Productive Germinal Center Responses Depend on the Nature of Stimuli Received by Anti-Insulin B Cells in Type 1 Diabetes–Prone Mice

Abstract

Islet autoantibodies, including those directed at insulin, predict type 1 diabetes (T1D) in mice and humans and signal immune tolerance breach by B lymphocytes. High-affinity insulin autoantibodies and T follicular helper cell involvement implicate germinal centers (GCs) in T1D. The VH125SD BCR transgenic model, in which 1–2% of peripheral B lymphocytes recognize insulin, enables direct study of insulin-binding B cells. Our prior studies showed that anti-insulin B cell receptor transgene site-directed to H chain locus mice fail to generate insulin Ab following T-dependent immunization, but it was unclear whether anti-insulin B cells were blocked for GC initiation, survival, or differentiation into Ab-secreting cells. Here, we show that insulin-binding B cells in T1D-prone anti-insulin B cell receptor transgene site-directed to H chain locus mice can spontaneously adopt a GC phenotype and undergo class switching to the IgG1 isotype, with little if any switching to IgG2b. T-dependent immunizations with insulin SRBC or insulin CFA drove anti-insulin B lymphocytes to adopt a GC phenotype, despite blunted insulin Ab production. Dual immunization against self (insulin) and foreign (4-hydroxy-3-nitrophenylacetyl hapten conjugated to keyhole limpet hemocyanin) Ags showed an anti-insulin (but not anti-4-hydroxy-3-nitrophenylacetyl) Ab block that tracked with increased expression of the apoptosis marker, activated caspase 3, in self-reactive GC B cells. Finally, T-independent immunization with insulin conjugated to Brucella abortus ring test Ag released immune tolerance to allow robust expansion of anti-insulin GC B cells and IgG-switched insulin Ab production. Overall, these data pinpoint GC survival and Ab-secreting cell differentiation as immune tolerance blocks that limit T-dependent, but not T-independent, stimulation of anti-insulin B cell responses.

Factors Governing B Cell Recognition of Autoantigen and Function in Type 1 Diabetes

“…Site-directed models were subsequently developed in which these anti-insulin BCR transgenes were introduced at the physiologic IgH and Igk locus, VH125 SD [31,118] and Vκ125 SD [36], respectively, which enabled B cells to undergo isotype switch, somatic hypermutation, and receptor editing. Anti-insulin B cells can spontaneously adopt a germinal center phenotype and undergo limited class switching in NOD.VH125 SD mice in vivo [35], which is dramatically enhanced by the presence of anti-insulin 8F10 T cells following co-transfer into Rag1-deficient NOD recipients [46]. In contrast to these studies indicating T1D dependence on germinal center formation, ~50% of NOD.SAP-deficient mice develop diabetes, despite showing a strong, albeit incomplete, reduction in germinal center B cells [119].…”