This study was aim to explore the method of establishing animal model of corneal endothelial injury and the potential of endothelial regeneration after injury.

30 normal axolotls of 2 weeks old were selected. They were divided into normal observation group, NaoH-injured group and mechanical-injured group with 10 axolotls each. In vivo confocal microscopy, histopathological examination, alizarin red-trichlorobenzene blue endothelial staining and gold chloride corneal nerve staining were used to observe. In the normal observation group, the normal structure of the eye and cornea was observed. Corneal endothelial cells were damaged by NaoH solution and mechanics, and the endothelial cell injury model was established to observe the regeneration of corneal endothelial cells. The corneal endothelial cell density at four time points was tested by normality test and homogeneity test of variance. The mean corneal endothelial cell density at different time points in different groups was compared by repeated measurement variance analysis. Mauchly spherical test was performed beforehand. According to the test results, if significant, the results of multivariate analysis of variance are adopted; otherwise, the correction free F test is selected.

The corneal thickness of axolotls was about (75.75±7.51) μm, and the corneal epithelial cell layer was about half of the corneal thickness. After the corneal endothelial cells were damaged by NaoH solution and mechanic, the density of corneal endothelial cells decreased significantly. The density of corneal endothelial cells in NaoH-injured group and mechanical-injured group at different time points was significantly different (F=31.38, 51.77; P<0.05). There was no significant difference between NaoH-injured group and mechanical-injured group at each time point (t= 1.37, 2.67, 0.70, 4.14; P>0.05). At 3 days after injury, the density of corneal endothelial cells in NaoH-injured group and mechanical-injured group were (128±14)/mm² and (113±11)/mm². And the density of corneal endothelial cells in NaoH-injured group was (157±20)/mm² and that in mechanical-injured group was (169±19)/mm² at 7 days after injury (t=19.39, 8.78; P<0.05). Compared with 3 days after injury, the density of corneal endothelial cells recovered with statistical significance (t=3.75, 8.07; P<0.05). At 14 days after injury, the density of corneal endothelial cells in NaoH injury group was (198±17)/mm², and that in mechanical injury group was (223±17)/mm². Compared with 3 days and 7 days after injury, there were significant differences (t=10.05, 8.07; P<0.05) and (t=4.94, 6.70; P<0.05). Over time, the density of corneal endothelial cells in both groups was gradually recovered. At 14 days after injury, the morphology, size and density of corneal endothelial cells almost turned back to normal.

Both NaoH solution and mechanic can successfully establish the animal model of corneal endothelial cell injury in axolotls. The preliminary results showed that the endothelial cells had a certain regeneration potential. This method could provide a new animal model for the study of corneal endothelial cell regeneration after injury.