VH GENE STRUCTURE PREDICTS A LARGE POTENTIAL ANTIINSULIN REPERTOIRE

The majority of insulin antibodies derived from immunization are IgG a ntibodies that cross-react extensively with the autologous hormone. To examine the relationship between VH genes expressed by such self-reac tive antibodies and their germline (non-rearranged) counterparts, we u sed the polymerase chain reaction (PCR) to amplify and isolate the ger mline progenitors of anti-insulin VH genes derived from BALB/c mice im munized with beef or human insulin. Results indicate that two anti-ins ulin mAbs (123 and 124) express VH genes which arise from a small subs et of the J558 gene family and are highly homologous to the VH gene us ed by the murine CD5 + B-cell tumor, BCL1. The anti-insulin IgG mAb 12 7 belongs to the VH-VIII (V gam 3.2) family and the amplification and isolation of germline VH genes from this small family precisely identi fied only two somatic mutation events in the CDRH2 of mAb 127. Another anti-insulin mAb, 133, also shows two replacement substitutions in th e CDRHs when compared to the germline encoded anti-dextran antibody 19 .1.2. These findings indicate that the IgG response to this small self -protein uses multiple VH genes which are largely germline encoded wit h only a low level of somatic mutation in their CDRHs. Additionally, a nalysis of N-segment additions in CDRH3s indicates anti-insulin B cell s may originate from both early (fetal) and adult repertoires. These d ata are consistent with the concept that the mechanisms of clonal aner gy or deletion do not regulate anti-insulin B cells and indicate that there is a large potential VH gene repertoire for insulin.