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AB0046 Cytokine profile in CD4+ FOXP3+ lymphocytes of RA patients treated with tocilizumab and clinical correlation (DAS28-CPR, rheumatoid factor
  1. F.J. Navarro-Blasco1,
  2. J.M. Sempere2,
  3. A. Romero3
  1. 1Reumatología, Hospital General Universitario Elche, Alicante
  2. 2Inmunology Division, Alicante University, Alicante
  3. 3Hematology and Cytometry, Hospital General Universitario Elche, Elche. Alicante, Spain


Background The main group of suppressor cells/regulators of the immune response (Treg cells), usually co-expressed different combinations of antigens CD4, CD25, and FoxP3, among other markers (1).A decrease in the number-function of this population in various autoimmune diseases, suggesting a role of it for potential use as a therapeutic target, diagnosis or prognosis, especially since their relationship or conversion to Th17, depending on the existing pattern of cytokines in their environment (2).

Objectives To identify the pattern of intracellular cytokines (IL6, IL10, IL17) and the membrane antigen CD152 (CTLA-4) in CD4+ FoxP3+ cells, in RA patients treated with tocilizumab (as usual protocol) and determine its correlation with the state of disease activity (DAS28-CRP) and rheumatoid factor (RF).

Methods Included 19 RA (fulfilling criteria ACR/EULAR) in standard treatment with tocilizumab (8mgr/kg iv/4w) for more than 3 months and 1 year of minimum evolution since diagnostic. We obtained whole blood samples in EDTA tubes immediately before dosing the corresponding monthly. We analyzed 6 samples of healthy individuals as control group. The analysis was performed by flow cytometry and direct immunofluorescence, using combinations of monoclonal anti-IL6-17-10-152-FoxP3 antiCD4 (BD Bioscience), combined with AX488, FITC, PE, and PerCPCy5.5. Descriptive analysis was performed, using t test bivariate tests and study of normal (normal null hypothesis p-values>0.05). Correlation between variables and scatter plot.

Results The patients (19 RA, 17 FR+, 15 DAS28 <2.3 and 4 DAS28>2.3) obtained mean percentage of CD4+ IL10+, CD4+ FoxP3+ IL10+ and CD4+ FoxP3+ IL10+ IL17+ > to that of healthy controls (37.12 vs. 25.6, 75.69 vs. 54.33, 59.8 vs. 43.78). The values shown inactive patients CD4+ FoxP3+ IL6+ and CD4+ FoxP3+ CD152+ IL6+ > to assets (76.31 vs. 49.52, 61.7 vs. 43.99). In patients FR+ vs. FR-, the difference is in the CD4+ IL17+ (35.8 vs. 31.1)

Conclusions Patients have a higher IL10 + Treg cells as a possible attempt to control the abnormal immune response. The increased CD4+ FoxP3+ IL17+ IL10+ in patients, would indicate a possible transformation of Treg in Th17 cells (involved in the pathogenesis of autoimmune diseases). Higher levels of CD4+ IL17+, in FR+ patients, would go along these lines. Higher levels of CD4 + FoxP3+ IL6+ and CD4+ IL6+ FoxP3+ CD152+ in inactive patients (DAS28 <2.3), which would indicate, a priori, that the immune mechanism in the few patients assets be led not only by IL-6. Meanwhile, higher levels in patients inactive CD152 (CTLA-4), an essential factor for exercise immunosuppression in Treg cells, would favor the recovery of suppressor function under treatment with tocilizumab. Our hypothesis is that Treg cells are involved in the autoimmune process and that the removal of some cytokines such as IL6, would in most cases to normalize its functioning activity. While a smaller proportion of ARs remain the evolving capacity of Treg cells to Th17 cells, maintaining the inflammatory activity.

  1. Valencia X et al. CD4 + CD25 + Foxp3 + regulatory T cell in autoimmune heatsink. Nat Clin Rheumatol Prat 2007, 3:619-26)

  2. KS Voo et al. Identification of IL17-producing FOXP3 + Regulatory T cells in humans.Proc Nati Acad Sci USA 2009, 106:4793-8

Disclosure of Interest F. Navarro-Blasco Grant/Research support from: RESEARCH SUPPORT ROCHE LAB., J. SEMPERE: None Declared, A. ROMERO: None Declared

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