树突状细胞介导CIK细胞抗恶性肿瘤效应的实验研究

目的 研究人脐血来源树突状细胞（DC）和同源细胞因子激活的杀伤细胞（CIK）,共孵育后,获得的DC CIK细胞对恶性肿瘤细胞的杀伤效应.方法 获取脐血单个核细胞,用不同细胞因子分别诱导DC及CIK细胞.按1：5比例将DC和CIK细胞一起培养.且以单独培养的脐血CIK细胞为对照.采用流式细胞技术分析检测细胞免疫表型,细胞扩增倍数用台酚蓝染色方法计算,杀伤率测定采用MTT法,细胞因子IL 12,IFN γ,TNF-α用ELISA方法检测.结果 来源于脐血CIK细胞扩增倍数低于同源DC CIK细胞（P＜0.05）;混合培养物DC CIK细胞中,CD3 CD8,CD3＋ CD56＋双阳性免疫细胞数量较单纯CIK细胞明显增高（P＜0.05）;脐血DC,CIK细胞共培养3天,其上清液中细胞因子（IL 12,IFN γ及TNF-d）含量比CIK细胞上清液中高（P＜0.01,＜0.05,＜0.05）;当效靶比为2.5∶1-20∶1时,DC-CIK细胞对HL60,K562白血病细胞、U266多发性骨髓瘤细胞株及SMMC 7721肝癌细胞杀伤率均高于CIK细胞（均P＜0.05）.结论 脐血DC可增强同源CIK细胞的增殖活性和抗肿瘤效应.该研究为脐血DC CIK细胞的免疫治疗提供了实验和理论依据.

Experimental Study on Anti-Malignant Tumor Effect Mediated by Dendritic Cells-CIK Cells

Objective Study on the cytotoxic effect against malignant tumor cells after co culture of human umbilical cord blood derived dendritic cells （DC） with homologous cytokine induced killer cells （CIK）.Methods DC and CIK cells were induced from cord blood mononuclear cells with different cytokines respectively.They were co cultured according to the ratio of 1 ∶ 5,and cord blood derived CIK cells alone as control.Immunophenotype changes were analyzed by flow cytometry; in creased number of cells were counted by trypan blue staining;the killing activity against malignant tumor cells was detected by MTT assay; the interferon γ （IFN γ）,tumor necrosis factor-α （TNF α）,and interleukin 12 （IL 12） levels of the cultured supernatants were detected by ELISA kits.Results The proliferation capability of cord blood DC CIK cells was significantly higher than that of CIK cells alone （P＜0.05） ; CD3 ＋ CD8＋,CD3＋ CD56＋ double positive cells in DC CIK co culture were significantly increased than that in pure CIK under the same conditions （P＜0.05） ;compared with CIK cells cultured alone,the level of IL 12,IFN γ,and TNFα in cultured supernatants of DC CIK cells increased noticeably on day 3 （P＜0.01 or P ＜0.05,P＜0.05）.With the effector-target ratio ranging from 2.5 ∶ 1 to 20 ∶ 1,the cytotoxic activity of the cord blood DC CIK cells against leukemia cell lines HL-60,K562,multiple myeloma cells U266 and hepatocellular carcinoma cells SMMC 7721 were much higher than that of CIK cells （P＜0.05）.Conclusion Umbilical cord blood derived DC can mediate en hanced proliferation and anti tumor effect of homologous CIK cells.This study provides experimental and theoretical basis for immunotherapy of cord blood DC-CIK cells.