靶向舌癌Tca8113细胞血管内皮细胞生长因子基因的siRNA表达载体构建及体外抑制作用  

VEGF-siRNA expression construction vector and its inhibition of vascular endothelial growth factor in Tca8113 cells

作　　者：姚志文[1] 刘唯[1] 程由勇[1] 李红玖[1] 李春华[1] 于大海[2] 

机构地区：[1]广州医学院附属深圳沙井医院口腔科,广东深圳518104 [2]广西医科大学口腔医学院,广西南宁530021

出　　处：《中国医药导报》2012年第3期28-30,共3页China Medical Herald

基　　金：国家自然科学基金资助项目(编号:30360111)

摘　　要：目的构建靶向舌癌Tca8113细胞血管内皮细胞生长因子(VEGF)基因的siRNA表达载体并在体外检测其对VEGF基因表达的抑制作用。方法设计合成靶向舌癌Tca8113细胞VEGF基因的siRNA cDNA序列并与Psilencer2.1-U6-neo连接,构建VEGF-siRNA表达载体(Pu-VEGF-siRNA),酶切鉴定和测序确认后,经脂质体Lipofectamine2000转染入Tca8113细胞,以空载体组为实验对照组(Pu-HK),未转染组为空白对照组。采用酶联免疫吸附(ELISA)和免疫组化方法检测染后Tca8113细胞VEGF蛋白表达差异。结果经过双酶切鉴定和测序分析,成功构建了靶向Tca8113细胞VEGF基因的siRNA表达载体。与实验对照组和空白对照相比较,转染Pu-VEGF-siRNA后Tca8113细胞的VEGF蛋白表达明显下降(P<0.05),但两对照组间差异无统计学意义(P>0.05)。结论成功构建了靶向舌癌Tca8113细胞VEGF基因的siRNA表达载体,且该载体在体外能有效抑制Tca8113细胞VEGF蛋白的表达。Objective To construct the vector carrying siRNA of human vascular endothelial growth factor（VEGF） and to evaluate its inhibitory effect on VEGF in Tca8113 cells.Methods Sequencer of short hairpin RNA targeting VEGF of Tca8113 cells was designed and synthesized.Psilencer 2.1-U6-neo-VEGF-siRNA（Pu-VEGF-siRNA） was constructed and transfected to Tca8113 cells by lipofectamine 2000.Eukaryotic expression vector（Pu-HK） as the experiment control group,non-transfection cell was used as negative control group,all of which were transfected to Tca8113 cells by lipofectamine 2000.The expression of VEGF protein in Tca8113 cells transfected with Pu-VEGF-siRNA was detected by immunohistochemistry and enzyme linked immunosorbent assay（ELISA） respectively.Results After evaluation and sequencing,Pu-VEGF-siRNA was successfully constructed.Compared to the negative and experiment control groups,the expression of VEGF protein were significantly decreased in the Pu-VEGF-siRNA group（P 0.05）;but there was no significant difference between the two control groups（P 0.05）.Conclusion Pu-VEGF-siRNA expression vector is successfully constructed and it can significantly inhibit VEGF gene expression in Tca8113 cells.

关 键 词：血管内皮细胞生长因子 RNA干扰 舌癌 

分 类 号：R329.28[医药卫生—人体解剖和组织胚胎学]