二藤合剂氯仿提取物对类风湿性关节炎T细胞的免疫调节作用

【目的】观察二藤合剂氯仿提取物对活化的类风湿性关节炎(RA)患者T细胞表达CD69以及Jurkat细胞周期的影响．推测其可能的细胞免疫调控机制。【方法】培养Jurkat细胞，加入二藤合剂各提取物培养72h，四甲基偶氮唑盐(MTT)法检测增殖率，筛选有效部位；无菌抽取RA患者全血，与不同浓度二藤合剂氯仿提取物或染料木黄酮预孵30min后，再加植物血凝素(PHA)培养24h，采用流式细胞术检测T淋巴细胞CD69的表达率；培养Jurkat细胞，加不同浓度的二藤舍剂氯仿提取物或氨甲喋呤(MTX)继续培养48h，流式细胞术检测细胞周期。【结果】二藤合剂各提取物中以氯仿提取物抑制Jurkat细胞增殖作用最强；10mg／L和20mg／L的二藤合剂氯仿提取物可显著抑制PHA刺激下RA患者的T细胞，尤其是CD4T细胞表达CD69；10mg／L浓度的氯仿提取物主要将细胞增殖周期抑制在G1期，与MTX作用于S期有明显区别。【结论】二藤合剂氯仿提取物可抑制RA患者T细胞尤其是CD4T细胞的早期活化，阻止T细胞能量和原料合成，从而阻断其DNA合成过程，减少细胞向G2期的转化，为其用于RA的治疗提供了实验依据。

Immunoregulation of Chloroform Component Extracted from Erteng Mixture on T Lymphocytes in Rheumatoid Arthritis

Objective] To investigate the effect of Erteng Mixture (EM) on activated T-cell CD69 expression and Jurkat cell cycle in rheumatoid arthritis (RA) and to explore its possible cellular immunoregulation mechanism. Methods] Jurkat cells were cultured with different kinds of components extracted from EM. After 72 hours, proliferation rate of Jurkat cell were detected with thiazolyl blue colorimetry (MTT) to screen the active component. Whole blood from RA patients were incubated in RPMI-1640 culture with various concentrations of chlorofonn component extracted from EM or with Genistein. Thirty minutes later, phytahematoagglutinin (PHA) were added successively into the culture. After 24 hours, CD69 expression rate of T lymphocytes activated by PHA was analyzed by flow cytometry. Jurkat cells were cultured with various concentrations of chloroform component extracted from EM and with methotrexate ( MIX) for 48 hours. After then, Jurkat cell cycle was analyzed by flow cytometry. Results ] Chloroform component extracted from EM had the strongest inhibitive effect on proliferation of Jurkat cells. Chloroform component (10 mg/L and 20 mg/L) could markedly inhibit CD69 expression on PHA-activated T lymphocytes. Chloroform component 10 mg/L blocked Jurkat cell cycle at G1 phase, which differed from MTX blocking Jurkat cell cycle at S phase. Conclusion] Chloroform component extracted from EM could significantly inhibit the early activation of CD4 T lymphocytes in RA and inhibit the energy and material synthesis in T-cell, thus hindering DNA synthesis and decreasing the cellular G2 phase transformation. This study will provide an experimental basis for clinical application of EM in treating RA.