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Title: The role of thioredoxin system in oxidative stress and cell aging
Authors: Lam, Suk-mi Levina
Degree: M.Phil.
Issue Date: 2006
Abstract: Aging is an inevitable part of the life natural process that is governed by decreasing in physiological functions that ultimately result in mortality. Reactive oxygen species (ROS) are believed to be one of the casual factors in aging. Consequently, the ability to respond appropriately to oxidative challenge is likely to be an important factor in combating diseases and disabilities of aging. Unique amongst others, mammalian erythrocytes are continuously subject to oxidative damage but devoid of protein synthesis machinery, which implies the repairing mechanism is most important for cell survival. To defend the attack, there are quite a number of pathways and mechanisms. Thioredoxin system, comprising that of the enzyme thioredoxin (Trx), thioredoxin reductase (TR) and NADPH is one of the best representative systems for its protective function against oxidative stress in various cells. While the existence and importance of thioredoxin system in erythrocytes are least understood, it would therefore be informative to use erythrocytes as a study model to find out the relationship of cell aging and oxidative stress. By means of their difference in density, young and old populations of red cells were separated, characterized and used in the analysis of thioredoxin system components. By immunoblotting, thioredoxin was found in young cells but not in old cells. The activity assay also revealed similar pattern that there was 0.19 U thioredoxin/mg total protein in young cells but no activity can be observed in old cells. Our findings suggest that the thioredoxin system may be involved in the aging process of erythrocyte. To further elucidate the role of thioredoxin system, young and old erythrocytes were subject to oxidative challenge with hydrogen peroxide. We found an increase of 63.5% thioredoxin system activity in young cells challenged with 4 mM H2O2. Though the level and activity of thioredoxin in old cells were not detectable, an enhancement of the whole thioredoxin system activity was noticed after oxidative challenge. Such an enhancement of the system activity implies a regulatory system of the thioredoxin system may exist, which can respond to oxidative stress but de novo protein synthesis is not required. It therefore becomes important to extend our study to nucleated cells to investigate if endogenous regulatory system of thioredoxin system does exist universally. HeLa and HT-29 cell cultures were employed to study the thioredoxin system activity upon oxidative challenge with and without prior treatment of cycloheximide, by which de novo protein synthesis of proteins would be inhibited. In both cell cultures, enhancement of thioredoxin system activity was observed upon oxidative challenge (8.2% and 19.7 % in HeLa cells; 13.5 % and 18.9 % increase in HT-29 cells, with and without the treatment of cycloheximide respectively). In this study, we have demonstrated enhancement of thioredoxin system activity upon oxidative challenge in both nucleated and anucleated cell models and that enhancement was not necessarily dependent on de novo protein synthesis. Our notion is that both de novo protein synthesis and a direct activation of the thioredoxin system may be involved in response to oxidative stress. Together with the yet to be characterized regulatory system, the full thioredoxin system appears to play important roles in cell aging and combat against oxidative stress.
Subjects: Hong Kong Polytechnic University -- Dissertations.
Cells -- Aging.
Oxidative stress.
Pages: vii, 78 leaves : ill. (some col.) ; 30 cm.
Appears in Collections:Thesis

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