天津医药

• 实验研究 •    

转染内皮祖细胞的脂质体与腺病毒载体的比较

孙宁   

  1. 天津医科大学第二医院
  • 收稿日期:2011-06-13 修回日期:2011-08-14 出版日期:2012-01-15 发布日期:2012-01-15
  • 通讯作者: 孙宁

Comparison Of Liposome And Adenovirus Vectors For HGF Gene Transfer To Endothelial Progenitor Cells

Sun Ning   

  • Received:2011-06-13 Revised:2011-08-14 Published:2012-01-15 Online:2012-01-15
  • Contact: Sun Ning

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摘要： 目的：比较脂质体和腺病毒作为肝细胞生长因子（HGF）基因转染内皮祖细胞载体的效力。方法：分离培养大鼠骨髓血管内皮祖细胞，并通过摄取DiL-acLDL、结合FITC-UEA-1及流式细胞术检测表面抗原CD133+进行鉴定。脂质体介导细胞转染：将细胞分为HGF质粒转染组（转染pIRES2-EGFP-HGF质粒）、空载质粒转染组（转染pIRES2-EGFP质粒）和空白对照组。病毒介导细胞转染：细胞分HGF病毒转染组（pAdxsi-GFP-HGF重组腺病毒转染）、空载病毒转染组（导入pAdxsi-GFP腺病毒）和空白对照组。荧光显微镜观察绿色荧光蛋白的表达，ELISA法检测HGF的表达，四甲基偶氮唑盐（MTT）检测细胞增殖能力。结果：脂质体和腺病毒载体均可成功将绿色荧光蛋白标记的HGF基因转入内皮祖细胞，其中腺病毒转染效率更高，最高可达80%以上，其表达HGF水平以及细胞增殖能力都明显强于脂质体转染。空载病毒转染组细胞增殖与空白对照组差异无统计学意义，而空载质粒转染组细胞增殖明显少于空白对照组。结论：腺病毒作为HGF基因转染内皮祖细胞的载体相对于脂质体载体具有更高效、低毒、高表达目的基因的优点。

关键词: 内皮祖细胞, 肝细胞生长因子, 脂质体, 腺病毒, 大鼠

Abstract: Objective: To compare the efficiency of liposome and adenovirus vector for hepatocyte Growth Factor (HGF) gene transfer to Endothelial Progenitor Cells (EPCs). Methods:1.Rat bone marrow-derived EPCs were separated by density gradient centrifugation and cultured, and identified by observing the experiment of uptaking DiL-acLDL and combining FITC-UEA-1 and analyzing the expression of CD133+ by Flow Cytometry. 2.Liposome mediated gene transfection: EPCs were divided to three group: pIRES2-EGFP-HGF group，pIRES2-EGFP group and blank control.3.Adenovirus mediated gene transfect- ion: EPCs were divided to three group: pAdxsi-GFP-HGF group, pAdxsi-GFP group and blank control. 4.The expression of green fluorescent protein (GFP) were observed by fluorescence microscope.The expression of HGF were detected by ELISA ,The proliferation were detected by MTT. Results: Liposome and adenovirus could deliver HGF gene marked by the green fluorescence protein (GFP) to EPCs. Adenovirus proved most effective with efficiencies of up to 80% with higher levels HGF expression and lower levels of cell death. Conclusion: Comparing the liposome, adenovirus vector for HGF gene transfer to EPCs has more efficiency, lower cell toxicity and higher HGF expression.

Key words: Endothelial Progenitor Cells, Hepatocyte Growth Factor, Liposome, Adenovirus, Rat