Simultaneous assessment of endothelial function, nitric oxide synthase activity, nitric oxide-mediated signaling, and oxidative stress in individuals with and without hypercholesterolemia

Abstract

BACKGROUND: Endothelial function is impaired in hypercholesterolemia and atherosclerosis. Based on mostly indirect evidence, this impairment is attributed to reduced synthesis or impaired biological activity of endothelium-derived nitric oxide (NO). It was the aim of this study to directly estimate and compare whole-body NO production in normo- and hypercholesterolemia by applying a nonradioactive stable isotope dilution technique in vivo. METHODS: We enrolled 12 normocholesterolemic and 24 hypercholesterolemic volunteers who were all clinically healthy. To assess whole-body NO synthesis, we intravenously administered L-[guanidino-(15N2)]- arginine and determined the urinary excretion of 15N-labeled nitrate, the specific end product of NO oxidation in humans, by use of gas chromatography-mass spectrometry. In addition, we measured flow-mediated vasodilation (FMD) of the brachial artery, expression of endothelial NOS (eNOS) in platelets, plasma concentration of the endogenous NOS inhibitor asymmetric dimethylarginine (ADMA), and urinary excretion of 8-isoprostaglandin F 2α (8-iso-PGF2α). RESULTS: After infusion of L-[guanidino-(15N2)]-arginine, cumulative excretion of 15N-labeled-nitrate during 48 h was 40% [95% CI 15%-66%] lower in hypercholesterolemic than normocholesterolemic volunteers [mean 9.2 (SE 0.8) μmol vs 15.4 (2.3) μmol/l, P = 0.003]. FMD was on average 36% [4%-67%] lower in hypercholesterolemic than normocholesterolemic volunteers [6.3 (4.0)% vs 9.4 (4.6)%, P = 0.027]. Normalized expression of NOS protein in platelets was also significantly lower in hypercholesterolemic volunteers, whereas there were no significant differences in plasma ADMA concentration or urinary excretion of 8-iso-PGF2α between the 2 groups. CONCLUSIONS: This study provides direct evidence for a decreased whole body NO synthesis rate in healthy people with hypercholesterolemia. © 2007 American Association for Clinical Chemistry.