Apoptosis in the corneal epithelium.
Apoptosis in the corneal epithelium.
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DOI：
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发表时间：
1996-05
影响因子：
4.4
通讯作者：
H. Ren;G. Wilson
中科院分区：
文献类型：
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作者：
H. Ren;G. Wilson
目的测定兔角膜上皮细胞的存活率及细胞凋亡情况。方法采用钙黄绿素-乙锭法检测上皮细胞的活性。在该双色荧光测定中，活细胞发出绿色荧光，而非活细胞的细胞核发出红色荧光。对正常角膜上皮表面上的非存活细胞的数量进行定量。还从剪切应力角膜检查表面和脱落细胞。TUNEL技术（TdT介导的dUTP缺口末端标记）用于检测凋亡细胞的DNA片段特征。结果钙黄绿素-乙锭活性测定显示正常上皮细胞表面主要由活细胞组成，非活细胞占整个上皮细胞的一小部分。有一个增加的密度从周边的上皮表面的中心，在flatmounts的角膜，TUNEL标记表明，一小部分的表面细胞表现出DNA片段化，而横截面显示，DNA片段化的细胞被发现只在上皮表面。来自剪切应力角膜的上皮细胞显示出凋亡细胞的数量增加。结论：正常上皮细胞表面主要由活细胞组成，只有少量的凋亡细胞和非活细胞。结果表明，无活力的上皮细胞脱落后，终末分化，而活细胞可以脱落的经典凋亡形成水泡。因此，从角膜表面去除细胞的机制似乎不止一种。
PURPOSE To determine the viability of corneal surface cells and to determine whether apoptosis is present in the corneal epithelium of the rabbit. METHODS The viability of epithelial surface cells was examined using a calcein-ethidium assay. In this two-color fluorescence assay, viable cells fluoresce green, and the nuclei of nonviable cells fluoresce red. The number of nonviable cells on the normal corneal epithelial surface was quantified. Surface and shed cells also were examined from shear-stressed corneas. The TUNEL technique (TdT-mediated dUTP nick-end labeling) was used to detect DNA fragmentation characteristic of apoptotic cells. RESULTS The calcein-ethidium viability assay revealed that the normal epithelial surface is composed mainly of viable cells, with nonviable cells making up a small percentage of the whole. There was an increase in the density of nonviable cells from the periphery of the epithelial surface to the center, In flatmounts of the cornea, TUNEL labeling demonstrated that a small percentage of surface cells exhibited DNA fragmentation, whereas cross-sections revealed that DNA-fragmented cells were found exclusively on the epithelial surface. Epithelial cells from shear-stressed corneas showed an increased number of apoptotic cells. CONCLUSIONS The normal epithelial surface consists mainly of viable cells, with only a small percentage of nonviable cells and apoptotic cells. The results suggest that nonviable epithelial cells are shed after terminal differentiation, whereas viable cells can be shed by classical apoptosis with the formation of blebs. Thus, there appears to be more than one mechanism for removal of cells from the corneal surface.