Analysis of the antigen‐specific T cell response in reactive arthritis by flow cytometry

Objective In reactive arthritis (ReA) a bacteria‐specific T cell response to the triggering microbe is detected in synovial fluid (SF), and an impaired Th1 cytokine response has been described. The recent identification of immunodominant bacterial proteins/peptides and new technologies make a more detailed analysis of the immune response possible. The aim of the present study was to use these new techniques to determine the antigen‐specific T cell frequency and the cytokine secretion pattern on stimulation with bacteria‐derived recombinant proteins in the peripheral blood (PB) and SF from patients with ReA. Methods In 3 patients with Chlamydia‐induced ReA and 2 patients with Yersinia‐induced ReA, the SF T cell response was investigated after stimulation with the Chlamydia‐derived proteins major outer membrane protein (MOMP) and heat‐shock protein 60 (Hsp60) and the Yersinia‐derived proteins 19‐kd protein and Hsp60. In 3 of these patients, the PB T cell response was investigated in parallel. T cells were stimulated in whole blood or whole SF with antigen plus anti‐CD28 for 6 hours, brefeldin A was added after 2 hours, and cells were fixed and stained with antibodies against the surface markers CD4 and CD69 and against the cytokines interferon‐γ (IFNγ), tumor necrosis factor α, interleukin‐10 (IL‐10), and IL‐4. Positive cells were quantified by flow cytometry. Results In the 3 patients with Chlamydia‐induced ReA, the antigen‐specific T cell frequency (percentage of IFNγ CD69 double‐positive CD4+ T cells) in response to MOMP (mean ± SD 1.2 ± 1.38%) and to Hsp60 (1.21 ± 1.45%) in SF was about the same. In the 2 patients with Yersinia‐induced ReA, the mean ± SD frequency was 0.66 ± 0.36% in response to the Hsp60 and 0.3% ± 0.22 in response to the 19‐kd protein. In the 3 patients whose PB was evaluated, the corresponding T cell response was ≥10 times lower. In 2 patients with Chlamydia‐induced ReA, antigen‐specific IL‐10–positive CD4+ T cells were detected in 0.10–0.23% of the CD4+ T cell subpopulation. Conclusion The frequency of antigen‐specific T cells to Chlamydia‐ and Yersinia‐derived antigens in the SF of ReA patients is between 1:200 and 1:50. Both the chlamydial Hsp60 and MOMP are dominant T cell antigens in Chlamydia‐induced ReA. In patients with Chlamydia‐induced ReA, we detected antigen‐specific IL‐10 secretion, which might mediate an inhibition of effective bacterial clearance.