The effects of different culture media, glucose, pyridine nucleotides and adenosine on the activity of 11β-hydroxysteroid dehydrogenase in rat Leydig cells

11 beta-Hydroxysteroid dehydrogenase (11 beta HSD) reversibly converts glucocorticoids into inert 11-ketosteroids. The direction of the reaction has been found to vary with the cell type and sub-cellular reparation used. We have investigated if the directionality of 11 beta HSD can be influenced by the nature of the culture medium and compounds added during incubation of rat testis Leydig cells.:We found that when the cells were cultured in Dulbecco's Modified Eagle Medium (DMEM) that the dehydrogenase (11 beta DH) activity was higher than the reductase (11KSR) activity (11 beta DH:11KSR ratio similar to 2:1). When glucose was omitted from the DMEM a higher 11 beta DH:11KSR ratio (similar to 33:1) was obtained. However, when the cells were cultured in a combination of DMEM/Ham's F12 (1:1, v/v), a ninefold increase in 11KSR activity was obtained whereas 11 beta DH activity was inhibited by 64% compared with cells incubated in DMEM alone. Consequently, the predominant activity changed from a dehydrogenase to a reductase (11 beta DH: 11KSR ratio 1:15). Addition of the individual components of the Ham's F12 medium to DMEM showed that only pyruvate and/or the amino acids were able to mimic the effects of DMEM/Ham's F12. Similar differential effects were found when NAD(+), NADH or adenosine were added to the Leydig cells incubated in DMEM (three to fivefold increases and 20-50% decreases in 11KSR and 11 beta DH activities, respectively). In contrast, NADP(+) was found to increase 11 beta DH activity (up to threefold) but NADPH had no effect on 11KSR activity. Cells incubated with DMEM/Ham's F12, NAD(+), NADP(+) and adenosine were found to have higher ATP levels (four to sixfold) than those incubated in DMEM alone. These results illustrate that the relative 11 beta DH and 11KSR activities of 11 beta HSD in Leydig cells are markedly and differentially altered by the nature of the incubation medium and compounds added. (C) 1999 Elsevier Science Ireland Ltd. All rights reserved.