目的：探讨脐带血和乳腺癌患者外周静脉血来源的CIK细胞程序性死亡分子-1（programmed cell death-1，PD-1）的表达及其对乳腺癌MCF-7细胞的杀伤作用。方法: 采集2015年6月至2015年12月在解放军第105医院住院的健康产妇脐带血和乳腺癌患者外周静脉血各5例，分离PBMC，体外培养、扩增CIK细胞。流式细胞术检测不同时间节点两种来源的CIK细胞PD-1表达情况，分别取培养第7、14、21、28天的CIK细胞与MCF-7细胞共培养，细胞计数（CCK-8）法测定CIK细胞对MCF-7细胞的杀伤率, 吖啶橙-溴化乙啶双染(AOEB)法观察共培养后CIK细胞凋亡变化，流式细胞术检测CIK细胞凋亡率。结果: 随着体外培养时间的延长，脐带血和乳腺癌患者静脉血来源的CIK细胞PD-1表达率均逐渐上升，第14天时脐带血组CIK细胞PD-1表达率低于乳腺癌组\[（38.42±4.76）% vs （50.54±3.50）%，P<0.05\]，至第21天后两组PD-1表达率均升高，但差异无统计学意义（P>0.05）。培养第7、14、21、28天两组CIK细胞对MCF-7细胞的杀伤率分别为(18.54±354）%和（21.74±427）%、（71.86±16.86）% 和（58.78±24.25）%、（44.32±26.87）%和（43.96±26.04）%、（43.24± 24.27）%和（40.28±23.69）%，以培养第14天的脐带血来源的CIK细胞的杀伤活性最强（P<0.05）。分析发现，两种来源的CIK细胞PD-1表达水平与CIK细胞的凋亡率呈正相关(r=0.971，r=0.900, 均P<0.01），与杀伤率呈负相关（r=-0.865，r=-0885, 均P<0.01）。结论: 活化的CIK细胞表面高表达PD-1，脐带血较乳腺癌患者静脉血来源的CIK细胞表面PD-1水平低；培养第14天的CIK细胞凋亡率均较低，其杀伤能力更强。

Objective:To detect the expressions of programmed cell death-1 (PD-1) on cytokine-induced killer (CIK) cells derived from umbilical cord blood or peripheral blood from breast cancer patients, as well as to investigate the cytotoxicity of CIK cells on MCF-7 cells. Methods: Umbilical cord blood from healthy pregnant women (n=5) and venous blood from breast cancer patients (n=5) were collected during June 2015 to December 2015 at the 105th Hospital of PLA. The PBMC was isolated, and CIK cells were differentiated and amplified  in vitro. The PD-1 expressions on CIK cells derived from two origins at different time points were detected by FCM; CIK cells at 7th, 14th , 21st and 28th days were used for the co-culture with MCF-7 cells, and the cytotoxicity of CIK cells on MCF-7 cells was determined by CCK-8 assay; CIK cell apoptosis after co-culture was observed by AOEB, and the apoptosis rate of CIK cells was determined by FCM. Results：Along with the extending of incubation time, the expressions of PD-1 on CIK cells of both groups increased gradually; PD-1 expression in CIK cells devived from umbilical cord blood at 14th day was lower than that devived from peripheral blood of breast cancer patients (\[38.42±4.76\]% vs \[50.54±3.50\]%,P>0.05), however, the expressions increased in both groups at 21st day and the difference between two groups was not statistically significant (P>005). Cytotoxicity rates of the CIK cells on the MCF-7 cell in the two groups at 7th ，14th ，21st and 28th days after co-culture were (18.54±3.54)% and (21.74±4.27）%，（71.86±16.86）% and（58.78±24.25）%，（44.32±2687）% and （43.96±26.04）% as well as （43.24±24.27）% and （40.28±23.69）% respectively, and among them the cytotoxicity of the CIK cells from umbilical cord blood at 14th day of the culturing was the highest (P<0.05). According to the analysis, there was a positive correlation between PD-1 expression and CIK cell apoptosis(r=0.971,r=0.900, all P<0.01), and a negative correlation between PD-1 expression and CIK cytotoxicity rate (r=-0.865，r=-0.885, all P<0.01). Conclusion：The activated CIK cells had high PD-1 expression，and CIK cells from patients with breast cancer had higher PD-1 expression than CIK cells from umbilical cord blood. The apoptosis rates of CIK cells at 14th day of culture were in both groups lower, and possessed higher cytotoxicities.

南京军区科技创新基金资助项目（No:15MS048）