17Beta-eEstradiol stimulates arachidonate release from human amnion-like WISH cells through a rapid mechanism involving a membrane receptor.
17beta-Estradiol (17beta-E(2)) greatly and dose-dependently stimulates [(3)H]arachidonic acid (AA) release from the human amnion-like Wistar Institute Susan Hayflick (WISH) cells. This action is abolished by the phospholipase A(2) inhibitor AACOCF(3), significantly reduced by the estrogen receptor (ER) antagonist ICI 182,780, and uninfluenced by cycloheximide. The estradiol-BSA conjugate E(2)coBSA, which binds putative membrane ERs and is unable to enter the cell, also highly stimulates [(3)H]AA release from WISH cells, although to a lesser extent compared with 17beta-E(2). The fluorescent conjugate E(2)coBSA-FITC specifically binds to the surface of a subset of intact WISH cells, and labeling intensity appears dose and time dependent. Cell permeabilization results in a dense intracellular staining, mainly in the peripheral cytoplasm. H-150, an antibody against the N terminus of human ERbeta, also labels the plasma membrane of intact WISH cells and the cytoplasm of permeabilized cells. Almost no labeling is observed using ER-21, an antibody against the N terminus of human ERalpha. RT-PCR evidences the presence of mRNA for ERbeta, not for ERalpha. Our data suggest that 17beta-E(2) stimulates [(3)H]AA release from WISH cells through an apparently nongenomic pathway and interaction with membrane binding sites. These last are, at least in part, similar if not identical to ERbeta.
Fiorini
Sara
S
Department of Biology, University of Ferrara, 44100-I Ferrara, Italy.
Ferretti
Maria E
ME
Biondi
Carla
C
Pavan
Barbara
B
Lunghi
Laura
L
Paganetto
Guglielmo
G
Abelli
Luigi
L
D000650:Amnion / Q000187:drug effects* / Q000378:metabolism*; D016718:Arachidonic Acid / Q000378:metabolism*; D001095:Arachidonic Acids / Q000494:pharmacology; D002460:Cell Line; D002462:Cell Membrane / Q000737:chemistry*; D002463:Cell Membrane Permeability; D015232:Dinoprostone / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004958:Estradiol / Q000031:analogs & derivatives* / Q000494:pharmacology*; D047628:Estrogen Receptor alpha; D047629:Estrogen Receptor beta; D005260:Female; D016650:Fluorescein-5-isothiocyanate; D005456:Fluorescent Dyes; D000077267:Fulvestrant; D006801:Humans; D010741:Phospholipases A / Q000037:antagonists & inhibitors; D011247:Pregnancy; D012333:RNA, Messenger / Q000032:analysis; D011960:Receptors, Estrogen / Q000037:antagonists & inhibitors / Q000235:genetics / Q000502:physiology*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012710:Serum Albumin, Bovine / Q000494:pharmacology; D014316:Tritium
D001095:Arachidonic Acids; D004791:Enzyme Inhibitors; D047628:Estrogen Receptor alpha; D047629:Estrogen Receptor beta; D005456:Fluorescent Dyes; D012333:RNA, Messenger; D011960:Receptors, Estrogen; C081565:arachidonyltrifluoromethane; D014316:Tritium; D000077267:Fulvestrant; D012710:Serum Albumin, Bovine; D016718:Arachidonic Acid; D004958:Estradiol; D010741:Phospholipases A; D016650:Fluorescein-5-isothiocyanate; D015232:Dinoprostone