Insulin-Like Growth Factor-I Selectively Stimulates Cholesterol Side-Chain Cleavage Expression in Ovarian Theca-lnterstitial Cells

Publisher Website
Google Scholar
Abstract
Evidence accumulating in the literature supports the concept that insulin-like growth factor I (IGF-I) may be an important local regulator of ovarian function. Recent studies have demonstrated that IGF-I synergistically augments LH stimulation of theca-interstitial cell (TIC) androgen biosynthesis. The purpose of the present studies was to begin to elucidate the molecular mechanisms of the interaction between IGF-I and LH. TIC were purified from ovaries of hypophysectomized immature rats by Percoll gradient centrifugation. When isolated TIC (5 .times. 106 viable cells per dish) were cultured (4 days) in serum-free medium, low amounts (<10 ng/ml) of androsterone were produced. Basal androsterone production was not changed by incubation with IGF-I (30 ng/ml). Treatment with LH (50 ng/ml) caused an 85-fold stimulation of androsterone synthesis that was further increased 2.1-fold by concomitant treatment with IGF-I. Immunoblot analysis demonstrated that untreated TIC contained low levels of 17.alpha.-hydroxylase/C17-20 lyase enzyme (P45017.alpha.) that were unchanged by incubation with IGF-I alone. LH treatment increased P-45017.alpha. content 5.5-fold and coincubation with LH plus IGF-I increased P45017.alpha. content 16-fold above control levels. Cholesterol side chain cleavage enzyme (P450scc) was readily detected in immunoblots from untreated TIC. P450scc content was increased 2.6-fold by LH treatment and 4.2-fold by LH plus IGF-I. Interestingly, IGF-I alone induced a 2-fold increase in P450scc. To determine if the increases in P450scc content were associated with increased enzyme activity, progesterone production was measured. Progesterone synthesis was increased 6-fold above basal levels by LH and 7.5 fold by LH plus IGF-I. Surprisingly, IGF-I treatment did not stimulate progesterone production. To determine if the P450scc induced by IGF-I was catalytically active, TIC were pretreated (2 days) with IGf-I to induce P450scc and then challenged (4 h) with LH, 8-bromo-cAMP, or 25-hydroxycholesterol. In each case P450scc activity was significantly greater in TIC pretreated with IGF-I compared with unprimed controls. There was no increase in TIC P45017.alpha. activity after any of the challenges with or without IGF-I pretreatment. Northern blot analysis showed that control TIC contained detectable amounts of P450scc mRNA which were increased 4-fold by incubation with IGF-I alone. LH caused a 50% decrease in P450scc mRNA while IGF-I plus LH increased mRNA 2-fold above control levels. P45017.alpha. mRNA was also detectable in untreated TIC. The P45017.alpha. mRNA was coordinately regulated with P450scc mRNA. These experiments demonstrated that IGF-I alone causes a selective increase in functional P450scc activity by stimulating increases in P450scc enzyme content and P450scc mRNA levels in the TIC, and that IGF-I augments the ability of LH to increase the content of P450scc and P45017.alpha. content in the TIC.