The Expression Of Tight Junction Protain In Corneal Epithelium After De-epithelialization With Alcohol

Objectives To investigate the expression of tight junction protein in corneal epithelium after chemical de-epithelialization with 20% ethanol solution in refraction operation. Methods Sixty Sprague-Dawley rats were randomly divided into two equal groups, ethanol group and mechanical group, each has 30 rats. In each group, the 30 rats were were randomly divided into six subgroups(named A,B,C,D,E,F), each has five ones. One eye of each rat was treated with de-epithelialization operation, while the other eye without any treatment(control eye). Of the six groups group A was used to observe the defect of corneal epitheliam at 24h , and we used other groups to detect the expression of tight junction protein claudin-1 and ZO-1 in corneal epithelium at 24,48,72,96,120h. Rats were executed by cordis sudden death, cornea of both operation and control eye were separated and were imbedded with optimum cutting temperature compound Tissue Freezing Medium. HE staining was used to observed histological changes of corneal tissue. We made freeze section to compare the expression of tight junction protein in corneal epithelium between ethanol group and mechanical group at identical time point though immunoflouresence method.Results1. Time of removing corneal epithelium: Removing corneal epithelium with 20% ethanol spent much less time than with mechanical group, and the difference between two groups was significant (P<0.05).2. The defact of corneal epithelium at 24h: The cornea of operated eyes of 60 rats all were not re-epithelialized 24h after operation under operating microscope. Though slit lamp photography analysising, the difference of the defect diamter of cornea between ethanol group and mechanical group was not significant. 48h after operation : under operating microscope, there was no defect on the cornea epithelium of all operated eyes, and the cornea seemed to be smooth.3. Histopathological changes of cornea: Both ethanol group and mechanical group, 48h after operation the area of which cornea epithelium was removed were re-epithelialized, but there were only 2-3 layers corneal epithelium, and cells seemed thin and flat, with paramorphia. After 96h the corneal epithelium recovered with normal appearance on the whole.4. The difference of the expression of ZO-1: ZO -1 was confined mainly to the superficial cell layer of the corneal epithelium. The expression optical density(OD) value at 24h and 48h in ethanol group showing a significant increase in comparison with mechanical group (P<0.05). But the difference in the expression OD value of ZO-1 at 72, 96, 120h was not observed(P>0.05).5. The difference of the expression of claudin-1: Claudin-1 was mainly expressed in the wing and basal cell layers of corneal epithelium. Compared the expression of claudin-1 in corneal epithelium of ethanol group with mechanical group at 24 and 48h, it has a statistical difference(P<0.05). But the difference in the expression of claudin-1 at 72, 96, 120h was not observed(P>0.05).Conclusion In corneal refraction operation, removing corneal epithelium with ethonal spent less time than removing mechanically, there was no signifigant difference in the time of re-epithelialization. But Chemical de-epithelialization with ethanol solution seem to be slower than mechanical de-epithelialization in restoring of tight junction in corneal epithelium. 20% ethanol affect the repairation of corneal epithelium in corneal refraction operation.