Inhibition of steroidogenesis in rat adrenal cells by 18-ethynyldeoxycorticosterone: evidence for an alternative pathway of aldosterone biosynthesis.
The effect of the mechanism-based inhibitor 18-ethynyldeoxycorticosterone (18-E-DOC) on the late steps of the aldosterone biosynthetic pathway was examined in freshly isolated cells of the zona glomerulosa (ZG) and fasciculata (ZF) from rat adrenal glands. ZG synthesis of aldosterone was inhibited by 18-E-DOC in a time- and concentration-dependent manner with a Ki of approximately 0.05 microM. The maximal degree of inhibition of ZG production of aldosterone and 18-hydroxycorticosterone (18-OH-B) was approximately 80%. ZF cells, perhaps surprisingly, were found to secrete 18-OH-B at levels approximately one-third to one-fourth those of ZG cells and the Ki of 18-E-DOC inhibition of 18-OH-B secretion was approximately 10 microM for ZF cells, 200-fold higher than for ZG cells. The inhibitor had no effect on the secretion of corticosterone by either ZG or ZF, and the secretion of 18-hydroxydeoxycorticosterone (18-OH-DOC) by both the ZG and ZF was inhibited only to a minor degree. 18-E-DOC inhibited the biosynthesis of aldosterone by ZG cells incubated with 10 microM added DOC or 18-OH-DOC by approximately 75%, similar to the degree of inhibition of aldosterone biosynthesis from endogenous substrate, whereas ZF biosynthesis of 18-OH-B from either substrate was inhibited by less than 40%. ZF cells do not express aldosterone synthase, the only enzyme known to convert 18-OH-DOC into 18-OH-B. Incubation of MA-10 cells stably transfected with the cDNA of the rat aldosterone synthase with 18-E-DOC resulted in a complete inhibition of the conversion of DOC to aldosterone with a Ki of approximately 0.02 microM. In addition, transfected cells expressing 11beta-hydroxylase convert DOC to 18-OH-B in very small quantities only and cannot convert 18-OH-DOC to 18-OH-B. These data suggest that neither 11beta-hydroxylase nor aldosterone synthase are responsible for the biosynthesis of 18-OH-B by ZF cells from DOC or 18-OH-DOC, that 20% of aldosterone synthesis appears not to be attributable to the actions of aldosterone synthase and that an unknown CYP11B enzyme is also involved in the biosynthesis of 18-OH-B.
Gomez-Sanchez
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CE
Harry S. Truman Memorial Veterans Hospital, Department of Internal Medicine, College of Medicine, University of Missouri-Columbia, 65201, U.S.A. intmdceg@showme.missouri.edu
Gomez-Sanchez
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EP
Foecking
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MF
Zhou
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MY
D000311:Adrenal Glands / Q000378:metabolism*; D000450:Aldosterone / Q000096:biosynthesis*; D000818:Animals; D002478:Cells, Cultured; D003900:Desoxycorticosterone / Q000031:analogs & derivatives* / Q000494:pharmacology; D051381:Rats; D017207:Rats, Sprague-Dawley
D003900:Desoxycorticosterone; D000450:Aldosterone; C071418:18-ethynyldeoxycorticosterone