hTERT诱导永生化口腔黏膜上皮细胞株的建立  

Establishment of immortalized oral mucosal epithelial cell strain by human telomerase reverse transcriptase

作　　者：陈之锋[1] 吕标[2] 郑超群[3] 胡逢春[1] 陶谦[1] 

机构地区：[1]中山大学光华口腔医学院附属口腔医院口腔医学研究所,广东广州510055 [2]江门市口腔医院,广东江门529020 [3]中山市人民医院口腔医疗中心,广东中山528403

出　　处：《中国口腔颌面外科杂志》2013年第2期91-96,共6页China Journal of Oral and Maxillofacial Surgery

基　　金：国家自然科学基金(81072227);广东省科技计划项目(2008B050100008)~~

摘　　要：目的:转染外源性人端粒酶反转录酶(human telomerase reverse transcriptase,hTERT)基因建立永生化口腔黏膜上皮(oral mucosal epithelial,OME)细胞株。方法:利用反转录病毒载体将外源性hTERT基因转入OME细胞,G418筛选出抗性细胞克隆后,进行免疫细胞化学(immunocytochemistry,ICC)鉴定细胞来源、细胞形态学观察、细胞增殖动力学研究、裸鼠成瘤实验以及hTERT基因和蛋白表达的检测。采用SPSS11.0软件包对数据进行统计学处理。结果:转染细胞与未转染细胞形态非常相似,均呈多边形、多角形,"铺路石"样排列;广谱角蛋白染色阳性,波丝蛋白染色阴性;未转染细胞体外培养36 d后停止生长,群体倍增数(population doublings,PDL)为9,转染细胞增殖活跃,PDL值超过80;RT-PCR和Western印迹结果均显示,hTERT在转染细胞中稳定表达,而在未转染细胞中不表达。结论:转染外源性hTERT基因的OME细胞呈稳定增殖的永生化状态。PURPOSE： To establish immortalized oral mucosal epithelial （OME） cell line by transfection with human telomerase reverse transcriptase （hTERT）. METHODS： Oral mueosal epithelial cells were infected with retroviral vector encoding hTERT. The infected cells were selected with G418 and checked by immunocytochemistry （ICC） for in vitro proliferation and the ability of tumorigenesis. RT-PCR and Western blot were used to detect the expression of hTERT. The data was processed with SPSSll.0 software package. RESULTS：Both infected and uninfected cells showed ＂flagstone＂ appearance; ICC confirmed the epithelial origin of the infected cells based on positive pan-cytokeratin and negative vimentin expression; Compared with uninfected cells, which arrested at the population doublings （PDL） of 9, the infected cells were more active by proliferated and reached at 80 to date; RT-PCR and Western blot showed hTERT expression. CONCLUSIONS： The infected oral mucosal epithelial cells were immortalized after transfection with hTERT and can serve as a genetically defined model of oral epithelial tumor study. Supported by National Natural Science Foundation of China（81072227） and Science and Technology Planning Project of Guangdong Province（2008B050100008）.

关 键 词：口腔黏膜上皮细胞 端粒酶反转录酶 转染 永生化 

分 类 号：R780.2[医药卫生—口腔医学]