INSULIN-LIKE GROWTH FACTOR BINDING PROTEINS FROM PRIMARY CULTURES OF RAT OVARIAN THECA-INTERSTITIAL (ROTI) CELLS: REGULATION BY LH AND IGF-I

Abstract

We and others have shown that insulin-like growth factor-I (IGF-I) and IGF binding proteins (IGFBPs) play an important role in ovarian function. In the present study, we tested the hypothesis that ROTI cells produce IGFBPs and that theca-interstitial cell IGFBP production is regulated by LH and IGF-I. Density gradient-purified ROTI cells were cultured in serum-free medium with selected concentrations of LH or IGF-I, alone and in combination. After 24 to 48 hours, supernatant media were removed and IGFBPs analyzed by Western ligand blotting and immunoblotting. Ligand blotting was carried out using equal amounts of radiolabeled IGF-I and IGF-II. Immunoblotting was carried out using anti rat IGFBP-3 antiserum and anti bovine IGFBP-2 antiserum (provided by Drs. Nicholas Ling and David Clemmons, respectively). Western ligand blotting of media obtained from cells cultured without added hormones (Control) revealed a predominant IGFBP of 29-30 kD with minor species of 28 and 35-40 kD. Immunoblotting with anti-IGFBP-2 detected the 29-30 kD species whereas immunoblotting with anti-IGFBP-3 failed to detect any IGFBP-3. High doses of LH (100 ng/ml) decreased IGFBP-2 expression while 100 ng/ml IGF-I increased IGFBP-2 levels 5 to 10-fold above control values. IGFBP-2 levels were increased by 24 hours and reached peak levels by 48 hours of incubation. We conclude that rat ovarian theca-interstitial cells produce several IGFBPs, primarily IGFBP-2. IGFBP-2 levels are increased by IGF-I and suppressed by high doses of LH. The production of IGFBP-2 by theca-interslitial cells and its regulation by LH and IGF-I may play an important role in ovarian androgen production.