表达新型干扰素的CIK细胞对结肠癌的治疗作用  

Inhibitory effect of CIK with neo-interferon on colon cancer

作　　者：陈艳[1] 刘辉[1] 于丽[2] 付玉华[2] 王敬晗[1] 钱其军[1,2] 

机构地区：[1]第二军医大学东方肝胆外科医院基因病毒治疗实验室,上海200438 [2]浙江理工大学生命科学学院新元医学与生物技术研究所

出　　处：《中华实验外科杂志》2011年第12期2158-2161,共4页Chinese Journal of Experimental Surgery

基　　金：基金项目：肝癌溶瘤病毒临床前/临床试验及中试生产工艺研究,艾滋病和病毒肝炎等重大传染病的防治重大专项（2008ZX10002-023）

摘　　要：目的探讨细胞因子诱导的杀伤细胞（CIK细胞）作为新型重组干扰素mix基因的载体治疗结肠癌的可行性。方法制备携带mix基因的腺病毒pDC359．mix＋F35；分离培养CIK细胞；腺病毒Ad5-EGFP、Adll-EGFP和Ad35．EGFP以感染复数（MOI）=100时感染CIK细胞24、48h后观察感染效率；腺病毒pDC3590mix＋F35以MOI=0、1、10、100、1000感染CIK细胞2、4、6、8d，酶联免疫吸附试验（ELISA）检测上清中干扰素mix表达量；磺酰罗丹明B（SRB）显色法检测浓度为0．001、0．01、0．1、1、10ug／L的干扰素mix和干扰素a-2b对结肠癌细胞株HT29和SW480的抑制作用；乳酸脱氢酶释放法（LDH）检测效靶比为3：1、10：1的CIK和CIK—mix对HT29和SW480的杀伤作用。结果腺病毒pDC359-mix＋F35成功制备，MOI=100，感染48h后对CIK细胞的感染效率可达80％以上；当MOI=100时，各时间段干扰素mix的表达量均达到（8．96±2．10）ng以上；当浓度为10ug／L，干扰素mix对结肠癌细胞株抑制率分别为72．8％和70．1％，而干扰素OL-2b分别为30．4％和28．6％；腺病毒pDC359-mix＋F35感染CIK后，CIK—mix对HT29和SW480的杀伤作用均比CIK增强了20％以上。结论表达干扰素mix的CIK细胞体外能够发挥联合的抗肿瘤效应。Objective： To investigate the feasibility of using cytokine-induced killer （CIK） cells as targeted vectors to deliver neo-interferon （mix） gene for colon cancer. Methods Replication-deficient adenovirus pDC359-mix ＋ F35 carrying mix gene was generated. CIK cells were isolated and cultured. The infection efficiency of Ad5-EGFP, Adll-EGFP and Ad35-EGFP to CIK for 24 h and 48 h at MOI = 100 was determined by fluorescence microscope. CIK ceils were infected by adenovirus pDC359-mix ＋ F35 at MOI =0, 1, 10, 100, 1000 for2, 4, 6, 8 h, then the mix concentration in the cuhure supernatant of CIK cells was determined by using enzyme linked immunosorbent assay （ELISA）. The inhibitory effect of mix and interferon a-2b at different concentrations of 0. 001, 0. 01, 0. 1, 1, 10 ug/L on colon cancer cells HT29 and SW480 was detected by Sulforhodamine B （SRB）. Cytotoxicity of CIK and CIK-mix in different ratio of efficiency to target 3：1 and 10：1 on HT29 and SW480 was tested by LDH release method in vitro. Results Replication-deficient adenovirus pDC359-mix ＋ F35 was successfully generated as high infection efficiency as 80% at MOI = 100 for 48 h. At MOI = 100, the mix concentration exceeded （8.96 ± 2. 10） ng level at different periods. At 10 ug/L, the inhibitory rate of interferon a-2b and mix on HT29 and SW480 cells was 30. 4% and 28.6%, and 72. 8% and 70. 1% respectively. The cytotoxicity of CIK-mix on HT29 and SW480 was stronger by 20% than CIK. Conclusion CIK cells with neo-interferon can exert synergistic antitumor effect.

关 键 词：新型重组干扰素mix CIK细胞 腺病毒 结肠癌 

分 类 号：R735.35[医药卫生—肿瘤]