Flt-1基因对BT325细胞VEGF蛋白表达及细胞增殖的影响  

Effect of Flt-1 gene on the expression of VEGF and proliferation in BT325 cells

作　　者：张鹏飞[1] 周建华[2] 周婉琦 张亚卓[3] 王红云[3] 

机构地区：[1]首都医科大学附属北京天坛医院神经外科,100050 [2]中国医学科学院药物研究所 [3]北京市神经外科研究所

出　　处：《中华神经外科杂志》2014年第9期941-944,共4页Chinese Journal of Neurosurgery

基　　金：北京市自然科学基金资助项目（7072015）;北京市科技新星计划资助项目（2007A042）

摘　　要：目的 探讨Flt-1基因对BT325胶质瘤细胞VEGF蛋白表达及细胞增殖的影响.方法 从人脐静脉内皮细胞中提取总RNA,逆转录聚合酶链式反应（RT-PCR）扩增Flt-1基因片段,基因重组技术将Flt-1基因片段插入真核表达载体pEGFP-C1中,双酶切及DNA测序鉴定pEGFP-C1-Flt-1重组质粒.pEGFP-C1-Flt-1重组质粒转染BT325胶质瘤细胞,转染后72hELISA法检测培养上清液中VEGF含量,MTT法检测细胞的增殖情况.结果 RT-PCR方法证实从人脐静脉血内皮细胞中成功获得1 384 bp Flt-1基因片段;经双酶切分析和DNA测序鉴定表明重组质粒pEGFP-C1-Flt-1构建成功;ELISA检测发现,BT325细胞被转染后72 h VEGF含量转染组为（56.3±41.2） pg/ml,空载体转染组为（95.5±35.3） pg/ml（P＜0.05）;MTT法检测发现,转染组和空载体转染组的生存率分别为（42.6±3.7）％和（92.8±6.6）％（P＜0.05）.结论 Flt-1基因可抑制BT325细胞VEGF的蛋白表达,抑制细胞增殖.Objective To study the Effects of Flt-1 gene on the expression of VEGF and proliferation in BT325 cells.Methods The total RNA was extracted from human umbilical vein endothelial cells as the template and Flt-1 gene was amplified by reverse transcription polymerase chain reaction（RT-PCR）.By using gene recombination technique,the Flt-1 cDNA was inserted into eukaryotic expression vector pEGFP-C1.The recombinant plasmid was identified by restriction enzyme digestion and sequence analysis.It was infected into BT325 cells.The quantity of VEGF protein in supernatant was detected by ELISA.The proliferation abilities were measured by MTT and FCM.Results The Flt-1 gene could be obtained by RT-PCR.The restriction enzyme digestion and PCR suggested that the recombinant plasmids were constructed successfully.72 hours after transfection,the VEGF protein levels were decreased （transfection group （56.3 ±41.2） pg/ml,untransfection group （95.5 ±35.3） pg/ml（P ＜0.05） and the proliferation rate of cells was inhibited transfection group （42.6 ± 3.7） ％ and untransfection group （92.8 ± 6.6） ％ （P ＜ 0.05）.Conclusions Flt-1 gene could inhibit the expression of VEGF and BT325 cells proliferation and induce their apoptosis in vitro.

关 键 词：Flt-1基因 基因重组 BT325细胞 

分 类 号：R739.41[医药卫生—肿瘤]