期刊
INVESTIGATIVE OPHTHALMOLOGY & VISUAL SCIENCE
卷 52, 期 13, 页码 9291-9297出版社
ASSOC RESEARCH VISION OPHTHALMOLOGY INC
DOI: 10.1167/iovs.11-8312
关键词
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资金
- Eye Bank Association
- National Eye Institute/National Institutes of Health [R01 EY20581]
- Research to Prevent Blindness
PURPOSE. This study compared susceptibility of Fuchs endothelial corneal dystrophy (FECD) and normal corneal endothelial cells (CECs) to oxidative stress, and studied the mechanism of oxidative-stress-induced apoptosis in FECD-affected endothelium. METHODS. For in vitro studies, immortalized normal and FECD human corneal endothelial cell lines (HCECi and FECDi, respectively) were exposed to tert-butyl hydroperoxide (tBHP). Apoptotic cell populations were distinguished using flow cytometry. Reactive oxygen species production was measured by a horseradish peroxidase assay. For ex vivo studies, CECs were exposed to tBHP. Oxidative DNA damage and apoptosis were assessed by anti-8-hydroxydeoxyguanosine antibody and TUNEL assay, respectively. p53 and phospho-p53 levels were assessed by Western blot and immunohistochemistry. RESULTS. Flow cytometry revealed a higher rate of apoptosis in FECDi than that in HCECi after exposure to 0.5 mM (P = 0.010) and 1.0 mM tBHP (P = 0.041). Further analysis showed increased production of H(2)O(2) by FECDi than that by HCECi. Oxidative DNA damage increased in both normal and FECD CECs after exposure to 0.5 mM tBHP (P = 0.031 and 0.022, respectively), leading to a 21% increase in TUNEL-positive CECs in FECD (P = 0.015) but no change in normal. Baseline p53 expression was twofold higher in FECD than that in normal endothelium (P = 0.002). Immunofluorescence revealed an increase in p53 and phospho-p53 levels in FECD compared with that in normal endothelium. CONCLUSIONS. FECD CECs are more susceptible to oxidative DNA damage and oxidative-stress-induced apoptosis than normal. Increased activation of p53 in FECD suggests that it mediates cell death in susceptible CECs. The authors conclude that p53 plays a critical role in complex mechanisms regulating oxidative-stress-induced apoptosis in FECD. (Invest Ophthalmol Vis Sci. 2011; 52: 9291-9297) DOI: 10.1167/iovs.11-8312
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