Production of insulin-like growth factor type I (IGF-I) by bovine ovarian cells in vitro

The aim of the work was to study regulators and mechanisms of insulin-like growth factor I (IGF-I) production by bovine ovarian cells cultured in vitro. The influence of stage of folliculogenesis, presence of serum, growth hormone (bGH) and cAMP agonist and antagonist was investigated. Cells were isolated in the periovulatory period by aspiration of small (2-4 mm in diameter) middle and large (5-15 mm in diameter) bovine ovarian follicles and cultured in DME/F-12 medium supplemented with 1 % antibiotic-antimycotic and 10 % fetal calf serum in a 24-well plate at 37 deg C and 5 % CO2 in air. IGF-I and cAMP content in samples was determined by radioimmunoassay. It was observed that bovine granulosa cells release IGF-I to the incubation medium at a maximum secretion level up to the fourth day of culture (Fig. 1). Cells cultured in a serum-supplemented medium produced significantly (P less than 0.05) lower amounts of IGF-I than those in a serum-free medium (Fig. 2). It had been found that the stage of folliculogenesis did not influence IGF-I production by cells cultured in a serum-free medium (Fig. 3). On the other hand, in the presence of serum, granulosa cells isolated from small follicles released IGF-I significantly more than the cells from middle and large follicles (Fig. 4). It was observed than GH given at physiological concentrations (1-100 ng per ml) significantly stimulated the production of both IGF-I (Fig. 5) and cAMP (Fig. 6). Moreover, dbcAMP (a cAMP agonist), similar to GH, had a stimulatory effect on IGF-I output (Fig. 7). Rp-cAMPS (a cAMP antagonist) when given alone had a minor or no influence on IGF-I production (Fig. 8) but one given together with GH, inhibited GH-induced IGF-I secretion (Fig. 5). These results suggest that 1) granulosa cells isolated from bovine ovarian follicles produce IGF-I in culture, 2) this IGF-I secretion depends on the follicular size and the presence of serum in a culture medium, 3) ovarian IGF-I production is stimulated by GH, and 4) cAMP may be a possible intracellular mediator of GH action on the IGF-I release in bovine ovary