目的:探讨硫酸软骨素(chondroitin sulfate,CS)在甘油冷冻保存角膜中对角膜内皮细胞(corneal epithelial cells,CEC)的保护作用。方法:Wistar大鼠角膜片40只随机分为实验组和对照组,对照组角膜片直接放入纯甘油中,实验组则在内皮面均匀黏附10g/L CS后再放入甘油中。-20℃保存2mo后行CEC台盼蓝-茜素红活性染色及电镜检查。取30只SD大鼠与保存后角膜片建立Wistar-SD同种异体穿透性角膜移植模型,测定术后不同时间点植片内TNF-α和IFN-γ的表达水平,检验保存效果。结果:台盼蓝-茜素红活性染色结果及术后植片内TNF-α和IFN-γ的表达结果显示,两组差异有显著性(P<0.05),实验组保存效果和角膜移植术后效果均优于对照组。电镜检查示,实验组CEC胞质内可见个别空泡,线粒体及内质网轻度肿胀;对照组CEC线粒体及内质网明显肿胀。结论:甘油冷冻能够保持CEC的活性,且CS能进一步增进保存效果。

AIM:To evaluate chondroitin sulfate on the glycerol cryopreservation of corneal endothelial cells(CEC).METHODS:Forty Wistar rats corneal pieces were divided into the experimental group:glycerol cryopreservation medium containing chondroitin sulfate(CS);and the control group:mere glycerol cryopre-servation medium.The two groups were saved at-20℃ for 2 months.All pieces were examined with trypan blue combined with alizarin red staining(viability staining) and electron-microscope examination for CEC.30 SD rats were taken to establish Wistar-SD rat model of allogenic penetrating keratoplasty.Slit lamp microscope detected the occurrence of corneal rejection,TNF-α and IFN-γ expression was examined by immunohistochemistry to test the effect of preservation.RESULTS:The staining results,electron-microscope examination and TNF-α,IFN-γ expression showed,the difference was significant in these two groups(P<0.05),the results of the experimental group was better than the control group after the saving and the corneal transplantation(P<0.05).The electron-microscope examination showed indivdual vacuoles in CEC cytoplasm,slight swelling of mitochondrial and endoplasmic reticulum in experimental group,obvious swelling of CEC mitochondrial and endoplasmic reticulum in control group.CONCLUSION:Glycerol cryopreservation can preserve the viability of CEC.Chondroitin sulfate can obviously improve the effect of preservation.

R779.65

中国辽宁省教育厅高等学校科学研究资助项目(No.2009A445)~~