腺病毒介导的靶向蛋白激酶B1和环氧合酶2短发夹RNA抑制人胃腺癌细胞的侵袭和转移  

Inhibitory effects of adenovirus-mediated shRNA targeting Aktl and COXo2 on invasion and metastasis of SGC-7901 cells

作　　者：张靖[1] 付彦超[1] 康春生[2] 张庆瑜[1] 王涛[1] 张洁[1] 

机构地区：[1]天津医科大学总医院消化科,300052 [2]天津医科大学总医院神经外科,300052

出　　处：《中华普通外科杂志》2009年第9期736-739,共4页Chinese Journal of General Surgery

基　　金：天津市自然科学基金资助项目（07JCZDJC07700）

摘　　要：目的构建靶向性蛋白激酶B1（Aktl）和环氧合酶2（COX-2）短发夹RNA（shRNA）腺病毒载体，研究其对SGC-7901人胃腺癌细胞侵袭和转移的抑制效果。方法构建腺病毒载体pGSadeno-Akt1＋COX-2（tad5-A＋C）并体外转染SGC-7901人胃癌细胞株后，用实时定量PCR和蛋白印迹分别检测对照组SGC-7901、空载组tAd5-HK和治疗组tAd5-A＋C中Akt1、COX-2 mRNA和蛋白质的表达，其中以对照组SGC-7901、空载组rAd5-HK作为阴性对照。用ELISA法分别检测它们的MMP-2和MMP-9含量；用Transwell法分析它们的侵袭和转移能力。结果构建的rAd5-A＋C重组腺病毒载体在转染SGC-7901细胞48h后呵显著抑制kktl和COX-2mRNA的表达，而治疗组rAd5-A＋C的Aktl和COX-2蛋白表达量与对照组和空载组相比分别下调68．4％和60．2％（均P〈0．01）；tad5-A＋C组中MMP-2和MMP-9含量分别为（39．7±1．7）ng／ml、（31．3±3．6）ng／ml，明显低于对照组SGC-7901（278．4±15．5）ng／ml、（225．4±15．1）ng／ml和空载组rAd5-HK（275．5±2．1）ng／ml、（226．0±23．3）ng／ml（P=0．01、P=0．021）。结论腺病毒介导的靶向性Aktl和COX-2shRNA可抑制人胃腺癌细胞的侵袭和转移。Objective To construct a short hairpin RNA （shRNA） adenovirus vector targeting Aktl （protein kinase B1, PKB1/Aktl ） and cyclooxygenase-2 （COX-2） and study its effects on the invasion and metastasis of SGC-7901 human gastric ade cells. Methods Aktl and COX-2 shRNA expression frames were subcloned to pGSadeno adenovirus vector by homologous recombination technology to construct pGSadeno-Aktl ＋ COX-2 （rAdS-A ＋ C ） vector. After screening and amplification, the recombinant adenovirus vector was digested with Pacl and transfected into SGC-7901 cells, the titer and transfection efficiency were detected by fluorescent microscopy. Aktl and COX-2 mRNA and protein expression was identified by real-time PCR and Western blot. MMP-2 and MMP-9 contents in control group SGC-7901 ,rAdS-HK and treatment group rAdS-A ＋ C were detected by ELISA assay and transwell assay analyzed cell invasion and metastasis ability. Results Adenovirus vector rAdS-A ＋ C was successfully constructed and it dramatically down-regulated Aktl and COX-2 mRNA and protein expression in SGC-7901 gastric cancer ceils. MMP-2 and MMP-9 contents in treatment group rAdS-A ＋ C were respectively （39. 7 ± 1.7） ng/ml, （31.3 ±3.6） ng/ml, and they were lower than those in control group SGC-7901 （278.4± 15.5） ng/ml,（225.4±15.1） ng/mlandrAd5-nK （275.5±2.1） ng/ml, （226.0 ±23.3） ng/ml （P= 0.01, P=0. 021 ）. Transwell assay showed treatment group rAd5-A ＋ C significantly inhibited the invasion and metastasis of SGC-7901 gastric adenocaeinoma cells. Conclusions Adenovirus-mediated targeting Aktl and COX-2 shRNA can inhibit the invasion and metastasis of SGC-7901 human gastric adenocarcinoma cells.

关 键 词：胃肿瘤 肿瘤转移 腺病毒 人 蛋白激酶类 环氧合酶2 

分 类 号：R686[医药卫生—骨科学]