N-亚硝基-N-甲基脲诱导视网膜变性小鼠模型的特征

Morphologic characteristics of N-Nitroso-N-methylurea-induced retinal degeneration in mice

目的 探讨N-亚硝基-N-甲基脲（MNU）诱导的小鼠视网膜退行性病变的形态和电生理特征.方法 实验研究.32只5周龄的成年C57/BL小鼠随机分为对照组和MNU造模组,分别腹腔注射0.9％氯化钠溶液和MNU （60 mg＆#183;kg-1）.在给药后3、7和14 d取视网膜,免疫荧光染色观察光感受器形态、氧化损伤情况和神经节细胞的数量.电镜观察细胞核、线粒体和带状突触（synaptic ribbon）的超微结构.Ganzfeld视网膜电图（ERG）检测暗适应最大混合反应.采用独立样本t检验进行数据分析.结果 免疫荧光显示：注射MNU后外核层（ONL）的厚度逐渐减小,OPN lSW标记的光感受器细胞的外节（0S）逐渐损伤,GFAP标记的M＆#252;ller细胞增生明显,ONL层硝基酪氨酸（Nitrotyrosine）着色增多提示MNU能导致氧化损伤.Bm-3a标记的神经节细胞数量减少不明显.扫描电子显微镜显示：MNU处理后,光感受器细胞的核呈浓染色,线粒体和带状突触消失.ERG结果显示：MNU处理后3d最大混合反应的a波和b波振幅下降.结论 MNU导致视网膜外核层和外丛状层的凋亡,电生理表现和视网膜色素变性疾病一致.

Objective To explore the characteristics of N-Nitroso-N-methylurea （MNU） induced retinal degeneration in mice.Methods Experimental study.Thirty-two C57/BL mice （5 weeks） in total were randomly divided into two groups：a control group and a MNU group.Sixty mg＆#183;kg-1 of MNU was intraperitonealy injected in mice,leading to damage of retinal photoreceptors,while the control group was given physiological saline.The eyes of each group were enucleated at 3 d,7 d and 14 d after injection.The slides were stained by immunofluorescence labeling to observe the morphological changes to the photoreceptors in MNU-induced retinal damage.Retinal fiat-mount immunofluorescence was performed to observe the change in ganglion cells.The ultrastructures of the nucleus,mitochondria and synaptic ribbon were observed under transmission electron microscope.Ganzfeld electroretinogram ERG was used to test the electrophysiological changes.The data were analyzed using independent t test.Results After MNU administration,immunofluorescence labeling showed a gradual decrease in the thickness of the outer nuclear layer （ONL）,the loss of photoreceptors labeled with anti-OPN1SW,reactive gliosis with anti-glial fibrillary acidic protein （GFAP） and oxidative damage in photoreceptors labeled with anti-nitrosylation.Brn-3a labeled ganglion cells did not appear to be lost two weeks after MNU administration.Electron microscopy showed concentrated coloring of the nucleus in the photoreceptor nucleus and the disappearance of mitochondria and metabolic synaptic ribbons after MNU administration.ERGs revealed that mice with MNU treatment showed a-waves and b-waves with reduced amplitudes.Conclusion MNU induced retinal apoptosis of ONL and outer piexiform layer.MNU administrated mice show similar electrophysiological results to retintis pigmentosa.