Progesterone receptor messenger ribonucleic acid and protein in luteinized granulosa cells of rhesus monkeys are regulated in vitro by gonadotropins and steroids

Diane M. Duffy, Theodore A. Molskness, Richard L. Stouffer

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    34 Scopus citations

    Abstract

    Previous studies in our laboratory indicated that the midcycle gonadotropin surge stimulates progesterone receptor (PR) expression in granulosa cells of the macaque preovulatory follicle. The current experiments were designed to determine whether gonadotropin or steroids continue to regulate PR in luteinized granulosa cells that contain these receptors after the LH surge. Luteinizing granulosa cells obtained from gonadotropin-treated rhesus macaques were cultured in chemically defined medium in the presence of low density lipoprotein (LDL; 100 μg/ml) with or without hCG (100 ng/ml) for up to 4 days. Cells were also cultured with various concentrations (0.25-250 ng/ml) of the 3β-hydroxysteroid dehydrogenase (3β-HSD) inhibitor trilostane to reduce progesterone (P) production in vitro. P and estradiol (E) in the media were assayed by RIA; PR mRNA was assessed by RNase protection assay, and cells expressing PR were identified by immunocytochemistry. Whereas hCG stimulated cellular P production through 4 days of culture, trilostane reduced hCG-stimulated P production in a dose-dependent fashion, with P levels decreasing more than 90% during incubation with 250 ng/ml trilostane (p < 0.05). When trilostane was removed from the media, P production returned to hCG-stimulated levels, indicating that trilostane action was reversible and nontoxic. Treatment with hCG increased (p < 0.05) PR mRNA levels in luteinized granulosa cells, whereas trilostane (250 ng/ml) alone did not alter levels compared to those in controls. 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 percentage of luteinized granulosa cells staining positive for PR but increased the intensity of PR staining. Trilostane treatment (25 ng/ml) in combination 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 cells retain some PR expression in the absence of luteotropic hormones, but that gonadotropin stimulates PR mRNA levels and enhances PR expression as assessed by intensity of nuclear PR staining. In the presence of gonadotropin, trilostane effectively inhibited P production and reduced the number of PR- positive cells, suggesting that P or a metabolite modulates PR expression in primate luteinized granulosa cells.

    Original languageEnglish (US)
    Pages (from-to)888-895
    Number of pages8
    JournalBiology of reproduction
    Volume54
    Issue number4
    DOIs
    StatePublished - Apr 1996

    ASJC Scopus subject areas

    • Reproductive Medicine
    • Cell Biology

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