Effect of Topically Applied Human Epidermal Growth Factor on Stromal Keratocyte Activity in Experimental Corneal Alkali Wounds

PURPOSE: The Influence of topically applied human epidermal growth factor(hEGF) on stromal keratocyte was evaluated in the experimental corneal alkali wounds. METHODS: Corneal alkali wounds were induced by applying a 5.5 mm round filter paper soaked in 1N NaOH onto the center of rabbit cornea for 60 seconds. The eyes were treated topically with 5 micrometer/ml hEGF(DaeWoong Pharma. Co. Ltd. Seoul, Korea) or PBS, 3 times per day. After 3- and 28-days treatment, the corneas were processed for transmission electron microscopy(TEM). RESULTS: After 3-days treatment, the basement membrane integrity was maintained and the stromal keratocyte remained damaged in both groups. However, after 28-days treatment, the basement membrane was disrupted and the keratocytes were activated in both groups. CONCLUSIONS: The findings of present study indicated that the optimal concentration of hEGF which is known to stimulate the epithelial healing was failed to affect the keratocyte activities as long as the basement membrane integrity was maintained.

Optimal Concentrations of hEGF for Epithelial and Endothelial Healing in Corneal Alkali Wounds: An Organ Cultured Study

Optimal concentrations of hEGF for epithelial and endothelial healing after alkali-injury were determined in an organ cultured system. By using a 5.5 mmround filter paper, corneal alkali wounds (1N NaOH, 60sec.) were induced in the rabbit eyes and then the whole cornea with 1 mmscleral rim were excised.The excised corneas were immersed in the Eagle`s minimum essential medium supplemented with 5% Dextran, 1%antibiotics and different concentrations of hEGF (DaeWoong Pharma.Co.Ltd. Seoul, Korea), i.e., O (control), 0.01, 0.1, 1.0, 10, 100ng/ml. After tryphan blue staining, morphometric analysis was performed at 1, 2, 3, 4 and 6 days time periods. The healing rates were calculated by using an image analysis system. After 7 days, the corneas were processed for an immunohistochemical observation of proliferating cell nuclear antigen (PCNA) positive cells in the re-surfaced epithelium. Epithelial re-surfacing was significantly enhanced by the hEGF treatment at a concentration of 1.0ng/ml. The thickness of regenerated epithelium appeared to be increased in a dose-dependent man-ner. The rate of endothelial healing was increased at the concentration of 100ng/ml. The results indicate that epithelial healing was stimulated at about 100 times lower concentration of hEGF than endothelial healing.