中华普通外科杂志
中華普通外科雜誌
중화보통외과잡지
CHINESE JOURNAL OF GENERAL SURGERY
2009年
9期
736-739
,共4页
张靖%付彦超%康春生%张庆瑜%王涛%张洁
張靖%付彥超%康春生%張慶瑜%王濤%張潔
장정%부언초%강춘생%장경유%왕도%장길
胃肿瘤%肿瘤转移%腺病毒%人%蛋白激酶类%环氧合酶2
胃腫瘤%腫瘤轉移%腺病毒%人%蛋白激酶類%環氧閤酶2
위종류%종류전이%선병독%인%단백격매류%배양합매2
Stomach neoplasms%Neoplasm metastasis%Adenovirus%human%Protein kinases%Cyclooxygenase-2
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-Aktl+COX-2(rAd5-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rAd5-HK和治疗组rAd5-A+C中Aktl、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48 h后可显著抑制Aktl和COX-2 mRNA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);rAd5-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和卒载组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.
目的 構建靶嚮性蛋白激酶B1(Aktl)和環氧閤酶2(COX-2)短髮夾RNA(shRNA)腺病毒載體,研究其對SGC-7901人胃腺癌細胞侵襲和轉移的抑製效果.方法 構建腺病毒載體pGSadeno-Aktl+COX-2(rAd5-A+C)併體外轉染SGC-7901人胃癌細胞株後,用實時定量PCR和蛋白印跡分彆檢測對照組SGC-7901、空載組rAd5-HK和治療組rAd5-A+C中Aktl、COX-2 mRNA和蛋白質的錶達,其中以對照組SGC-7901、空載組rAd5-HK作為陰性對照.用ELISA法分彆檢測它們的MMP-2和MMP-9含量;用Transwell法分析它們的侵襲和轉移能力.結果 構建的rAd5-A+C重組腺病毒載體在轉染SGC-7901細胞48 h後可顯著抑製Aktl和COX-2 mRNA的錶達,而治療組rAd5-A+C的Aktl和COX-2蛋白錶達量與對照組和空載組相比分彆下調68.4%和60.2%(均P<0.01);rAd5-A+C組中MMP-2和MMP-9含量分彆為(39.7±1.7)ng/ml(31.3±3.6)ng/ml,明顯低于對照組SGC-7901(278.4±15.5)ng/ml(225.4±15.1)ng/ml和卒載組rAd5-HK(275.5±2.1)ng/ml、(226.0±23.3)ng/ml(P=0.01、P=0.021).結論 腺病毒介導的靶嚮性Aktl和COX-2shRNA可抑製人胃腺癌細胞的侵襲和轉移.
목적 구건파향성단백격매B1(Aktl)화배양합매2(COX-2)단발협RNA(shRNA)선병독재체,연구기대SGC-7901인위선암세포침습화전이적억제효과.방법 구건선병독재체pGSadeno-Aktl+COX-2(rAd5-A+C)병체외전염SGC-7901인위암세포주후,용실시정량PCR화단백인적분별검측대조조SGC-7901、공재조rAd5-HK화치료조rAd5-A+C중Aktl、COX-2 mRNA화단백질적표체,기중이대조조SGC-7901、공재조rAd5-HK작위음성대조.용ELISA법분별검측타문적MMP-2화MMP-9함량;용Transwell법분석타문적침습화전이능력.결과 구건적rAd5-A+C중조선병독재체재전염SGC-7901세포48 h후가현저억제Aktl화COX-2 mRNA적표체,이치료조rAd5-A+C적Aktl화COX-2단백표체량여대조조화공재조상비분별하조68.4%화60.2%(균P<0.01);rAd5-A+C조중MMP-2화MMP-9함량분별위(39.7±1.7)ng/ml(31.3±3.6)ng/ml,명현저우대조조SGC-7901(278.4±15.5)ng/ml(225.4±15.1)ng/ml화졸재조rAd5-HK(275.5±2.1)ng/ml、(226.0±23.3)ng/ml(P=0.01、P=0.021).결론 선병독개도적파향성Aktl화COX-2shRNA가억제인위선암세포적침습화전이.
Objective To construct a short hairpin RNA (shRNA) adenovirus vector targeting Aktl (protein kinase B1, PKBI/Aktl) and cyclooxygenase-2 (COX-2) and study its effects on the invasion and metastasis of SGC-7901 human gastric adenocarcinoma 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 PacI 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、rAd5-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 cells. MMP-2 and MMP-9 contents in treatment group rAd5-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/ml and rAd5-HK (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 adenocacinoma cells. Conclusions Adenovirus-mediated targeting Aktl and COX-2 shRNA can inhibit the invasion and metastasis of SGC-7901 human gastric adenocarcinoma cells.
