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Intracellular regulation of estradiol and progesterone production by cultured bovine granulosa cells
Author(s) -
Legault Sylvie,
Bailey Janice L.,
Fortier Michel A.,
Rouillier Paul,
Guilbault Louis A.
Publication year - 1999
Publication title -
molecular reproduction and development
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.745
H-Index - 105
eISSN - 1098-2795
pISSN - 1040-452X
DOI - 10.1002/(sici)1098-2795(199912)54:4<371::aid-mrd7>3.0.co;2-z
Subject(s) - forskolin , protein kinase c , endocrinology , biology , medicine , genistein , tyrosine kinase , phorbol , activator (genetics) , adenylate kinase , granulosa cell , protein kinase a , intracellular , signal transduction , stimulation , receptor , kinase , ovary , microbiology and biotechnology , biochemistry
The objective of the present study was to investigate the implication of protein kinase A (PKA), protein kinase C (PKC), and receptor protein tyrosine kinase (R‐PTK) pathways in the regulation of estradiol (E2) and progesterone (P4) production by bovine granulosa cells. Cells were harvested from bovine follicles (8–15 mm diameter) and cultured without serum for an initial 3 days (37°C; 5% CO 2 in air; D1–D3). On the fourth day of culture (D4), E2 and P4 production were stimulated with FSH (1–6 ng/ml) or forskolin (FSK) in the presence or absence of intracellular effectors of PKA, PKC, and R‐PTK. Culture medium was collected and replaced each day. Stimulation of granulosa cell adenylate cyclase activity with FSK (0.06–3.75 μM) mimicked FSH, inducing a quadratic increase ( P < 0.001) of E2 production and a continuous elevation of P4 ( P < 0.01). Inhibition of R‐PTK activity with genistein (25–50 μM) increased the sensitivity of cells to FSH as demonstrated by a leftward shift in the dose response curve ( P < 0.001). Treatment with transforming growth factor‐α (TGFα; 0.1 ng/ml) abolished the FSH‐induced E2 production ( P < 0.001) and this effect was not reversed ( P < 0.001) by FSK or by genistein. Furthermore, the inhibitory effect of TGFα on FSH‐induced E2 production was reproduced by phorbol 12‐myristate 13‐acetate (PMA; 1.25–2.5 μM), a PKC activator ( P < 0.001). Interestingly, genistein inhibited P4 production ( P < 0.05). From these results, we conclude that E2 production by bovine granulosa cells is mediated by intracellular factors and can be stimulated downstream from the FSH receptor. The results also suggest that stimulation of R‐PTK and/or PKC activities, as probably occurs with TGFα, negatively affects the PKA pathway, thus decreasing E2 production. Furthermore, inhibition of R‐PTK leads to an increase production of E2 and may limit luteinization of bovine granulosa cells. Mol. Reprod. Dev. 54:371–378, 1999. © 1999 Wiley‐Liss, Inc.