The effects of reduced o 2 and antioxidants on steroidogenic capacity of cultured rat leydig cells

Abstract

We have examined the effects of reduced O 2 tension and the antioxidant dimethylsulfoxide (DMSO) to determine if O 2-derived free radicals are the cause of decreased steroidogenic capacity (testosterone and progesterone production) of cultured rat Leydig cells. Rat Leydig cells were initially cultured under standard conditions of 5% CO 2, 95% air (19% O 2) with or without DMSO. Addition of DMSO resulted in increased basal testosterone production on days 2, 3 and 4 of culture. hCG (10 mIU)-stimulated testosterone secretion was 2–3 times greater on days 2 and 3 in the presence of DMSO. Lowering the O 2 concentration to 5% in the presence of DMSO resulted in even greater hCG-stimulated testosterone production on days 1 to 3. However, the effect of DMSO or low O 2 and DMSO were not seen after 5 days. The reduced O 2 concentration resulted in an increase in hCG (10 mIU)-stimulated progesterone synthesis throughout the culture, particularly on days 4 to 8. Also, when total steroid (progesterone and testosterone) was determined, cells cultured under reduced O 2 conditions responded with increased steroid production on days 1 to 8 in comparison to controls (19% O 2). These results demonstrate that lowered O 2 concentration and DMSO provide a protective effect resulting in the maintenance of testosterone production and an increase in progesterone synthesis. These findings suggest that free radical-mediated damage of enzymes may result in decreased steroidogenic capacity of cultured Leydig cells.