L-4F, an apolipoprotein A-1 mimetic, restores nitric oxide and superoxide anion balance in low-density lipoprotein-treated endothelial cells

Background-Low-density lipoprotein (LDL) impairs endothelial cell function by uncoupling endothelial nitric oxide synthase (eNOS) activity, which allows superoxide anion (O-2(.-)) to be generated rather than nitric oxide (.NO). Recent reports indicate that apolipoprotein (apo) A-1 mimetics inhibit the development of atherosclerotic lesions in LDL receptor-null mice. Here we hypothesize that L-4F, an apoA-1 mimetic that inhibits atherosclerosis induced by hypercholesterolemia, protects endothelial cell function by preventing LDL from uncoupling eNOS activity. Methods and Results-Bovine aortic endothelial cells were incubated with LDL +/- L-4F, and changes in A23187-stimulated .NO and O-2(.-) generation were determined by ozone chemiluminescence and superoxide dismutase inhibitable ferricytochrome c reduction, respectively. Western analysis of eNOS immunoprecipitates was used to determine effects of LDL and L-4F on heat shock protein 90 (hsp90) interactions with eNOS. LDL decreased .NO production and increased eNOS-dependent O-2(.-) generation. Pretreatment of LDL with L-4F increased .NO and decreased O-2(.-) generation. By itself, L-4F had no effect on O-2(.-) but did increase .NO generation. Stimulation of endothelial cells incubated with LDL decreased the association of hsp90 with eNOS. Pretreatment of LDL with L-4F prevented a decrease in hsp90 association with eNOS and often enhanced association on stimulation. Conclusions-These data demonstrate that L-4F protects endothelial cell function by preventing LDL from uncoupling eNOS activity. L-4F allows endothelial cell to maintain coupled eNOS activity to generate .NO even in the face of atherogenic concentrations of LDL.