文档介绍：SV40大T抗原对人皮肤成纤维细胞生物学行为的影响
作者:王新文,金岩,刘源,王军琳,赵宇,董蕊
【关键词】大T抗原
【Abstract】 AIM: To investigate the role of SV40 large T (LT) antigen on human skin fibroblasts transformation and to observe the biological alteration of the cells. METHODS: Human skin firbroblasts were cultured and transfected with eukaryotic expressing plasmid psv3neo which containing the large T. After selection by G418 to stabilize the transfection, the large T mRNA and phenotypical changes of the transformants were examined. RESULTS: LT cDNA transformed cell colonies were isolated and were subcultured. LT mRNA expressed within the cytoplasm in most transformed cells when determined by in situ hybridization. After a period of time, however, the transfected cells gradually entered crisis and died. CONCLUSION: The ability of LT to enable the normal human skin fibroblasts immortalized is limited. So it is impossible to establish immortalized human skin fibroblast cell line only by large T.
【Keywords】 SV40; large T antigen; skin; fibroblasts
【摘要】目的: 研究SV40大T抗原(LT)基因对人正常皮肤成纤维细胞体外转化作用, 以及转化后细胞生物学特性的改变. 方法: 采用脂质体介导的方法,利用含有LT cDNA的质粒psv3neo转染人正常皮肤成纤维细胞. G418筛选后,挑选阳性克隆扩大培养,观察基因及蛋白在细胞内的表达,以及对细胞增殖等生物学特性的改变情况. 结果: 分离获得转化细胞克隆,原位杂交显示LT mRNA在转染细胞胞质中表达. 免疫组化结果显示,细胞质中有SV40大T抗原表达. 细胞计数、BrdU、流式细胞仪检测表明,经过一段时间的旺盛生长,在转染后第16代,细胞即进入危机期,以一种比衰老细胞更快的速度死亡. 结论: LT 可以促进正常皮肤成纤维细胞增殖,但这种能力是有限的,在转染后期甚至会促进细胞死亡. 所以要使人的正常皮肤成纤维细胞永生化,单纯利用LT