THE EFFECTS OF INSULIN ON 3-BETA-HYDROXYSTEROID DEHYDROGENASE EXPRESSION IN HUMAN LUTEINIZED GRANULOSA-CELLS

Objective: We determined the relative effects of insulin and FSH on pr ogesterone accumulation as well as activity, protein content, and mRNA expression of 3beta-hydroxysteroid dehydrogenase (3betaHSD) in human luteinized granulosa cells. Methods: Luteinized granulosa cells obtain ed from women undergoing in vitro fertilization were plated and grown to near confluence and treated with FSH, insulin, or a combination of insulin and FSH. Progesterone production as well as enzyme activity, p rotein content, and mRNA expression for 3betaHSD were evaluated. Resul ts: Progesterone production was not affected by insulin alone but incr eased threefold in the presence of FSH (50 ng/muL) alone. The presence of FSH plus insulin (100 nmol/L) caused a significant increase in pro gesterone accumulation greater than that of FSH alone. The already hig h basal levels of 3betaHSD activity were unaffected by insulin alone b ut increased 1.7-fold in the presence of FSH. The combination of FSH ( 50 ng/mL) and insulin (100 nmol/L) increased activity 1.3-fold over FS H alone (P < .02). Insulin (greater than 100 nmol/L) alone increased 3 betaHSD protein content as measured by Western analysis 1.8-2-fold ove r basal levels, whereas FSH alone increased protein content 2.8-fold, and was further augmented by the addition of insulin in a dose-related fashion up to 3.5-fold over basal levels. Insulin increased 3betaHSD mRNA twofold over basal levels; FSH alone increased mRNA expression of 3betaHSD 3.2-fold. In the presence of insulin plus FSH, 3betaHSD mRNA expression increased 7.6-fold over basal levels. For comparison, insu lin alos stimulated cytochrome P450 aromatase activity, P450 aromatase protein, and mRNA but to a greater degree than that seen for 3betaHSD . Conclusion: Insulin is a regulator of both 3betaHSD and aromatase ex pression in human granulosa cells. Elevated insulin levels could there fore affect steroid production in human granulosa cells and presumably alter the menstrual cycle and fertility.