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1701 Curli amyloid/DNA complexes from bacterial biofilms break tolerance in murine lupus using T cell-independent and T cell-dependent modalities
  1. Michael H Lee1,2,
  2. Sarah A Tursi2,
  3. Lauren K Nicastro2,
  4. Benjamin L Green1,2,
  5. Roberto Caricchio3,
  6. Çagla Tükel2 and
  7. Stefania Gallucci1,2
  1. 1Laboratory of Dendritic Cell Biology
  2. 2Center for Microbiology and Immunology, Department of Microbiology, Immunology and Inflammation, Lewis Katz School of Medicine, Temple University
  3. 3Division of Rheumatology, Department of Medicine, Lewis Katz School of Medicine, Temple University, Philadelphia, Pennsylvania 19140, USA


Background Epidemiological studies suggest that bacterial infections promote SLE disease in predisposed individuals, but the underlying mechanisms remain unknown. We have found that a subset of SLE patients has asymptomatic bacteriuria associated with markers of inflammation and flares, suggesting that chronic exposures to microbial products may trigger flares in lupus. Our labs have shown that the bacterial amyloid curli, expressed in multicellular communities (biofilms) by many bacteria including E. coli, plays a major role in triggering lupus autoimmunity during infection. Curli amyloid/DNA complexes strongly activate dendritic cells and macrophages. When given systemically, curli/DNA complexes and infections with curli-expressing E. coli trigger production of anti-dsDNA and anti-chromatin autoantibodies in lupus prone mice and in wild type mice. This stimulation is diminished in TLR2 or TLR9 deficient mice, suggesting a TLR-mediated activation of innate immunity. We have now focused on the effects of curli/DNA complexes on B cells.

Methods Young wild type C57BL/6 mice, lupus prone Sle1,2,3 mice, 3H9 mice and CD40L-/- mice were injected with curli/DNA complexes from biofilms or infected with amyloid for short and long-term studies. Splenic B cells were stained by flow cytometry ex vivo. For in vitro experiments, B cells were sorted by positive selection with CD45R (B220), supplemented with anti-CD43Ab-Biotin. B cell purity (>98%), proliferation, activation markers and signaling molecules were measured by Flow cytometry, Western Blot and qRT-PCR. Autoantibodies were measured by ELISA.

Results In vitro, curli/DNA complexes could induce class switch to IgG, in the absence of T cell help, in wildtype B cells, and even more in Sle1,2,3 and 3H9 B cells, which recognize DNA, suggesting an antigen-specific activation of B cells by curli/DNA. Curli/DNA induced non-canonical NFκB activation and transcription of aicda, the master regulator of class switch recombination. In vivo, exposure to curli/DNA broke tolerance to DNA in 3H9 mice. Moreover, it induced autoantibodies in CD40L-/- mice, though at lower levels than in WT mice.

Conclusions The induction of non-canonical NFκB activation, aicda, and class switch recombination, in the absence of T cells help in vitro, suggests that the fibrillar structure of curli/DNA complexes can cross-link BCRs, some recognizing DNA, and can also trigger a second pathway which substitutes T cell help to induce isotype switching. The lower levels of autoantibodies elicited by curli/DNA in mice deficient of T cell help suggests that curli/DNA complexes break tolerance to DNA with T cell-independent and T cell-dependent modalities.

Acknowledgments This work was supported by the NIH, NIAID grant R21-AI119947 and the Lupus Research Institute, now Lupus Research Alliance, Innovative Grant (SG); NIH NIAMS grant R01-AR061569, R56-AR072115, Lupus Research Alliance, TIL (RC); NIH, NIAID R21-AI125429, R21-AI132996, R21-AR072115 (CT).

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