REGULATION OF INSULIN-LIKE GROWTH-FACTOR (IGF) BINDING PROTEIN-2 SYNTHESIS AND DEGRADATION BY PLATELET-DERIVED GROWTH-FACTOR AND THE IGFS IS ENHANCED BY SERUM DEPRIVATION IN VASCULAR SMOOTH-MUSCLE CELLS

Growth factors such as platelet-derived growth factor (PDGF) and insulin-like growth factor (IGF-I) stimulate proliferation and migration of vascular smooth muscle cells (SMC). IGF-I bioactivity is modulated by high-affinity binding proteins (IGFBP) which are important regulators of these processes. Porcine vascular SMC synthesize IGFBP-2 and IGFBP-4 in vitro. In the present study, levels of lGFBP-2 in conditioned media (CM) were increased approximately 1.6 to 2.2-fold when cells were exposed to PDGF (20 ng/ml) or insulin (5 mu g/ml) for 24 hr following a 24 hr incubation in serum-free media, or following a 72 hr exposure to either growth factor. Similar increases in IGFBP-2 mRNA levels were observed. Exposure of cells to PDGF for 24 hr without prior serum deprivation resulted in smaller (47 +/- 11%) increases in IGFBP-2 protein levels but failed to alter mRNA levels. IGF-I, FGF, TGF-beta and EGF failed to increase IGFBP-2 using either experimental paradigm. in contrast, IGFBP-2 protein levels were consistently decreased (75 +/- 14%) after 72 hr of exposure to IGF-II without corresponding decreases in IGFBP-2 mRNA levels. Immunoprecipitation of [S-35] methionine-labeled ICFBP-2 indicated that this decrease was not due to a decrease in synthesis of IGFBP-2. Immunoblot analysis of CM from cells treated with IGF-II indicated that the decrease in intact protein corresponded with an increase in two non-IGF binding IGFBP-2 fragments of 22 and 14 kD. Increased abundance of these fragments was also observed following IGF-I exposure, although corresponding decreases in intact IGFBP-2 were not usually observed. The relative abundance of these fragments did not appear to be affected by treatment with PDGF or insulin. in contrast to IGFBP-2, regulation of the levels of IGFBP-4 in CM did not appear to be altered by serum deprivation. Insulin consistently increased IGFBP-4 mRNA and protein levels under all situations. PDGF tended to increase IGFBP-4 protein levels, although this effect was less consistent and not as great as the increase observed with insulin. Treatment with IGF-I or -II consistently decreased IGFBP-4 levels in CM but tended to increase their mRNA levels under all situations. These data indicate that insulin, PDGF, and the IGFs regulate both IGFBP-2 and IGFBP-4. While PDGF and insulin stimulate IGFBP-2 and 4 synthesis, the IGFs appear to activate protease(s) which regulate IGFBP-2 and -4 levels post-translationally. The regulation of IGFBP-2 levels by each of these mechanisms appears to be amplified by serum deprivation, but th is is not observed with ICFBP-4. (C) 1995 Wiley-Liss, Inc.