Interferon-alpha and interferon-gamma modulate Fas-mediated apoptosis in mitomycin-C-resistant human Tenon's fibroblasts.
BACKGROUND
The aim of the study was to investigate, using a native mitomycin-C-resistant human Tenon's fibroblast cell line, the possibility that interferon-alpha and gamma could be used with Fas agonists as an alternative anti-fibrotic strategy to mitomycin-C in trabeculectomy.


METHODS
A clinically resistant and in vitro verified mitomycin-C-resistant human Tenon's fibroblast cell line was pretreated with interferon-alpha and interferon-gamma for 48 h before stimulation with an agonistic Fas antibody (CH11) for 2 days to induce cell death. Cell death assays were undertaken. Changes in apoptosis-related proteins were determined by flow cytometry and Western blot.


RESULTS
Pretreatment with interferon-alpha or interferon-gamma for 48 h increased Fas, Fas-associated protein with death domain and caspase-8 expression. Protein expression was further increased by combined exposure to interferon-alpha and gamma. Pretreatment with cytokines had no effect on Fas-L and Bcl-2. Interferon-alpha alone did not change the rate of induced cell death. A combination of interferon-alpha and gamma synergistically increased the sensitivity of mitomycin-C-resistant human Tenon's fibroblast cell line to induced cell death. An antagonistic anti-Fas antibody (ZB4) completely blocked induced cell death. Broad caspase inhibitors specific for caspases-8 and -3 reduced induced deaths in interferon pretreated mitomycin-C-resistant human Tenon's fibroblast cell line in a dose-dependent manner.


CONCLUSIONS
Interferon-alpha and interferon-gamma render mitomycin-C-resistant human Tenon's fibroblast cell line sensitive to Fas-mediated apoptosis. The mechanism involves increased death-inducing signalling complex formation by upregulation of Fas, Fas-associated protein with death domain and caspase-8 expression.
Centre for Vision Research, Westmead Millennium Institute, The University of Sydney, Sydney, New South Wales, Australia.
Xiao Yang
XY
Wang
Jonathan G
JG
Crowston
Andrew J R
AJ
White
Hans
H
Zoellner
Paul R
PR
Healey
D017304:Annexin A5 / Q000378:metabolism; D000998:Antiviral Agents / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects*; D015153:Blotting, Western; D002460:Cell Line; D004351:Drug Resistance; D053222:Fas Ligand Protein / Q000378:metabolism*; D005347:Fibroblasts / Q000378:metabolism / Q000473:pathology; D005434:Flow Cytometry; D019084:Fluorescent Antibody Technique, Indirect; D006801:Humans; D016898:Interferon-alpha / Q000494:pharmacology*; D007371:Interferon-gamma / Q000494:pharmacology*; D016685:Mitomycin / Q000494:pharmacology*; D020411:Oligonucleotide Array Sequence Analysis; D058475:Tenon Capsule / Q000378:metabolism / Q000473:pathology*
D017304:Annexin A5; D000998:Antiviral Agents; D053222:Fas Ligand Protein; D016898:Interferon-alpha; D016685:Mitomycin; D007371:Interferon-gamma
Fas; apoptosis; glaucoma; interferon; trabeculectomy