SAT0030 CITRULLINE-REACTIVE B CELLS ARE PRESENT IN INFLAMED GINGIVAL TISSUE AND DISPLAY CROSS-REACTIVITY BETWEEN BACTERIAL AND HUMAN ANTIGENS
Authors: Natalia Sherina, Vijay Joshua, Radha Thyagarajan, Natalie Sippl, Lena Israelsson, Heidi Wähämaa, Ragnhild Stålesen, Kaja Eriksson, Tülay Yucel-Lindberg, Aase Hensvold, Caroline Grönwall, Anca Catrina, Vivianne Malmström, Antonio Lanzavecchia, Luca Piccoli, Khaled Amara, Karin Lundberg
Journal: Annals of the Rheumatic Diseases (2019)

<h3>Background:</h3> Antibodies targeting citrullinated proteins (ACPA) are highly specific for rheumatoid arthritis (RA). However, the etiology and molecular basis for ACPA production is still unclear. Based on an epidemiological association between RA and periodontitis (PD), and the unique ability of the oral pathogen <i>P.gingivalis</i> (<i>Pg</i>) to express a PAD enzyme that can citrullinate both bacterial and human proteins, it has been hypothesised that the ACPA response may be triggered in the gum mucosa in response to <i>Pg</i>. <h3>Objectives:</h3> The main purpose of this study was to investigate if citrulline-reactive B cells reside in inflamed gingival tissue, and to examine ACPA cross-reactivity between citrullinated bacterial and human epitopes on a monoclonal level. <h3>Methods:</h3> Using a single-cell antibody cloning approach, 55 recombinant monoclonal antibodies (mAbs) were generated from gingival tissue (GT) CD19+ B cells (n=1 ACPA+ RA/PD patient). Citrulline reactivity was determined using the anti-CCP2 kit (EuroDiagnostica AB), and in-house peptide ELISAs (including a citrullinated peptide derived from <i>Pg</i> PAD (CPP3) and citrullinated peptides derived from human α-enolase, fibrinogen, vimentin, fillaggrin and histone 4). Reactivity against CCP2 and CPP3 was also investigated in: 19 mAbs from bronchoalveolar lavage (BAL) CD19+ B cells (n=2 ACPA+ RA patients); 29 mAbs from bone marrow (BM) plasma cells (n=4 ACPA+ RA patients); 142 mAbs from synovial fluid (SF) plasma cells (n=5 ACPA+ RA patients); and 36 mAbs from peripheral blood memory/plasma cells (n=4 ACPA+ RA patients). Predicted germline versions were produced for two of the mAbs. <h3>Results:</h3> Among 55 GT mAbs, 14 unique clones (25%) were reactive to the bacterial CPP3-peptide. We also detected CPP3-reactive mAbs from BAL (n=9/19), BM (n=3/29), SF (n=1/142) and peripheral blood (n=1/36). Interestingly, 11 out of 28 (39%) CPP3-reactive clones also bound citrullinated peptides derived from human proteins. Notably, three of these clones were positive in the clinical anti-CCP2 test, and when converted back to the predicted germline sequence, these clones became CCP2 negative, while maintaining reactivity against the bacterial CPP3 peptide. <h3>Conclusion:</h3> Our data show that B cells reactive with a citrullinated peptide derived from <i>Pg</i> PAD are present in gingival tissue, lungs, bone marrow, blood and the inflamed joint of ACPA+ RA patients. Moreover, the finding that a number of these clones are cross-reactive with citrullinated peptides derived from human proteins as well as the gold standard CCP2 test suggests mechanisms of molecular mimicry in the generation of ACPA. Importantly, the germline versions of these ACPA were <i>Pg</i>-reactive but not autoreactive, supporting the hypothesis of a bacterial origin for this ACPA response. <h3>Disclosure of Interests:</h3> Natalia Sherina: None declared, Vijay Joshua: None declared, Radha Thyagarajan: None declared, Natalie Sippl: None declared, Lena Israelsson: None declared, Heidi Wähämaa: None declared, Ragnhild Stålesen: None declared, Kaja Eriksson: None declared, Tülay Yucel-Lindberg: None declared, Aase Hensvold: None declared, Caroline Grönwall: None declared, Anca Catrina Grant/research support from: Yes, but not for the presented study., Vivianne Malmström: None declared, Antonio Lanzavecchia: None declared, Luca Piccoli: None declared, Khaled Amara: None declared, Karin Lundberg: None declared