title: The effect of 17 beta-estradiol on intracellular calcium homeostasis in human endothelial cells.
abstract: The cardiovascular effects of estrogen are mediated in part by augmenting the function of endothelial nitric oxide synthase. Endothelial nitric oxide synthase activity is dependent on many cofactors including Ca(2+). Hence, we investigated the effect of chronic 17 beta-estradiol treatment on the intracellular Ca(2+) concentration and endothelial nitric oxide synthase protein expression in the human endothelial cell line, EA.hy926, using spectrofluorometry and Western blot, respectively. Inhibiting the sarco(endo)plasmic reticulum Ca(2+) ATPase with thapsigargin caused an increase in the intracellular Ca(2+) concentration, which was higher in chronically 17 beta-estradiol-treated (1muM, 24h) cells loaded with Fura-2-acetoxymethyl ester compared to vehicle-treated cells, suggesting a higher endoplasmic reticulum Ca(2+) content in 17 beta-estradiol-treated cells. An enhanced Ca(2+) influx pathway in chronically 17 beta-estradiol-treated cells was also observed. In addition, 17 beta-estradiol-treated cells expressed higher levels of endothelial nitric oxide synthase protein in comparison to vehicle-treated cells. The chronic effect of 17 beta-estradiol on Ca(2+) homeostasis and endothelial nitric oxide synthase expression was attenuated with the nonselective estrogen receptor inhibitor, ICI 182,780 (10muM, 7alpha, 17beta-[9-[(4,4,5,5,5-Pentafluoropentyl)sulfinyl]nonyl] estra-1,3,5(10)-triene-3,17-diol). Furthermore, analysis of the thapsigargin-evoked Ca(2+) response in chronically 17 beta-estradiol-treated estrogen receptor alpha-knockdown cells showed no significant difference in Ca(2+) response compared to vehicle-treated estrogen receptor alpha-knockdown cells, indicating that the regulation of Ca(2+) homeostasis by 17 beta-estradiol is mediated through an estrogen receptor alpha-dependent pathway. These data revealed an estrogen receptor alpha-dependent modulation of Ca(2+) homeostasis accompanying the enhancement of endothelial nitric oxide synthase expression in 17 beta-estradiol-treated human endothelial cells.
authors.affiliation: Department of Physiology & Pharmacology, Thomas J. Long School of Pharmacy and Health Sciences, 751 Brookside Rd, University of the Pacific, Stockton, CA 95211, USA.
authors.forename: Der
authors.initials: D
authors.lastname: Thor
authors.forename: James A
authors.initials: JA
authors.lastname: Uchizono
authors.forename: Geoff P
authors.initials: GP
authors.lastname: Lin-Cereghino
authors.forename: Roshanak
authors.initials: R
authors.lastname: Rahimian
mesh_terms: D002118:Calcium / Q000378:metabolism*; D002460:Cell Line; D042783:Endothelial Cells / Q000187:drug effects*; D004958:Estradiol / Q000494:pharmacology*; D004967:Estrogens / Q000494:pharmacology*; D006706:Homeostasis / Q000187:drug effects*; D006801:Humans
chemical_list: D004967:Estrogens; D004958:Estradiol; D002118:Calcium