体外转染SV40T抗原对鼠内皮细胞周期素D1、P53和P21表达的影响  

Expression of Cyclin D1,P53 and P21 in Rat Endothelial Cells Transformed by SV40 T Antigen in Vitro

作　　者：贾俊[1] 赵怡芳[2] 张文峰[2] 何三纲[2] 

机构地区：[1]武汉大学口腔医学院口腔生物医学工程教育部重点实验室,湖北武汉430079 [2]武汉大学口腔医学院口腔颌面外科,湖北武汉430079

出　　处：《武汉大学学报（医学版）》2007年第1期40-43,共4页Medical Journal of Wuhan University

基　　金：湖北省科技厅课题(编号:2002P1206)

摘　　要：目的:探讨SV40 T抗原促进鼠内皮细胞增殖动力改变的机制,为基因治疗转化细胞提供理论依据。方法:用含有SV40 T抗原的表达质粒(pRNS-1)转染大鼠主动脉内皮细胞,经G418加压筛选,获得阳性克隆后扩增培养。在光镜下观察转染前后细胞的形态和生长特性;流式细胞仪检测细胞增殖周期;PCR检测外源基因的整合;免疫组化蛋白水平检测SV40 T抗原的表达;Western blot检测周期素(Cyclin)D1、P53和P21的表达。结果:转染后的鼠内皮细胞体外生存期延长,大量细胞由G1期进入S期,对血清和生长因子的依赖性降低。与转染前的细胞相比,转染后的鼠内皮细胞表达更多的P53和Cyclin D1,P21表达量降低。结论:SV40 T抗原可能通过调节Cyclin D1、P53和P21的表达来影响鼠内皮细胞的增殖。Objective： To study the control mechanisms which determine the proliferative switch of rat endothelial cells transformed by SV40 tumor antigen. Methods： The plasmid pRNS-1 containing SV40T antigen was transduced into the purified rat aortic endothelial cells （RAECs）. The positively transduced cell clones were expanded by the selection by G418. The original and transduced cells were examinated by phase-contrast microscope for growth rate and morphological features, by flow cytometry for cell proliferative dynamics. The integration of SV40 T gene was determinded by PCR, and expression of SV40 T antigen was studied by immunohistochemistry. In addition, the expression of CyclinD1, P53 and P21 was examinated by Western blot. Results： Endothelial cells transduced with SV40 T antigen exhibited longer life span in vitro and depended less on sera and growth factors. After being transduced, more cells entered into S phase from G1 phase. Compared with the untransduced endothelial cells, the transduced ones expessed more P53 and Cyclin D1, but less P21. Conclusion： SV40 T antigen may affect the proliferative switch of rat endothelial cells by regulating the expression of CyclinD1, P53 and P21.

关 键 词：细胞培养 细胞周期 细胞周期蛋白质 蛋白质P53 蛋白质P21 

分 类 号：R54[医药卫生—心血管疾病]