REGULATION OF INHIBIN ALPHA-SUBUNIT AND BETA(A)-SUBUNIT MESSENGER-RIBONUCLEIC-ACID LEVELS BY CHORIONIC-GONADOTROPIN AND RECOMBINANT FOLLICLE-STIMULATING-HORMONE IN CULTURED HUMAN GRANULOSA-LUTEAL CELLS

Citation
M. Eramaa et al., REGULATION OF INHIBIN ALPHA-SUBUNIT AND BETA(A)-SUBUNIT MESSENGER-RIBONUCLEIC-ACID LEVELS BY CHORIONIC-GONADOTROPIN AND RECOMBINANT FOLLICLE-STIMULATING-HORMONE IN CULTURED HUMAN GRANULOSA-LUTEAL CELLS, The Journal of clinical endocrinology and metabolism, 79(6), 1994, pp. 1670-1677
Citations number
55
Categorie Soggetti
Endocrynology & Metabolism
ISSN journal
0021972X
Volume
79
Issue
6
Year of publication
1994
Pages
1670 - 1677
Database
ISI
SICI code
0021-972X(1994)79:6<1670:ROIAAB>2.0.ZU;2-H
Abstract
We studied the effects of recombinant human FSH (rhFSH) and purified h CG on the steady state messenger ribonucleic acid (mRNA) levels of inh ibin alpha- and beta(A)-subunits in cultured granulosa-luteal cells of preovulatory ovarian follicles obtained from women undergoing in vitr o fertilization. Specific mRNA transcripts for the alpha- and beta(A)- subunits were detected in Northern and dot blot filter hybridization a nalyses, and the levels of these mRNAs were induced by rhFSH and hCG i n a distinct concentration- and time-dependent manner. The basal and h CG-stimulated alpha-subunit mRNA levels were first determined at 2- to 3-day intervals over a 3- to 10-day culture period after the initiati on of the cultures. Both the basal and hCG-stimulated alpha-subunit mR NA levels declined steadily during culture, but the maximal relative s timulatory effect of hCG was observed on day 7 of culture. All subsequ ent experiments, therefore, were performed on days 6-8 of culture. Bot h gonadotropins induced alpha-subunit mRNA levels with slower kinetics than those of the beta(A)-subunit. Varying between experiments, rhFSH and hCG increased the expression of the alpha-subunit with a maximal effect of 2.5- to 5.7-fold and 1.7- to 7.2-fold, respectively, above b asal levels 24-48 h after stimulation. rhFSH and hCG induced beta(A)-s ubunit mRNA levels with 3.0- to 5.8-fold and 2.3- to 8.6-fold increase s above basal levels, respectively, at 2 h; thereafter, only moderate or no stimulation of the beta(A)-subunit mRNA levels could be detected at 7-48 h. Treatment of the cells with the RNA synthesis inhibitor ac tinomycin-D prevented the induction of alpha-subunit mRNA levels by hC G, and no significant differences were detected in the stability of al pha-subunit mRNA transcripts in hCG-treated cells vs. untreated cultur es. This indicates that hCG induces transcription of the alpha-subunit gene rather than maintains the levels of preexisting transcripts. As the kinetics of induction of alpha- and beta(A)-subunit mRNAs by gonad otropins were different, we examined how the inhibition of protein syn thesis affects the induction of alpha- and beta(A)-subunit mRNAs by hC G. Cycloheximide had no effect on basal alpha-subunit mRNA levels at 2 or 24 h. However, it inhibited at 24 h the induction of the alpha-sub unit by hCG. On the other hand, cycloheximide had no effect on basal o r hCG-stimulated beta(A)-subunit mRNA levels at 2 h, but at the 24 h p oint, it markedly induced both basal and hCG-stimulated beta(A)-subuni t mRNA levels. Thus, the slower induction of the alpha-subunit mRNA ma y require de novo synthesis of protein factors for the mediation of th e effect of hCG. By contrast, protein synthesis inhibition did not aff ect the rapid induction of beta(A)-subunit by hCG, but it may prevent the degradation of the beta(A)-subunit mRNA seen after the initial tra nsient induction of its levels. Taken together, our results suggest th at both gonadotropins are potent inducers of inhibin alpha- and beta(A )-subunit mRNAs in human granulosa-luteal cells and that the induction of the mRNAs of these two subunits is regulated through different mec hanisms.