Regulation of 11 beta-hydroxysteroid dehydrogenase type 1 gene expression by LH and interleukin-1 beta in cultured rat granulosa cells

Granulosa cells from preovulatory follicles show increased expression of 11 beta-hydroxysteroid dehydrogenase type 1 (11 beta HSD1) at the time of ovu lation. As ovulation may be an inflammatory process, this may be a mechanis m of local enhancement of the activity of anti-inflammatory glucocorticoids . In this study, we examined direct effects of LH, the proinflammatory cyto kine, interleukin-1 beta (IL-1 beta), and pharmacological activators of pro tein kinase A (PKA) (forskolin and dibutyryl (db) cAMP) and PKC (LH-releasi ng hormone and phorbol 12-myristate 13-acetate (PMA)) signalling on the exp ression of 11 beta HSD1 mRNA in vitro. Granulosa cells from immature female rat ovaries were cultured (pretreatment) in serum-free medium 199 containi ng recombinant human (rh) FSH (1 ng/ml) for 48 h to induce responsiveness t o LH. Cell monolayers were then washed and cultured (test treatment) for a further 12 h in the presence of rhLH (0-100 ng/ml), IL-1 beta (0-50 ng/ml), or both. Total RNA was extracted from granulosa cell monolayers and taken for quantitative ribonuclease protection analysis of 11 beta HSD1 mRNA. The low level of 11 beta HSD1 mRNA detectable in un-stimulated (control) cultu res was increased approximately twofold by the 48-h pretreatment with rhFSH , Subsequent exposure to rhLH (1-100 ng/ml) for a further 12 h dose-depende ntly increased 11 beta HSD1 mRNA expression by an additional two- to threef old. Forskolin (10 mu M), db-cAMP (2 mM), LH-releasing hormone (LHRH; 1 mu M) and PMA (200 nM) were also stimulatory. IL-1 beta (0.05-50 ng/ml) stimul ated 11 beta HSD1 mRNA expression in a dose-related manner, both in the: ab sence and in the presence of rhLH (3 ng/ml). The interaction between IL-1 b eta (5 ng/ml) and rhLH (3 ng/ml) was additive. Co-treatment with a 50-fold excess of IL-1 receptor antagonist fully reversed the action of IL-1 beta. We conclude that 11 beta HSD1 mRNA expression in functionally mature granul osa cells is directly stimulated by gonadotrophins and IL-1 beta in vitro, potentially involving post-receptor signalling via PKA- and PKC-mediated pa thways. Thus both LH and IL-1 beta may serve physiological roles in the upr egulation of 11 beta HSD1 gene expression by granulosa cells in ovulatory f ollicles.