婴儿双歧杆菌介导HSV-TK/GCV对裸鼠肾癌组织细胞凋亡及p-Stat3、Bcl-xL表达的影响  

Effects of Bifidobacterium infantis-mediated HSV-TK/GCV suicide gene therapy system on apoptosis and p-Stat3 and Bcl-xL expression in renal carcinoma cells transplanted nude mice

作　　者：任劲[1] 肖啸[1] 蒋立[1] 杨柒[1] 唐伟[1] 

机构地区：[1]重庆医科大学附属第一医院泌尿外科,重庆400016

出　　处：《第三军医大学学报》2014年第4期375-380,共6页Journal of Third Military Medical University

基　　金：重庆市医学科学技术研究重点项目(2011-1-012)~~

摘　　要：目的评价婴儿双歧杆菌介导单纯疱疹病毒胸苷激酶(HSV-TK)/丙氧鸟苷(GCV)自杀基因治疗系统对裸鼠肾癌的疗效,初步探讨其诱导肾癌组织细胞凋亡及对肾癌组织p-Stat3、Bcl-xL表达的影响。方法采用肾癌786-O细胞悬液皮下注射法建立裸鼠肾癌模型,将模型裸鼠分为4组,分别经尾静脉注射生理盐水(生理盐水组)、婴儿双歧杆菌(婴儿双歧杆菌组)、携带空质粒pGEX-5X-1的婴儿双歧杆菌(空质粒组)、携带重组质粒pGEX-TK的婴儿双歧杆菌(重组质粒组),再联合腹腔注射GCV。治疗4周结束后,称取各组裸鼠质量及荷瘤质量。TUNEL染色法检测各组肾癌组织细胞凋亡情况。免疫组化染色法检测各组肾癌组织p-Stat3、Bcl-xL蛋白表达情况。Real-time PCR检测各组肾癌组织Stat3、Bcl-xL mRNA表达水平。Western blot检测各组肾癌组织p-Stat3、Bcl-xL蛋白表达水平。结果治疗后,重组质粒组较空质粒组、婴儿双歧杆菌组和生理盐水组裸鼠质量增加(P<0.05),荷瘤质量降低(P<0.05)。TUNEL染色结果显示重组质粒组凋亡指数明显高于空质粒组、婴儿双歧杆菌组和生理盐水组(P<0.01)。免疫组化染色结果显示重组质粒组肾癌组织p-Stat3、Bcl-xL蛋白表达明显低于空质粒组、婴儿双歧杆菌组和生理盐水组(P<0.01)。重组质粒组肾癌组织Stat3、Bcl-xL mRNA及p-Stat3、Bcl-xL蛋白表达水平较空质粒组、婴儿双歧杆菌组和生理盐水组明显下调(P<0.01)。结论婴儿双歧杆菌介导HSV-TK/GCV自杀基因治疗系统明显抑制裸鼠肾癌生长,可能通过抑制Stat3活化进而阻断Stat3信号转导通路,下调其下游抗凋亡蛋白Bcl-xL的表达来诱导裸鼠肾癌组织细胞凋亡,发挥抑瘤作用。Objective To evaluate the curative effect of Bifidobacterium infantis （BI）-mediated herpes simplex virus thymidine kinase （HSV-TK）/ganciclovir （GCV） suicide gene therapy system on renal carcinoma in nude mice and investigate the effects of HSV-TK/GCV on renal carcinoma cells apoptosis and p-Stat3 and Bcl-xL expression.MethodsThe renal carcinoma model was established by subcutaneous injection of human renal carcinoma cell line 786-O into nude mice. The model mice were randomly divided into 4 groups, which were treated by intraperitoneal injection of GCV in combination with tail vein injection of normal saline （normal saline group）, BI （BI group）, BI carrying empty plasmid BI-pGEX-5x-1 （empty plasmid group）, and BI carrying recombinant plasmid BI-pGEX-TK （recombinant plasmid group）. After 4 weeks treatment, tumor weights and nude mice weights were recorded. The apoptosis of renal carcinoma tissue cells was determined by TUNEL assay.The expression of p-Stat3,Bcl-xL protein was determined by immunohistochemical staining. The expression levels of Stat3,Bcl-xL mRNA and expression levels of p-Stat3,Bcl-xL protein were detected by real-time fluorescent quantitative PCR（real-time PCR） and protein imprinting method （Western blotting） respectively.ResultsAfter treatment, comparing with the empty plasmid group,the bifidobacterium infants group and the normal saline group,the nude mice weights of recombinant plasmid group increased（P〈0.05）, the tumor weights decreased（P〈0.05）. TUNEL assay showed the apoptosis index of recombinant plasmid group was obviously higher than that of the empty plasmid group, the bifidobacterium infants group and the normal saline group（P〈0.01）. Immunohistochemical staining showed that the expression of p-Stat3, Bcl-xL protein of recombinant plasmid group is significantly lower than that of the empty plasmid group, the bifidobacterium infants group and the normal saline group（P〈0.01）. The expression levels of Stat3, Bcl-xL mRNA of recombinan

关 键 词：婴儿双歧杆菌 HSV-TK GCV 基因治疗 肾癌 P-STAT3 Bcl-xL 

分 类 号：R37[医药卫生—病原生物学] R73-362[医药卫生—基础医学]