Hydrogen peroxide stimulates apoptosis in cultured human retinal pigment epithelial cells.
OBJECTIVE
To determine whether hydrogen peroxide (H2O2), a physiological mediator of oxidative stress induces apoptosis in retinal pigment epithelial (RPE) cells.


METHODS
To demonstrate that oxidatively stressed retinal pigment epithelial cells undergo apoptosis consequential to mitochondrial dysfunction, biochemical parameters of apoptosis were determined in cultured cells after treatment with 50-200 mM H2O2 for different times. Caspase-3 protease activity was determined from hydrolysis of DEVD-rho-nitroanilide. Expression of the anti-apoptotic protein, bcl-2 and the pro-apoptotic proteins p53 and p21 were analyzed by western blotting.


RESULTS
Caspase-3 activity significantly increased in cells exposed to H2O2. Also, the expression of bcl-2 in cells treated with 200 microM H2O2 was diminished, whereas expression of p53 and p21waf-1 was increased compared to the controls.


CONCLUSIONS
Exposure of retinal pigment epithelial cells to concentrations of H2O2 that cause in vitro mitochondrial DNA damage also promotes apoptosis.
Jin
G F
GF
Department of Ophthalmology and Visual Sciences,University of Texas Medical Branch, Galveston, Texas 77555-0787, USA.
Hurst
J S
JS
Godley
B F
BF
D017209:Apoptosis / Q000187:drug effects*; D015153:Blotting, Western; D053148:Caspase 3; D020169:Caspases / Q000378:metabolism; D002478:Cells, Cultured; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins / Q000378:metabolism; D006801:Humans; D006861:Hydrogen Peroxide / Q000494:pharmacology*; D008297:Male; D018384:Oxidative Stress / Q000187:drug effects; D010857:Pigment Epithelium of Eye / Q000187:drug effects* / Q000378:metabolism; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D013997:Time Factors; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism
C493956:CDKN1A protein, human; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins; D019253:Proto-Oncogene Proteins c-bcl-2; D016159:Tumor Suppressor Protein p53; D006861:Hydrogen Peroxide; C505407:CASP3 protein, human; D053148:Caspase 3; D020169:Caspases