Detection of autoantibodies to citrullinated BiP in rheumatoid arthritis patients and pro-inflammatory role of citrullinated BiP in collagen-induced arthritis

Hirofumi Shoda,Kazuhiko Yamamoto,Keishi Fujio,Akiko Okamoto,Mihoko Shibuya,Shuji Sumitomo,Tomohisa Okamura,Tetsuji Sawada

doi:10.1186/ar3520

Abstract

IntroductionAnti-citrullinated protein/peptide antibodies (ACPAs) are highly specific to rheumatoid arthritis (RA) patients and are thought to have a close relationship with the pathogenesis of arthritis. Several proteins, including fibrinogen, vimentin, and alpha-enolase, were reported as ACPA-target antigens, and their importance in RA pathogenesis was widely proposed. We identified citrullinated immunoglobulin binding protein (citBiP) as another ACPA target in RA patients and examined its pro-inflammatory role in arthritis.MethodsWe measured the levels of anti-citBiP, anti-BiP, and anti-cyclic citrullinated peptide (CCP) antibodies in the serum of RA patients (n = 100), systemic lupus erythematosus (SLE) patients (n = 60), and healthy controls (n = 30) using ELISA and immunoblotting. Epitope mapping was performed using 27 citBiP-derived peptides. In the mouse study, after DBA/1J mice were immunized with BiP or citBiP, serum titers of ACPAs were measured by ELISA and immunohistochemistry. The development of collagen-induced arthritis (CIA) was observed in BiP- or citBiP-pre-immunized mice.ResultsThe serum levels of anti-BiP and anti-citBiP antibodies were significantly increased in RA patients, although only anti-BiP antibodies were slightly increased in SLE patients. Interestingly, anti-citBiP antibody levels were higher than anti-BiP antibody levels in 72% of RA patients, whereas no significant increase in anti-citBiP antibody levels was detected in SLE patients and healthy controls. The serum levels of anti-CCP antibodies were correlated with those of anti-citBiP antibodies in RA patients (R2 = 0.41). Several citrulline residues of citBiP were determined to be major epitopes of anti-citBiP antibodies, one of which showed cross-reactivity with CCP. Immunization of DBA/1J mice with citBiP induced several kinds of ACPAs, including anti-CCP and anti-citrullinated fibrinogen antibodies. Pre-immunization with citBiP exacerbated CIA, and anti-CCP antibody levels were increased in citBiP-pre-immunized CIA mice.ConclusionsCitBiP is a newly described ACPA target that may play a pro-inflammatory role in arthritis.

Highlights

Anti-citrullinated protein/peptide antibodies (ACPAs) are highly specific to rheumatoid arthritis (RA) patients and are thought to have a close relationship with the pathogenesis of arthritis
Anti-citrullinated immunoglobulin binding protein (citBiP) antibodies in the serum of RA patients We first measured the serum levels of anti-binding protein (BiP) and anticitBiP antibodies in RA patients, systemic lupus erythematosus (SLE) patients, and healthy controls (Figure 1A)
Seventy-two percent of RA patients showed higher anti-citBiP antibody levels than anti-BiP antibody levels, whereas there was no significant difference in the serum titers of anti-BiP and anti-citBiP antibodies in the non

Summary

Introduction

Anti-citrullinated protein/peptide antibodies (ACPAs) are highly specific to rheumatoid arthritis (RA) patients and are thought to have a close relationship with the pathogenesis of arthritis. Several proteins, including fibrinogen, vimentin, and alpha-enolase, were reported as ACPA-target antigens, and their importance in RA pathogenesis was widely proposed. We identified citrullinated immunoglobulin binding protein (citBiP) as another ACPA target in RA patients and examined its pro-inflammatory role in arthritis. Citrullinated filaggrin is not present in the inflammatory synovium of RA patients, several citrullinated auto-antigens, including citrullinated fibrinogen, vimentin, type II collagen, and alpha-enolase, have been reported as target antigens of ACPAs in the synovia of RA patients [9,10,11,12,13,14]. In one hypothesis for the pathogenesis of RA, ACPAs bind to these citrullinated auto-antigens in the synovial tissues and form immune complexes that induce inflammatory processes [15]. Auto-antigen citrullination and ACPA development are considered as important steps in the pathogenesis of RA

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