Superoxide Production in Vascular Smooth Muscle Contributes to Oxidative Stress and Impaired Relaxation in Atherosclerosis

Abstract

The endothelium is a source of reactive oxygen species in short-term models of hypercholesterolemia and atherosclerosis. We examined a chronic model of atherosclerosis for increased vascular production of superoxide (O2-.) and determined whether endothelial overexpression of superoxide dismutase (SOD) would improve endothelium-dependent relaxation. Superoxide generation was 3 times higher in isolated aortas from Watanabe heritable hyperlipidemic (WHHL) rabbits (2 to 4 years old) compared with aortas from New Zealand White (NZ) rabbits (43+/-10 versus 14+/-2 relative light units x min(-1) x mm(-2), n=9, P<0.05). After in vitro transduction with adenovirus containing the gene for CuZn-SOD (AdCMVCuZn-SOD) or extracellular SOD (AdCMVEC-SOD), endothelial O2-. levels in WHHL aortas were significantly reduced. Gene transfer of SOD to WHHL aortas, however, failed to improve the impaired relaxation to acetylcholine or calcium ionophore. By use of the oxidative fluorescent dye hydroethidine, an in situ assay indicated markedly increased generation of O2-. throughout the wall of WHHL aorta, especially within layers of smooth muscle. This finding was confirmed by demonstrating increased O2-. levels in smooth muscle cells cultured from WHHL aorta. We conclude that elevated O2-. levels in atherosclerotic vessels are not confined to the endothelium but occur throughout the vascular wall, including smooth muscle cells. Reduction in endothelial O2-. levels is not sufficient to improve endothelium-dependent relaxation. Generation of reactive oxygen species within the media may contribute to vasomotor dysfunction in atherosclerosis.

Cytochrome P450 1B1 Contributes to the Development of Angiotensin II–Induced Aortic Aneurysm in Male Apoe−/− Mice

“…21 Briefly, aorta sections were incubated in PBS for 30 minutes at 37 C and then encircled with the hydrophobic pen. DHE (5 mmol/L) was topically applied, followed by incubation at 37 C in the dark for 30 minutes.…”

Mechanisms of H2O2-induced oxidative stress in endothelial cells

“…Incubation was continued for an additional 30 minutes in the presence of 2 µM dihydroethidium (Carter et al 1994;Miller et al 1998) (DHE, Molecular Probes). After incubation, PAEC were washed, placed in phenol-red-free and serum-free M199, and imaged.…”

“…PAEC were incubated for 30 minutes with 2 µmol/L dihydroethidium ( (Carter et al 1994;Miller et al 1998) DHE, Invitrogen) after 1 hour incubation with 60 µmol/L H 2 O 2 in M199 or M199 as a control. After incubation with DHE, PAEC were washed in phenol-red-free M199 and imaged with a Zeiss fluorescent microscopy utilizing a rhodamine filter cube.…”

Gene Transfer of Endothelial NO Synthase and Manganese Superoxide Dismutase on Arterial Vascular Cell Adhesion Molecule-1 Expression and Superoxide Production in Deoxycorticosterone Acetate-Salt Hypertension

“…7,8 In angiotensin II (Ang II)-induced hypertension, Ang-II stimulates vascular NADPH oxidase to produce superoxide, which not only inactivates NO and impairs vasomotor function 9,10 but also contributes to atherogenesis by the activation of VCAM-1. 11 The elevated superoxide is present throughout the atherosclerotic vessel wall, 12 and a major source of superoxide is from the electron transport chain in mitochondrial respiration. 13 Two recent studies have indicated that there is also a substantial increase of vascular superoxide with reduced NO release in deoxycorticosterone acetate (DOCA)-salt hypertension, 14,15 a model well known for its suppressed plasma renin and angiotensin levels.…”