Effect Of TNF-α Antagonists On Functional Status Of T Helper Cells In Rheumatoid Arthritis

【Background】Rheumatoid arthritis (RA) is a chronic, inflammatory poly-arthritis and synovitis with frequent progression to joint destruction and disability. The immunol abnormality of RA includes dysfunction of T cell subsets and abnormal activation of T cells. There are numerous evidences showed that T lymphocytes are involved in the pathogenesis of RA, including CD4+,CD8+T cells that present in synonial. Although it is still unable to understand the mechanism of autoimmune disease, it has been proved that T lymphocyte immune dysfunction, CD4+T helper cell subsets imbalance closely related. Thl and Th2 imbalance on the considerable research, but a recent study found that different from Thl and Th2 cell subsets called Th17 cells, the cell group characterized as secretion of interleukin (IL)-17 was past found in the synovial fluid of RA synovium and the high level of IL-17 expression in vitro and IL-17 in synovial cells ,which stimulate the release of TNF-α,IL-6 and other inflammatory mediators, such elements may be made in the development of RA occurred and the dominant the role of view. It is reported that the expression of Th17 cell was found in the synovial fluid of RA patients and the IL-17 in mice. However, no studies have reported RA patients before and after treatment in the peripheral blood of Th1 and Th2 and Th17 expression and changes. In the past decades, biological agents has become a major breakthrough in the treatment of RA point. Although the biological agents has a rapid development, but in RA treatment mechanisms need further study and exploration, in the light of such as cytokine production, promotion high promote IL-2, CSF and TNF-αaffinity IL-2 receptor expression, and promote T cell activation, the topics studied in patients with RA in the peripheral blood before and after biological agents treatment Th cell subsets and significance of the changes.【Objectives】The first purpose of our study is to explore the percentages of Th1,Th2,Th17 cells in peripheral blood from patients with rheumatoid arthritis (RA), and to study their correlations with the activity of the disease. Second, to ascertain the effect of TNF-αantagonists—etanercept treatment on the T-helper cells subsets (including Th1, Th2, Th17) in rheumatoid arthritis (RA). Last to analyze the relations between the changes of Th1,Th2,Th17 cells of RA patients and the disease activity, who received MTX single and etanercept combined MTX therapy. so as to in-depth study of TNF-αand Th cell subsets in the pathogenesis of RA role in providing clues.【Methods】1. Peripheral blood lymphocytes were obtained from the patients with active RA(n=10), from individuals with stable, well-controlled RA(n=20), and from healthy controls(n=10). The frequencies of CD4+IFN-γ+Th1,CD4+IL-4+Th2,CD4+IL-17+Th17 were quantified by using flow cytometry(FCM). The level of paired cytokines: IFN-γ, IL-4 and IL-17 were accessed by ELISA in that group of RA and control patients.2. Use FCM to examine the expression of IL-4, IFN-γand IL-17 on CD4+T cells and enzyme-linked immunosorbent assay (ELISA) to examine the Production of IFN-γ, IL-4 and IL-17 in PBMC between the MTX therapy group and etanercept combined therapy group.3. Records the morning stiffness, using visual analog gauge (VAS, 10cm) Evaluation of rest pain (0 to 10 cm→is no tenderness pain); counting several joint tenderness and swelling of joints (including the proximal joint hands, metacarpophalangeal joint, wrist joints, elbow, shoulder and knee, a total of 28 joints); value disease activity index (DAS28); record health assessment questionnaire (HAQ) average; to investigate the level of Anti-CCP antibody, CRP, ESR and RF isotype between the MTX therapy group and etanercept combined therapy group, and to analyze the correlation between the changes of Th1,Th2,Th17,Th1/Th2 and activities of the desease.【Results】The percentage of Th1 and Th17, intensity of Th17 and value of Th1/Th2 were higher than the healthy controls (P<0.05); After etanercept treatment the percentage and intensity of Th17 were significantly decreased (P<0.01), as well as the percentage of Th1 and value of Th1/Th2 (P<0.05), the mean fluorescence intensity(MFI) of Th1 cells before and after treatment there was no significant difference; Th2 cells positive rate and the MFI has no statistically significant difference between before and after. Th1/Th2 ratio also declined after treatment. The MTX treatment group before and after the changes in Th cell subsets was no significant difference, level of IL-17 decreased significantly but IFN-γ,IL-4 had no significant difference; The percentage of IL-17+Th17 on CD4+ T cells correlate positively to CRP and DAS28.To analyse the relation between expresstion of Th17 cell,Th1/Th2 ratio and CRP, RF subtype, anti-CCP antibody levels DAS28. The results showed that Th17 cells of RA patients with positive rate and CRP, DAS28 level was positively correlated ,but with patients in the RF subtype, anti-CCP antibody level of correlation was not significant (P>0.05), Th1/Th2 ratio of the above-mentioned clinical indicators and there is no statistically significant correlation (P>0.05).【Conclusions】 Th17 cells in Peripheral blood of active RA patients was significantly higher than that of normal controls, and the disease activity is related to Th17 suggested that the incidence of RA may be related to RA development and Th17 may be an important factor in RA development.Th17 cells may play a role in the pathogenesis of RA, the down regulation of Th17 and Th1/Th2 is significant for the evaluation of the curative effect of etanercept.This study established a Th cell subsets in particular Th17 in FCM and ELISA methods, in order to further study Th17 role in the pathogenesis of RA and the pathogenesis of RA-depth study and provide a theoretical basis and experimental basis.