HBsAg基因修饰的树突状细胞体外诱导抗HepG2.2.15特异性细胞毒作用  

Cytotoxicity Induced by HBsAg Gene Modified Dendritic Cells against Hepatocellular Carcinoma Cell HepG2.2.15

作　　者：杨静悦[1] 任军[1] 白俊[1] 刘都户[1] 范黎[1] 斯小明[1] 腾增辉[2] 杨文涛[2] 

机构地区：[1]第四军医大学西京医院肿瘤科,陕西西安710032 [2]第四军医大学药理教研室,陕西西安710032

出　　处：《癌症》2004年第8期914-917,共4页Chinese Journal of Cancer

基　　金：高等学校优秀青年教师教学科研奖励基金(TRAPOYT99-16);军队杰出人才基金(OIJO16)~~

摘　　要：背景与目的目前缺少一种针对乙型肝炎病毒感染相关肝癌的有效免疫治疗手段。以树突状细胞(dendriticcell,DC)为基础的肿瘤特异性免疫治疗方法,为免疫治疗乙型肝炎病毒感染相关的肝癌提供了新方法。本实验通过体外负载乙型肝炎表面抗原基因(HBsAg)的重组质粒转染DC,制备HBsAg-DC瘤苗,评价HBsAg-DC瘤苗体外诱导抗HepG2.2.15的细胞毒性T淋巴细胞反应。方法将已构建含HBsAg基因的重组质粒pCR3.1-S转染培养第5天的DC;Westernblot和免疫荧光法鉴定转染基因表达;以MTT法测定DC诱导的抗HepG2.2.15特异性细胞毒作用。结果细胞表型鉴定结果显示诱导5天的DCCD1a、CD11c、CD86、CD80和HLA-DR表达量分别为55.0%、98.6%、86.1%、66.1%和88.9%;采用免疫荧光和Westernblot法研究表明HBsAg基因能在转染的DC中表达;MTT法检测特异性细胞毒作用结果显示不同效靶比,pCR3.1-S转染DC组均显示对HepG2.2.15肝癌细胞高效特异的杀伤活性,杀伤率分别为(52.3±2.8)%(E∶T为5∶1)、(64.6±2.4)%(10∶1)、(78.8±2.6)%(20∶1)、(82.1±2.4)%(40∶1),显著高于pCR3.1-DC组和DC组(P<0.05,n=4)。结论重组质粒转染的DC能够有效表达HBsAg;并在体外诱导出特异性CTL反应。BACKGROUND &OBJECTIVE: Up to now,there is no efficient immunotherapy for hepatocellular carcinoma (HCC). Dendritic cell (DC) vaccine could be a potential tool for HCC immunotherapy. This study was to evaluate the effect of dendritic cells (DCs) transfected with recombinant plasmid bearing hepatitis B virus surface antigen (HBsAg) gene, and the capability of generating specific cytotoxic T lymphocytes (CTL) response against HepG2.2.15 in vitro, which were induced by genetically modified DCs. METHODS: After cultured for 5 days, the DCs were transfected with pCR3.1 S by liposome. The HBsAg gene expression on pCR3.1 transfected DCs was identified by Western blot analysis, and immunofluorescence methods. The cytotoxicity against HepG2.2.15, which were induced by DCs, was tested by MTT assay. RESULTS: DCs up regulated the expression of CD1a (55.0%), CD11c (98.6%), CD86 (86.1%), CD80 (66.1%), and HLA DR (88.9%) after cultured for 5 days. Indirect immunofluo rescence,and Western blot analysis showed that HBsAg gene was expressed on transfected DCs. The death rates of HepG2.2.15 cells induced by DCs transfected with pCR3.1 S were (52.3±2.8)%(E∶T=5∶1), (64.6±2.4)%(10∶1), (78.8±2.6) (20∶1), (82.1±2.4)%(40∶1), while the pCR3.1 transfected and non transfected DCs only induced relatively lower cytotoxicity (P< 0.05, n=4). CONCLUSION: DCs transfected with recombined plasmid expressed HBsAg efficiently, and the genetically modified DCs evoke a higher CTL response in vitro.

关 键 词：树突状细胞 T淋巴细胞 免疫治疗 肝肿瘤 

分 类 号：R735.7[医药卫生—肿瘤]