Conversion of 5(10)-oestrene-3β,17β-diol to 19-nor-4-ene-3-ketosteroids by luteal cells in vitro: possible involvement of the 3β-hydroxysteroid dehydrogenase/isomerase

Abstract

ABSTRACT We have previously suggested that in porcine granulosa cells, a putative intermediate, 5(10)-oestrene-3,17-dione is involved in 4-oestrene-3,17-dione (19-norandrostenedione; 19-norA) and 4-oestren-17β-ol-3-one (19-nortestosterone: 19-norT) formation from C19 aromatizable androgens. In this study, luteal cells prepared from porcine, bovine and rat corpora lutea by centrifugal elutriation were used as a source of 3β-hydroxysteroid dehydrogenase/isomerase in order to investigate the role of this enzyme in the biosynthesis of 19-norsteroids. Small porcine luteal cells made mainly 19-norT and large porcine luteal cells 19-norA from 5(10)-oestrene-3β,17β-diol, the reduced product of the putative intermediate 5(10)-oestrene-3,17-dione. However, neither small nor large cells metabolized androstenedione to 19-norsteroids. Serum and serum plus LH significantly stimulated formation of both 19-norA and 19-norT from 5(10)-oestrene-3β,17β-diol, compared with controls. Inhibitors of the 3β-hydroxysteroid dehydrogenase/isomerase (trilostane and cyanoketone) significantly reduced formation of 19-norT in small porcine luteal cells and 19-norA in large porcine luteal cells, although they were effective at different concentrations in each cell type. In parallel incubations, formation of [4-14C]androstenedione from added [4-14C]dehydroepiandrosterone was also inhibited by cyanoketone in both small and large porcine luteal cells in a dose-dependent manner; however, trilostane (up to 100 μmol/l) did not inhibit androstenedione formation in large porcine luteal cells. In addition, the decrease in progesterone synthesis induced by trilostane and cyanoketone (100 μmol/l each) was accompanied by a parallel accumulation of pregnenolone in both cell types. These results suggest that 3β-hydroxysteroid dehydrogenase/isomerase, or a closely related enzyme, present in small and large porcine luteal cells can convert added 5(10)-3β-hydroxysteroids into 19-nor-4(5)-3-kestosteroids in vitro. In the porcine ovarian follicle, therefore, formation of 19-norA from androstenedione can be envisaged as a two-step enzymatic process: 19-demethylation of androstenedione to produce the putative intermediate 5(10)-oestrene-3,17-dione, and subsequent isomerization to 19-norA. In contrast to granulosa cells, porcine luteal cells synthesized 19-norA or 19-norT only when provided with the appropriate substrate. Unfractionated rat luteal cells also metabolized 5(10)-oestrene-3β,17β-diol to a mixture of 19-norA and 19-norT; conversion was inhibited by trilostane. In addition, small bovine luteal cells synthesized mainly 19-norT and formation was also inhibited by trilostane and cyanoketone. In addition to 19-norA, an unknown metabolite, formed in low amounts by large porcine luteal cells, appears to be related to another steroid which accumulated at high inhibitor concentrations; it may represent 5(10)-oestrene-3,17-dione postulated as a putative intermediate formed during 19-norsteroid biosynthesis. Journal of Endocrinology (1991) 129, 233–243