兔角膜缘上皮细胞羊膜种植及生物学特性研究

Planting and Biological Character of Rabbit Corneal Epithelial Cells on Amniotic Membrane

研究兔角膜缘上皮细胞（含干细胞）在完整羊膜和去上皮羊膜上种植及生物学特征，探讨组织工程技术体外重建角膜上皮良好的方式和方法。将兔角膜缘上皮细胞分别种植于去上皮羊膜基底膜和完整羊膜上进行体外培养扩增。从倒置显微镜下的生长特性、光镜下的组织结构、超微结构、免疫组织化学检测等方面对培养获得的复合组织进行观察。结果表明兔角膜缘上皮细胞在完整羊膜上不易贴壁、生长缓慢、很难汇合成片。而在去上皮羊膜上细胞能黏附生长并增殖形成由羊膜基底膜和4～5层上皮细胞组成的复合组织，基底层细胞与羊膜之间有半桥粒连接，细胞之间可见桥粒连接，细胞表达角蛋白3。该复合物可作为组织工程化角膜上皮组织。

This study was aimed at investigating the cultivation and biological character of corneal epithelial cells （ECs） planted on intact and denued amniotic membrane（AM） as a substrate and trying to find out satisfactory methods for the reconstruction of corneal epithelium using tissue engineering. Rabbit corneal epithelial cells were planted on denuded AM and intact AM respectively. The cultivated corneal epithelial sheet was examined by use of inverted microscope, HE staining pathologyical section and transmission electron microscopy （ TEM ）, and was also detected immunohistochemically.The results revealed that rabbit corneal ECs grew slowly and were difficult to stick and converge on intact AM, whereas they were easy to grow and proliferate on denuded AM. The cultivated corneal ECs showed four to five layers of stratification composed of the basement membrane of AM and multiple layers of corneal ECs showed the presence of CK3. TEM unveiled that the multiple layers of corneal ECs had numerous desmosomal junctions attaching to the basement membrahe with hemidesmosomes. Therefore, the above cultivated corneal epithelial sheet can be used as engineering tissue for ocular surface reconstruction.