Background/Purpose: TNF inhibitors are widely used for inflammatory diseases but often induce ANAs that sometimes progress to overt SLE. TNF deficient mice fail to generate germinal centers (GCs) but are still able to mount effective humoral responses to exogenous antigens. A similar loss of GCs occurs in the tonsils of RA patients treated chronically with TNF inhibitors. The goal of these studies was to analyze the mechanism of induction of ANAs by TNF inhibition and the nature of the checkpoint between ANA induction and clinical SLE in a mouse model. 

Methods: TNF deficient autoimmune prone Sle1 mice were generated and the 3H9 IgVH transgene that confers anti-DNA and anti-cardiolipin specificity was introduced. Mice were followed clinically and sacrificed at the age of 52 weeks. Spleen cells were analyzed by single cell PCR for the repertoire of Vk light chains associated with the 3H9 heavy chain. Selected 3H9/Vk combinations were transfected into 293 cells and supernatants were tested for autoreactivity by ELISA.

Results: TNF deficient Sle1 mice did not spontaneously develop clinical SLE or proteinuria and survived until at least 52 weeks of age. Surprisingly, serum levels of both anti-chromatin and anti-dsDNA antibodies were decreased in TNF deficient mice compared with their TNF sufficient controls, with similar results in the 3H9 TNF deficient mice.  By contrast, TNF deficient mice developed high titers of IgG anti-cardiolipin autoantibodies. Similar results were observed in Sle1.TNFR1 deficient but not Sle1.TNFR2 deficient mice. Single cell analysis of the follicular B cell repertoire of 3H9.Sle1 TNF deficient mice revealed a loss of 3H9/Vk12-46.Jk2 encoded B cells that can bind to histones and chromatin.

We have previously shown a preferential selection of Vk5 light chains into the GCs of 3H9 SLE prone mice; these light chains confer anti-chromatin activity in their germline configuration and acquire high affinity anti-DNA and anti-cardiolipin activity as a result of somatic mutations.  Since GCs are not present in TNF deficient mice we analyzed the light chain repertoire of splenic class switched B cells and plasma cells. 3H9.Sle1 mice had marked overrepresentation of Vk5 encoded light chains among their class switched B cells and plasma cells. By contrast, 3H9.Sle1 TNF deficient mice had few Vk5 encoded plasma cells and instead had a vast overrepresentation of Vk4-57-1. Co-transfection of this light chain with 3H9 into 293 cells revealed that this combination had anti-cardiolipin but not anti-DNA or anti-chromatin activity. Further analysis of the Vk4-57-1 encoded light chains from class switched B cells of TNF deficient mice revealed that they were all germline encoded.

Conclusion: TNF deficiency has significant effects on the Ig repertoire of mature and antigen activated B cells.  The loss of GCs in TNF deficient Sle1 mice alters the spontaneously autoreactive Ig repertoire and is associated with a decrease in somatic mutations in autoreactive B cells suggesting that these cells have matured in an environment that is not exposed to cognate T cell help. The data further suggest that a “second hit” that bypasses the germinal center defect is required for TNF inhibition to induce pathogenic autoimmunity.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/the-effect-of-tnf-inhibition-on-the-autoreactive-b-cell-repertoire-in-sle-prone-mice/