Jie Wang1, Andrew Lin, Luo Lu. 1. Division of Molecular Medicine, Department of Medicine, David Geffen School of Medicine, University of California Los Angeles, Torrance, California 90502, USA.
Abstract
PURPOSE: Epidermal growth factor (EGF) stimulates migration in corneal epithelial wound healing. The purpose of this study was to investigate the effect of EGF-induced alpha-tubulin deacetylation through activating HDAC6 on migration in corneal epithelial wound healing. METHODS: Human corneal epithelial (HCE) cells were cultured in DMEM/F12 medium containing 10% FBS in a 37 degrees C incubator supplied with 5% CO(2). Western blot analysis was used to determine protein expression. Activity of HDAC6 was suppressed by trichostatin A (TSA) and by siRNA specific to HDAC6. Corneal epithelial cell migration was measured by using scratch-induced directional migration assay in cultured cells and by corneal epithelial debridement using a mouse whole-eye organ culture model. RESULTS: The authors found EGF stimulated corneal epithelial cell migration in wound healing by enhancing HDAC6 activity, resulting in the deacetylation of alpha-tubulin. EGF stimulated HDAC6 enzymatic activity and protein translocation toward the leading edge of the cell. Inhibition of HDAC6 activity by TSA significantly suppressed EGF-induced cell migration and delayed EGF-induced wound healing in epithelially debrided mouse corneas. In the meantime, knockdown of HDAC6 mRNA with specific siRNA effectively abolished EGF-induced deacetylation of alpha-tubulin, resulting in the inhibition of cell migration. CONCLUSIONS: These results reveal an important mechanism that involves EGF-induced HDAC6 activation and alpha-tubulin deacetylation, subsequently affecting corneal epithelial migration in the wound-healing process.
PURPOSE:Epidermal growth factor (EGF) stimulates migration in corneal epithelial wound healing. The purpose of this study was to investigate the effect of EGF-induced alpha-tubulin deacetylation through activating HDAC6 on migration in corneal epithelial wound healing. METHODS:Human corneal epithelial (HCE) cells were cultured in DMEM/F12 medium containing 10% FBS in a 37 degrees C incubator supplied with 5% CO(2). Western blot analysis was used to determine protein expression. Activity of HDAC6 was suppressed by trichostatin A (TSA) and by siRNA specific to HDAC6. Corneal epithelial cell migration was measured by using scratch-induced directional migration assay in cultured cells and by corneal epithelial debridement using a mouse whole-eye organ culture model. RESULTS: The authors found EGF stimulated corneal epithelial cell migration in wound healing by enhancing HDAC6 activity, resulting in the deacetylation of alpha-tubulin. EGF stimulated HDAC6 enzymatic activity and protein translocation toward the leading edge of the cell. Inhibition of HDAC6 activity by TSA significantly suppressed EGF-induced cell migration and delayed EGF-induced wound healing in epithelially debrided mouse corneas. In the meantime, knockdown of HDAC6 mRNA with specific siRNA effectively abolished EGF-induced deacetylation of alpha-tubulin, resulting in the inhibition of cell migration. CONCLUSIONS: These results reveal an important mechanism that involves EGF-induced HDAC6 activation and alpha-tubulin deacetylation, subsequently affecting corneal epithelial migration in the wound-healing process.
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