PROGESTERONE-RECEPTOR MESSENGER-RIBONUCLEIC-ACID AND PROTEIN IN LUTEINIZED GRANULOSA-CELLS OF RHESUS-MONKEYS ARE REGULATED IN-VITRO BY GONADOTROPINS AND STEROIDS

Previous studies in our laboratory indicated that the midcycle gonadot ropin surge stimulates progesterone receptor (PR) expression in granul osa cells of the macaque preovulatory follicle. The current experiment s were designed to determine whether gonadotropin or steroids continue to regulate PR in luteinized granulosa cells that contain these recep tors after the LH surge. Luteinizing granulosa cells obtained from gon adotropin-treated rhesus macaques were cultured in chemically defined medium in the presence of low density lipoprotein (LDL; 100 mu g/ml) w ith or without hCG (100 ng/ml) for up to 4 days. Cells were also cultu red with various concentrations (0.25-250 ng/ml) of the 3 beta-hydroxy steroid dehydrogenase (3 beta-HSD) inhibitor trilostane to reduce prog esterone (P) production in vitro. P and estradiol (E) in the media wer e assayed by RIA; PR mRNA was assessed by RNase protection assay, and cells expressing PR were identified by immunocytochemistry. Whereas hC G stimulated cellular P production through 4 days of culture, trilosta ne reduced hCG-stimulated P production in a dose-dependent fashion, wi th P levels decreasing more than 90% during incubation with 250 ng/ml trilostane (p < 0.05), When trilostane was removed from the media, P p roduction returned to hCG-stimulated levels, indicating that trilostan e action was reversible and nontoxic. Treatment with hCG increased (p < 0.05) PR mRNA levels in luteinized granulosa cells, whereas trilosta ne (250 ng/ml) alone did not alter levels compared to those in control s. Before culture, 68 +/- 11% of luteinizing granulosa cells expressed PR; intense nuclear staining was typically observed. After 2 days of culture, 78 +/- 3% of cells remained PR-positive, but nuclear staining was more heterogeneous. Incubation with hCG did not alter the percent age of luteinized granulosa cells staining positive for PR but increas ed the intensity of PR staining. Trilostane treatment (25 ng/ml) in co mbination with hCG significantly reduced the percentage of PR-positive cells (54 +/- 9%) when compared with hCG treatment (83 +/- 2%, p < 0. 05). These in vitro data suggest that macaque luteinized granulosa cel ls retain some PR expression in the absence of luteotropic hormones, b ut that gonadotropin stimulates PR mRNA levels and enhances PR express ion as assessed by intensity of nuclear PR staining. In the presence o f gonadotropin, trilostane effectively inhibited P production and redu ced the number of PR-positive cells, suggesting that P or a metabolite modulates PR expression in primate luteinized granulosa cells.