May 2003
Volume 44, Issue 13
Free
ARVO Annual Meeting Abstract  |   May 2003
Hydrogen Peroxide Selectively Protects Lens Epithelial Cells from Apoptosis by Activating Phosphatidylinositol-3 kinase/Akt Signaling Pathway
Author Affiliations & Notes
  • D. Sailaja
    Department of Ophthalmology and Neuroscience Center, LSU Health Sciences Center, New Orleans, LA, United States
  • G. Chandrasekher
    Department of Ophthalmology and Neuroscience Center, LSU Health Sciences Center, New Orleans, LA, United States
  • Footnotes
    Commercial Relationships  D. Sailaja, None; G. Chandrasekher, None.
  • Footnotes
    Support  NIH/NEI Ro1 EY 12701
Investigative Ophthalmology & Visual Science May 2003, Vol.44, 310. doi:
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      D. Sailaja, G. Chandrasekher; Hydrogen Peroxide Selectively Protects Lens Epithelial Cells from Apoptosis by Activating Phosphatidylinositol-3 kinase/Akt Signaling Pathway . Invest. Ophthalmol. Vis. Sci. 2003;44(13):310.

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

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Abstract

Abstract: : Purpose: Growth factor-mediated phosphatidylinositol-3 kinase (PI-3K)/Akt signaling is involved in protecting several cell types from apoptosis. Previous studies from our laboratory showed that, similar to growth factors, hydrogen peroxide (H2O2) can also stimulate PI-3K/Akt pathway in lens epithelial cells ( ARVO Meeting 2002, Abs.No. 2371). The purpose of the current study was to investigate whether PI-3K/Akt pathway activated by H2O2 can exhibit an anti-apoptotic effect in lens epithelial cells. Methods: Rabbit lens or corneal epithelial cells derived from primary cultures were stimulated with H2O2 (150-250 µM). Epithelial cells were subjected to programmed cell death using staurosporin (10 ng/ml). Cell apoptosis was determined by identifying characteristic DNA fragmentation in agarose gels and DNA condensation in live cells by Hoechst staining. Percentages of apoptotic cells in cultures were determined using Image-Proplus computer software program. Activation of anti-apototic factor Akt was determined by identifying phospho-Akt by SDS-PAGE followed by immunoblotting. Results: Treatment of cells with staurosporin produced significant apoptosis as measured by DNA laddering and Hoechst staining. H2O2 did not induce apoptosis in epithelial cells. Incubation of lens epithelial cells with H2O2 before and during treatment with staurosporin almost completely blocked the DNA fragmentation and reduced the Hoechst stain-positive cell count by more than 50%. Presence of PI-3K inhibitors wortmannin (200 nM) and LY294002 (12.5 µM) reduced the ability of H2O2 to block DNA fragmentation and condensation. Wortmannin and LY294002 inhibited Akt activation. Unlike in lens epithelial cells, H2O2 did not show anti-apoptotic properties in corneal epithelial cells and H2O2 treatment had no inhibitory effect on DNA fragmentation during the staurosporin-induced apoptosis process. Conclusions: H2O2, which has been implicated in cataract formation, did not cause lens epithelial cell apoptosis; rather, it rescues these cells from apoptosis. Absence of this protective function of H2O2 in other ocular tissue such as corneal epithelium highlights the unique property of lens epithelial cells to adapt to H2O2-oxidative stress environment and to use this for survival signaling. Our studies also demonstrated that PI-3K/Akt signaling pathway activated by H2O2 plays an important role in preventing lens epithelial cell apoptosis.

Keywords: signal transduction • apoptosis/cell death • oxidation/oxidative or free radical damage 
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