稳定表达人VEGF165的NIH/3T3细胞株的筛选与鉴定

 目的 培养NIH/3T3细胞,并进行慢病毒载体介导的VEGF165基因转染,为建立转基因小鼠血管瘤模型奠定基础.方法 采取贴壁培养法分离培养NIH/3T3细胞,细胞随机分为3组,分别为Lenti-VEGF165-EGFP 重组慢病毒载体转染组、Lenti-EGFP慢病毒转染组和未转染组,转染后72 h荧光显微镜观察细胞绿色荧光蛋白的表达,流式细胞仪分选后用ELISA方法检测VEGF165外分泌.结果 ELISA 检测转染后小鼠NIH/3T3细胞,Lenti-VEGF165-EGFP 重组慢病毒载体感染组、Lenti-EGFP慢病毒转染组和未转染组培养上清中VEGF165浓度分别为(205±15)、0、0 ng/L(P<0.05) 结论 NIH/3T3细胞获得慢病毒载体介导的VEGF165表达基因修饰且稳定传代,为下一步血管瘤动物模型研究奠定了基础.

Screening and identificating of NIH/3T3 cell strains to VEGF165 mediated by lentiviral vectors

Objective To isolate and culture NIH/3T3 cells and transfect the cells with lentiviral vectors. Methods The cells were randomly divided into 3 groups （lenti - VEGF165 - EGFP transfection group, lenti - EGFP transfection group, and non - transfection group） to observe green fluorescent proteins and infection efficiency after 72 hours under the inverted fluorescent microscope. The external secretion of VEGF was measured by ELISA. Results After mouse NIH3T3 cell transfection, the concentrations of VEGF in supernatant in the lenti - VEGF165 - EGFP transfecti0n group, lenti - EGFP transfection group, and non - transfection group were 205± 15,0,0 ng/L, respectively（ P 〈 0. 05 ）. Conclusions Mouse NIH3T3 cells have been transfected by lentivirus, serving as a potential animal model of hemangioma by mouse NIH3T3 cell VEGF gene implantation.