经角膜基质细胞诱导的大鼠骨髓间充质干细胞在人羊膜上构建角膜上皮移植片

Reconstruction of a corneal epithelial graft with amnion and induced marrow mesenchymal stem cells

目的:研究将骨髓间充质干细胞(mesenchymal stem cells,MSC)诱导分化后在羊膜表面培养构建角膜上皮移植片的可行性。方法:取SD大鼠骨髓间充质干细胞和角膜基质细胞,分别培养并传2代后进行共培养,取共培养7d后的MSC覆载于新鲜人羊膜,再培养7d。对MSC及诱导后MSC进行免疫荧光染色和扫描电镜检查;对覆载细胞的羊膜进行HE染色及免疫荧光染色。结果:MSC在体外培养条件下贴壁生长,免疫荧光染色CD29和CD44阳性,CK12阴性。经角膜基质细胞诱导分化后,MSC细胞CK12染色转为阳性。诱导后MSC接种到羊膜表面,迅速贴壁生长,组织学特性无明显改变,CK12染色仍为阳性。结论:经角膜基质细胞诱导的MSC表现出角膜上皮细胞特征,在羊膜上生长后保持不变。

AIM：To investigate the feasibility of reconstruction of a corneal epithelial graft with amnion and rat marrow mesenchymal stem cells（MSC）after co-cultured with mouse cornea stroma cells（CSC）.METHODS：MSC and CSC cells were isolated from SD rats and cultured for two generations respectively.MSC and CSC were then cultured together.After co-cultured for 7 days,the MSC cells were seeded onto the human amnion and cultured for additional 7 days.The MSC cells of passage 2 and the co-cultured MSC cells were detected by immunofluorescence staining and scanning electronic microscopy.The amnion with induced MSC cells were examined with HE and immunofluorescence staining.RESULTS：The MSC cells in vitro adhered,grew and expressed positive staining to CD29 and CD44,but negative to CK12.The co-cultured MSC cells induced by CSC expressed positive staining to CK12.The induced cells grew rapidly on the amnion,and showed unchanged cytological characteristics with positive CK12 staining.CONCLUSION：The MSC cells induced by CSC cells show the corneal epithelial cell characteristics and maintain unchanged on the amnion,indicating that it can be a corneal epithelial graft for further use of reconstruction of the ocular surface.