三种实验动物角膜活体共焦显微镜检测的比较解剖学研究

Anatomical study of the corneal structures of three experimental animal models by in vivo confocalmicroscopy

背景海德堡视网膜厚度分析仪和角膜模块的结合实现了对眼表活体组织结构的非侵入性检查，利用共焦显微镜对常用实验动物角膜结构进行比较研究可为相关研究提供依据。目的利用活体共焦显微镜比较新西兰大白兔、Lewis大鼠、Swiss小鼠的角膜结构，建立实验动物角膜的活体组织图像资料，为共焦显微镜的实验研究提供依据。方法利用海德堡视网膜厚度分析仪（HRT—II）的Rostoek角膜模块对新西兰大白兔、Lewis大鼠、Swiss小鼠的角膜进行活体分析，角膜的每一层各采集20张共焦显微镜图片，比较分析实验动物角膜各层的形态学特点及角膜内皮细胞密度。结果共焦显微镜下3种实验动物角膜表层上皮细胞表现为高反光或低反光的多形细胞，基底上皮细胞表现为暗的细胞质，细胞核不可见，细胞间排列紧密、规则；前弹力层均表现为含有丰富上皮下神经丛的无定形物质。兔角膜基质层在黑色背景中散布着高反光物质，即为角膜基质细胞核，后基质层细胞密度高于前基质层；大鼠和小鼠的角膜基质层仅观察到大量反光的星形结构，无明显的细胞核。3种实验动物的角膜内皮细胞形态相似，均表现为高反光的胞体，边界较暗且细胞排列成蜂窝状。新西兰兔前基质角膜细胞密度中位数为387．5个／mm^2，后基质角膜细胞密度中位数为223，5个／mm^2，明显少于前基质的细胞密度（U=0．000，P=0．000）；新西兰兔、Lewis大鼠、Swiss小鼠角膜内皮细胞密度中位数分别为2192．5、1936．0、1565．0个／mm^2，总体差异有统计学意义（H=49．940，P=0．000），兔角膜内皮细胞密度明显高于大鼠和小鼠，差异均有统计学意义（疋。=0．000，P=0．000；x^2=0．000，P=0．000），大鼠和小鼠的角膜内皮细胞密度差异亦有统计学意义（x^2=0．000，P=0．000）。结论共焦显微镜下新西兰大白兔、Lewis大鼠、Swiss小鼠角膜各层的细胞形态相似，但内皮细胞密度和基质细胞形态之间存在明显差异。HRT-Ⅱ的Rostock角膜模块可为动物实验提供角膜各层次的高分辨率图像。

Background Noninvasive methods such as in vivo confocal microscopy and Orbscan Ⅱ corneal topography have been used to examine ocular surface structure at the cellular level. However, very few domestic reports about the corneal structures of experimental animals investigated by confocal microscopy are available. Objective This study was to compare the anatomical differences of the corneal structures of three frequently used experimental animals presented by in vivo confocal microscopy, and to offer a database on the information provided by the in vivo study of the corneal structures of these animals. Methods Bilateral corneas of 3 clean adult male New Zealand rabbits,3 clean adult male Lewis rats and 3 clean adult male Swiss mice were examined by in vivo confocal microscopy. The morphological characteristics of every layer of the corneas and the endothelial cell densities were analyzed and compared. Results Superficial epithelium cells of the three animal models were characterized as polygon cells with high or low reflective border. The arrangement of the basal epithelial cells was regular with tight contacts but these cells lacked visible nuclei. The Bowman＇ s layer of cornea presented as an amorphous sheet containing abundant subepithelial plexus. In the rabbits, a highly reflective structure in the corneal stroma wasconfirmed as the nucleus, and the cell density of the posterior stroma was significantly lower than that of anterior stroma（387.5 cells/mm^2 versus 223.5 cells/mm^2 ） （ U = 0. 000, P = 0. 000 ）. Massive light-reflecting astreoids were displayed in the stroma of the rats and the mice. Corneal endothelial cells （CECs） of the three animal models had similar shapes and arrangements, presenting with high refractive cell bodies with dark borders and honeycomb-like arrangements. The CECs densities were 2192.5,1936.0,1565.0 cells/ram2 in the New Zealand rabbits, Lewis rats and Swiss mice, respectively, showing a statistically significant difference among them （ H = 49. 940, P = 0. 000 ） , and that of the rabbits was significantly higher than that in the rats and mice （,,v2 = 0.000, P = 0.000;X2 = 0.000, P=O. 000）. Significant difference was also seen between the rats and the mice in the CECs densities （X2 = O. 000, P=0. 000）. Conclusions The CECs of the three animal modes are similar in morphology. But the structures of their stromal cells and endothelial cell densities are different. The combination of in vivo confocal microscopy and Orbscan Ⅱ corneal topography offers high-resolution imaging for each layer of the cornea.