Comparative analysis of the effects of gonadotropin-releasing hormone agonist on the proliferative activity, apoptosis, and steroidogenesis in cultured porcine granulosa cells at varying stages of follicular growth

This study was conducted to analyze comparative effects of gonadropin-relea sing hormone (GnRH) agonist on the proliferation, apoptosis, and differenti ated function of cultured porcine granulosa cells from varying follicle sta ges. Comparative analyses of porcine granulosa cells from varying follicle stages to respond to GnRH agonist were performed in terms of proliferating cell nuclear antigen (PCNA) expression, occurrence of apoptosis, and 17 bet a-estradiol (E-2) and progesterone (P) secretion. PCNA expression was exami ned by the avidin/biotin immunoperoxidase method with a monoclonal antibody to PCNA, and apoptosis was assessed by in situ DNA 3'-end labeling method and DNA fragmentation analysis. E-2 and P were measured by radioimmunoassay s, The PCNA positive rate of granulosa cells cultured in the presence of Gn RH agonist (10(-9) M) was lower compared with that of cells cultured in the absence of GnRH agonist, However, the apoptosis positive rate was higher, and E-2 and P secretion by cultured granulosa cells was lower in the presen ce of GnRH agonist (10(-9) M) compared with that in the absence of GnRH ago nist, The inhibitory effect of GnRH agonist on PCNA positive rate of cultur ed cells was prominent in granulosa cells from small and medium but not fro m large follicles, By contrast, the inhibitory effect of GnRH agonist on E- 2 and P secretion by cultured cells was prominent in granulosa cells from l arge but not small and medium follicles. The stimulatory effect of GnRH ago nist on apoptosis positive rate of cultured cells was, however, uniform reg ardless of the stages of follicular growth. These results demonstrate that GnRH agonist exerts diverse actions on granulosa cells over the course of f ollicular growth, One downregulates granulosa proliferation in immature fol licles as well as steroidogenesis in mature follicles, and the other upregu lates apoptosis of granulosa cells regardless of the stages of follicular g rowth.