INSULIN-LIKE GROWTH-FACTOR-I (IGF-1) STIMULATES THE IGF BINDING-PROTEIN SYSTEM IN RAT THECA INTERSTITIAL-CELLS

There has been considerable interest in rat ovarian insulin-like growt h factor binding proteins IGFBPs because they are potent inhibitors of FSH action. In situ, IGFBP-2 and -4 and IGFBP-3 mRNAs are expressed i n rat theca interstitial (TIC) and theca lutein cells respectively. Al though much is known about IGFBPs in rat TIC at the mRNA level, the sy nthesis and regulation of IGFBP proteins remain poorly understood. The purpose of this study was to identify the species of IGFBPs produced by TIG and to determine the effects of LH and IGF-1 on their expressio n. This was accomplished by culturing rat TIG for 2 days in serum-free medium with graded doses of LH and/or ICF-I, and measuring IGFBP mRNA s in the cells and IGFBP proteins in the conditioned media by RT-PGR a nd Western immunoblotting respectively. The RT PGR analysis identified strong bands for IGFBP-2 and -4 mRNAs in TIG. In some treatments, the mRNAs for IGFBP-3 and -6 were also identified, but transcripts for IG FBP-1 and -5 were undetectable. Two species of IGFBPs were detected in the conditioned media of control (untreated) TIG, the 31 kDa IGFBP-2 and the 24 kDa (non-glycosylated) and 28 kDa (glycosylated) forms of I GFBP-4. There was no detectable IGFBP-5 and barely detectable amounts of IGFBP-3 and -6 in the conditioned media. Treatment with LH (0.2-20 mu U/ml) caused no significant changes in the levels of the 31 kDa IGF BP-2 and the 24 kDa and 28 kDa IGFBP-4 bands, and there was no detecta ble IGFBP protease activity. In contrast, IGF-I (100 ng/ml) stimulated the expression of IGFBP-2 IGFBP-4 and a 17.5kDa IGFBP-4 fragment. The immunoreactive IGFBP-4 fragment suggests the media contained an IGFBP -4 protease. The IGF-I effects were dose dependent (ED(50) = 12.4 +/- 3.3 ng/ml). Go-treating TIG with LH (0.2-20 mu U/ml) caused no signifi cant change in the activity of IGF-I in stimulating the expression of IGFBP-2, IGFBP-4 and IGFBP-4 protease. We have demonstrated that ICF-I acts directly on rat TIC to stimulate the expression of the intrinsic IGFBP system. LH, either alone or together with ICF-I, did not signif icantly change the expression of TIG IGFBP proteins. Therefore, we hyp othesize that ICF-I, but not LH, may be a physiologically important re gulator of the IGFBP system in rat TIG. Because IGF-I is a potent stim ulator of theca function, changes in the expression of this intrinsic IGFBP system could have new implications for ovarian androgen producti on, both at the physiologic and pathophysiologic levels.