【摘要】 目的研究调节性T细胞对外周血来源的DC-CIK细胞增殖及杀伤活性的影响。方法无菌分离外周血单个核细胞,体外诱导分化为DC-CIK细胞。实验分2组:DC-CIK组为常规单个核细胞诱导分化的DC-CIK;DCCIK-Tregdel组为培养前利用免疫磁珠分选去除CD4+CD25+T细胞的外周血单个核细胞诱导分化的DC-CIK。细胞扩增倍数法检测2组细胞的增殖能力;MTT法检测肿瘤细胞的杀伤活性;流式细胞仪检测免疫表型;ELISA法检测细胞因子分泌水平。结果培养14 d的DC-CIK-Tregdel组细胞的扩增倍数高于DC-CIK组,并且CD3+CD8+、CD3+CD56+双阳性细胞高于DC-CIK组(P<0.05);在5∶1、10∶1、20∶1效靶比范围内,培养14 d的DCCIK-Tregdel组细胞对SGC-7901胃癌细胞株的杀伤率明显高于DC-CIK组(P<0.05),并且这种杀伤作用随着效靶比的增高而增强;DC-CIK-Tregdel组培养上清液TGF-β水平低于DC-CIK组,而IFN-γ分泌水平明显高于DC-CIK组(P<0.05)。结论 CD4+CD25+T细胞对DC-CIK抗肿瘤效应细胞具有免疫抑制作用,去除CD4+CD25+T细胞能增加DC-CIK细胞增殖能力和抗肿瘤活性,为DC-CIK-Tregdel细胞有效治疗肿瘤提供了实验依据。更多还原

【Abstract】 Objective The purpose was to investigate the effect of regulatory T cells on proliferation and killing activity of dendritic cells and cytokine-induced killer cells( DC-CIK) from peripheral blood. Methods DC-CIK cells were prepared from peripheral blood mononuclear cells of healthy human. The experiment was divided into two groups: DC-CIK cells group( conventional mononuclear cells were induced to DC-CIK cells) and DC-CIK-Tregdelcells group( the peripheral mononuclear cells without CD4+CD25+regulatory T cell were induced to DC-CIK cells). The proliferation ability and killing activity of the two groups cells were detected by cell expansion in multiples method and MTT assay; phenotype of regulatory T cells were detected with flow cytometry. The TGF-β and IFN-γ were detected with ELISA in the cells culture supernatant of the two groups. Results The cell proliferation of the DC-CIK-Tregdelgroup cultured for 14 days was obviously higher than that of the DC-CIK group( P < 0. 05). Flow cytometry showed that the percentages of CD3+CD8+and CD3+CD56+T cells in DC-CIK-Tregdelgroup were significantly increased as compared with DC-CIK group( P < 0. 05). In the range of 5∶ 1,10∶ 1 and 20∶ 1 effector to target ratio,the killing rate of the DC-CIK-Tregdelgroup on gastric cancer cell line SGC-7901 was respectively increased than that of the DC-CIK group( P < 0. 05) and this killing effect was enhanced with increased effector to target ratio. ELISA showed that the level of TGF-β in supernatant liquid of the DC-CIKTregdelgroup was lower than that of the DC-CIK group,while the level of IFN-γ in supernatant liquid of the DC-CIK-Tregdelgroup was higher than that of the DC-CIK group( P < 0. 05). Conclusion CD4+CD25+T cells have immunosuppressive effect to anti-tumor activity of DC-CIK cells,and removing CD4+CD25+T cells can increase the DCCIK cell proliferation and anti-tumor activity,which provides an experimental basis for the effective treatment of tumor cells with the DC-CIK-Tregdelcells.更多还原