Dietary Correction of Hypercholesterolemia in the Rabbit Normalizes Endothelial Superoxide Anion Production

BACKGROUND: We have shown that hypercholesterolemia increases vascular superoxide anion (O2-) production, which could be responsible for augmented inactivation of endothelium-derived vascular relaxing factor. We sought to determine whether this increased vascular O2- production is due to infiltration of macrophages into the intima and whether dietary treatment of hypercholesterolemia normalizes O2- production. METHODS AND RESULTS: A specific and sensitive assay for O2- based on chemiluminescence of lucigenin was used; the amount of O2- produced by vascular ring segments was quantified based on known quantities of O2- produced by xanthine-xanthine oxidase standards. O2- production of aortic segments from normal rabbits (n = 9), cholesterol-fed rabbits (1% cholesterol diet for 1 month, n = 7), and rabbits fed a 1% cholesterol diet for 1 month followed by a normal diet for 1 month (regression rabbits, n = 5) was measured. At the end of these diets, serum cholesterol levels were 1.5 +/- 0.2, 26.0 +/- 3.9, and 1.8 +/- 0.5 mmol/L (58 +/- 6, 1000 +/- 150, and 71 +/- 19 mg/dL) in the normal, cholesterol-fed, and regression animals, respectively. Vessels from normal rabbits with endothelium produced 0.32 +/- 0.06 nmol O2-/mg dry wt per minute, whereas those without endothelium produced approximately twice as much O2- (0.66 +/- 0.12 nmol O2- mg dry wt per minute. Vessels with endothelium from cholesterol-fed rabbits produced 4.5-fold more O2- than vessels from normal animals. This increased production of O2- was normalized by endothelial removal. This increased production of O2- was not due to infiltration of macrophages in the intima, because there was no correlation between vascular O2- production and macrophage infiltration assessed by immunohistochemistry with use of a specific antibody against rabbit macrophage. O2- production by vessels from regression rabbits was similar to that observed in normal animals, and as in the normal rabbits, endothelial removal increased O2- production. Aortic rings from these animals also were studied in organ chambers. Dietary lowering of cholesterol dramatically improved vasodilator responses to acetylcholine and A23187 (P < .05 versus cholesterol-fed rabbits). CONCLUSIONS: Dietary lowering of cholesterol not only improves endothelium-dependent vascular relaxation but also normalizes endothelial O2- production. Decreases of O2- production by dietary lowering of cholesterol not only may improve vasomotor control but also may improve other aspects of vascular integrity in atherosclerosis.