The aim is to examine histopathological changes and expression of epidermal growth factor receptor (EGFR) in tracheal epithelia caused by application of topical mitomycin-C (MMC) in rabbit model after the tracheotomy procedure. The conventional tracheotomy was performed in 16 rabbits. They were randomly divided into two equal groups. The first group was applied MMC at a concentration of 0.4 mg/ml around tracheotomy for 5 min, and the other group was not taken a treatment as a control. The animals were sacrificed at the end of 4 weeks. Their tracheas were evaluated with H&E and Masson's trichrome histochemically, and with antiepidermal growth factor receptor immunohistochemically. Results showed that there was no significant difference between MMC and control group for inflammatory cells (P=0.09). The numbers of fibroblasts and subepithelial tissue thickness in the group exposed to MMC were significantly lower than the control group (P<0.05). In contrast, the percentage of EGFR in the application of MMC group was significantly higher than the control group (P<0.05). The application of topical MMC on airway epithelia after tracheotomy showed significant elevation in the levels of epithelial EGFR expression compared to controls in a rabbit model. The activation of epithelial EGFR may facilitate epithelial healing, but further studies are needed to assess the effect of topical MMC on respiratory epithelia.