体外角膜上皮细胞凋亡的研究

The study of corneal epithelial cell apoptosis, in vitro

为观察离体角膜上皮细胞经传代培养后细胞死亡的规律及确定其性质。本文应用流式细胞仪 ( FACStarplus)检测凋亡细胞的亚 2倍体 DNA以及细胞凋亡的形态学观察 ,对体外培养的每一代兔角膜缘上皮细胞的凋亡情况进行了观察及分析。结果显示 :角膜缘原代上皮细胞被检测出 7.0 0± 2 .2 3 %的凋亡细胞 ;从第 6代开始 ,角膜上皮细胞凋亡数目显著升高 ( P<0 .0 1) ;之后细胞凋亡数目急剧增加 ,第 8～ 9代后细胞出现集体“自杀”现象。透射电镜观察到这些细胞出现了核浓缩、胞膜发泡、凋亡小体等典型细胞凋亡的形态学改变。结论 :离体角膜缘上皮细胞因外部生长环境的改变 ,在培养到第 6代时出现了细胞凋亡现象。

Objective:To study the death of corneal epithelial cells while culturing in vitro.Methods:Degraded DNA(<2n DNA) in corneal epithelial cells of every passages cultured in vitro was analyzed by flow cytometry and transmission electron microscopy was used for morphological analysis of apoptosis.Results:Flow cytometric analysis of propidium iodide stained(PI) cells indicated that 7 00±2 23% of the cells contained <2n DNA content in primary cultured limbal cells.The index of PI stained <2n DNA in limbal cells was increased after the 6 th passage(P<0 01).All cells committed suicide after the 8 th to 9 th passage.Examination by transmission electron microscopy revealed typical morphological featuresof those cells undergoing apoptosis,including chromatin condensation,vacuolation of the cytoplasm,apoptosis bodies etc.Conclusion:Apoptosis was undergoing in the 6 th passage of limbal cells while culturing in vitro,as a result of changing the extracellular environment for cell growth.