INSULIN-LIKE GROWTH-FACTOR BINDING PROTEIN-3 PROTEOLYSIS IN CHILDREN WITH INSULIN-DEPENDENT DIABETES-MELLITUS - A POSSIBLE ROLE FOR INSULININ THE REGULATION OF IGFBP-3 PROTEASE ACTIVITY

Limited proteolysis of serum insulin-like growth factor (IGF) binding protein (IGFBP)-3 has been described in various conditions and may inc rease the bioavailability of IGFs. The physiological regulators of ser um IGFBP-3 protease activity are unknown. To characterize the relation ship between insulin and IGFBP-3 protease activity, we have examined s erum IGFBP-3 proteolysis in children with untreated in sulin-dependent diabetes mellitus (IDDM) and have followed the effect of insulin ther apy on serum IGFBP-3 proteolysis at 1 day, 1 week, and 1 month after t he initiation of insulin therapy. Ligand blot analysis of sera from un treated children with LDDM showed that intact IGFBP-3 was 50 +/- 98 of the age-matched control pool. After the initiation of insulin treatme nt, IGFBP-3 did not change significantly at 1 day after treatment but increased dramatically at 1 week (90 +/- 13%) and 1 month after treatm ent (102 +/- 13%). In contrast, when measured by immunoradiometric ass ay (which detects both intact and fragments of IGFBP-3), IGFBP-3 level s were 70% of the control pool before insulin therapy and did not incr ease significantly until 1 month after treatment. Immunoblot analysis demonstrated that intact IGFBP-3 doublet was diminished to 41 +/- 78 o f controls, whereas the major IGFBP-3 fragment (30 kDa) was increased in IDDM sera before insulin therapy. After insulin, intact IGFBP-3 inc reased and the 30-kDa fragment decreased to values comparable to those observed in controls. In vivo IGFBP-3 proteolysis, which implies prea ssay exposure of serum IGFBP-3 to proteases, was estimated by immunobl ot analysis. IGFBP-3 proteolysis was increased before insulin therapy (160 +/- 9%) and decreased to 81 +/- 9% at 1 week and to 71 +/- 11% at 1 month after insulin treatment. Residual serum IGFBP-3 protease acti vity was estimated by a I-125-IGFBP-3 degradation assay. Serum IGFBP-3 protease activity increased significantly in untreated diabetics, com pared with activity in controls (128 +/- 5% vs. 99 +/- 11%). During in sulin therapy, serum IGFBP-3 protease activity decreased gradually to 91 +/- 5% of control values at 1 month. Molecular sizes of the IGFBP-3 proteolytic fragments (30 kDa, 24 kDa, and 19 kDa) and inhibition pro file of IGFBP-3 protease were similar in IDDM and pregnancy sera, indi cating that similar proteases (cation-dependent serine proteases) were active in both conditions. These results suggest an important role of insulin in the regulation of IGFBP-3 protease activity. Increased IGF BP-3 proteolysis in the sera of children with IDDM may serve to counte ract the catabolic state induced by insulin deficiency.