Secretion of IGF-1 by ovine granulosa cells: effects of growth hormone andfollicle stimulating hormone

Insulin-like growth factor-1 (IGF-1) is implicated in follicle development and is considered to mediate the actions of growth hormone (GH) and gonadot rophins at the ovarian level. However, the expression and secretion of IGF- 1 by the ovary are controversial, partly because of species and cell-type s pecificity. The present study investigated whether IGF-1 is produced by ovi ne granulosa cells and whether its production is regulated by GH and follic le stimulating hormone (FSH). Follicles (greater than or equal to 4.0 mm) w ere obtained from ewes during seasonal anoestrus. Granulosa cells were cult ured for a total period of 96 h in Dulbecco's modified Eagle's medium (DMEM )/Ham's F-12 medium supplemented with BSA (0.1%, w:v), transferrin (0.5 mu g/ml) and testosterone (100 ng/ml). In the first set of experiments, cells were incubated in the presence of bovine calf serum (BCS) (2.5%) for the in itial 48 h of culture. The cells were then cultured for the next 48 h in me dium without BCS, but containing either GH (0, 2, 20, and 200 ng/ml) or FSH (0, 20, 200, and 2000 ng/ml). The medium was assayed for oestradiol (E), p rogesterone (P) and IGF-1. There were six wells per treatment and the exper iment was carried out four times. Control granulosa cells maintained both I GF-1 and E secretion, with only low levels of progesterone output. In all e xperiments, both GH and FSH produced significant (P < 0.001) dose-related i ncreases in E, IGF-1 and P secretion into the medium. The maximum responses to GH (20 or 200 ng/ml) were 402% for E and 528% for IGF-1 compared with c ontrols. The maximum responses to FSH (200 or 2000 ng/ml) were 460% for E a nd 514% for IGF-1. The objective of the second set of experiments was to de termine the effect of the progestogenic status of cells on IGF-I production . Granulosa cells were cultured both in the presence and absence of BCS (2. 5% in the medium) during the initial 48 h of culture. For the next 48 h, ce lls were cultured in serum-free medium. Addition of BCS to the medium durin g the initial 48 h of culture stimulated progesterone production. However, it did not affect either IGF-1 or oestradicol secretion between 49 and 96 h of culture, or the cell numbers at the end of culture. In conclusion, (1) IGF-1 is secreted by granulosa cells irrespective of their progestogenic st atus and (2) concomitant increases in E and IGF-1 production by granulosa c ells as a result of GH and/or FSH treatment suggest a role for GH and FSH i n the regulation of ovarian function. (C) 2000 Elsevier Science B.V. Ail ri ghts reserved.