目的： 比较脐血来源的细胞因子诱导杀伤（cytokine-induced killer，CIK）细胞和肿瘤患者外周血来源CIK（peripheral blood-derived CIK，PB-CIK）细胞的增殖、激活型和抑制型表面标志物、耐药基因ABCG2以及干细胞转录因子表达的差异，探讨脐血来源CIK（umbilical cord blood-derived CIK，CB-CIK）细胞应用于肿瘤过继细胞免疫治疗的优越性。 方法: 采用Ficoll-plaque淋巴细胞分离液分离脐血及肿瘤患者外周血单个核细胞，加入细胞因子诱导培养CIK细胞。流式细胞术检测CIK细胞的增殖、免疫表型、激活型表面标志物CD28、CD27和抑制型表面标志物PD-1的表达，RT-PCR检测耐药基因ABCG2和干细胞转录因子 c-Myc、Nanog、Oct-4、Sox2的表达。 结果: CB-CIK细胞与PB-CIK细胞体外培养第4天时均开始增殖，第10天时CB-CIK细胞的增殖指数明显高于PB-CIK细胞 \[（251.52±16.76）% vs （158.00±43.19）%，P<0.05\]。体外诱导培养13 d后CB-CIK细胞中CD3+CD56+细胞比例较PB-CIK细胞高\[（21.20±4.82）% vs （10.06±3.46）%，P<0.05\]；激活型CD4+CD28+、CD4+CD27+和 CD8+CD27+细胞比例在CB-CIK细胞中也明显升高\[（32.40±16.81）% vs （18.65±9.23）%，（2748±13.53）% vs (0.98±055)%，（41.76±13.98）% vs （2.58±2.10)%；P<0.05或P<0.01\]；而抑制型CD8+PD-1+细胞比例明显降低\[(3.25±2.13)% vs (8.05±9.23)%，P<0.01\]。此外，CB-CIK细胞与PB-CIK细胞均表达干细胞转录因子c-Myc、Nanog、Oct-4、Sox2 ，但两者之间无显著差异（P>0.05）；而仅CB-CIK细胞表达高水平的耐药基因 ABCG2。 结论: CB-CIK细胞较PB-CIK细胞增殖速度快、激活型表面标志物表达水平高、抑制型表面标志物表达水平低、高表达耐药基因 ABCG2，应用于肿瘤过继细胞免疫治疗有一定的优势。

Objective:To analyze the proliferation and differential expressions of activated and inhibitory surface markers, drug-resistance gene ABCG2 and stem cell transcription factors in umbilical cord blood-derived cytokine induced killer (CIK) cells or peripheral blood-derived CIK (PB-CIK) cells in cancer patients, and to explore the superiority of umbilical cord blood-derived CIK (CB-CIK) cells in adoptive cellular immunotherapy. Methods: Cord blood mononuclear cells and peripheral blood mononuclear cells in cancer patients were isolated using Ficoll-plaque density gradients and incubated with cytokines. The proliferation, immunophenotypes, activated surface markers (CD28, CD27) and inhibitory surface marker (PD-1) were analyzed by flow cytometry, and the drug-resistance gene ABCG2 or stem cell transcription factors ( c-Myc, Nanog, Oct-4 and Sox2 ) were determined by RT-PCR. Results: CB-CIK cells and PB-CIK cells started to proliferate on the fourth day in vitro, and the proliferation index of CB-CIK cells was significantly higher than that of PB-CIK cells on day 10 (\[251.52±16.76\]% vs \[158.00±43.19\]%, P<005\]. Thirteen days after incubation, the proportion of CD3+CD56+ cells in CB-CIK cells was higher than that in PB-CIK cells (\[21.20±4.82\]% vs \[10.06±346\]%, P<0.05). When detecting the status of CIK cells, the proportions of activated CD4+CD28+, CD4+ CD27+ and CD8+CD27+ cells were significantly higher in CB-CIK cells than in PB-CIK cells (\[32.40±16.81\]% vs \[18.65±923\]%; \[27.48±13.53\]% vs \[0.98±0.55\]%; \[41.76±13.98\]% vs \[2.58±2.10\]%，P<0.05 or P<0.01), while the proportion of inhibitory CD8+PD-1+ cells was significantly lower in CB-CIK cells (\[3.25±2.13\]% vs \[8.05±9.23\]%, P<0.01). The stem cell transcription factors ( c-Myc, Nanog, Oct and Sox2 ) were expressed both in CB-CIK cells and PB-CIK cells. Moreover, no significant differences were found between the two kinds of CIK cells. Furthermore, only CB-CIK cells expressed a high level of drug-resistance gene ABCG2 . Conclusion: Compared with the PB-CIK cells, the CB-CIK cells showed increasing proliferation rates, higher expressions of activated surface markers, and lower expression of inhibitory surface marker. Besides, drug-resistant gene ABCG2 is highly expressed in CB-CIK cells, showing several advantages in applying to adoptive cellular tumor immunotherapy.

国家自然科学基金资助项目（No. 81171985，No. 81171986，No. 812111102）