摘要
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 with autologous immune cells has been developed as a major adjuvant therapy for malignant tumors, but its mechanism of action has not been elucidated. OBJECTIVE： To investigate the relationship between cytokine-induced killer cell secretion and apoptosis in human liver cancer stem cells. METHODS： Human liver cancer stem cells, Hep G2 cells, were isolated and enriched using serum-free suspension method. The peripheral blood mononuclear cells from patients with liver cancer were induced by γ-interferon, CD3 monoclonal antibody and recombinant human interleukin-2 to form killer cells. Passage 1 liver cancer stem cells were divided into control group（culture alone） and experimental group（co-culture of cytokines-induced killer cells and human liver cancer stem cells）. At 48 hours after culture, apoptosis in human liver cancer stem cells was detected using flow cytometry, and expression of caspase-3 mR NA and protein was detected using RT-PCR and western blot, respectively. RESULTS AND CONCLUSION： The apoptotic rate in the control group was significantly lower than that in the experimental group（P 〈0.05）. The expressions of caspase-3 at m RNA and protein levels were both higher in the experimental group than the control group（P〈 0.05）. Experimental findings show that cytokines-induced killer cells can significantly promote apoptosis in human liver cancer stem cells, and up-regulate the caspase-3 mR NA and protein expressions dramatically.
出处
《中国组织工程研究》
CAS
北大核心
2016年第14期2033-2039,共7页
Chinese Journal of Tissue Engineering Research
