论文部分内容阅读
目的:探讨患者冻存外周血干细胞体外诱导CIK细胞的增殖能力、表型特征及其对B细胞淋巴瘤细胞系的杀伤效应。方法:复苏患者冻存外周血干细胞悬液,Ficoll密度梯度方法分离单个核细胞及从健康人新鲜外周血分离单个核细胞,经IFNγ、IL-2、CD3McAb诱导培养获得CIK细胞。细胞计数板观察CIK细胞增殖能力;流式细胞术检测CIK的表型;以CFSE标记靶细胞区分效应细胞,PI染色,FACS检测靶细胞死亡率。结果:患者冻存外周血干细胞来源的CIK细胞较健康人增殖速度慢,最大增殖倍数低,但CD3+CD56+细胞比例与健康人差异无统计学意义,分别为(16.14±8.49)%和(17.52±3.30)%,P=0.744。患者及健康人CIK细胞与相应的诱导培养前单个核细胞相比,杀伤活性明显增强,在效靶比10:1时患者及健康人CIK细胞对Daudi细胞的杀伤活性为(29.23±8.85)%、(20.13±2.36)%,与培养前相比P值分别为0.004、0.001;患者CIK细胞杀伤活性强于健康人,P=0.05。结论:患者冻存外周血干细胞可以诱导生成具有抗肿瘤活性的CIK细胞,为其临床应用提供了实验依据。
OBJECTIVE: To investigate the proliferation, phenotypic characteristics and cytotoxicity of CIK cells induced by cryopreserved peripheral blood stem cells in vitro in patients with B-cell lymphoma. Methods: Peripheral blood stem cell suspension was cryopreserved in peripheral blood of patients. Fibronuclear mononuclear cells were isolated by Ficoll density gradient centrifugation and mononuclear cells were isolated from fresh peripheral blood of healthy volunteers. CIK cells were induced by IFNγ, IL-2 and CD3McAb. The proliferation of CIK cells was observed by cell counting plate. The phenotype of CIK was detected by flow cytometry. The effector cells were differentiated by CFSE labeled cells, PI staining and FACS were used to detect the target cell death rate. Results: CIK cells derived from cryopreserved peripheral blood stem cells had slower proliferation rate and lower maximum multiplication rate than healthy individuals, but there was no significant difference in the proportion of CD3 + CD56 + cells between the two groups (16.14 ± 8.49% vs. (17.52 ± 3.30)%, P = 0.744. Compared with the corresponding pre-culture mononuclear cells, the cytotoxicity of CIK cells was significantly increased in patients and healthy people compared with that of CIK cells (29.23 ± 8.85%) at the effective target ratio of 10: 1 , (20.13 ± 2.36)%, respectively. Compared with the pre-culture, the P values were 0.004 and 0.001 respectively. The cytotoxic activity of CIK cells in healthy volunteers was higher than that in healthy volunteers (P = 0.05). Conclusion: Cryopreserved peripheral blood stem cells can induce the generation of CIK cells with anti-tumor activity, which provides an experimental basis for its clinical application.