Bcl-2 overexpression increases survival in human retinal pigment epithelial cells exposed to H(2)O(2).
The integrity of the retinal pigment epithelium, especially that of the macula is essential for the preservation of vision into old age. The chronic exposure to sunlight and peroxidized lipids from phagocytized photoreceptor outer segments imposes a high level of oxidative stress on the retinal tissues, which increases with age as antioxidant protection declines and therefore could accelerate apoptosis. Bcl-2 known to facilitate mitochondrial DNA repair and cellular survival in other tissues was overexpressed in a single clone of human retinal pigment epithelium cells after stable transfection with humanbcl-2 in rhoSFV-neoexpression factor. Near confluent cells (2nd-4th generation permanently bcl-2 transfected) were protected from mitochondrial dysfunction after exposure to H(2)O(2) up to 150 microM. With 200 microM H(2)O(2), function in transfected cells declined by only 25% control activity as determined by MTT reduction assays, compared to wild type and vector only transfected cells expressing normal bcl-2 levels. Similarly the bcl-2 -transfected cells were more resistant to mitochondrial DNA damage after H(2)O(2) treatment than the other groups and suffered 50% less damage after exposure to 200 microM H(2)O(2), as assayed by quantitative polymerase chain reaction assays. These data suggest that bcl-2 overexpression protects human RPE cells from mitochondrial respiratory dysfuction, mitochondrial DNA damage and promotes cellular survival in response to oxidative stress induced by H(2)O(2).
Godley
Bernard F
BF
Retina Foundation of the Southwest, Dallas, TX 75321, USA. bgodley@retinafoundation.org
Jin
Gui-Fang
GF
Guo
Yan-Shi
YS
Hurst
John S
JS
D015153:Blotting, Western; D018929:Cell Culture Techniques; D002470:Cell Survival / Q000187:drug effects / Q000502:physiology; D004249:DNA Damage; D004272:DNA, Mitochondrial / Q000235:genetics; D004305:Dose-Response Relationship, Drug; D019254:Genes, bcl-2; D006801:Humans; D006861:Hydrogen Peroxide / Q000494:pharmacology*; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism; D010084:Oxidation-Reduction; D018384:Oxidative Stress / Q000502:physiology; D010857:Pigment Epithelium of Eye / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000502:physiology*; D014162:Transfection
D004272:DNA, Mitochondrial; D019253:Proto-Oncogene Proteins c-bcl-2; D006861:Hydrogen Peroxide