April 1999
Volume 40, Issue 5
Free
Articles  |   April 1999
Oxidant-induced apoptosis in cultured human retinal pigment epithelial cells.
Author Affiliations
  • J Cai
    Department of Biochemistry, School of Medicine, Emory University, Atlanta, Georgia 30322, USA.
  • M Wu
    Department of Biochemistry, School of Medicine, Emory University, Atlanta, Georgia 30322, USA.
  • K C Nelson
    Department of Biochemistry, School of Medicine, Emory University, Atlanta, Georgia 30322, USA.
  • P Sternberg, Jr
    Department of Biochemistry, School of Medicine, Emory University, Atlanta, Georgia 30322, USA.
  • D P Jones
    Department of Biochemistry, School of Medicine, Emory University, Atlanta, Georgia 30322, USA.
Investigative Ophthalmology & Visual Science April 1999, Vol.40, 959-966. doi:
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    • Get Citation

      J Cai, M Wu, K C Nelson, P Sternberg, D P Jones; Oxidant-induced apoptosis in cultured human retinal pigment epithelial cells.. Invest. Ophthalmol. Vis. Sci. 1999;40(5):959-966.

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

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Abstract

PURPOSE: To determine the mechanism of oxidant-induced cell death in cultured human retinal pigment epithelium (hRPE). METHODS: Cultured hRPE cells were treated with different concentrations of a chemical oxidant, t-butylhydroperoxide (tBH), for different periods of time. Apoptosis was determined with terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick-end labeling (TUNEL) and flow cytometry. Mitochondrial membrane potential (mtdelta psi) was measured by rhodamine 123 staining and subsequent flow cytometry. Release of mitochondrial cytochrome c (cyt c) and cleavage of procaspase 3 and caspase substrates were determined by western blot analysis. RESULTS: t-Butylhydroperoxide caused time- and dose-dependent activation of apoptosis in hRPE, indicated by characteristic morphologic changes; TUNEL-positive labeling; phosphatidylserine (PS) exposure; and procaspase 3, poly(ADP-ribose)polymerase, lamin, and tubulin cleavage. An early decrease of mtdelta psi was observed before caspase activation, together with the release of mitochondrial cyt c. CONCLUSIONS: Results indicate that tBH can induce apoptosis in hRPE, probably by triggering the mitochondrial permeability transition, which results in swelling and release of mitochondrial intermembrane proteins.

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