The role of mitochondrial oxidative damage in ageing: a chemical intervention approach

During ageing the human body becomes less able to do what it once could. The reason is that damage accumulates to the molecules of which cells are made up and to whole tissues. One common form of damage comes from oxygen, which is essential for breathing and respiration, but is also a danger to biological molecules. In cells, the energy-storage molecule ATP is produced in the mitochondria, by using oxygen to burn food supplies such as sugar and fats. When demand for energy is high, as for example during exercise, the mitochondria are not much of a problem. But when they have nothing to do because energy is not being consumed, they become much more likely to interact with oxygen to produce dangerous reactive molecules. These can go on to damage important biological molecules such as DNA and protein and the lipids in biological membranes, and this damage may be an important part of the ageing process. The damage is particularly likely to happen in the mitochondria themselves. So mitochondria may be key players in the ageing process, both as the site at which damaging molecules are produced and then as the main target of the damage that they cause. In this project we plan to develop and use chemicals that are targeted to the mitochondria to test these ideas and to try and intervene in the damage process. There is a charge across the surface of the mitochondria. We shall use a charged chemical 'tug' to drag biologically active molecules into the mitochondria. Some of these molecules act as reporters, for instance of the rate at which the mitochondria are generating the dangerous reactive molecules, the amount of damage that they are causing and the size of the charge across the surface of the mitochondria. Others protect against dangerous reactive molecules by detoxifying them. We shall test the efficacy of these molecules in a short-lived species, the fruit fly Drosophila, by adding them to the food. One of the advantages of Drosophila for this kind of work is that we can make long-lived strains, by feeding the flies on a restricted diet or making mutations in genes. We can therefore assess the role of mitochondrial damage in slowing down the ageing process in these long-lived strains. Ultimately, the goal of this work is to improve the health and quality of life of people as they age, and the aim of this project is to pave the way to the production of drugs that could achieve this goal.

The leading candidate mechanism for the generation of molecular damage during ageing is the free radical theory. It postulates that reactive oxygen and nitrogen species generated by the interaction of oxygen with electrons from the mitochondrial electron transport chain cause damage to DNA, protein and lipid, particularly in the mitochondria themselves. Despite abundant correlative evidence consistent with the free radical theory, critical experimental tests have proved elusive, and details of the molecular processes at work await elucidation. Progress has been hampered by the lack of chemical reagents to report on and modify critical aspects of the postulated mechanisms, particularly in vivo. In this project, we propose to develop chemical probes and reagents that are targeted to the mitochondria in vivo. These will be designed to: (a) block oxidative damage, (b) measure oxidation of protein thiols, (c) measure production of reactive oxygen and nitrogen species, (d) measure mitochondrial membrane potential. These chemicals will be validated using the fruit fly, Drosophila by making measures of oxidative damage to DNA, protein and lipid in mitochondria and elsewhere, of membrane potential, and of effect on lifespan in oxidatively stressed and normal flies. We shall also use the chemical reporters to determine how different forms of damage and mitochondrial membrane potential are altered in flies that are long-lived through dietary restriction or reduced activity of the insulin/IGF signalling pathway.