三种基因转导方法在不同代龄复制性衰老细胞中的比较研究

以增强型绿色荧光蛋白 (EGFP)为报告基因 ,检测腺病毒、逆转录病毒和脂质体对 2 0代和 33代WI 38细胞的转导效率、EGFP的表达强度、对细胞增殖能力和生物学行为影响 ,比较病毒及非病毒方法对不同代龄成纤维细胞 (WI 38)细胞基因转导的特性 .结果显示 :对 2 0代和 33代WI 38细胞,腺病毒转染效率均最高 ,逆转录病毒其次 ;EGFP表达强度腺病毒最高 ,逆转录病毒最低 ;各方法2 0代细胞转导效率及表达强度均高于 33代细胞 (P <0 0 5 ) .逆转录病毒对细胞繁殖和SA β gal染色没有影响 ,腺病毒和脂质体均能导致细胞增殖能力下降 ,SA β gal染色阳性率上升 ,尤以脂质体明显 (P <0 0 5 ) .表明逆转录病毒介导基因对WI 38细胞衰老进程几乎没有影响 ,且转导效率较高 ,适于衰老细胞转基因研究

Investigation of Methods of Virus and Non-virus Gene Transfer into Human Fibroblasts (WI 38)

Using enhanced green fluorescent protein (EGFP)as a reporter, observed the transfer efficiency ,the influence upon aging process and cell biological behavior in human fibroblasts cells (WI 38 cells) of 20 and 33 passage transfected by adenovirus, retrovirus and liposome mediated methods, compare the gene transfer characteristic in WI 38 of 20 and 33 passage using methods of virus and non virus. The results showed that gene transfer efficiency was highest in adenovirus and lowest in liposome. The EGFP expression intensity was also highest in adenovirus but lowest in retrovirus. In all the three methods, the transfer efficiency and EGFP expression intensity were higher in younger cells ( P <0 05). Retrovirus has almost no influence upon cell proliferation ability and SA β gal staining positive rate, but the retrovirus and liposome decreased the proliferation ability and increased positive rate ( P <0 05).Those results suggest that retrovirus gene transfer method should be the first choice of transfer gene into aging cell, which has comparable high efficiency and nearly no negative effect.