Early determinants of H2O2-induced endothelial dysfunction

Reactive oxygen species (ROS) can stimulate nitric oxide (NO center dot) production from the endothelium by transient activation of endothelial nitric oxide synthase (eNOS). With continued or repeated exposure, NO center dot production is reduced, however. We investigated the early determinants of this decrease in NO center dot production. Following an initial H2O2 exposure, endothelial cells responded by increasing NO center dot production measured electrochemically. NO center dot concentrations peaked by 10 min with a slow reduction over 30 min. The decrease in NO center dot at 30 min was associated with a 2.7-fold increase in O-2(center dot-) production (p < 0.05) and a 14-fold reduction of the eNOS cofactor, tetrahydrobiopterin (BH4, P < 0.05). Used as a probe for endothelial dysfunction, the integrated NO center dot production over 30 min upon repeated H2O2 exposure was attenuated by 2.1-fold (P = 0.03). Endothelial dysfunction could be prevented by BH4 cofactor supplementation, by scavenging O-2(center dot-) or peroxynitrite (ONOO-), or by inhibiting the NADPH oxidase. Hydroxyl radical ((OH)-O-center dot) scavenging did not have an effect. In summary, early H2O2-induced endothelial dysfunction was associated with a decreased BH4 level and increased O-2(center dot-) production. Dysfunction required O-2(center dot-), ONOO-, or a functional NADPH oxidase. Repeated activation of the NADPH oxidase by ROS may act as a feed forward system to promote endothelial dysfunction. (c) 2006 Elsevier Inc. All rights reserved.