Direct demonstration that receptor crosslinking or aggregation is important in insulin action

Abstract

Exposure of adipocytes to antibodies to the insulin receptor resulted in a blockade of 125I-labeled insulin binding, stimulation of glucose oxidation, and many more insulin-like effects. Allowing for differences in purity, antireceptor antibody was equipotent with insulin on a molar basis. Both the bivalent F(ab'')2 and monovalent FAB'' fragments of the antireceptor antibody were fully active in inhibiting 125I-labeled insulin binding. Bivalent F(ab'')2 also retained its insulin-like effects. The monovalent Fab'' lost almost all ability to stimulate glucose oxidation and acted as a competitive antagonist of insulin-stimulated glucose oxidation. Addition of anti-F(ab'')2 antisera, which crosslink the Fab''-receptor complexes, resulted in a restoration of the insulin-like activity of the antibody. When cells were exposed to submaximal doses of insulin, addition of anti-insulin antibodies at low concentration enhanced the biological activity of insulin. Receptor occupancy by ligand was not sufficient for signal generation and the insulin-like effects of antireceptor antibody (and perhaps insulin itself) require receptor aggregation or clustering. This aggregation appears to be independent of microfilaments or microtubules because the insulin-like effects of antireceptor antibody and of insulin itself, are unaffected by agents that disrupt these structures. Porcine insulin, guinea pig anti-insulin serum and human and goat antibodies were used.