组织工程口腔黏膜上皮治疗角膜缘干细胞缺陷症的实验研究  

Management of corneal limbal stem cell deficiency with engineering autologous oral mucosal epithelium in rabbit

作　　者：赵晓玉[1] 吕岚[1] 韩斌[1] 蔺琪[1] 张旭[1] 邱波[1] 

机构地区：[1]首都医科大学附属北京同仁医院北京同仁眼科中心,北京100730

出　　处：《眼科研究》2007年第8期568-573,共6页Chinese Ophthalmic Research

摘　　要：目的探讨体外培养的自体组织工程口腔黏膜上皮重建兔角膜上皮的可行性。方法制作兔角膜缘干细胞缺陷模型32只,实验组Ⅰ～Ⅲ以自体口腔黏膜上皮细胞为种子细胞制作组织工程上皮,移植到实验组模型兔角膜表面,分别观察2周、1个月、3个月,对照组移植空白载体膜,观察3个月。术后裂隙灯显微镜下观察,以角膜新生血管、混浊度及上皮染色评分评价移植效果。用组织病理、免疫组织化学和印迹细胞技术评价角膜上皮重建的可能性。结果模型兔角膜混浊,有大量新生血管和杯状细胞。实验组移植后角膜透明,印迹细胞检查PAS(-)。实验组与对照组术后角膜总评分差异有统计学意义(P=0.000),p63表达阳性,角膜上皮的组织特点及角蛋白的表达与正常角膜上皮相似。结论组织工程口腔黏膜上皮在角膜基质微环境的诱导下可分化为角膜样上皮细胞,有重建角膜上皮的作用。Objective To investigate the feasibility of the reconstruction of corneal epithelium using engineering autologous oral mucosal epithelial cells in the eyes of limbal stem cell deficiency. Methods Models of corneal limbal stem cell deficiency were made in 32 New Zealand white rabbits by the resection of corneal epithelium and superficial stroma extending 1 mm outside the limbus. Epithelial cells from autologous oral mucosa were cultured and amplified in vitro and was transplanted with fibrin gel together to the corneal surface of rabbits models for 2 weeks（ group Ⅰ ）, 1 month（ group Ⅱ ） and 3 months（ group Ⅲ ）. Fibrin gel without cells was transplanted in control group. Therapeutical effect was evaluated by the total scoring of corneal neovascularization, opacity and fluorescence staining. HE staining was performed to observe the histological change. Immunohistochemistry was used for p63 staining and location and study of keratins after transplantation. Immunohistochemistry of Muc5AC and corneal impression cytology were performed to assess the phenotype of corneal epithelium. Results A great deal of corneal neovascularization was found in rabbit model at 1 month after surgery. Corneal impression cytology indicated the presence of numerous goblet cells. After the transplantation of engineering autologous oral mucosal epithelial sheet,the cornea was clear with a little neovascularization. The total scoring of cornea in group Ⅰ - Ⅲ was significantly less than that of control group. Expression of p63 was positive. The histology of corneal epithelium,PAS reaction and expression of keratins in experiment groups were similar to normal corneal epithelium at 3 months after transplantation. Cornea of control group showed conjunctiva phenotype with revascularization and goblet cells. Conclusion Engineering autologous oral mucosal epithelial sheet can successfully reconstruct the corneal epithelium of limbal stem cell deficiency disease. The stem cells of oral mucosal epithelium can transdifferentiate in

关 键 词：角膜缘干细胞缺陷症 口腔黏膜上皮 成体干细胞 横向分化 

分 类 号：R779.6[医药卫生—眼科]