Estradiol increases rat aorta endothelium-derived relaxing factor (EDRF) activity without changes in endothelial NO synthase gene expression: possible role of decreased endothelium-derived superoxide anion production

Objectives: Estradiol is known to exert a protective effect against atheros clerosis, but the mechanism(s) whereby this protection is mediated is/are u nclear. However, estradiol-treated castrated animals exhibit increased acti vity of endothelium-derived relaxing factor (EDRF), which could contribute to vasculoprotection. In the present work, we investigated the molecular me chanism(s) of the enhancement of EDRF activity in the thoracic aorta of oop horectomized female rats given 17 beta-estradiol (E-2, 2 or 40 mu g/kg/day) compared to those given a placebo. Methods and Results: The abundance in t he thoracic aorta of NO synthase I, II and III mRNA (using RT-PCR) and of N O synthase I, II and III immunoreactive protein (using Western blotting) wa s unaltered by E-2. NO synthase activity (based on arginine/citrulline conv ersion) in thoracic aorta homogenates did not differ significantly among th e three groups, suggesting that NO production was not enhanced by E-2. In c ontrast, lucigenin-enhanced chemiluminescence of aorta from the E-2 group w as decreased compared to that of the placebo group. Desendothelialization a nd exogenously added superoxide dismutase suggested that this difference wa s due to a decrease in extracellular endothelium-derived production of supe roxide anion (O-2(-.)). Experiments in cultured bovine aortic endathelial c ells confirmed a decreased extracellular production of O-2(-.) In response to ethinylestradiol (1 nM) using both lucigenin-enhanced chemiluminescence and ESR spectroscopy. Luminol-enhanced chemiluminescence revealed that ethi nylestradiol-treated cultured endothelial cells generated less peroxynitrit e (the byproduct of NO. and O-2(-.) interaction) than control cells. Conclu sion: Estradiol increases rat aorta EDRF activity in the absence of changes in endothelial NO synthase gene expression. The decreased endothelium-deri ved generation of O-2(-.) in response to estrogens could account for enhanc ed EDRF-NO bioactivity and decreased peroxynitrite release. All of these ef fects could contribute to the vascular protective properties of estrogens. (C) 1999 Published by Elsevier Science B.V. All rights reserved.