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Relative influence of pro- and antioxidant enzymes on the cellular oxidative damage in man. - AN0401

Description
Diet influences the activities of pro- and antioxidant enzymes in both animals and humans. The direct consquences of such changes on cellular oxidative damage is not clear. A sub-cellular system will be developed based upon the microsomal fraction ( membrane vesicles containing the significant enzymes) isolated from various soft tissues and human cell lines qualitatively and quantitatively engineered with respect to key enzymes. Changes in the composition and content of pro- and antioxidant enzymes in subcellular fractions will be effected by a combination of the following: a) direct addition of cytosolic enzymes; b) induction of certain enzymes through exposure to drugs or plants protective factors. Data obtained from this system will form the basis for developing a predictive model for determining the nature/level of cellular oxidative damage in humans.
Objective
1. Purification of target enzymes. 2. Production of microsomal enzyme systems form human cell lines expressing human membrane-bound target enzymes and from human tissues. 3. Characterise the integrity, enzyme levels and metabolite levels of microsomal system. 4. Set up assays to measure oxidative damage of microsomal systems and of whole cells and use to monitor oxidative damage. Eventual assay choice to be made in consultation with other workers from the antioxidant programme. 5. Set up subcellular models using techniques and material developed om 1-4. 6. Assess the relative importance and contribution of target enzymes in subcellular model subjected to oxidative stress. 7. Develop a predictive model for determining the level/nature of cellular oxidative damage utilising marker enzyme activity data. Incorporate putative protective fractions and determine their effect on oxidative damage. 8. Validate this using enzyme activity data from associated research.
Time-Scale and Cost
From: 1992

To: 1995

Cost: £534,229
Contractor / Funded Organisations
IFR - Institute of Food Research (BBSRC)
Keywords