IL-12对CIK细胞体外增殖及体内外杀瘤活性影响的实验研究

The Effect of IL-12 on the proliferation in vitro and anti-tumor effects of CIK cells in vivo and in vitro

目的动态观察IL-12对细胞因子诱导的杀伤细胞(cytokine-induced killer cells,CIK)体外增殖,体内外的细胞毒活性的影响。方法采用三种不同的细胞因子组合扩增CIK细胞,即IL-2组:IFN-γ、CD_3单抗、IL-1和IL-2;IL-2加IL-12组:IFN-γ、CD_3单抗、IL-1、IL-2和IL-12;IL-12组: IFN-γ、CD_3单抗、IL-1和IL-12。流式细胞术分析细胞表型,细胞计数法测定细胞的增殖。MTT法测定CIK体外细胞毒活性,并通过做扫描和透射电镜,对CIK细胞杀伤的肿瘤细胞进行形态学观察。研究CIK细胞对BGC-823荷瘤裸鼠的体内抗肿瘤作用。结果三种方法均可使细胞增殖能力显著增加,对CD_3^+CD_(56)^+细胞的诱导作用也无明显差异,IL-12与IL-2联合作用组促增殖能力和细胞毒性作用明显强于其他两组(P＜0．05)。被CIK细胞杀伤的肿瘤细胞的死亡形式是坏死和凋亡。动物实验证实CIK有较强的体内抗瘤活性,可延长荷瘤鼠的生存期,联合IL-12与IL-2诱导的CIK细胞疗效较好。结论IL-12同样可以用于CIK细胞的诱导,而联合IL-12与IL-2诱导的CIK细胞其增殖能力和体内外抗肿瘤活性均增强。

Objective To investigate the effect of IL-12 on the proliferation profile in vitro and cytotoxicity of CIK（cytokine induced killer） cells in vivo and in vitro.Methods Adopt three different cytokine- combination to expand the CIK.Tne same inducing agents in the three groups were IFN-γ,CD3McAb and IL-1. In addition, IL-2 was added in group IL-2, and IL-12 was added in group IL-12,and both IL-2 and IL-12 were added in group IL-12 ＋ IL-2 as well.Surface antigen expression was detected by flow cytometry. The cellular prolifelation and cytotoxic activity were analyzed by cell counting and MTT assay. The morphology of the tumor target cells killed by CIK was observed through scanning electron microscope and transmission electron microscope. The anti-tumor effects on BGC-823 bearing nude mice in vivo of CIK was tested. Results CIK cells generated by three methods showed high proliferation activity, and no significance in inducing CD3 ＋ CD56 ＋ cells was found among three groups.The combination of IL-2 ＋ IL-12 evidently enhanced both the proliferation rates and the cytotoxicity of the CIK cells compatred with the other two groups. Death patterns of tumor target cells were necrosis and apoptosis. CIK cells had greater anti-tumor effects in mice bearing BGC,-823 tumor and could prolong the mice＇ survival time. The CIK cells induced by both IL-2 and IL-12 had better effect. Conclusion IL-12 could be used to induced the CIK cells as well as IL-2. CIK cells induced by combining IL-12 with IL-2 had strong proliferative ability and higher cytotoxicity to tumor cells in yitro and in vivo, which could be used as a potential antitumor adoptive immunotherapy in clinic.