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目的:检测连接粘附分子-1(junction adhesion molecule,JAM-1)在正常人角膜上皮各层中的表达及分布特点并与咬合蛋白(occludin)进行比较。方法:培养人角膜上皮细胞提取细胞总RNA。以逆转录后获得的cDNA为模板PCR扩增目的基因JAM-1及occludin。流式细胞仪检测JAM-1蛋白表达。双重免疫荧光观察JAM-1与occludin在正常人角膜上皮组织的原位表达。结果:通过RT-PCR在培养人角膜上皮细胞中检测到JAM-1与occludin扩增片段;流式细胞仪检测到JAM-1蛋白表达;双重免疫荧光结果显示occludin染色主要位于表层上皮层细胞之间;而JAM-1荧光染色不仅见于上皮表层,在整个上皮层细胞之间均可见其荧光反应。结论:Occludin主要位于正常人角膜上皮表层细胞之间,JAM-1在正常人角膜上皮的全层中均有表达。
OBJECTIVE: To detect the expression and distribution of junction-adhesion molecule-1 (JAM-1) in normal human corneal epithelial layers and to compare with occludin. Methods: Cultured human corneal epithelial cells to extract total cellular RNA. The target gene JAM-1 and occludin were amplified by RT-PCR. The expression of JAM-1 protein was detected by flow cytometry. Double immunofluorescence was used to observe the expression of JAM-1 and occludin in normal human corneal epithelium. Results: The amplified fragments of JAM-1 and occludin were detected by RT-PCR in human corneal epithelial cells. The expression of JAM-1 protein was detected by flow cytometry. The results of double immunofluorescence showed that occludin staining was mainly located in epithelial cells Between the epithelial cells, the fluorescence of JAM-1 was not only found in the epithelial layer, but also in the whole epithelial cells. Conclusion: Occludin is mainly located in normal human corneal epithelial cells, and JAM-1 is expressed in all layers of normal human corneal epithelium.