The influence of beta-very low density lipoprotein on nitric oxide synthase expression and activity in vitro and in vivo

The objective of this study was to determine whether β-VLDL might modulate endothelial nitric oxide synthase (eNOS) expression and nitric oxide (NO) production. Human umbilical vein endothelial cells (HUVEC) were treated with increasing concentrations of β-VLDL (30–250 mg cholesterol/dl) for various durations (6–48 h). Semiquantitative RT-PCR and Western blot analyses indicate that both eNOS mRNA and protein expression were downregulated by high levels of β-VLDL after 48h incubation. Total nitrite (NO _x) production in experimental media and DPBS after HUVEC were treated for 48 h with various concentrations of β-VLDL was assessed by Griess assay. NO_x production was increased in experimental media for all concentrations of β-VLDL compared to control. The amount of NO_x detected in DPBS was increased with 60 mg cholesterol/dl of β-VLDL, and decreased with 250 mg cholesterol/dl of β-VLDL compared to control, respectively. These findings suggest a potential mechanism whereby high concentrations of β-VLDL may induce endothelial dysfunction.; Another objective of this study was to examine the effect of vitamin E on eNOS and iNOS mRNA expression in hypercholesterolemic rabbits. In the present study, the rabbits were divided into three experimental groups: control diet, 0.5% cholesterol diet, and 0.5% cholesterol + 0.2% vitamin E diet for 4 weeks. Semiquantitative RT-PCR analysis of thoracic aorta indicated that eNOS mRNA expression was significantly upregulated in cholesterol-fed rabbits compared to control rabbits. The supplementation of vitamin E significantly reduced eNOS expression as compared to the cholesterol-fed rabbits. The expression of iNOS mRNA was not significantly changed in cholesterol- and cholesterol + vitamin E-fed rabbits compared to control rabbits, respectively. β-VLDL isolated from vitamin E-fed rabbits resisted copper-induced oxidation, as assessed by thiobarbituric acid reactive substance (TBARS) assay. Moreover, western blotting of plasma for nitrotyrosine protein content was significantly enhanced in cholesterol-fed rabbits as compared to control rabbits, while vitamin E supplementation produced an insignificant lowering in the cholesterol-fed rabbits. These results suggest that eNOS may be, in part, regulated by oxidative stress and vitamin E may be detrimental under hypercholesterolemic conditions.