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背景:采用自体免疫细胞进行免疫治疗已发展为治疗恶性肿瘤的主要辅助手段之一,但其作用机制尚未阐明。目的:探讨细胞因子诱导的杀伤细胞分泌因子对人肝癌干细胞凋亡的影响。方法:采用无血清悬浮细胞培养法富集获得人肝癌干细胞;以γ-干扰素、CD3单克隆抗体和重组人白细胞介素2诱导肝癌患者外周血单个核细胞产生细胞因子诱导的杀伤细胞。将第1代肝癌干细胞分为2组,对照组单纯进行肝癌干细胞培养,实验组将细胞因子诱导的杀伤细胞与肝癌干细胞共培养,培养48 h后用流式细胞仪检测肝癌干细胞的凋亡情况,RT-PCR和Western blot检测凋亡相关基因caspase-3 mR NA与蛋白的表达。结果与结论:(1)对照组的肝癌干细胞凋亡率显著低于细胞因子诱导的杀伤细胞组(P<0.05)。(2)实验组的促凋亡相关基因caspase-3 mR NA和蛋白表达显著高于对照组(P<0.05)。(3)实验结果表明,细胞因子诱导的杀伤细胞会明显促进肝癌干细胞的凋亡,并显著上调肝癌干细胞促凋亡相关基因caspase-3mR NA和蛋白的表达水平。
BACKGROUND: Immunotherapy using autoimmune cells has been developed as one of the main aids for the treatment of malignant tumors, but its mechanism of action has not yet been elucidated. Objective: To investigate the effect of cytokine-induced killer cell secretory factors on the apoptosis of human hepatoma stem cells. METHODS: Human hepatocellular carcinoma (HCC) stem cells were enriched by serum-free suspension cell culture method, and cytokine-induced killer cells were induced from peripheral blood mononuclear cells of hepatocellular carcinoma patients induced by γ-interferon, CD3 monoclonal antibody, and recombinant human interleukin-2. The first generation liver cancer stem cells were divided into two groups. The control group was simply cultured with hepatoma stem cells. In the experimental group, cytokine-induced killer cells were co-cultured with hepatoma stem cells, and the apoptosis of hepatoma stem cells was detected by flow cytometry after 48 hours of culture. , RT-PCR and Western blot were used to detect the expression of apoptosis-related gene caspase-3 mR NA and protein. RESULTS AND CONCLUSION: (1) The apoptosis rate of HCC stem cells in the control group was significantly lower than that of the cytokine-induced killer cells group (P < 0.05). (2) The expression of caspase-3 mR NA and protein in the experimental group was significantly higher than that in the control group (P<0.05). (3) The experimental results show that cytokine-induced killer cells can significantly promote the apoptosis of hepatoma stem cells, and significantly up-regulate the expression of caspase-3 mRNA and caspase-3 mRNA in liver cancer stem cells.