Role of transforming growth factor-β1 in gene expression and activity of estradiol and progesterone-generating enzymes in FSH-stimulated bovine granulosa cells

Abstract

Survival and inhibitory factors regulate steroidogenesis and determine the fate of developing follicles. The objective of this study was to determine the role of transforming growth factor-β1 (TGFB1) in the regulation of estradiol-17β (E2) and progesterone (P4) secretion in FSH-stimulated bovine granulosa cells. Granulosa cells were obtained from 2 to 5 mm follicles and cultured in serum-free medium. FSH dose (1 and 10 ng/ml for 6 days) and time in culture (2, 4, and 6 days with 1 ng/ml FSH) increased E2secretion and mRNA expression of E2-related enzymes cytochrome P450 aromatase (CYP19A1) and 17β-hydroxysteroid dehydrogenase type 1 (HSD17B1), but notHSD17B7. TGFB1 in the presence of FSH (1 ng/ml) inhibited E2secretion, and decreased mRNA expression of FSH receptor(FSHR),CYP19A1, andHSD17B1, but notHSD17B7. FSH dose did not affect P4secretion and mRNA expression of 3β-hydroxysteroid dehydrogenase (HSD3B) and α-glutathioneS-transferase (GSTA), but inhibited the amount of steroidogenic acute regulatory protein(STAR)mRNA. Conversely, P4and mRNA expression ofSTAR, cytochrome P450 side-chain cleavage(CYP11A1),HSD3B, andGSTAincreased with time in culture. TGFB1 inhibited P4secretion and decreased mRNA expression ofSTAR,CYP11A1,HSD3B, andGSTA. TGFB1 modified the formation of granulosa cell clumps and reduced total cell protein. Finally, TGFB1 decreased conversion of androgens to E2, but did not decrease the conversion of estrone (E1) to E2and pregnenolone to P4. Overall, these results indicate that TGFB1 counteracts stimulation of E2and P4synthesis in granulosa cells by inhibiting key enzymes involved in the conversion of androgens to E2and cholesterol to P4without shutting down HSD17B reducing activity and HSD3B activity.