Abstract and Introduction
Abstract
Introduction: The purpose of this study was to quantitatively evaluate the contribution of synovial lymphoid aggregates to autoantibody (rheumatoid factor [RF] and anti-cyclic citrullinated peptide [anti-CCP]) and total immunoglobulin (IgG and IgM) production in rheumatoid arthritis (RA) patients and the effect thereon of the B-cell-depleting antibody, rituximab, in the ARISE (Assessment of Rituximab's Immunomodulatory Synovial Effects) trial.
Methods: Autoantibodies as well as total IgM and IgG were quantified by enzyme-linked immunosorbent assay in extracts of synovial tissues and matched serum from patients with RA or osteoarthritis (OA). Synovial biopsies and serum were obtained at baseline and 8 weeks following rituximab therapy in 14 RA patients. A synovial/serum index (SSI) was calculated as the ratio of synovial to serum antibody/albumin, with values above 1 representing synovial enrichment. Lymphoid aggregates were evaluated histologically.
Results: Anti-CCP IgG, but not RF-IgM, was significantly enriched in RA synovia compared with serum. Total IgM and IgG were also enriched in RA, but not in OA. SSI correlated significantly with mRNA content for both IgM and IgG, demonstrating that it reflected synovial immunoglobulin production. RA synovia with lymphocyte aggregates contained significantly elevated RF-IgM and anti-CCP IgG compared with tissues with diffuse lymphoid infiltration. Rituximab treatment did not affect synovial autoantibody or total immunoglobulin SSI overall. However, in aggregate-containing tissues, rituximab significantly reduced total IgM and IgG SSI as well as IgM and IgG1 mRNA. Surprisingly, RF-IgM and anti-CCP IgG SSIs were unchanged by rituximab in aggregate-containing synovia.
Conclusions: Combined with earlier observations that synovial lymphoid aggregates are unaltered by rituximab treatment, these data suggest that lymphoid aggregates may provide a protective niche for autoantibody-producing cells.
Trial Registration: The ARISE trial is registered at ClinicalTrials.gov as number NCT00147966.
Introduction
Rheumatoid arthritis (RA) is associated with the presence of certain circulating autoantibodies, such as rheumatoid factors (RFs) and anti-cyclic citrullinated peptide (anti-CCP). The latter has received recent attention because elevated levels can precede development of joint symptoms and because it acts synergistically with the shared HLA-DR epitope to enhance the risk of developing RA. A contribution of B cells and their products to the pathogenesis of RA is supported by the clinical success of rituximab, a B-cell-depleting antibody targeting CD20. Whereas long-lived plasma cells are unaffected by rituximab, circulating B cells are nearly completely depleted and modest, albeit significant, decreases in circulating RF and anti-CCP antibodies are observed. The effect of rituximab on the rheumatoid synovium is just beginning to be characterized. Recently, we and others reported that, following rituximab treatment, synovial B cells are depleted less effectively, and more variably, than their circulating counterparts. In the subset of patients with synovial lymphoid aggregates, rituximab treatment did not alter the number or size of these aggregates. Because such aggregates are associated with elevated synovial immunoglobulin synthesis, as determined by mRNA levels for IgG constant regions, and perhaps also autoantibody synthesis, we sought to determine the effect of rituximab treatment on synovial autoantibody production.
The local synthesis of immunoglobulins and autoantibodies by rheumatoid synovium is well appreciated but its contribution to the circulating pool is poorly understood. Explants of rheumatoid synovial tissue are capable of synthesizing immunoglobulins, RF, and anti-CCP IgG. Similarly, dispersed cells from rheumatoid synovia synthesize immunoglobulins and RF, and synovial fluid-derived mononuclear cells secrete anti-CCP antibodies. Although these techniques are valuable for the understanding of the contribution of local antibody synthesis to the pathogenesis of RA, their applicability in interventional biopsy-based clinical trials is limited. Synovial tissues obtained by arthroscopy or needle biopsy typically do not yield enough tissue to recover a sufficient amount of dispersed cells, and the viability of synovial biopsies for explant cultures might be compromised when samples have to be transported from clinical sites to the laboratory. With this in mind, we developed and validated a novel set of techniques that can be used on frozen specimens for the measurement of autoantibodies and immunoglobulins in paired synovial biopsies and sera obtained prior to, and following, an intervention. These methods were used to evaluate the effect of rituximab treatment on synovial autoantibody and immunoglobulin production and the role of lymphoid architecture on this effect.