Activity of 3β-hydroxysteroid dehydrogenase associated with progesterone production in bovine granulosa cells cultured under different concentrations of serum, insulin-like growth factor I, and gonadotropin

Abstract

Three-b-hydroxysteroid dehydrogenase (3b-HSD) is the enzyme responsible for progesterone production. This study aimed to determine whether 3b-HSD activity can be shown to reflect progesterone production by granulosa cells cultured under different serum conditions and follicle stimulating hormone (FSH), luteinising hormone (LH), and insulin-like growth factor I (IGF-I) concentrations. Large bovine follicles were dissected from abattoir ovaries to recover granulosa cells. Cells were washed, stained for viability, and plated for 48 h in basic medium with or without 5% foetal calf serum (FCS). Subsequently, cells were exposed to FSH (1 ng/mL), LH (10 or 100 ng/mL), or FSH (1 ng/mL) + IGF-I (1 or 10 ng/mL) in a serum-free medium for another 96 h to predict degree of luteinisation. Before and after incubation, granulosa cells were stained for 3b-HSD activity. The high dose of IGF-I (10 ng/mL) increased (P < 0.05) progesterone secretion over 2.5-fold compared with FSH alone or the low dose of IGF-I (1 ng/mL) in cells preincubated with FCS. This was clearly reflected by darker 3b-HSD staining than in cells exposed to FSH and low dose IGF-I.