Testosterone Stimulates Progesterone Production and mRNAs Expression of Steroidogenic Acute Regulatory Protein (StAR), P450 Cholesterol Side-Chain Cleavage (P450scc), and Luteinizing Hormone Receptor (LH-R) in Hen (Gallus domesticus) Granulosa Cells.

The chicken ovary is organized into a hierarchy of yellow yolky follicles that ovulate on successive days, and their steroid secretion is dependent on the developmental degree of the follicle. Testosterone has been recently described to have an important role in follicle ovulation. Active or passive immunization of laying hens against testosterone blocks ovulation without affecting follicle development. Further, testosterone stimulates progesterone production in a paracrine fashion within the ovary. We suggest that testosterone may play a role in preovulatory follicle maturation by stimulation of granulosa progesterone production, via the induction of granulosa cell StAR, P450scc and LH-R mRNAs expression in the hen F1 follicles. F1 granulosa cells were collected and cultured with testosterone (0, 10 and 100 ng/ml) for 3 h. Six cultures were performed with 8 replicates per treatment, with 100,000 viable cells per well in 250 μl of culture medium. Total RNA was extracted from cells pooled after 3h of culture, reverse transcription was performed and StAR, P450scc and LH-R mRNAs were quantified by real time PCR. Number of mRNA copies within the cells was estimated by regression analysis of CT values against a cDNA standard curve prepared for each gene. Progesterone production was measured in spent culture media by ELISA. Testosterone at 10 or 100 ng/ml significantly increased progesterone production and mRNA expression of StAR, P450scc and LH-R. The total increase of progesterone production was 34%, while for mRNA expression was 88.4% and 97.2% for StAR when 10 and 100 ng/ml of testosterone were added respectively, 99.5% and 150% for P450scc, and 142.1% and 184.4% for LH-R. In conclusion, testosterone stimulates granulosa cell progesterone production in hen pre-ovulatory hierarchical follicles due to stimulation of StAR, P450sccc and LH-R mRNA expression. We suggest that a stimulatory paracrine effect of thecal testosterone may play a role in increasing progesterone production by granulosa cells in the maturing pre-ovulatory follicle. Granulosa cell progesterone production in the F1 follicle may be enhanced by the stimulatory action of increasing circulating testosterone originating from less mature follicles in the hierarchy, to initiate the pre-ovulatory release of progesterone. This in turn stimulates the positive feedback action of progesterone on LH release to generate the respective pre-ovulatory surges. Rangel PL was a grantee of the International Foundation for Science (Stockholm, Sweden).

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