Impaired early B-cell tolerance in patients with rheumatoid arthritis

Purpose

Autoantibody production is a characteristic of most autoimmune diseases including rheumatoid arthritis (RA), but the stage of B-cell development at which B-cell tolerance is broken remains unknown. We previously established in healthy donors that most autoreactive developing B cells were removed either in the bone marrow or in the periphery, revealing two B-cell tolerance checkpoints. We aimed to determine whether the autoreactive B cells in RA evade one or both of these checkpoints. In addition, analysis of the antibody repertoires in RA might lead to potential insights about the mechanisms that may explain loss of B-cell tolerance.

Methods

We obtained peripheral blood from four patients with clinically and serologically documented active RA, who had not yet received steroids, disease-modifying anti-rheumatic drugs or biologic therapy, or who had been off such regimens for at least 4 months. After isolating their peripheral B cells, we sorted, cloned and expressed in vitro antibodies from single new emigrant (CD19⁺CD10⁺IgM⁺CD27^-) and mature naive (CD19⁺CD10^-IgM⁺CD27^-) B cells. In total, we examined by ELISA the specificity of 232 recombinant antibodies.

Results

The antibody sequences in two of the patients showed a significant increase in the proportion of 11 amino-acid long CDR3s in IgK chains (previously described in RA synovium and blood), while the other two patients demonstrated an increase in Jκ1 usage along with a shift towards more downstream Vκ segments. This skewed IgK repertoire suggests that receptor editing, one of the mechanisms that ensures B-cell tolerance, is defective in those two patients. Despite the different antibody sequence patterns of our RA patients, all four patients showed significantly increased proportions of polyreactive antibodies in the new emigrant compartment (23.1% versus 8.2% in controls, P = 0.0094), autoreactive antibodies in the mature naïve compartment (45.3% versus 20.4% in controls, P = 0.0001), and anti-CCP antibodies in both compartments (13.8% versus 0% in controls, P = 0.019). This demonstrates that autoreactive B cells in RA patients fail to be removed at both checkpoints of early B-cell tolerance.

Conclusions

RA patients exhibit defective central and peripheral B-cell tolerance, which may favor the development of autoimmunity. We have also identified two distinct groups of RA patients based on their antibody sequences that suggest divergent mechanisms within the RA population – yet lead to a common phenotype.