类风湿关节炎患者滑膜抗环瓜氨酸肽表位表达与Thl7／调节性T细胞的失衡和滑膜炎症的相关性分析

Association of synovial cyclic citrullinated peptide expression with Thl7/Treg imbalance and synovitisin rheumatoid arthritis patients

目的分析类风湿关节炎（RA）患者滑膜抗环瓜氨酸肽（CCP）抗体表位的表达情况与Thl7／调节性T细胞的失衡、滑膜炎症和临床表现的相关性。方法抗CCP抗体单链抗体（ScFv）通过噬菌体蛋白展示技术构建。以6xHisoTag免疫组织化学法检测39例RA患者和35例对照组关节滑膜抗CCP抗体的表达情况。流式细胞术（FCM）检测外周血单个核细胞（PBMCs）中Thl7闹节性T细胞的百分比，酶联免疫吸附试验（ELISA）法检测Thl7／调节性T细胞相关细胞因子。RA滑膜病理及临床病情活动性评分纳入分析。主要采用，检验、单因素方差分析进行统计学处理。结果①抗CCP抗体表位在RA组与对照组表达率分别为76．9％和11．4％，差异有统计学意义X^2=31．9，P〈0．01）；②滑膜抗CCP抗体表位表达阳性的RA组患者外周血Thl7细胞，Thl7／i）~节性T细胞比值，Thl7相关细胞因子白细胞介素（IL）－6，IL．17a，IL．23，肿瘤坏死因子（TNF）α，调节性T细胞相关细胞因子转化生长因子（TGF）-β_1，以及外周血和滑液中抗CCP抗体水平均高于滑膜抗CCP抗体表位表达阴性的RA组；③滑膜抗CCP抗体表达阳性RA患者病情活动性DAS28以及病理滑膜细胞增生、淋巴细胞聚集和弥漫性淋巴细胞浸润评分高于抗CCP抗体表达阴性的RA患者。结论抗CCP抗体表位表达与Thl7／调节性T细胞的失衡和滑膜炎症存在密切联系，可能在RA发病机制中发挥作用。

Objective To assess the association of synovial cyclic citrullinated peptide（CCP）expres- sion with T helper 17 （Th 17 ） cells/Regulatory T cells （Treg） imbalance, the histological and clinical features of synovitis in rheumatoid arthritis （RA）. Methods CCP expression in synovial specimens from 39 patients with RA and 35 controls was detected by immunohistochemistry assay（IH） using 6xHis tagged anti-CCP single chain fragment V （ScFv） antibodies, which were generated by pHEN2 phagemid recombinant antibodies display system. The frequencies of Thl7/Treg cells in the peripheral blood mononuclear cells （PBMCs） were determined by flow cytometry （FCM）. Thl7frreg cells associated cytokines were analyzed by enzyme-linked immunosorbent assay （ELISA）. The histological scores and clinical features of synovitis were included in the study. Chi-square test and ANOVA were used for statistical analysis. Results （j~） The prevalences of synovial CCP expression were significantly different between RA group and the control（76.9% and 11.4% respectively, X2=31.9, P〈0.01）. （~） The frequencies of Thl7 cells, Thl7/Treg ratio, Thl7 cells associated cytokines as IL-6, IL-17a, IL-23, TNF-o~, and the Treg cells associated cytokines TGF-[3~, the serum and synovial fluid antiCCP＇antibodies in the RA patients with synovial CCP positive expression were significantly higher in RA patients with CCP positive than those with CCP negative. Disease activity score DAS28 index and histological features quantified variations of the synovial biopsy specimens （synoviocyte hyperplasia, focal aggregates oflymphocytes, and diffuse infiltrates of lymphocytes） in RA were higher in synovial CCP positive expression patients than in the negative. Conclusion Synovial CCP expression is strongly associated with the Thl7/Treg imbalance and synovitis, which may play a crucial role in the pathogenesis of RA.