整合素α_v重组腺病毒的构建和鉴定及对人肝癌细胞黏附特性的影响  

Construction of a recombinant adenoviral vector carrying integrinα_v and the influence of integrinα_v on the adhesion characteristics of SMMC7721 cells

作　　者：李旭艳[1] 刘靖华[1] 李志杰[1] 莫永炎[1] 刘亚伟[1] 刘志锋[1] 邓鹏[1] 姜勇[1] 

机构地区：[1]南方医科大学病理生理学教研室,广东省功能蛋白质组学重点实验室,广东广州510515

出　　处：《中国危重病急救医学》2006年第7期413-416,F0005,共5页Chinese Critical Care Medicine

基　　金：国家"973"计划重点基础研究项目(2002CB513005);广东省科技计划项目(A1090202);广州市科技计划项目(2001Z035011);广东省自然科学基金重点项目(13058)

摘　　要：目的构建表达人整合素αv亚基(整合素αv)重组腺病毒,并在人肝癌细胞株SMMC7721细胞中表达,探讨整合素αv对肝癌细胞黏附特性的影响。方法将人整合素αvcDNA克隆到腺病毒穿梭质粒pAd-Track巨细胞病毒(CMV)启动子下游,与腺病毒骨架质粒pAdEasy1在细菌内同源重组,在HEK293A细胞中包装成整合素αv重组腺病毒。以重组腺病毒感染SMMC7721细胞24h后,用蛋白质免疫印迹法(West-ernblot)检测整合素αv的表达;通过黏附实验研究其对SMMC7721细胞黏附的影响。结果构建了整合素αv重组腺病毒载体并制备出高滴度重组腺病毒。重组腺病毒感染SMMC7721细胞后,整合素αv能够正确表达,并且SMMC7721细胞黏附率显著高于对照组和空白组(P<0.01)。结论整合素αv基因在SMMC7721细胞的黏附中起重要作用,是肿瘤转移的重要分子基础。Objective To establish human integrin-αv adenovirus expressed in human liver cancer SMMC-7721 cells and analyze the characteristics of integrin-αvadhesion to liver cancer cells. Methods Human integrin-αv cDNA was cloned into the plasmid pAdTrack-CMV. Then, pAdTrack-integrin-αv. was cotransformed with adenoviral backbone vector into E. coli strain BJ5183. The recombinant adenoviral plasmid was identified by restriction analysis with Pac I and transfected into HEK-293A cells to package and amplify recombinant adenovirus particles. Western blot showed that integrin-αv gene was exactly transcripted and expressed in SMMC-7721 cells. Twenty-four hours after transfection, the effect of integrin-αv on adhesion of SMMC-7721 cells was detected by adhesion experiment. Results Recombinant aenovirus vector of integrin-αv gene was successfully constructed and high titers of recombinant adnovirus was obtained. Adhesion cell count was significantly higher in integrin-αv-transfected cells than in untransfected and Ad-null-transfected cells （P〈0. 01 ）. Conclusion Over-espression of integrinαv promotes adhesion of SMMC-7721 cells, is an important molecular mechanism of tumor metastasis.

关 键 词：整合素 细胞黏附 腺病毒载体 

分 类 号：R735.7[医药卫生—肿瘤] R394[医药卫生—临床医学]