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目的:明确通痹合剂抑制T淋巴细胞活化的活性部位,研究其对T细胞双信号分子的影响,阐明通痹合剂免疫调控作用的机制。方法:分离大鼠淋巴细胞,加入通痹合剂不同提取部位,ConA刺激48 h,流式细胞术(FCM)检测CD3阳性细胞表达CD71;加入不同浓度通痹合剂乙酸乙酯部位和甲氨蝶呤(MTX),ConA刺激48 h,FCM检测T细胞表达TCR、CD28和ICOS。结果:ConA刺激后CD3阳性细胞表达CD71显著上调至69.7%,通痹合剂乙酸乙酯部位(TBHJ)可抑制CD3+CD71+表达(32.5%);ConA刺激T细胞表达TCR、CD28和ICOS明显升高,与阴性对照组(Control)有非常显著性差异(P<0.001),TBHJ可明显抑制T细胞表达TCR、CD28和ICOS,尤其抑制ICOS作用较强,呈浓度依赖效应;MTX可明显抑制T细胞表达TCR、CD71。结论:TBHJ可明显抑制T淋巴细胞活化,其对T细胞活化的双信号分子途径均有下调作用,尤其以抑制ICOS明显,提示TBHJ可通过阻断CD28-ICOS第二信号途径,诱导免疫耐受。本研究为通痹合剂治疗类风湿性关节炎等自身免疫性疾病提供了实验依据。
OBJECTIVE: To clarify the effect of Tongbi Mixture on inhibiting the activation of T lymphocytes and to study the effect of Tongbi Mixture on T-cell dual signal molecules, and to elucidate the mechanism of Tongbi Mixture’s immune regulation. Methods: Lymphocytes were isolated from rats and added with different extraction sites of Tongbai Mixture. After stimulated by ConA for 48 h, CD71 positive cells were detected by flow cytometry (FCM), and CD71 was expressed. Ethyl acetate and methotrexate (MTX), ConA stimulation 48 h, TCR, CD28 and ICOS were detected by FCM. Results: The expression of CD71 in CD3 positive cells was significantly up-regulated to 69.7% after ConA stimulation, and the expression of CD3 + CD71 + was down-regulated by TBHJ in ConA-stimulated cells (P <0.05) (P <0.001). TBHJ could significantly inhibit the expression of TCR, CD28 and ICOS in T cells, especially in a concentration-dependent manner. Inhibition of T cells by MTX Expression of TCR, CD71. CONCLUSION: TBHJ can significantly inhibit the activation of T lymphocytes, which has a downregulation of the dual signaling pathways that activate T cells, especially ICOS inhibition, suggesting that TBHJ can induce immune tolerance by blocking the second signal pathway of CD28-ICOS . This study provides an experimental basis for the treatment of autoimmune diseases such as rheumatoid arthritis by Tongbi Mixture.