December 2002
Volume 43, Issue 13
ARVO Annual Meeting Abstract  |   December 2002
Hyperoxia Induces Retinal Endothelial Cells Apoptosis through Formation of Peroxynitrite
Author Affiliations & Notes
  • X Gu
    Medical College of Georgia Augusta GA
    Cellular Biology and Anatomy
  • SE Brooks
    Vascular Biology Center
    Medical College of Georgia Augusta GA
  • RB Caldwell
    Vascular Biology Center Cellular Biology and Anatomy
    Medical College of Georgia Augusta GA
  • Footnotes
    Commercial Relationships   X. Gu, None; S.E. Brooks, None; R.B. Caldwell, None. Grant Identification: NIH Grant EY04618, EY11766, RPB
Investigative Ophthalmology & Visual Science December 2002, Vol.43, 2739. doi:
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      X Gu, SE Brooks, RB Caldwell; Hyperoxia Induces Retinal Endothelial Cells Apoptosis through Formation of Peroxynitrite . Invest. Ophthalmol. Vis. Sci. 2002;43(13):2739.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract: : Purpose: Vascular endothelial cell apoptosis is a major event in the pathogenesis of hyperoxia-induced vaso-obliteration during oxygen-induced retinopathy. Our previous studies suggest that hyperoxia induces endothelial cell apoptosis through formation of the nitric oxide (NO)- and superoxide (O2.-)-derived oxidant peroxinitrite (ONOO-). We have now directly tested this hypothesis and determined the effects of ONOO-on the PI3 kinase/AKT intracellular signaling pathway that controls endothelial cell survival. Methods: Cultured bovine retinal vascular endothelial cells (BREC) were exposed to 40% oxygen (hyperoxia) or 21% oxygen (normoxia). Apoptosis was determined by an assay for caspase-3 activity and by immunoblot analysis of cleaved PARP. Relative quantitative RT-PCR and flurometric DAN reaction were used to measure NOS expression and nitrite formation. Formation of O2.- and ONOO- was measured by using the oxidative fluorescent dye hydroethidine (HE) and by immunoblotting with anti-nitrotyrosine antibody. The NOS inhibitor L-NAME and the O2.- and ONOO- scavengers PEG-SOD and uric acid were tested for effects in blocking hyperoxia-induced apoptosis. Immunoblotting and AKT kinase assay were used to detect the effect of ONOO- on growth factor-induced activation of AKT. Results: BREC exposure to hyperoxia induced apoptosis. This effect was associated with increased formation of nitric oxide, superoxide and peroxynitrite. Hyperoxia also upregulated expression of endothelial NOS. The NOS inhibitor and O2.- and ONOO- scavengers blocked the hyperoxia-induced apoptosis. Analyses of the PI3 kinase/AKT cell survival pathway showed that ONOO- blocks VEGF- or bFGF-induced activation of AKT. Conclusion: Taken together, these studies suggest that the NO derived oxidant ONOO- plays a key role in hyperoxia-induced BREC apoptosis, possibly by blocking growth factor induced activation of the PI3 kinase/AKT survival signaling pathway. It is recognized that ONOO- may also exert its detrimental effect on endothelial cells by altering other pathways. Further studies will be necessary to fully clarify this issue.

Keywords: 323 apoptosis/cell death • 504 oxidation/oxidative or free radical damage • 614 vascular cells 

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