摘要
目的:探讨PEG-PEI/Fe3O4纳米磁流体-TK对肝癌细胞HepG2凋亡的影响。方法:构建重组质粒PEGFP-AFP-TK并通过纳米磁流体分别转染入AFP表达阳性的肝癌细胞HepG2及AFP表达阴性的肝癌细胞SMMC7721。转染12h后,于荧光显微镜下动态观察;48h后,RT-PCR和Western blot分析HepG2细胞中HSV-TK基因的表达情况,用MTT法检测HepG2细胞的存活,用流式细胞分析HepG2细胞的凋亡。结果:纳米磁流体能将重组质粒PEGFP-AFP-TK转入HepG2细胞,并且其转染效率高于脂质体;RT-PCR及Western blot证明转染的HepG2细胞有HSV-TK基因的表达;MTT及流式细胞分析说明HSV-TK基因发挥杀伤细胞作用。结论:纳米磁流体能将重组质粒PEGFP-AFP-TK转染HepG2细胞并获得表达,PEG-PEI/Fe3O4纳米磁流体-TK可显著抑制HepG2细胞增殖,可望成为肝癌基因治疗的新型生物制剂之一。
Objective： To investigate the effect of PEG-PEI/Fe3O4 Nano-magnetic fluid-TK on Apoptosis of liver cancer cells（HepG2）in vitro. Methods： Construcr eeombinant plasmid PEGFP-AFP-TK was constructed, which was delivered by nano-magnetic fuilds into AFP positive HepG2 cells and AFP negative SMMC7721 cells. The fluorescence was detected in order to evaluate the transfection rate 12h after transfeetion RT-PCR and Western blot were used to detect expression of HSV-TKgene at 48 h after transfeetion. MTT assay was used to evaluate the effect of HSV-TK on the proliferation of HepG2 cells, Flow cytometry was used to analyze the apoptosis of HepG2 cells. Results ： Nano-Magnetic fluids delivered plasmid PEGFP-AFP-TK into HepG2 cells and HSV-TK gene was successfully expressed. The transfection eficacy of nano-magnetic fluids was superior than lipofectamine in HepG2 ceils. RT-PCR and Western blot demonstrated that TK gene was expressed in HepG2 cells after being transfected with nano-magnetic fluids/PEGFP-AFP-TK. MTT and flow eytometry showed HSV-TK gene exerts a cell-killing effect. Conclusions：Nano-Magnetic fluids could successfully deliver PEGFP-AFP-TKinto HepG2 cells and induce expression of HSV-TK,it may become a promising gene vector for liver cancer gene therapy. AFP enhancer can specifically enhance the expression of target gene within the cell with positive AFP.
出处
《重庆医科大学学报》
CAS
CSCD
北大核心
2009年第6期661-664,共4页
Journal of Chongqing Medical University
基金
国家自然科学基金资助项目(30672047)
中国博士后科学基金资助项目(20060390884)
湖南省博士后科学基金资助项目(2006FJ4240)
