Background/Purpose: A central mediator of Systemic Lupus Erythematosus (SLE) pathogenesis is interferon-alpha (IFNα), which is elevated in the serum of SLE patients. IFNα has been shown to enhance B cell signaling and promote survival. It also plays a major role in the induction of B cell activating factor (BAFF). While past studies have explored the effects BAFF on B cell tolerance, little work has focused on how IFNα itself directly affects this process. We hypothesize that elevated levels of IFNα directly contribute to the breach of B cell tolerance in SLE. To address this question, we have obtained an adenoviral vector encoding mouse IFNα (mDEF201), which we are using to induce sustained elevation of serum IFNα in a mouse model of B cell tolerance (3H9).

Methods: 6-8 week old 3H9 transgenic mice, which contain a knock-in Ig heavy chain derived from a DNA-specific hybridoma, were injected IV with 10⁷ PFU of Ad-mIFNα (mDEF201) or Ad-dI70-3 (empty vector). At 2 weeks post-treatment immune cell populations in the spleen and bone marrow were examined by flow cytometry, and anti-DNA antibody production was measured by ELISA. Serum levels of IFNα and BAFF were quantified by ELISA and IFN-induced gene expression was assessed by qRT-PCR.

Results: Mice administered with mDEF201 showed elevation of serum IFNα from 48h to 2 weeks post infection. At 2 weeks post infection, the mRNA expression of several IFN-inducible genes, but not BAFF, was also elevated in infected mice. There was only a 0.5 fold increase in BAFF expression at the protein level. mDEF201 infection resulted in a marked increase in the level of anti-ss/dsDNA IgM^a autoantibodies (OD450 PBS=0.32, Ad-di70-3=0.29, mDEF201=1.46 for ssDNA, and PBS=0.13, Ad-di70-3=0.10, mDEF201=0.56 for dsDNA, p<0.0001) signifying a potential breach of B cell tolerance. Consistent with this idea, mDEF201 infected mice displayed altered B cell activation and homeostasis, with a significant increase in the frequency of CD86⁺ B cells as well as total number of mature B cells. mDEF201 infection also resulted in increased frequency of plasmablasts, CD138⁺ plasma cells and IgM^a+/IgG2^a+ germinal center (GC) B cells. Despite the increase in IgG2^a+ GC B cells, anti-ss/dsDNA IgG2^a antibodies were not increased with elevated IFNα at 2 weeks post infection. To assess the effect of IFNα on B cell anergy we examined the Igλ1⁺ population, a well characterized dsDNA-specific anergic B cell population in 3H9 mice. Igλ1⁺ B cells also displayed increased activation (CD86⁺) and entry into germinal centers; however, anti-ss/dsDNA Igλ1⁺ levels were not increased. Infection with mDEF201 resulted in a mild increase in T cell activation (CD69⁺); although, no difference was seen in the total number of T follicular helper cells.

Conclusion: Taken together, these data suggest that IFNα may be a major contributing factor in breaches of B cell tolerance in SLE not only through the induction of BAFF, but also through direct effects on autoreactive B cells. Elevation of IFNα may promote the activation and differentiation of autoreactive B cells towards an antibody producing state; however, in the absence of other immune defects, some peripheral tolerance mechanisms seem to remain partially intact preventing a whole-cell breach of tolerance.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/interferon-alpha-disrupts-dna-specific-b-cell-tolerance-in-3h9-mice/