人视网膜色素上皮细胞脱分化模型的建立  

Establishment of human retinal pigment epithelium( RPE) cell EMT model

作　　者：王敏利[1,2,3] 吕立夏[1,2,3] 徐国彤[1,2,3] 

机构地区：[1]同济大学附属第十人民医院眼科,上海200072 [2]同济大学医学院眼科研究所,上海200092 [3]同济大学医学院再生医学系,上海200092

出　　处：《同济大学学报（医学版）》2014年第6期1-5,28,共6页Journal of Tongji University(Medical Science)

基　　金：国家重点基础研究发展973计划(2013CB967501)

摘　　要：目的建立人视网膜色素上皮(retinal pigment epithelium,RPE)细胞脱分化的模型,体外模拟增生性玻璃体视网膜病变(proliferative vitreoretinopathy,PVR)中RPE的上皮-间质转化(epithelial-mesenchymal transition,EMT)过程。方法将ARPE-19细胞培养在无血清、含N2和NEAA的petri dish中,建立RPE的EMT模型,采用倒置荧光显微镜观察细胞形态的转变,采用基因芯片和定量PCR来检测EMT相关基因的表达,采用Western blotting和ELISA来检测相关蛋白的表达。结果 ARPE-19细胞在建模后形态明显发生了改变,基因芯片和Q-PCR检测显示一些与EM T相关的因子(如TGF-β1,Vimentin,CD46)及炎症因子(IL-15)和细胞因子(EGF)的表达上调,ELISA在蛋白水平上显示建模后细胞表达的TGF-β1,IL-15,和EGF蛋白明显增多,Western blotting显示Vimentin蛋白的表达也显著上调。结论 ARPE-19在N2和NEAA的petri dish中培养可以诱导EMT的发生。Objective To establish an epithelial mesenchymal transition（ EMT） model of human retinal pigment epithelium（ RPE） cells. Methods Human retinal pigment epithelium ARPE-19 cells were treated with N2 and nonessential amino acids（ NEAA） in un-coating petridish. Cell morphology was observed with phase-contrast microscope; the expression TGF-β1,vimentin,CD46 genes was measured by gene chip and quantitative PCR; the expression levels of TGF-β1,vimentin,CD46 proteins and inflammatory factors IL-1,IL-15,EGF were detected by Western blotting and ELISA.Results After treatment of N2 and NEAA,the morphology of ARPE-19 cells changed significantly.Gene Chip and Q-PCR confirmed that the expression of EM T related genes TGFβ-1,vimentin,CD46 and inflammatory factors IL-1,IL-15,and EGF were significantly increased. Western blotting analysis confirmed the increase expression of vimentin protein. ELISA showed that the protein levels of TGF-β1,IL-15,and EGF were also significantly increased. Conclusion Treatment with N2 and NEAA in un-coating petridish can induce EMT of ARPE-19 cells.

关 键 词：上皮-间质转变 视网膜色素上皮细胞 增生性玻璃体视网膜病变 

分 类 号：R774.1[医药卫生—眼科]