转染T-bet基因的人胃癌SGC-7901细胞分泌IFN-γ的作用  

Adeno-associated Virus Mediated T-bet Gene Transfer into SGC-7901 Cell to Regulate IFN-γ Production

作　　者：邱谷风[1] 王锁英[2] 王胜军[1] 邵启祥[1] 马洁[1] 杨敏[1] 许小朋[1] 毛朝明[1] 苏兆亮[1] 黄新祥[1] 许化溪[1] 

机构地区：[1]江苏大学检验医学研究所免疫研究室,镇江212013 [2]江苏大学附属医院儿科,镇江212001

出　　处：《生物医学工程学杂志》2009年第3期606-609,619,共5页Journal of Biomedical Engineering

基　　金：国家自然科学基金资助项目(30471627,30300169);江苏省社会发展项目资助(BS2007041)

摘　　要：采用基因克隆与腺相关病毒包装技术，构建人Thl细胞特异性转录因子T-bet基因的腺相关病毒载体，并转染入胃癌细胞系SGC-790I细胞。检测转染细胞IFN-γ的分泌功能及其对SGC-7901细胞生物学作用的影响。结果：（1）获得含有T-bet基因的重组腺相关病毒（rAAV—eGFP-T—bet）；（2）用rAAV-eGFP—T-bet感染SGC-7901细胞后，约30％～40％的细胞出现绿色荧光。对被感染细胞进行RT—PCR，扩增出1670bp的T-bet，并经Westernblot证实；（3）转染后的细胞测得388bp的IFN-γ目的条带和IFN-γ蛋白的表达。由此表明，人T-bet基因的腺相关病毒载体rAAV—eGFP—T-bet，可成功转染SGC-7901，并致使转染细胞分泌IFN-γ。为进一步开展T-bet基因干预治疗肿瘤的研究奠定了基础。In order to investigate the effect of T-bet on malignant cells, we selected SGC-7901, a kind of human gastric carcinoma cell line, and used gene clone technique and adeno-associated virus （AAV） packing technology, thus obtaining a recombinant rAAV-eGFP-T-bet and T-bet gene-transfected SGC-7901 cells. Then the function of T- bet gene-infeeted SGC-7901 cells was researched by detecting the levels of IFN-γ and T-bet production. The results showed： （1）It was verified that rAAV-T-bet＇s packing was completed;（2）After SGC-7901 cells was transfected by rAAV-eGFP-T-bet, a green fluorescence was found in about 30%-40% SGC-7901s, and the gene of 1 670 bp （T- bet） and 388 bp （IFN-γ） were generated from SGC-7901s cells; （3） The proteins of IFN-γ and T-bet secreted by SGC-7901 cells were also detected. These reveal that SGC-7901 cell is efficiently infected by rAAV encoding T-bet, which can induce transfected cells to secret IFN-γ. It may be useful in the researches on cancer immune therapy of transfecting T-bet gene.

关 键 词：T-BET基因 基因克隆 重组质粒 腺相关病毒载体 IFN-Γ 

分 类 号：R735.2[医药卫生—肿瘤]