Background/Purpose:

Mechanisms regulating the autoimmune response in rheumatoid arthritis (RA) are not well understood. However, it is known that T CD4⁺ lymphocytes play a pivotal role initiating and perpetuating the synovial and systemic chronic inflammation found in the disease. Methotrexate (MTX) is the most commonly used Disease-modifying antirheumatic drugs (DMARD) in RA patients. Regrettably, less than thirty percent of these patients do not respond to MTX and need to initiate additional treatments. The potential role of CD4⁺ T lymphocytes in the MTX clinical response in early RA remains unknown.

The objective of this study is to evaluate the relevance of the pattern of IFNγ, IL-4 and IL-17A production by naïve (T_N), central memory (T_CM), non-terminated effector memory (T_NTEM) and terminated effector memory (T_TEM) CD4⁺ T cells for the clinical response to MTX in recently diagnosed DMARD naïve RA patients.

Methods:

The number of IFNγ, IL-4, and IL-17A producing CD4⁺ T lymphocytes, and in their T_N, T_CM, T_NTEM and T_TEM subsets in fourty DMARD naïve recently diagnosed RA patients were assayed using a multiparametric flow cytometry. The patients were treated with weekly MTX and the clinical response to the treatment was established after six month of MTX follow up. The patients were classified as responders or non-responders. We also studied twenty-five age and sex-matched healthy subjects as controls. Peripheral blood mononuclear cells (PBMC) were obtained and stimulated during six hour with phorbol-myristate-acetate and ionomycin. To study the intracellular cytokine production by CD4+ T lymphocyte subsets, we used monoclonal antibodies specific for the surface antigens CD3, CD4, CD45RA, CD27 and cells were fixed and permeabilized, and simultaneously stained with IFNγ, IL-4, and IL-17 intracellular cytokines. We acquired in a FacsAria-II flow cytometer and analyzed by FacsDiva and Flow-Jo software.

Results:

At basal pretreatment conditions, MTX non-responder RA patients showed a significant increased number of CD4⁺IL-17⁺T lymphocytes with respect to responder RA patients, explained by an expansion of the CD4⁺IL-17⁺T_N and CD4⁺IL-17⁺T_CM subsets. The number of CD4⁺IFNγ⁺ T lymphocytes was significantly increased in MTX non-responder RA patients with respect to that of responders. This elevated CD4+IFNγ+T lymphocyte number was due to an increase in the CD4⁺IFNγ⁺ T_N, CD4⁺IFNγ⁺ T_CM and CD4⁺IFNγ⁺ T_NTEM subset numbers found in non-responder RA patients. There were not significant differences in the CD4⁺IL-4⁺T lymphocyte numbers between MTX responder and non-responder AR patients at basal conditions.

Conclusion:

Recently diagnosed DMARD naïve RA patients who eventually do not respond to MTX treatment show an abnormal increased numbers of the circulating IL-17A and IFNγ producing T_N and T_CM CD4⁺ lymphocytes with respect to MTX responder RA patients. These IL-17A and IFNg abnormalities in naïve T CD4 lymphocytes might be a good biomarker for the clinical response to MTX in DMARD naïve recently diagnosed RA patients.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/the-pattern-of-proinflamatory-cytokine-expression-by-cd4-t-lymphocytes-segregates-the-clinical-response-to-methotrexate-in-recently-diagnosed-rheumatoid-arthritis-patients/