Effects of anti-insulin antibody on insulin binding to liver membranes: evidence against antibody-induced enhancement of insulin binding to the insulin receptor.
In the presence of anti-insulin antibody, 2-to 3-fold enhancement of 125I-insulin binding to liver membranes was observed when binding was estimated by the radioactivity of 125I-insulin bound to the membrane pellets. However, after 125I-insulin was covalently cross-linked to liver membranes using disuccinimidyl suberate in the presence of anti-insulin antibody, sodium dodecyl sulfate-polyacrylamide gel electrophoresis and autoradiography showed that 125I-insulin bound to the alpha-subunit of the insulin receptor was inhibited by anti-insulin insulin antibody in an dose-dependent manner. More importantly, at an anti-insulin antibody dilution range between 1:50 and 1:5,000, sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed two 125I-labelled bands of mol wt 62,000 and 27,000, while only one band of mol wt 130,000 was revealed in the absence of anti-insulin antibody. These Mr = 62,000 and Mr = 27,000 bands were found to be the heavy and the light chain of anti-insulin IgG molecules respectively. Pepsin digested anti-insulin serum had only an inhibitory effect on 125I-insulin binding to liver membranes. Non-immunized guinea pig serum or IgG completely abolished the enhanced effect of anti-insulin antibody. Further, this enhanced effect was inhibited by Fc fragment-specific anti-IgG serum or H&L-chain-specific anti-IgG serum in a dose-dependent manner. Protein A also inhibited the effect of anti-insulin antibody. In IM-9 lymphocytes and human red blood cell ghosts, which have no Fc gamma receptors, enhancement of insulin binding was not observed in the presence of anti-insulin antibody.(ABSTRACT TRUNCATED AT 250 WORDS)
D000818:Animals; D000937:Antigen-Antibody Reactions; D002462:Cell Membrane / Q000378:metabolism; D003432:Cross-Linking Reagents; D007328:Insulin / Q000276:immunology* / Q000378:metabolism; D008099:Liver / Q000378:metabolism*; D008297:Male; D008970:Molecular Weight; D051381:Rats; D011972:Receptor, Insulin / Q000378:metabolism*; D011961:Receptors, Fc / Q000378:metabolism
D003432:Cross-Linking Reagents; D007328:Insulin; D011961:Receptors, Fc; D011972:Receptor, Insulin