EPO通过上皮间充质转化促进皮肤角质细胞增殖和迁移的体外研究  

Research of erythropoietin on skin keratinocyte proliferation and migration via epithelial-mesenchymal transformation in vitro

作　　者：李雨阳 彭浩然 赵越 任双双[1] 苗雷英[1] LI Yu-yang;PENG Hao-ran;ZHAO Yue;REN Shuang-shuang;MIAO Lei-ying(Department of Cariology and Endodontics,Nanjing Stomatological Hospital,Medical School of Nanjing University,Nanjing 210008,China)

机构地区：[1]南京大学医学院附属口腔医院·南京市口腔医院牙体牙髓病科,南京210008

出　　处：《中华老年口腔医学杂志》2022年第6期327-333,共7页Chinese Journal of Geriatric Dentistry

基　　金：江苏省自然科学基金青年项目(项目编号:BK20200665);南京市卫生科技发展医学重点科技发展项目(项目编号:ZKX19031)。

摘　　要：目的:初步探讨促红细胞生成素(Erythropoietin,EPO)通过上皮间充质转化(Epithelial-Mesenchymal Transition,EMT)对人永生皮肤角质细胞H aCat增殖和迁移的影响。方法:体外培养H aCat细胞,使用不同浓度的EPO(0,1,10,50,100IU/mL)处理细胞,CCK-8检测其增殖变化,流式细胞术探究其凋亡变化;采用划痕和transwell实验观察细胞迁移能力的改变;使用实时定量PCR(Real-time quantitative PCR,RT-qPCR)和Western blot检测EMT及趋化相关基因的改变情况。结果:当EPO≥50IU/mL时,可以显著促进H aCat细胞的增殖(P<0.05),且细胞的凋亡率没有明显增加;当EPO≥50IU/mL时,细胞的水平移动和垂直移动能力均显著增加(P<0.05);qPCR显示,随着EPO浓度增加,上皮标志物E-cadherin的mRNA表达逐渐下降,间质标志物N-cadherin和Fibronectin表达逐渐升高,且趋化因子CCL2、CCL8和CCL21的mRNA均逐渐上调。Western blot结果显示,E-cadh erin的蛋白表达逐渐下降,N-cadh erin和Vimentin表达逐渐升高。结论:EPO能够诱导皮肤细胞H aCat上皮间充质转化的发生并能促进其增殖和迁移。Objective:To investigate the effect of EPO on the proliferation and migration of human immortal keratinocyte HaCat through EMT.Methods:HaCat cells were cultured in vitro and treated with different concentrations of EPO(0,1,10,50,100 IU/m L).The proliferative changes were detected by CCK-8,and the apoptotic changes were investigated by flow cytometry.The changes of cell migration ability were observed by scratch and transwell assay.Real-time quantitative PCR(RT-qPCR)and Western blot were used to detect the changes of EMT and chemotactic related genes.Result:Cell proliferation was promoted when EPO concentration≥50 IU/m L(P<0.05)and the apoptosis rate was not significantly increased.When EPO concentration≥50 IU/m L,the abilities of cell horizontal and vertical migrations were strengthened(P<0.05).The results of qPCR showed that with the EPO concentration increasing,epithelial marker E-cadherin expression was gradually decreased and mesenchymal markers N-cadherin and fibronectin were down-regulated,also,m RNA level of chemokine for CCL2,CCL8 and CCL21 showed up-regulation for the same tendency.Furthermore,Western blot showed the protein level of E-cadherin was gradually decreased whereas the N-cadherin and Vimentin were down-regulated.Conclusion:EPO could induce HaCat for its EMT activity and promote cell proliferation and migration.

关 键 词：促红细胞生成素 上皮间充质转化 人永生角质形成细胞 细胞增殖 细胞凋亡 细胞迁移 趋化因子 

分 类 号：R782.4[医药卫生—口腔医学]