健康人和肿瘤患者CIK细胞的生物学特性比较

The comparison of biological characteristics on CIK cells from healthy donors and tumor patients

目的比较健康人和肿瘤患者外周血来源的CIK细胞的体外扩增速度、细胞表型和杀伤活性；观察健康人CIK细胞对裸鼠体内移植瘤的预防和治疗作用。方法常规无菌采集健康人和肿瘤患者外周血单核细胞各10份，定向诱导成CIK细胞，然后直接活细胞计数法比较两者的扩增速度；流式细胞仪检测比较两者细胞表型；MTT法比较两者对K562和LOVO细胞株的杀伤活性。取30只裸鼠分为3个组，预防组：裸鼠先尾静脉注射CIK细胞，连续5天，第6天背部皮下接种A549细胞。治疗组：裸鼠于第一天接种A549细胞，次日，分为3组，第一组局部（接种A549部位）注射CIK细胞；第二组尾静脉注射CIK细胞；第三组局部（接种A549部位）及尾静脉均注射CIK细胞，均连续治疗5天。对照组：裸鼠，第一天背部皮下接种A549细胞，第二天开始在尾静脉注射生理盐水，连续5天。四周后处死全部裸鼠，测量肿瘤大小，称重，计算肿瘤体积，抑瘤率，并做病理检查。结果健康人CIK细胞的扩增速度显著高于肿瘤患者CIK细胞（P〈0．05）；流式细胞仪检测显示健康人CIK细胞CD3^＋CD8^＋、CD3^＋CD56^＋细胞百分比显著高于患者CIK细胞（P〈0．05）；健康人CIK细胞对K562，LOVO的杀伤活性强于患者CIK细胞（P〈0．05）。对照组各裸鼠背部皮下移植瘤出现较早，并且以相近速率快速生长，体积较大；CIK细胞预防组及治疗组中局部注射组和尾静脉注射组的移植瘤出现较晚，生长较慢，体积较小，联合治疗组治疗效果更佳（P〈0．05）。结论体外实验中，健康人CIK细胞体外扩增快，CD3^＋CD8^＋、CD3^＋CD56^＋细胞比例高，对肿瘤细胞的杀伤活性强于肿瘤患者CIK细胞。动物实验中，健康人CIK细胞对裸鼠移植瘤有预防和治疗作用。

objective To compare the proliferation in vitro, phenotype and cytotoxicity of CIK cells from peripheral blood of healthy donors and tumors patients. To observe the function of prevention and treatment of CIK cells from healthy donors on transplanted tumor of nude mouse in vivo. Methods 10 shares of peripheral blood of healthy donors and tumors patients were respectively harvested. The lymphocytes were isolated by density gradient centrifugation and suspended to prepare CIK cells. The proliferation of the ceils were compared by cell counting. The phenotype of CIK cells were analysed by flow cytometry. The cytotoxic activities to K562 cells and LOVO cells were determined by MTT method. Thirty nude mice were divided into 3 groups. Prevention group： 6 nude mice injected CIK cells into caudal vein every day, continuous 5 days, at the 6th day, A549 cells were injected in back subcutaneously of the mice. Treating group： 18 nude mice were injected A549 ceils in back subcutaneously at the 1st day, the 2na day, separated the nude mice into 3 groups,,6 nude mice in each group, all groups were treated for 5 days. A： Injected CIK cells into part of injected A549 cells every day. B： Iniected CIK cells into caudal vein every day. C： Iniected CIK cells into caudal vein and on the local part every day. Control group： 6 nude mice were injected A549 cells in back subcutaneously at the 1st day, on the 2nd day, injected saline into caudal vein every day for 5 days. All nude mice were killed after four weeks, measuring tumor size, weighing, calculating the tumor volume and inhibitory rates, and having pathological examinations. Results The proliferation speed of CIK cells from healthy donors were significantly higher than those from tumors patients （P〈0.05）; The phenotype percentage of CD3＋CD8＋,CD3＋CD56＋ of CIK cells from healthy donors were significantly higher than those from tumors patients （P 〈0.05）; the anti-tumor effect on K562 and LOVO cell line of CIK ceils from healthy donors were significantly higher than those from tumors patients （P〈0.05）. The transplantation tumor of mice appear early , grow repid and the volume is bigger in control group, the transplantation tumor of mice appear late and grow slow in preventiom group, the transplantation tumor of treating group appear later, grow slower, volume smaller, the treatment effect of combination treatment group is the best （P〈0.05）. Conclusion In vitro experiment, the proliferation the percentage of CD3＋CD8＋, CD3＋CD56~ and the cytotoxicity of CIK cells originated from of healthy donors are higher than the CIK cells from tumor patients. In animal experiment, CIK cells from healthy donors have the precaution and therapeutical effect to transplantation tumor that translated in nude mice.