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Abstract

Initial predominance of the oxidative activity of type I 11 β-hydroxysteroid dehydrogenase in primary rat Leydig cells and transfected cell lines 
Ge,Renshan; Hardy,Matthew P.
Journal of Andrology 21(2): 303-310
Publication date: 2000


Glucocorticoids suppress testosterone production in Leydig cells. The levelof glucocorticoid action is set within the Leydig cell by the number ofglucocorticoid receptors and by the activity of 11beta-hydroxysteroiddehydrogenase (11betaHSD). This enzyme acts either as an oxidaseinactivating glucocorticoid or as a reductase amplifying its action. It iscurrently unknown whether extracellular conditions might cause 11betaHSDoxidative and reductive activities in Leydig cells to change inversely orindependently. The aim of the present study was to determine whetherextracellular conditions set in vitro by various culture time and mediacomponents, such as glucose and pyruvate, affect the relative rates of11betaHSD oxidation and reduction. Primary rat Leydig cells and cell lines(COS1 and CHOP cells) transfected with 11betaHSD-I complementary DNA(cDNA), were incubated with 25 nmol/L (physiologic range) or 500 nmol/L(stress range) concentrations of radiolabeled substrates, corticosterone or11-hydrocorticosterone, for 0 to 24 hours. Oxidative activity predominatedover reductive activity under initial conditions when product formationincreased linearly with time. For example, in Dulbecco's modified Eaglemedium/F12 medium (containing 5.5 mmol/L glucose), the peak ratio ofoxidation to reduction (with 1 denoting equivalence of oxidative andreductive activities) was 5.5 in rat Leydig cells, 19.7 in COS1 cells, and20.8 in CHOP cells. Glucose stimulated reductive activity but did notchange the predominant direction of 11betaHSD catalysis at earlier times.In COS1 cells transfected with 11betaHSD-I cDNA, oxidative activity rapidlyincreased during the first 2 hours of the incubation, then graduallydecreased while reductive activity increased steadily. The relative ratioof oxidation to reduction rapidly declined to less than 0.5 at 6 hours, andthus the favored direction of catalysis changed from oxidation toreduction. However, in transfected CHOP cells, 11betaHSD oxidative activityrapidly increased during the first 2 hours and continued to increase for 24hours. The ratio of oxidative to reductive activity rapidly declined butkept above 1 in CHOP cells for 24 hours, and the favored direction ofcatalysis remained predominantly oxidative. These results revealed that11betaHSD-I is a predominant oxidase initially in Leydig cells and 2 celllines, and that the oxidative activity is gradually lost over time. Thedata suggest that type I 11betaHSD is a predominant oxidase in Leydig cellsin vivo.