目的：检测外周血滤泡辅助性T细胞(follicular T helper cells , Tfh cells)的比例及其表面标志，分析与类风湿关节炎(rheumatoid arthritis , RA)患者疾病活动度的关系。方法： 收集40例类风湿关节炎患者及20例正常对照组的外周血，分离外周血单个核细胞(peripheral blood mononuclear cells,  PBMC)及血清，流式细胞术检测PBMC中CD4+CXCR5+Tfh细胞(CXCR5, 趋化因子受体, C-X-C chemokine receptor type 5)的比例及其表面程序性细胞死亡1(programmed death-1, PD-1)、可诱导的协同刺激性因子(inducible co-stimulator, ICOS)、CD40配体(CD40 ligand, CD40L)及白介素21受体（interleukin-21  receptor, IL-21R）的表达情况，实时荧光定量PCR方法检测B 细胞淋巴瘤-6(B-cell lymphoma 6, Bcl-6)、白介素-21(interleukin-21, IL-21)及IL-21R的表达，酶联免疫吸附法(enzyme-linked immunosorbent assay , ELISA)检测血清中IL-21和B 细胞趋化因子CXC配体13 (B cell-attracting chemokine CXC ligand 13, CXCL13)水平。结果： （1）流式细胞术结果提示，与正常对照组比较，RA患者组外周血CXCR5+ CD4+Tfh细胞表达与正常对照组相比明显升高（16.75±3.92  vs. 7.49±1.84，P＜0.001）；低疾病活动度或缓解组RA外周血CXCR5+ CD4+Tfh细胞表达与正常对照组相比明显升高（16.62±3.43 vs. 7.49±1.84，P＜0.001）；中疾病活动度组RA外周血CXCR5+ CD4+Tfh细胞表达与正常对照组相比明显升高(16.82±3.07 vs. 7.49±1.84, P＜0.001)；高疾病活动度组RA外周血CXCR5+ CD4+Tfh细胞表达与正常对照组相比明显升高(16.87±5.50 vs. 7.49±1.84, P＜0.001)；RA患者外周血ICOS、PD-1、IL-21R在CD4+CXCR5+细胞的表达与正常对照组相比显著升高 (ICOS+ CXCR5+CD4+细胞, 8.37±4.28 vs. 3.72±1.81, P＜0.001; PD-1+ CXCR5+CD4+细胞, 1.57±1.10 vs. 0.24±0.30, P=0.035; IL-21R+ CXCR5+CD4+细胞, 4.60±4.05 vs. 0.20±0.19, P=0.006)，而CD40L在CD4+CXCR5+细胞的表达在两组间差异无统计学意义(3.38±3.71 vs. 0.54±0.34, P=0.135)。（2）RA患者组较正常对照组外周血中IL-21R mRNA升高(5.00±4.94 vs. 0.74±0.55, P<0.001)，而转录因子Bcl-6 mRNA ［4.54(3.33, 7.23) vs. 5.31(2.81, 7.44), P=0.329］以及IL-21 mRNA ［0.72(0.26, 3.45) vs. 0.56(0.27, 3.71) P=0.195］表达差异无统计学意义（P>0.05）；Bcl-6、IL-21和IL-21R mRNA的表达在不同组RA患者之间差异也无统计学意义（P>0.05）。（3）RA患者血清中IL-21、CXCL13水平明显高于正常对照组［IL-21, (200.49±154.56) ng/L vs. (8.21±5.95) ng/L, P<0.001; CXCL13, (43.09±1.28)ng/L vs. (93.72±49.72) ng/L,P<0.001］；相关分析显示IL-21、CXCL13浓度与RA疾病活动度正相关（r>0.4），而血清类风湿因子(rheumatoid factor, RF)和抗环瓜氨酸多肽抗体（anti-citrullinated protein antibody, anti-CCP）与RA疾病活动度无相关性(r<0.2)。结论： 外周血Tfh细胞及其相关细胞因子在RA患者中显著升高，说明Tfh细胞可能参与RA的发病机制，阻断Tfh细胞的产生，有望成为治疗RA患者的新的作用靶点。

Objective： To detect cell frequency and surface markers of peripheral CD4+CXCR5+ follicular helper T (Tfh) cells and analyze the correlation between CD4+CXCR5+Tfh cells and rheumatoid arthritis (RA) disease activity. Methods: Forty RA patients meeting the American College of Rheumatology classification criteria for RA and twenty healthy controls (HC) were included. The peripheral blood mononuclear cells and sera were collected. The expressions of CD4+CXCR5+Tfh cells (CXCR5, C-X-C chemokine receptor type 5) and inducible T cell costimulator (ICOS), programmed death 1 positive (PD-1), interleukin-21 receptor (IL-21R) and CD40 ligand (CD40L) positive on CD4+CXCR5+Tfh cells were analyzed by flow cytometry. The transcript levels of B-cell lymphoma 6 (Bcl-6), as well as IL-21 and IL-21R, were measured by real-time polymerase chain reaction. Besides, serum IL-21 and CXCL13 concentrations were determined by enzyme-linked immunosorbent assay. The potential association between Tfh cells and RA disease activity was detected. Results: The cell surface marker of CXCR5+ on CD4+ cells was significantly increasingly higher across the following groups versus HC: total RA patients (16.75±3.92 vs.7.49±1.84, P＜0.001); RA patients with low disease activity or remission (16.62±3.43 vs. 7.49±1.84, P＜0.001); RA patients with moderate disease activity (16.82±3.07 vs. 7.49±1.84, P＜0.001) and RA patients with high disease activity (16.87±5.50 vs. 7.49±1.84, P＜0.001). Besides, the percentages of ICOS+, PD-1+, IL-21R+ on CD4+CXCR5+Tfh cells in the RA patients were significantly higher than that of HC (ICOS+CD4+CXCR5+cells, 8.37±4.28 vs. 3.72±1.81, P＜0.001; PD-1+CD4+CXCR5+cells, 1.57±1.10 vs. 0.24±0.30, P=0.035; IL-21R+CD4+CXCR5+cells, 4.60±4.05 vs. 0.20±0.19, P=0.006). But the percentage of CD40L+ on CXCR5+CD4+Tfh cells in the RA patients was not significantly higher than that of HC (3.38±3.71 vs. 0.54±0.34, P=0.135). The IL-21R mRNA expression was elevated significantly (5.00±4.94 vs. 0.74±0.55, P<0.001) in the RA patients but not in Bcl-6 mRNA［4.54(3.33, 7.23) vs. 5.31(2.81, 7.44), P=0.329］or IL-21 mRAN［0.72(0.26, 3.45) vs. 0.56(0.27, 3.71), P=0.195］. Additionally, the serum interleukin-21 (IL-21)and CXCL13 levels in the RA patients were higher than in the healthy controls ［IL-21, (200.49±154.56) ng/L vs. (8.21±5.95) ng/L, P<0.001; CXCL13, (93.72±49.72) ng/L vs. (43.09±1.28) ng/L, P<0.001］ and were both positively correlated with RA disease activity indexes, including erythrocyte sedimentation rate, the disease activity score in 28 joints (ESR-based or CRP-based), clinical disease activity index, and simplified disease activity index. However, none of the Tfh cells, anti-citrullinated protein antibody or rheumatoid factor had any relationship with RA disease activity. Conclusion: Peripheral Tfh cells and their relevant cytokines are higher in RA patients than healthy controls, indicating Tfh cells may participate in the pathogenesis of RA, therefore, blocking the pathway of Tfh cells may be reasonable cellular targets for therapeutic intervention.