oxidase, xanthine oxidase, and the mitochondrial electron transport chain (3,35,48,56). In addition, under some conditions, nitric oxide synthase (NOS) can become a source of O 2 - (15,31,36,43,49,50). Moreover, nitric oxide (NO) produced from NOS can react with O 2 -to generate the potent oxidant, peroxynitrite (ONOO -) (2, 46, 51, 54, 55). Endothelial NOS (eNOS) is responsible for the enzymatic generation of NO within the endothelium. eNOS is a homodimeric heme-containing enzyme that converts l-arginine and oxygen to l-citrulline, NO, and water, using NADPH as a source of reducing equivalents (36). This process involves the calcium/calmodulin-regulated transfer of electrons from NADPH through a flavin adenine dinucleotide (FAD)-and flavin mononucleotide (FMN)-containing reductase domain to the heme of the l-arginine-binding oxygenase domain, which also binds tetrahydrobiopterin (BH 4 ) (6, 30). In the absence of BH 4 , NO synthesis is abrogated, and instead, O 2 -is generated (43,50).NO is involved in a number of roles within the endothelium, regulating vascular tone, vascular growth, platelet aggregation, and modulation of inflammation (23,29,32). As such, endothelial dysfunction produced by a decrease in bioavailable NO is a characteristic feature in many disease states, and one of the mechanisms underlying this dysfunction is th...