title: Isoforms of 11beta-hydroxysteroid dehydrogenase in human granulosa-lutein cells.
abstract: To date, two isoforms of 11beta-hydroxysteroid dehydrogenase (11betaHSD) have been characterized: a low affinity, NADP+-dependent isoform (11betaHSD1) and a high affinity, NAD+-dependent isoform which metabolizes dexamethasone and is inhibited by cortisone (11betaHSD2). Having previously reported a relationship between ovarian 11betaHSD activities and conception in women undergoing in vitro fertilization (IVF-ET), the objective of the present study was to identify which isoforms of 11betaHSD metabolize glucocorticoids in cultures of human granulosa-lutein cells. In both intact cells and cell homogenates, two distinct 11betaHSD activities were identified with differing affinities for cortisol (Km = 490 nM and 2.6 microM). Even at low concentrations, cortisol oxidation was preferentially supported by NADP+ and was independent of NAD+. Although inhibited by the hemisuccinate ester of glycyrrhetinic acid, carbenoxolone, the predominant 11betaHSD activity in intact cells was resistant to end-product inhibition. Intact cells were also able to reduce [3H]cortisone (Km = 190 nM) but did not metabolize [3H]dexamethasone. 11BetaHSD1 mRNA was expressed in 23 of 28 cell cultures whereas 11betaHSD2 mRNA was not expressed in any of the 22 independent cultures studied by reverse transcriptase-polymerase chain reaction (RT-PCR). We conclude that human granulosa-lutein cells express both type 11betaHSD and a novel isoform of this enzyme. While the low affinity 11beta-dehydrogenase and 11-ketosteroid reductase activities exhibit properties consistent with 11betaHSD1, the high affinity 11beta-dehydrogenase differs from 11betaHSD2 in that it is NADP+-dependent, does not metabolize dexamethasone and is resistant to end-product inhibition.
authors.affiliation: Department of Biochemistry and Molecular Biology, Royal Free Hospital School of Medicine, University of London, UK.
authors.forename: A E
authors.initials: AE
authors.lastname: Michael
authors.forename: M
authors.initials: M
authors.lastname: Evagelatou
authors.forename: D P
authors.initials: DP
authors.lastname: Norgate
authors.forename: R J
authors.initials: RJ
authors.lastname: Clarke
authors.forename: J W
authors.initials: JW
authors.lastname: Antoniw
authors.forename: B A
authors.initials: BA
authors.lastname: Stedman
authors.forename: A
authors.initials: A
authors.lastname: Brennan
authors.forename: R
authors.initials: R
authors.lastname: Welsby
authors.forename: I
authors.initials: I
authors.lastname: Bujalska
authors.forename: P M
authors.initials: PM
authors.lastname: Stewart
authors.forename: B A
authors.initials: BA
authors.lastname: Cooke
mesh_terms: D042842:11-beta-Hydroxysteroid Dehydrogenases; D002478:Cells, Cultured; D005260:Female; D005938:Glucocorticoids / Q000378:metabolism*; D006107:Granulosa Cells / Q000201:enzymology*; D006801:Humans; D006913:Hydroxysteroid Dehydrogenases / Q000032:analysis* / Q000378:metabolism; D007527:Isoenzymes / Q000032:analysis* / Q000378:metabolism; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000032:analysis
chemical_list: D005938:Glucocorticoids; D007527:Isoenzymes; D012333:RNA, Messenger; D006913:Hydroxysteroid Dehydrogenases; D042842:11-beta-Hydroxysteroid Dehydrogenases