Vitamin E binds to specific binding sites and enhances prostacyclin production by cultured aortic endothelial cells

We evaluated the effect of d-α-tocopherol (vitamin E) on the production of prostacyclin (PGI₂) by cultured bovine aortic endothelial cells. Vitamin E at physiological doses significantly enhanced the production of PGI₂ by aortic endothelial cells when added to the culture simultaneously with histamine, the Ca²⁺ ionophore A23187 (A23187), plasma-derived serum (PDS), or arachidonic acid. This effect was found to occur in a time- and dose-dependent manner, and the maximal enhancement was produced by 9.28μM of vitamin E for 1h incubations. Significantly lower amounts of lipid peroxides were measured in endothelial cells stimulated by 10% PDS with 9.28μM of vitamin E than in those stimulated without vitamin E for over 24h, although the stimulation during the initial 1 to 12h period did not have a significant effect on lipid peroxide formation in cultured aortic endothelial cells. We also demonstrated that bovine aortic endothelial cells have specific binding sites for [³H]vitamin E that exhibited time- and temperature-dependent saturability. At 4°C, the nonspecific binding was 8-12% of the total binding, and the specific binding reached equilibrium by 2h. Specific binding increased with the concentration of [³H]vitamin E and became saturated at concentrations between 1.5μM and 2.0μM per 2.0x10⁵ cells. Raising the unlabeled vitamin E concentration from 97.7nM to 1,000μM reduced the specific binding of 2.0μM [³H]vitamin E. The Scatchard plot of [³H]vitamin E binding to the endothelial cells shows two classes of binding sites: one with a high affinity (K(a1) 2.48±0.32x10⁷M^-1, n=6) and a low capacity (n₁ 1.20±0.34x10⁷ sites/cell) and the other with a low affinity (K(a2) 1.18±0.32x10⁵M^-1) and a high capacity (n₂ 3.39±0.53x10⁹ sites/cell). Our results suggest that the endothelial cells binding sites for vitamin E may play some roles in vascular homeostasis in vivo, and that vitamin E may prevent the development of atherosclerotic changes due in part to the enhancement of PGI₂ production by the vascular wall and its action as an antioxidant in vascular endothelial cell.