Immune Cell SR对冻存PBMNC诱导培养CIK的效果研究  

Efficiency of Inducing CIK from Cryopreserved PBMNC by Using Immune Cell SR

作　　者：马冬磊 林科佳 陈晨 魏宗科 魏志璋 罗晓玲 王宇环 MA Dong-Lei;LIN Ke-Jia;CHEN Chen;WEI Zong-Ke;WEI Zhi-Zhang;LUO Xiao-Ling;WANG Yu-Huan(Shenzhen Hornetcorn Biotechnology Co. Ltd ,Shenzhen 518045 , Guangdong Province, China)

机构地区：[1]深圳市合一康生物科技股份有限公司,技术研发中心,广东深圳518045

出　　处：《中国实验血液学杂志》2018年第3期894-899,共6页Journal of Experimental Hematology

基　　金：深圳市科技创新委员会资助-技术攻关(JSGG20160226161357949)

摘　　要：目的:探讨Immune Cell SR(血清替代物)在冻存复苏后PBMNC诱导CIK细胞培养过程中的效果,为免疫细胞的工业化生产提供一个新的策略。方法:提取健康志愿者外周血单个核细胞,经短期冻存后复苏并诱导培养CIK细胞,采用台盼蓝拒染法检测细胞活率,流式细胞术分析免疫细胞亚群,并用Calcein-AM/PI双染法检测其杀伤活性;结果:冻存后PBMNC中在对照组未能正常扩增,与新鲜组相比,2%SR组细胞扩增倍数较低,且差异显著(P<0.05),5%SR组、10%AP组与新鲜组比较,差异均无统计学意义(P>0.05);冻存前后CD3^+、CD3^+CD8^+、CD3^+ CD56^+细胞亚群比较均无明显差异(P>0.05);细胞培养后,各组间CD3^+、CD3^+CD4^+、CD3^+CD8+、CD3^+CD56^+、CD3^-CD56^+亚群及体外杀伤活性比较均无明显差异(P>0.05);结论:采用5%SR替代自体血浆可以很好地诱导冻存的PBMNC培养CIK细胞,且无外源性蛋白,安全可靠,为免疫细胞冻存技术在细胞治疗中的应用提供条件。Objective： To investigate the efficiency of inducing CIK from peripheral blood mononuclear cells（ PBMNC）by using immune cell serum replacement（ immune cell SR）,so as to provide a new strategy for the industrialized production of immune cells.Methods： The PBMNC of healthy volunteers were collected,and these cells were thaw ed after short-term cryopreservation and cultured to induce CIK cells.The cells viability was measured by trypan blue exclusion,the phenotypes were analyzed by flow cytometry,and the cytotoxicity was determined by Calcein-AM/PI double staining.Results： In cryopreserved PBMNC,the control group cells failed to normally proliferate.Cell proliferation ratio was low in 2% SR group in comparison with the fresh group,and the difference was significant（ P〉0.05）,however,differences were not statistically significant between 5% SR and fresh group or between 10% AP and fresh group.CD3^＋,CD3^＋CD8^＋ and CD3^＋CD56^＋ cell subsets were not significantly different before and after cryopreservation（ P〉0.05）.After being cultured,CD3^＋,CD3^＋CD4^＋,CD3^＋CD8^＋,CD3^＋CD56^＋ and CD3^-CD56^＋ subsets and the cytotoxicity in vitro were not significantly different among all group（ P〉0.05）.Conclusion： 5% SR without the protein of animal origin can be safely used as a substitute for autologous plasma in CIK induced from cryopreserved PBMNC by culture,thus providing a basis for the application of cryopreservation technique of immune cells to cell therapy.

关 键 词：外周血单个核细胞 细胞因子诱导的杀伤细胞 冻存 血浆替代物 

分 类 号：R392.12[医药卫生—免疫学]