Hormonal control of ovarian cell production of insulin-like growth factor binding proteins

To determine if the hormonal effects on insulin-like growth factor binding protein (IGFBP) production differed between granulosa and thecal cells. bot h cell types were collected and cultured in serum-free medium with various hormone treatments, arranged in three experiments. Following treatment, cel ls were enumerated and media were collected. concentrated 10-fold and subje cted to ligand blotting, Experiment 1 revealed that greater than or equal t o 1.5 x 10(5) viable cells at plating were needed for maximal IGFBP product ion by granulosa and thecal cells. The major forms of IGFBPs produced were a 27-34-kDa IGFBP (IGFBP-2 and -5), and a 20-22-kDa IGFBP (IGFBP-4) by the granulosa cells and a 40-44-kDa IGFBP (IGFBP-3), 34-kDa IGFBP (IGFBP-2), 27 -29-kDa IGFBP (IGFBP-5) and a 20-22-kDa IGFBP (IGFBP-4) by the thecal cells . In Experiment 2A, insulin stimulated production of IGFBP-5 by thecal cell s. and basic fibroblast growth factor (bFGF) inhibited the insulin-induced increase in IGFBP-5 production: epidermal growth factor (EGF) and luteinizi ng hormone were without effect. The small amounts of IGFBP-2 and -3 produce d by thecal cells of Experiment 2A were not affected by treatment. Producti on of IGFBP-2/-5 by granulosa cells in Experiment 2B was inhibited by insul in, with EGF and bFGF further enhancing insulin's inhibitory effect; follic le-stimulating hormone was without effect. In Experiment 3A, insulin enhanc ed production of IGFBP-5 by thecal cells whereas glucagon blocked insulin's stimulatory effect. In contrast, insulin or glucagon alone had no effect o n production of the IGFBP-4 by thecal cells but when combined inhibited IGF BP-4 production. The small amounts of IGFBP-2 and -3 produced by thecal cel ls of Experiment 3A were not affected by treatment. In Experiment 3B. produ ction of IGFBP-2/-5 by granulosa cells was attenuated in the presence of co rtisol with or without insulin and insulin plus glucagon; glucagon and cort isol decreased production of IGFBP-4 by granulosa cells. These results sugg est that production of IGFBP-2. -4. and -5 by granulosa and thecal cells ar e differentially affected by hormonal stimuli. and that IGFBP-3 is more con sistently produced by thecal cells than granulosa cells of cattle although its production was not hormonally regulated. (C) 2001 Elsevier Science Irel and Ltd. All rights reserved.