文章摘要
鲁 悦,杨 航,袁 林,徐国平,姚志峰,刘永彪.多聚乙烯亚胺介导肿瘤细胞基因转染[J].南京医科大学学报,2012,(12):1684~1689
多聚乙烯亚胺介导肿瘤细胞基因转染
The polyethylenimine-mediated gene transfection in tumor cells
投稿时间:2012-07-03  
DOI:10.7655
中文关键词: 多聚乙烯亚胺  转染  细胞周期  Hela细胞  A549细胞
英文关键词: polyethyleneimine  transfection  cell cycle  Hela cell  A549 cell
基金项目:国家自然科学基金资助(30870741)
作者单位
鲁 悦 1南京医科大学附属肿瘤医院放疗科,江苏 南京 210009
2南京医科大学第一附属医院放疗科,江苏 南京 210029 
杨 航 南京医科大学第一附属医院放疗科,江苏 南京 210029 
袁 林 南京医科大学第一附属医院放疗科,江苏 南京 210029 
徐国平 南京医科大学第一附属医院放疗科,江苏 南京 210029 
姚志峰 南京医科大学第一附属医院放疗科,江苏 南京 210029 
刘永彪 南京医科大学第一附属医院放疗科,江苏 南京 210029 
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中文摘要:
      目的:利用阳离子多聚物多聚乙烯亚胺(PEI)纳米凝胶将增强型绿色荧光蛋白(enhanced green fluorescence protein,EGFP)转入同步化的肿瘤细胞中并检测绿色荧光蛋白的表达率,探讨PEI作为载体介导肿瘤细胞基因转染中细胞周期对转染效率的影响?方法:通过6 MV X线照射及药物阻滞使Hela及A549细胞同步化,在5个PEI/DNA梯度比(2/2?4/2?6/2?8/2?10/2 ?滋g/?滋g)下利用PEI介导EGFP报告基因转染进入上述同步化细胞,流式细胞仪和荧光显微镜下测定绿色荧光蛋白表达率,通过梯度试验分析细胞周期时相与表达率之间的关系?结果:在不同增殖状态及PEI/DNA比值梯度下,一定剂量-时间的X射线和顺铂可以使Hela细胞同步化于S期,一定剂量-时间的艾素可以使A549细胞同步化于G2/M期,而血清饥饿法培养则可以使细胞阻滞于G0/G1期,在不同细胞周期时相下EGFP的转染效率不同,两种同步化细胞均表现出S期及G2/M期细胞的转染率显著高于对照组(P < 0.05)和G0/G1期细胞(P < 0.0001)?结论:PEI导入基因的表达效率受细胞周期制约,PEI介导的基因转染受到宿主细胞摄取?转运入核及转录能力的影响,而该进程都是细胞周期依赖性的?
英文摘要:
      Objective:To investigate the feasibility of gene transfection of Hela and A549 cells mediated by cationic polymer-polyethylenimine(PEI) and evaluate the effect of cell cycle status on efficiency of PEI-mediated gene delivery. Methods: Hela and A549 cells were synchronized into different phases of the cell cycle by 6 MV X ray radiation or drugs or serum deprivation. The synchronized cells were transfected using PEI carrier at various PEI/DNA ratio(P/D=2/2,4/2,6/2,8/2,10/2?滋g/?滋g),and the transient transfection efficiency of enhanced green fluorescence protein (EGFP) gene in these cells was determined by flow cytometry and fluorescent microscopy. The relationship between cell cycle status and the efficiency of transgene expression was analyzed. Results:Hela cell could be synchronized in S phase by X ray radiation or incubation with Cisplatin(PDD),and A549 cell could be synchronized in G2/M phase by incubation with Docetaxel(TXT) at certain dose-time condition. Both Hela and A549 cells were synchronized in G0/G1 phase serum deprivation. Transfection efficiency in synchronized cells was different at different phases of the cell cycle. PEI showed the minimal transfection efficiency at G0/G1 phase and the medium at G2/M phase and the maximum at S phase(P < 0.05,S,G2/M phase group vs control; P < 0.0001,S,G2/M phase group vs G0/G1 phase). Conclusion:PEI is an effective and safe carrier for gene delivery. Cell cycle status restricted the expression efficiency of transfection mediated by PEI. Transfection using PEI is influenced by cellular uptake,transportation into nucleus and transcription,and those processes are cell-cycle-dependent.
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