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目的探讨通过补肾解毒和健脾解毒两种不同药物血浆体外作用慢性乙型肝炎(chronic hepatitis B,CHB)患者树突状细胞(dendritic cells,DCs)后对DCs功能的改善,比较哪种药物血浆更能促进DCs激活T淋巴细胞的功能提高。方法选择门诊CHB患者30例,根据其免疫状态,分为免疫耐受期组15例,免疫清除期组15例,抽取每例患者外周血60mL,分离培养出成熟的DCs,在培养第7天加入含补肾解毒(六味甘露饮)与健脾解毒(四君甘露饮)的复方中药血浆促进DCs成熟,另设无药物血浆作为对照。培养至第9天,将DCs负载HBVcore18-27核心肽后与自身的T淋巴细胞共培养72h,收集T淋巴细胞,应用流式细胞技术检测T细胞表面分子CD3、CD28、CD4、CD8及程序型坏死因子-1(programmed death-1,PD-1)的表达水平。结果与无中药血浆的共培养比较,免疫耐受期组和免疫清除期组的患者经补肾解毒和健脾解毒药物血浆干预后的DCs与T细胞共培养,在一定程度上都能提高CD3、CD4、CD28的表达水平,降低CD8、PD-1的表达水平,差异均有统计学意义(P<0.05),且补肾解毒药物血浆对提高免疫耐受组期患者CD28分子表达率、减低T细胞表面PD-1分子的表达率,效果均优于健脾解毒血浆的干预,差异均有统计学意义(P<0.05)。结论通过补肾解毒、健脾解毒药物血浆对DCs体外干预,并与自身T淋巴细胞共培养的方法,能够促进CHB患者DCs激活T细胞的功能提高和T细胞表面分子的表达,且补肾解毒法对慢性乙型肝炎病毒(hepatitis B virus,HBV)感染免疫耐受期患者影响更为显著。
Objective To investigate the effects of dextran and detoxification of kidney on the function of DCs after dendritic cells (DCs) in vitro treated by two kinds of drugs, Can promote the function of DCs to activate T lymphocytes more. Methods Thirty outpatients with CHB were enrolled in this study. According to their immune status, they were divided into immunologically tolerated group (n = 15) and immunosuppressive group (n = 15). Peripheral blood samples of 60 mL were collected from each patient and mature DCs were isolated and cultured. Join with kidney detoxification (Liu Wei Man Lu Yin) and the spleen and detoxification (Quadruple Manlu drink) plasma of Chinese medicine to promote DCs maturity, the other with no drug plasma as a control. After cultured for 9 days, DCs were loaded with HBVcore18-27 core peptide and co-cultured with their own T lymphocytes for 72h. T lymphocytes were collected. Flow cytometry was used to detect CD3, CD28, CD4, CD8, The expression of programmed death-1 (PD-1) Results Compared with the co-culture of Chinese medicine-free plasma, DCs and T cells co-cultured with T lymphocytes in the immunocompromised and immunosuppressed groups were able to increase CD3, CD4, CD28 and CD8, PD-1 expression were significantly different (P <0.05). The effect of Bushen Jiedu Poisoning on the expression of CD28 in immunosuppressive group and the decrease of T cell The expression and efficiency of PD-1 on the surface were better than those of spleen and detoxifying plasma, the difference was statistically significant (P <0.05). Conclusion The method of Bushen detoxication, intervention of spleen and detoxification drug plasma on DCs in vitro and co-culture with T lymphocytes can promote the function of DCs activated T cells and the expression of T cell surface molecules in CHB patients. Patients with chronic hepatitis B virus (HBV) infection are more affected by immune tolerance.