【摘要】 ①目的研究Twist基因转染人胃癌细胞株MKN28对血管内皮生长因子(VEGF)表达的影响。②方法利用DNA的限制性内切酶双酶切技术鉴定重组质粒PCDNA3.1-Twist、脂质体介导转染技术和G418筛选得到稳定表达Twist蛋白的Twist-MKN28(PCD-NA3.1-Twist转染的MKN28胃癌细胞株)细胞模型;采用Western blot检测细胞中Twist的表达;采用ElISA法检测细胞上清液中VEGF的蛋白表达量;采用半定量逆转录多聚酶链反应RT-PCR技术检测细胞中VEGF的mRNA表达量。③结果阳性质粒组Twist-MKN28细胞的Twist蛋白表达强于空白质粒组PCDNA3.1-MKN28和未转染组MKN28;Twist-MKN28组细胞上清液VEGF蛋白分泌明显增加,与PCDNA3.1-MKN28组和MKN28组比较差异有显著性(P<0.05),Twist-MKN28组细胞VEGF mRNA半定量结果和MKN28组、PCD-NA3.1-MKN28组比较,其表达量分别增加28.3%和26.5%,而PCDNA3.1-MKN28组和MKN28组比较无明显改变。④结论 Twist可促进人胃癌细胞株MKN28中VEGF的表达。更多还原

【Abstract】 Objective To study the effect of transfected Twist gene on expression of VEGF in human gastric carcinoma cells.Methods The re-constructed plasmid PCDNA3.1-Twist was proved by restriction endonuclease technology.MKN28 cells,a human gastric carcinoma cell line,were transfected with PCDNA3.1-Twist plasmid using lipofectanine transfecting technique.Transfected cells were selected with G418.Expression of Twist protein was detected by Western blot in cells transfected Twist gene.VEGF protein of cell culture supernatants was dectected by ElISA methed.The expression of VEGF mRNA was deteced by RT-PCR method.Results MKN28 cells transfected with PCDNA3.1-Twist plasmid showed stronger intracellular expression of Twist protein than MKN28 cells transfected with PCDNA3.1 and MKN28 cells without the transfection.The secrection of VEGF protein in cell culture supernatants of MKN28 cells transfected with PCDNA3.1-Twist plasmid was signifient increased compared with control(P<0.05).The VEGF mRNA expression of MKN28 cells transfected with PCDNA3.1-Twist plasmid was higher compared with cells transfected with PCDNA3.1 plasmid and MKN28 cells without transfection.They increased 28.3% and 26.5% respectively.Conclusion Twist can increase expression of VEGF in MKN28 cells.更多还原