Mechanisms of H2O2-induced oxidative stress in endothelial cells exposed to physiologic shear stress.
Hydrogen peroxide (H2O2) is produced by inflammatory and vascular cells and induces oxidative stress, which may contribute to vascular disease and endothelial cell dysfunction. In smooth muscle cells, H2O2 induces production of O2 by activating NADPH oxidase. However, the mechanisms whereby H2O2 induces oxidative stress in endothelial cells are not well understood, although O2 may play a role. Recent studies have documented increased O2 in endothelial cells exposed to H2O2 via uncoupled nitric oxide synthase (NOS) and NADPH oxidase under static conditions. To assess responses to H2O2 in porcine aortic endothelial cells (PAEC) under shearing conditions, a constant flow rate of 24. 4 ml/min was applied to produce physiologically relevant shear stress (8. 2 dynes/cm). Here we demonstrate that treatment with 100 muM H2O2 increases intracellular O2 levels in PAEC. In addition, we demonstrate that l-NAME, an inhibitor of NOS, and apocynin, an inhibitor of NADPH oxidase, reduced O2 levels in PAEC treated with H2O2 under physiologic shear suggesting that both NOS and NADPH oxidase contribute to H2O2-induced O2 in PAEC. Co-inhibition of NOS and NADPH oxidase also reduced intracellular O2 levels under shear. We conclude that H2O2-induced oxidative stress in endothelial cells exhibits increased intracellular O2 levels through NOS and NADPH oxidase under shear. The inhibition of NOS and NADPH with H2O2 exposure is nonlinear, suggesting some interdependent or compensating system within endothelial cells. These findings suggest a complex interaction between H2O2 and oxidant-generating enzymes that may contribute to endothelial dysfunction in cardiovascular diseases.
Coyle
Christian H
CH
Cell and Synthetic Interface Engineering Laboratory, Department of Biomedical Engineering, University of Iowa, Iowa City, Iowa 52242, USA.
Kader
Khalid N
KN
D000818:Animals; D002478:Cells, Cultured; D042783:Endothelial Cells / Q000187:drug effects* / Q000378:metabolism; D006801:Humans; D006861:Hydrogen Peroxide / Q000633:toxicity*; D019255:NADPH Oxidases / Q000502:physiology; D019331:NG-Nitroarginine Methyl Ester / Q000494:pharmacology; D052250:Nitric Oxide Synthase Type III / Q000502:physiology; D018384:Oxidative Stress; D013314:Stress, Mechanical; D013481:Superoxides / Q000378:metabolism; D013552:Swine
D013481:Superoxides; D006861:Hydrogen Peroxide; D052250:Nitric Oxide Synthase Type III; D019255:NADPH Oxidases; D019331:NG-Nitroarginine Methyl Ester