Effects of recombinant bovine somatotrophin, insulin-like growth factor-I and insulin on the proliferation of bovine granulosa cells in vitro

Abstract

ABSTRACT Treatment of heifers with recombinant bovine somatotrophin (BST) significantly increases the population of small ovarian follicles and peripheral concentrations of somatotrophin, insulin-like growth factor-I (IGF-I) and insulin. To investigate the possible mechanism(s) involved in the action of BST on ovarian follicles, the effects of BST, IGF-I and insulin, given alone or in combination with either FSH or LH, on the proliferation of bovine granulosa cells in vitro were examined using a serum-free culture system. Bovine granulosa cells were obtained from antral follicles classified into three size categories according to diameter: small <5 mm; medium-sized 5–10 mm and large >10 mm. The proliferation of granulosa cells was assessed by the incorporation of [3H]thymidine into the cultured cells. Both FSH and LH (1–1000 ng/ml) inhibited the proliferation of bovine granulosa cells obtained from all three size classes of follicles in a dose-dependent manner. BST, at doses ranging from 1 to 1000 ng/ml, had no effect on the proliferation of granulosa cells from small and medium-sized follicles, but inhibited the division of granulosa cells from large follicles in a dose-dependent manner. Treatment with either IGF-I (10–3000 ng/ml) or insulin (0·5–1000 ng/ml) stimulated, in a dose-dependent manner, the proliferation of granulosa cells obtained from all three size categories of follicles. No synergistic interaction between BST (30 ng/ml) and either FSH (50 ng/ml) or LH (5 ng/ml) was observed in granulosa cells from all three size classes of follicles. In contrast, physiological concentrations of both IGF-I (100 ng/ml) and insulin (1 ng/ml) acted in synergy with both FSH (50 ng/ml) and LH (5 ng/ml) to stimulate the proliferation of granulosa cells from small follicles, whilst no such synergistic interactions were observed in granulosa cells from medium-sized and large follicles. It was concluded that the increase in the number of small ovarian follicles induced by BST treatment in heifers may be mediated by increased peripheral concentrations of IGF-I and/or insulin, possibly acting in synergy with gonadotrophins. Furthermore, insulin probably acts through its own receptor rather than acting via the type-I IGF receptor, as it can stimulate the proliferation of bovine granulosa cells at physiological concentrations. Journal of Endocrinology (1993) 139, 67–75