title: 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.
abstract: OBJECTIVE
  Estradiol is known to exert a protective effect against atherosclerosis, but the mechanism(s) whereby this protection is mediated is/are unclear. However, estradiol-treated castrated animals exhibit increased activity of endothelium-derived relaxing factor (EDRF), which could contribute to vasculoprotection. In the present work, we investigated the molecular mechanism(s) of the enhancement of EDRF activity in the thoracic aorta of oophorectomized female rats given 17 beta-estradiol (E2, 2 or 40 micrograms/kg/day) compared to those given a placebo.
  
  
  RESULTS
  The abundance in the thoracic aorta of NO synthase I, II and III mRNA (using RT-PCR) and of NO synthase I, II and III immunoreactive protein (using Western blotting) was unaltered by E2. NO synthase activity (based on arginine/citrulline conversion) in thoracic aorta homogenates did not differ significantly among the three groups, suggesting that NO production was not enhanced by E2. In contrast, lucigenin-enhanced chemiluminescence of aorta from the E2 group was decreased compared to that of the placebo group. Desendothelialization and exogenously added superoxide dismutase suggested that this difference was due to a decrease in extracellular endothelium-derived production of superoxide anion (O2-.). Experiments in cultured bovine aortic endothelial cells confirmed a decreased extracellular production of O2-. in response to ethinylestradiol (1 nM) using both lucigenin-enhanced chemiluminescence and ESR spectroscopy. Luminol-enhanced chemiluminescence revealed that ethinylestradioltreated cultured endothelial cells generated less peroxynitrite (the byproduct of NO-. and O2-. interaction) than control cells.
  
  
  CONCLUSIONS
  Estradiol increases rat aorta EDRF activity in the absence of changes in endothelial NO synthase gene expression. The decreased endothelium-derived generation of O2-. in response to estrogens could account for enhanced EDRF-NO bioactivity and decreased peroxynitrite release. All of these effects could contribute to the vascular protective properties of estrogens.
authors.affiliation: INSERM U397 et Laboratoire de Physiologie, Institut Louis Bugnard, CHU Rangueil, Toulouse, France.
authors.forename: M A
authors.initials: MA
authors.lastname: Barbacanne
authors.forename: J
authors.initials: J
authors.lastname: Rami
authors.forename: J B
authors.initials: JB
authors.lastname: Michel
authors.forename: J P
authors.initials: JP
authors.lastname: Souchard
authors.forename: M
authors.initials: M
authors.lastname: Philippe
authors.forename: J P
authors.initials: JP
authors.lastname: Besombes
authors.forename: F
authors.initials: F
authors.lastname: Bayard
authors.forename: J F
authors.initials: JF
authors.lastname: Arnal
mesh_terms: D000818:Animals; D001013:Aorta, Thoracic; D015153:Blotting, Western; D004730:Endothelium, Vascular / Q000187:drug effects / Q000378:metabolism*; D004958:Estradiol / Q000494:pharmacology*; D005260:Female; D066298:In Vitro Techniques; D008163:Luminescent Measurements; D009566:Nitrates / Q000378:metabolism; D009569:Nitric Oxide / Q000378:metabolism*; D019001:Nitric Oxide Synthase / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000032:analysis; D051381:Rats; D017207:Rats, Sprague-Dawley; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013481:Superoxides / Q000378:metabolism*
chemical_list: D009566:Nitrates; D012333:RNA, Messenger; D013481:Superoxides; C039642:peroxynitric acid; D009569:Nitric Oxide; D004958:Estradiol; D019001:Nitric Oxide Synthase