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Abstract #19645 Published in IGR 9-3
Interferon-α and interferon-γ sensitize human tenon fibroblasts to mitomycin-C
Wang XY; Crowston JG; Zoellner H; Healey PRInvestigative Ophthalmology and Visual Science 2007; 48: 3655-3661
PURPOSE: To investigate the effect of interferon (IFN)-α and IFN-γ pretreatment on mitomycin C (MMC)-induced cell death in human Tenon fibroblasts (HTFs) and the mechanisms by which IFN-α and IFN-γ modulate the susceptibility of HTFs to MMC. METHODS: HTFs were pretreated with IFN-α and IFN-γ for 48 hours before 5-minute application of 0.4 mg/mL MMC. Cell death after 48 hours was determined by Annexin V/propidium iodide (PI) staining and lactate dehydrogenase (LDH) release assay. Fas, Fas-ligand, and Bcl-2 expression were determined by flow cytometry. Fas associated death domain (FADD), Bax, cytochrome c, and caspase expression were determined by Western blot analysis and immunofluorescence staining. RESULTS: MMC treatment increased cell death and upregulated Fas and FADD expression, but had no effect on Fas-Ligand, Bax, Bcl-2, or cytochrome c. Neither IFN-α nor IFN-γ alone induced HTF death, but each increased cell death 2 days after MMC treatment in a dose-dependent fashion. Combination IFN-α and IFN-γ had a synergistic effect. IFN-α and IFN-γ pretreatment increased Fas expression. Fas upregulation was associated with increased sensitivity to MMC. IFN pretreatment increased procaspase-8, procaspase-9, and procaspase-3 expression, and caspase-3 activation. Caspase-8, caspase-3, and broad caspase inhibitors, but not caspase-9 inhibitor, inhibited MMC-induced cell death in nonpretreated and IFN-pretreated cells. CONCLUSIONS: IFN-α and IFN-γ enhance the susceptibility of HTFs to MMC-induced cell death through a Fas-mediated and a caspase-3-dependent pathway. Pretreatment with IFN primed HTFs to MMC, providing a potential means for initially slowing the healing response with IFN and subsequently terminating fibroblast activity through MMC-induced cell death.
Dr. X.Y. Wang, Center for Vision Research, University of Sydney, Westmead, Hospital Sydney, NSW, Australia. Xy_wang@wmi.usyd.edu.au
Classification:
12.8.10 Woundhealing antifibrosis (Part of: 12 Surgical treatment > 12.8 Filtering surgery)
3.5 Molecular biology incl. SiRNA (Part of: 3 Laboratory methods)