Purchase this article with an account.
D. Li, W. Ma, F. Sun, A. Spector; The Combined Protective Effect of Two Antioxidative Defense Genes . Invest. Ophthalmol. Vis. Sci. 2003;44(13):322.
Download citation file:
© ARVO (1962-2015); The Authors (2016-present)
Purpose: Since oxidative stress is an initiating or early event in the development of cataract, murine immortal lens epithelial cell lines have been developed which survive various peroxide stresses. Analysis of the gene expression of 12,422 genes and ESTs in these conditioned cell lines indicated a small group of antioxidative defense genes which had significantly amplified expression. Are all of these genes required for defense against oxidative stress or is the cellular response indicative of biological redundancy and/or a change in control of gene expression? To begin to answer this question, two genes, catalase and GSH-S-transferase alpha 2 (GSTα2) have been transfected into control murine and human immortal lens epithelial cell lines to determine the degree of protection they provide to the cells when subjected to a number of oxidative stresses and to compare their behavior to the oxidative stress resistant lines. Methods: αTN4-1 murine immortal lens epithelial cells were conditioned to survive H2O2 stress (H), (tertiary butylhydroperoxide) (T) and a combination of both H2O2 and TBHP (HT) by gradually increasing the peroxide concentration in the medium over a period of six to eight months. Such cells, as well as unconditioned αTN4-1 and human immortal lens epithelial cells (HLE-B3) were then compared to cell lines transfected with catalase and GSTα2. In some cases, a vector carrying both cDNAs was utilized. This vector contains the cytomegalovirus immediate early promoter and elongation factor 1α(EF-1α) promoter to facilitate simultaneous expression of catalase and GSTα2. The resistance of the various cell lines to a variety of oxidative stresses can then be measured by examining cell viability, metabolic function, DNA damage and membrane integrity. Results: Transfection of both αTN4-1 and HLE-B3 cells resulted in positive Western blots for both genes in both systems. Preliminary enzyme activity measurements were significantly elevated. The ability of these cells to resist oxidative stress will be reported. Conclusions: If these cell lines with enriched catalase and GSTα2 are shown to protect the cell systems from oxidative stress, it will offer a basis for utilizing such enrichment in animal systems to possibly provide the necessary protection to prevent cataract formation.
This PDF is available to Subscribers Only