脐血来源的CIK细胞大容量制备方法的建立  

Establishment of large-capacity culture method of CIK cells from cord blood

作　　者：李翠萍[1] 陈宝安[2] 陈津[3] 周敏[2] 王晓波[4] 傅强[3] 仲伟云[3] 张益红[3] 

机构地区：[1]江苏省血液中心 [2]南京东南大学附属中大医院血液科,210009 [3]南京红十字血液中心,210003 [4]江苏省肿瘤医院内科,210009

出　　处：《临床肿瘤学杂志》2005年第3期291-292,296,共3页Chinese Clinical Oncology

摘　　要：目的:探讨脐血来源的CIK细胞大容量培养方法,研究一种运用于临床治疗的细胞制剂,以期用于恶性血液病及实体瘤的免疫治疗。方法:脐血取自妊娠足月正常分娩的产妇。两步离心法加Ficoll分离出脐血单个核样细胞,在含5%正常人AB血清的液体培养体系中加入γINF、CD3单克隆抗体、IL2诱导CIK细胞扩增,流式细胞仪检测CIK细胞对K562、Raji细胞的杀伤活性。结果:每份脐血分离后可获得(0.8～1.2)×108单个核样细胞,加细胞因子扩散10天后,CIK细胞增殖最高,达60～100倍以上,30天后趋缓。CD3+CD56+细胞双表达由培养前1.0%～1.2%到30.0%～80.1%。对K562、Raji细胞的杀伤活性在效、靶比为5∶1时达50.0%～71.7%,培养14天时最高,30天后逐渐降低。结论:采用药用细胞因子及正常人血清培养脐血来源的CIK细胞,具有增殖旺盛、对K562、Raji细胞的杀伤活性强的特点,可以安全地应用于临床治疗。Objective:This study was conducted to establish the large-capacity culture method of cytokine-induced killer (CIK) cells for clinical therapy and assess its effect on the function of cell-mediated immunity following CIK cells reinfusion .Methods:Mononuclear cells (MNCs)harvested from cord blood from women after normal period of gestation by re-centrifuging and Ficoll separating ,co-cultured on 5% human serum supplemented with interferon-gamma (γ-IFN),monoclonal anti-CD_3 antibodies and interleukin (IL-2) to induce CIK cells to expand. The cytotoxic activity against K562,Raji was shown using flow cytometric analysis.Results:(0.8～1.2)×10~8 MNCs were harvested from a pack of umbilical cord blood.The CIK cells were expanded by priming with r-IFN followed by anti-OCD3 monoclonal antibody and IL-2.The total amount of expanded CIK cells exceeded 60 to 100-fold after 10 days. The percentage of CD^+_3 CD^+_(56) cells generated following culture from 1.0%～1.2% to 30.0%～80.1%.This effect was significantly enhanced by co-culturing with cytokines. The cytotoxic activity of CIK cells was enhanced to 50.0%～71.7% at the lower effector to target ratio of 5∶1.The maximal cytotoxic activity was shown at day 14, reduce gradually after 63 days.Conclusion:The results showed that the large-capacity culture method of CIK cells is a simple and efficient way of expanding CIK cells, a simple and safe therapy for treating the hematological malignances and tumors. CIK cells showed high lytic activity against K526,Raji cells, which could be enhanced by co-culturing with human serum and cytokines used in clinical.

关 键 词：脐血 细胞因子 CIK细胞 制备 

分 类 号：R392-33[医药卫生—免疫学]