树突状细胞诱导抗胃癌效应的实验研究

Empirical study of anti-gastric cancer effects induced by dendritic cells

目的探讨树突状细胞(DC)抗胃癌效应的作用。方法分离正常人外周血单个核细胞(PBMC),加入GMCSF和IL-4培养,于第5d加入TNF-α继续培养,诱导分离DC。将淋巴细胞与DC混合培养4d后,加入胃癌细胞MKN45、MKN28及SGC7901抗原培养至10d,诱导产生肿瘤特异性的CTL。ELISA检测IL-12和IFN-γ的水平,MTT法检测DC诱导的CTL对MKN45、MKN28、SGC7901和A549体外杀伤效应,流式细胞术检测CTL对肿瘤细胞周期、凋亡的影响。结果培养10d后,IL-12、IFN-γ的分泌量明显提高,DC对同种不同分化类型的胃癌细胞株MKN45、MNK28和SGC7901均有强烈的杀伤效应,杀伤活性显著高于A549(P<0.01)。与A549比较,MKN45、MKN28和SGC7901的细胞周期显著抑制、凋亡率增加(P<0.01)。结论 DC体外可诱导出高效而特异的抗胃癌效应,显著抑制胃癌细胞分裂增殖,促进其凋亡。

Objective To investigate the anti - gastric cancer effects of dendritic cells.（DC） in vitro. Methods Peripheral blood mononuclear cells （PBMC）were isolated from normal human person and cultured with granulocyte - macrophage colony - stimulating factor（ GM - CSF）and IL - 4 in medium. On day 5, TNF - α was added to induce PBMC into DC which were eventually isolated. After DC were mixed and cultured with T lymphocytes for 4 d, they were treated with the tumor antigens of MNK45, MKN28 and SGC 7901 until day 10 to obtain the cytotoxic T lymphocytes（CTL） with tumor specificity. IL - 12 and IFN - γ levels were detected by ELISA. The killing activity of DC - induced CTL on four gastric cancer cell lines of MNK45, MKN28, SGC 7901 and A549 in vitro were detected by MTT method. The inhibitive effects of CTL on tumor cell cycle and apoptosis were detected by flow cytometry. Results The secretory volume of IL - 12 and IFN - γ in the supernatant of the lymphocytes increased significantly after 10 d of cuhure. DC - induced CTL had much stronger cytotoxicity to different differentiated gastric cancer cell lines including MKN45, MNK28 and SGC 7901 than they did to A549 （ P 〈 0.01 ）. Compared with A529, the cell cycles of other three cell lines were significantly inhibited and apoptotic ratios increased significantly （ P 〈 0.01 ）. Conclusion DC treated with tumor antigens can induce high efficiency and specificity of anti - tumor effects. The T lymphocytes activated by these DC can significantly inhibit the division and growth of gastric cancer cells as well as promote their apoptosis.