Preexposure of IM-9 lymphocytes to the somatomedin peptide insulin-like growth factor-I (IGF-I) results in a time- and concentration-dependent reduction in specific receptors for IGF-I. Since insulin and proinsulin are structurally homologous to IGF-I, we investigated the ability of insulin analogues to compete for occupancy and to directly modulate IGF-I receptor concentrations.
IGF-I binds rapidly and reversibly to IM-9 cells at 15°C, with half-maximal displacement of 125I-I-IGF-I at IGF-I concentrations of 3.6 × 10−9 M and insulin concentrations of 5 × 10−7 M. Preexposure of cells at 37°C to either IGF-I or insulin produced a concentration-dependent reduction in binding of 125I-IGF-I. A 50% decrease in binding was observed following preincubation of cells with IGF-I at 2.5 × 10−9 M and insulin at 2 × 10−7 M. At higher insulin concentrations (10−6–10−5 M), up to 70% reduction in 125I-IGF-I binding occurred. Bovine proinsulin and guinea pig insulin competed less potently than porcine insulin for the IGF-I receptor, and produced receptor loss in proportion to their ability to occupy the IGF-I receptor. Scatchard analysis indicated that at all insulin concentrations, the decrease in binding was secondary to loss of available IGF-I receptors, with no change in affinity. Receptor loss was evident following 1–2 h preexposure to insulin, with a t½ of 4 h and maximal receptor loss within 10 h. Similarly, IGF-I and IGF-II competed for occupancy of the IM-9 insulin receptor, with 50% displacement of 125I-insulin occurring at peptide concentrations of 3.5 × 10−9 M (insulin), 3.5 × 10−8 M (IGF-II), and 3 × 10−7 M (IGF-I). Preexposure of cells to these peptides at 37°C for 20 h resulted in a concentration-dependent reduction in binding of 125I-insulin, with the order of analogue effectiveness being insulin > IGF-II > IGF-I.
These data emphasize the structural and functional homology of insulin and the somatomedin peptides, IGF-I and II, as well as their respective receptors. Additionally, the data support the conclusion that the insulin and somatomedin peptides not only bind to both receptors, but downregulate each receptor in proportion to their ability to occupy that receptor.