Catalase enrichment using recombinant adenovirus protects alphaTN4-1 cells from H(2)O(2).
Since oxidative stress has been implicated in the development of numerous diseases including cataract, this laboratory has created and investigated the stress response of murine immortal lens epithelial cell lines (alphaTN4-1) conditioned to withstand lethal peroxide concentrations. Two of a group of antioxidative defense (AOD) enzymes found in such cells to have markedly enhanced activity are catalase (CAT) and GSH S-transferase alpha2 (GST). In order to determine if enrichment of one or both of these AODs is sufficient to protect alphaTN4-1 cells from lethal H(2)O(2) levels, these cells were infected with adenovirus vectors capable of expressing these AODs at a high level. With this system, gene enrichment and increased enzyme activity were observed with both CAT and GST vectors. The percentage of cells infected ranged from about 50 to 90% depending on the multiplicity of infection (MOI). CAT but not GST protected the cells from H(2)O(2) stress. The CAT activity was increased from 15- to 150-fold and even at the lower levels protected the cells from H(2)O(2) concentrations as high as 200 microM or more (H(2)O(2) levels which rapidly kill non-enriched cells). Even when only about 50% of the cell population is infected as judged by GFP infection, the entire population appeared to be protected based on cell viability. The CAT enrichment appears to protect other intracellular defense systems such as GSH from being depleted in contrast to non-enriched cell populations where GSH is rapidly exhausted. The overall results suggest that enriching the cellular CAT gene level with an appropriate recombinant viral vector may be sufficient to protect in vivo systems from peroxide stress.
Ma
Wanchao
W
Department of Ophthalmology, Columbia University, 630 West 168th Street, New York, NY, USA.
Nunes
Irene
I
Young
C S Hamish
CS
Spector
Abraham
A
D000256:Adenoviridae / Q000201:enzymology* / Q000235:genetics; D000818:Animals; D002374:Catalase / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D019610:Cytoprotection / Q000235:genetics; D004351:Drug Resistance / Q000235:genetics; D004847:Epithelial Cells / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism*; D018014:Gene Transfer Techniques; D017930:Genes, Reporter; D005822:Genetic Vectors / Q000235:genetics / Q000378:metabolism; D005978:Glutathione / Q000187:drug effects / Q000378:metabolism; D006861:Hydrogen Peroxide / Q000378:metabolism / Q000633:toxicity*; D007908:Lens, Crystalline / Q000166:cytology; D051379:Mice; D011995:Recombination, Genetic
D006861:Hydrogen Peroxide; D002374:Catalase; D005978:Glutathione