应用羊膜上皮干细胞微环境培养人角膜内皮细胞的研究  

Microenvironment of amniotic epithelium cells enhances the proliferation of human corneal endothelial cells

作　　者：王忠浩[1] 陈玮[1] 宋莉[2] 沙翔垠[2] 梁轩伟[1] 

机构地区：[1]眼科学国家重点实验室,中山大学中山眼科中心,广州市510060 [2]广州医学院第二附属医院眼科,510120

出　　处：《实用医学杂志》2013年第6期857-861,共5页The Journal of Practical Medicine

基　　金：广东省医学科研基金(编号:A2010185);广东省自然科学基金(编号:10151018201000012);广东省科技计划项目资助(编号:2011B080701033)

摘　　要：目的:建立一种利用羊膜上皮干细胞(human amniotic membrane epithelial cell,HAEC)微环境培养人角膜内皮细胞(human corneal endothelial cells,HCEC)的方法。方法:制备羊膜上皮干细胞微环境培养HCEC,并探讨诱导HCEC增殖的最佳培养微环境,倒置相差显微镜和透射电镜观察培养过程中细胞的形态学变化,MTT和Giemsa染色观察细胞增殖情况,Hoechst33342检测凋亡细胞比例。结果:在HCEC基本培养液(corneal endothelial cell medium,CEM)的基础上添加20%HAEC上清、HAEC-HCEC的微环境可促进HCEC的增殖,减少凋亡,细胞传代能力显著增强,HAEC-HCEC组传至4代仍保持多角形的内皮细胞形态。结论:羊膜上皮干细胞微环境培养可有效提高HCEE的增殖能力,更好地维持HCEC的形态,并能抑制其凋亡进程。Objective To establish an effective method to enhance the proliferation of human corneal endothelial cells （HCECs）. Methods The culture conditions of HCEC were optimized by utilizing the totipotent characteristics of human amniotie membrane epithelial stem cells （HAEC） to establish the optimal culture microenvironment of HAEC to promote the proliferation of HCEC. The morphology of HCEC was observed by using phase-contrast microscope and transmission electron microscope. MTF assay and Giemsa staining were performed to detect the proliferation of HCEC. The rate of apoptotic cells was investigated by using Hoechst33342 staining assay. Results Compared to the corneal endothelial cells medium （CEM）, the microenvironment containing 20% HAEC-eonditioned medium and HAEC-HCEC co-culture mieroenvironment could promote the proliferation of HCEC and could reduce the apoptosis of HCEC. The cells in HAEC-HCEC microenvironment group could be passaged 4 times without lossing their polygonal appearance. Conclusion The HAEC microenvironment could effectively enhance the proliferation of HCEC, maintain the morphology of HCEC, and inhibit the process of apoptosis of HCEC.

关 键 词：人角膜内皮细胞 人羊膜干细胞 增殖 凋亡 

分 类 号：R77[医药卫生—眼科]