Background/Purpose: Anti-DNA B cells are a primary cause of pathology in individuals with systemic lupus erythematosus (SLE), producing autoantibodies that deposit in diverse tissues and cause extensive inflammatory damage. Congenic mice generated through the introgression of New Zealand Black (NZB) chromosome 1 (C1) susceptibility loci onto the non-autoimmune C57BL/6 (B6) strain are an effective way of modelling lupus and parsing its pathogenesis. Our laboratory has previously provided evidence that a B cell defect instrumental in autoantibody production localizes to the C1d (170.8-181 Mb) region of the NZB C1 and we have further shown that additional T cell and dendritic cell (DC) defects map to the C1b-d (124-181 Mb) locus. Since B cells from both C1d and C1b-d mice with transgenes for hen egg lysozyme (HEL)-specific immunoglobulin and soluble HEL demonstrated a breach of anergy, we hypothesized that the C1d interval alone would be sufficient to breach tolerance to nuclear antigen. Here, we have used a knock-in model of anergy to ssDNA to address this question.

Methods: Genes for the 3H9 heavy chain (IgH^a) and Vκ8 light chain were knocked into their proper loci in the C1d and C1b-d congenic strains in order to generate mice with homogeneous, ssDNA-specific B cell repertoires. Female B6, C1d and C1b-d Vκ8/3H9 and wild type mice were aged to 8 months, at which time sera, splenocytes and bone marrow were collected. Serum levels of anti-nuclear autoantibody production were measured by ELISA, while T and B cell activation and localization were assessed using flow cytometry and immunofluorescence microscopy.  

Results: C1d.Vκ8/3H9 mice produce significantly higher levels of anti-ssDNA IgG2a^a autoantibodies compared to their B6 counterparts. In line with this increased IgG2a^a production, these mice also have a significant increase in the proportion of germinal centre B cells and plasma cells compared to B6 Vκ8/3H9 mice. As anticipated, C1b-d.Vκ8/3H9 mice show an even stronger breach of anergy, with significant increases in anti-ssDNA IgG, IgM and IgG2a^a autoantibodies as well as significantly higher proportions of germinal centre B cells than their B6 counterparts. These changes mirror the observed increase in the number of follicular helper and IFNγ-producing T cells, which are thought to provide support for germinal centre B cell development and subsequent autoantibody production. C1d and C1b-d mice also show evidence of defective receptor editing, with significantly lower proportions of B cells with the Igλ1 light chain in the bone marrow compared to B6 counterparts. In agreement with our previous observations of increased survival and peripheral expansion of autoreactive B cells in the HEL model, the proportions of Igλ1⁺ B cells normalize in the spleen, further supporting the observed breach of anergy.

Conclusion: Our results confirm the presence of a B cell defect to nuclear antigen in the C1d interval and further imply that this defect involves perturbation of both central and germinal centre tolerance.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/breach-of-b-cell-anergy-in-new-zealand-black-congenic-mice/