Association of smoking with the constitution of the anti–cyclic citrullinated peptide response in the absence of HLA–DRB1 shared epitope alleles

Objective

Smoking is a risk factor for anti–cyclic citrullinated peptide (anti‐CCP) antibody–positive rheumatoid arthritis (RA) in patients with HLA–DRB1 shared epitope (SE) alleles. It is unknown whether smoking influences not only the presence of these antibodies, but also other characteristics of the anti‐CCP response, such as isotype usage. The aim of this study was to determine the influence of smoking on anti‐CCP isotypes in RA patients, and to determine whether this influence is observed in the presence and/or absence of SE alleles.

Methods

IgA, IgM, and IgG subclasses of anti‐CCP antibodies were measured by enzyme‐linked immunosorbent assay in serum obtained at the first visit to the Leiden Early Arthritis Clinic from 216 patients with anti‐CCP–positive RA whose smoking habits were also assessed. HLA genotyping data were available for 202 of these patients.

Results

IgA and IgM anti‐CCP were more frequent in RA patients who were smokers than in those who were nonsmokers (odds ratio 2.8 and 1.8, respectively). In addition, levels of all isotypes of anti‐CCP, except IgG3, were significantly higher (P < 0.05) in smokers. The number of anti‐CCP isotypes was higher in smokers compared with nonsmokers, both in SE‐negative RA (P = 0.04) and in SE‐positive RA (P = 0.07).

Conclusion

Patients with anti‐CCP–positive RA who have a current or former tobacco exposure display a more extensive anti‐CCP isotype usage in general, and IgA and IgM in particular, compared with patients with anti‐CCP–positive RA who have never smoked. In contrast to its influence on the incidence of anti‐CCP positivity, the influence of tobacco exposure on the constitution of the anti‐CCP response is significant in SE‐negative RA. These findings suggest a differential effect of tobacco exposure on the induction as compared with the propagation of the anti‐CCP antibody response.

     

Spondyloarthritis mass cytometry immuno-monitoring: a proof of concept study in the tight-control and treat-to target TiCoSpA trial

Objective

Mass cytometry (MC) immunoprofiling allows high-parameter phenotyping of immune cells. We set to investigate the potential of MC immuno-monitoring of axial spondyloarthritis (axSpA) patients enrolled in the Tight Control SpondyloArthritis (TiCoSpA) trial.

Methods

Fresh, longitudinal PBMCs samples (baseline, 24, and 48 weeks) from 9 early, untreated axSpA patients and 7 HLA-B27⁺ controls were analyzed using a 35-marker panel. Data were subjected to HSNE dimension reduction and Gaussian mean shift clustering (Cytosplore), followed by Cytofast analysis. Linear discriminant analyzer (LDA), based on initial HSNE clustering, was applied onto week 24 and 48 samples.

Results

Unsupervised analysis yielded a clear separation of baseline patients and controls including a significant difference in 9 T cell, B cell, and monocyte clusters (cl), indicating disrupted immune homeostasis. Decrease in disease activity (ASDAS score; median 1.7, range 0.6–3.2) from baseline to week 48 matched significant changes over time in five clusters: cl10 CD4 T_nai cells median 4.7 to 0.02%, cl37 CD4 T_em cells median 0.13 to 8.28%, cl8 CD4 T_cm cells median 3.2 to 0.02%, cl39 B cells median 0.12 to 2.56%, and cl5 CD38⁺ B cells median 2.52 to 0.64% (all p<0.05).

Conclusions

Our results showed that a decrease in disease activity in axSpA coincided with normalization of peripheral T- and B-cell frequency abnormalities. This proof of concept study shows the value of MC immuno-monitoring in clinical trials and longitudinal studies in axSpA. MC immunophenotyping on a larger, multi-center scale is likely to provide crucial new insights in the effect of anti-inflammatory treatment and thereby the pathogenesis of inflammatory rheumatic diseases.

CD25+ cell depletion hastens the onset of severe disease in collagen‐induced arthritis

Objective

CD4+,CD25+ T regulatory cells may offer opportunities to intervene in the course of autoimmune disease. We wished to evaluate their potential for influencing systemic and chronic joint inflammation by investigating their involvement in collagen‐induced arthritis (CIA).

Methods

We depleted DBA/1 mice of CD25+ regulatory cells by injection of a depleting monoclonal antibody specific for CD25 14 days before a single immunization with type II collagen (CII) in Freund's complete adjuvant. CD4+,CD25+ T cells were adoptively transferred to some groups of mice during immunization. Mice were then scored for signs of arthritis, and blood was taken periodically to measure the amounts of CII‐specific antibodies. Splenocytes of treated mice were examined in vitro to determine the effects of depletion on proliferation to CII and control antigens.

Results

CD25+ cell–depleted DBA/1 mice had significantly more severe disease than control mice following collagen immunization. The magnified severity was also accompanied by higher antibody titers against collagen, and in vitro tests showed increased proliferation of collagen‐specific T cells. Adoptively transferring CD4+,CD25+ T cells into depleted mice was shown to reverse the heightened severity. Control mice, which were depleted and immunized with the neoantigen keyhole limpet hemocyanin (KLH), had neither an increased antibody response toward KLH nor an augmented proliferative response, indicating that CD25+ cell depletion preferentially affects immunity against self antigen.

Conclusion

These results establish a link between CD4+,CD25+ regulatory cells and CIA and provide a rationale for investigating CD4+,CD25+ T regulatory cells in the treatment and prevention of arthritis.