利用皮肤干细胞的横向分化重建角膜上皮的初步研究  

A pilot study on transdifferentiation of skin stem cell in reconstructing corneal epithelium

作　　者：黄冰[1] 王智崇[1] 葛坚[1] 陈系古[2] 刘敬波[1] 范志刚[1] 陶靖[1] 刘炳乾[1] 郭芬芬[2] 

机构地区：[1]中山大学中山眼科中心,广州510060 [2]中山大学实验动物中心

出　　处：《中华医学杂志》2004年第10期838-842,共5页National Medical Journal of China

基　　金：国家"九七三"重点基础研究发展规划基金资助项目 (G19990 5 43 0 9 6) ;广东省科技计划基金资助项目 (2 0 0 3A3 0 2 0 40 1) ;教育部博士点基金资助项目 (2 0 0 3A0 5 5 80 74)

摘　　要：目的　观察皮肤干细胞在角膜基质上的分化发育情况 ,探讨皮肤干细胞重建角膜上皮的可能性。方法　将体外分离培养的 1～ 4代人和兔皮肤干细胞种植在兔角膜基质上 ,在体外和体内观察皮肤干细胞的分化发育情况。实验标本作HE染色及用人上皮细胞角蛋白K19单克隆抗体和角膜上皮细胞特异性标志蛋白K3/K12抗体AE5进行免疫组织化学鉴定。结果　皮肤干细胞在角膜基质的调控下可以横向分化发育成表达角膜上皮细胞特异性标志蛋白K3/K12的细胞 ,呈现AE5抗体阳性 ;并可形成类似角膜上皮外观透明的多层细胞结构。结论　利用皮肤干细胞有可能重建角膜上皮 ,进行自体生物角膜的构建。Objective To observe the differentiation and development of skin stem cells on corneal stroma and to discuss the possibility of reconstructing corneal epithelium with skin stem cells. Methods Pieces of human and rabbit skin were obtained during operation. Rabbit eye balls were taken,and pieces of corneal stroma without epithelium were prepared. Skin stem cells from the rabbit skin and human skin were cultured. The human skin stem cells of the first generation to 4 th generation were implanted on the rabbit corneal stroma and cultured. Three rabbits underwent autotransplantation of the rabbit skin stem cells of the first generation to 4 th generation on the pieces of corneal stroma with the superficial lamina removed and then fed for 100～114 days. Another 3 rabbits underwent allotransplantation of the rabbit skin stem cells of first to 4 th generation on the pieces of corneal stroma with the superficial lamina removed and then fed for 100 days. Then the rabbits were killed and their eye balls taken out. The rabbit corneas implanted with human or rabbit epithelial cells and the rabbit corneas with the autogeneous or heterogeneous epithelial cells were sliced and underwent immunohistochemistry with human AE5 antibody corresponding to the specific surface marker keratin K3/K12 common to humankind and rabbit,and human epithelial cell keratin K-19 monoclonal antibody. Results Since the 3 rd day of transplantation the transplanted human epithelial cells formed multiplayer and were human AE5 antibody and human K19 monoclonal antibody positive. The autotransplanted corneas remained basically transparent without obvious vascular hyperplasia till the cornea specimens were taken. Histological examination showed intact multiplayer epithelium and immunohistochemistry showed human AE5 positive. The allotransplanted\rabbit corneas showed congestion since the 9 th day. Histological examination showed that the corneas were nor so transparent as the autotransplanted ones and the epithelium was nor intact with a lot of lymp

关 键 词：皮肤干细胞 细胞横向分化 功能重建 角膜上皮 免疫组织化学 组织工程 

分 类 号：R779.65[医药卫生—眼科]