Dietary polyphenols as modulators of redox signalling pathways to reduce chronic inflammation in the elderly

A great deal of evidence indicates that consumption of diets rich in fruit and vegetables helps maintain health by protecting against age-related disorders including some cancers and cardiovascular diseases. Numerous studies have been undertaken to try and understand the mechanisms by which these diets exert beneficial effects on health. Many previous studies identified the potentially protective components of fruit and vegetables as 'antioxidants' that were thought to prevent a generalised unwanted oxidation of cells and tissues that increased with ageing. However, despite extensive studies, formal trials have shown no beneficial effect of 'antioxidants' when they are given as supplements. In recent years it has also become recognised that oxidation processes, mediated by highly reactive free radicals, are not necessarily deleterious to cells, but may be very localised within sub-cellular compartments of cells and that they normally regulate intracellular signalling processes and mediate many key physiological effects. Thus the original idea that 'antioxidants' in the diet act to scavenge all oxidants in cells and tissues and therefore produce benefit has been questioned. During ageing there is a breakdown of highly regulated cell signalling pathways due to changes in oxidation in very specific parts of cells and this leads to the production of deleterious substances that promote inflammation and which increase the susceptibility of the elderly to many chronic disorders. It is clear from internationally-based studies of diets consumed by healthier elderly populations that a group of compounds found in fruit and vegetables, called polyphenols are beneficial against many age-related disorders, but we currently do not understand how they act to produce these health benefits. Polyphenols were originally thought to have beneficial effects because they are 'antioxidants' and our hypothesis is that they are protective against chronic age-related disorders by targeting specific parts of the cell to prevent oxidation of key components of the beneficial intracellular signalling pathways. New developments in analytical techniques now permit analysis of the potential effects of dietary polyphenols on local oxidation and cell signalling processes at key sub-cellular sites and the proposed study will utilise these new techniques. As part of the study, we will develop a cell culture-based test to identify which specific dietary polyphenols can exert beneficial effects, and undertake studies with groups of elderly subjects to determine whether any polyphenols identified as beneficial in the cell culture test can reduce markers of inflammation. Thus the project will potentially lead: 1) to identification of those dietary polyphenols that act to minimise the pro-inflammatory state in the elderly and therefore provide a basis for the food industry to make logical and justified cases to the public for consumption of specific foods. 2) to identification of polyphenols that act to minimise the pro-inflammatory state and which may provide the basis for novel formulations of polyphenol-enriched food products that could be targeted at maintenance and imporvement of health in the elderly. 3) to provision, in the longer term, of the underpinning data that leads to a diet-based improvement in immune function and a delay in the onset of frailty in the elderly.

The major thiol/disulphide couples: glutathione, thioredoxins (TRx) and cysteine/cystine are key regulators of redox-signalling pathways. These become oxidised during ageing and cause activation of redox-sensitive transcription factors leading to increased pro-inflammatory cytokines. Polyphenols from fruit and vegetables protect against age-related disorders and gene array studies indicate that they modify cellular thiols. The project will determine whether polyphenols and metabolites affect key thiol couples in cell culture and in intervention studies examining subjects of different ages. Lymphocytes from subjects aged 20, 40, 60 and 80+ will be analysed for reduced and oxidised glutathione, cysteine and cystine, nuclear and cytosolic TRx1 and mitochondrial TRx2 redox status, activation of NFkappa-B and cytokine production. Plasma cytokines, reduced and oxidised glutathione, cysteine and cystine will be analysed. Jurkat cells transfected with glutaredoxin1 (Grx1) fused to redox-sensitive green fluorescent protein (roGFP) to monitor sub-cellular reduced/oxidised glutathione couples will be used as a model to identify polyphenols that modify thiol couples. TRx1 and 2 redox couples and cellular cysteine/cystine will also be determined. Activation of redox-sensitive transcription factors and release of cytokines will be examined for those polyphenols found to affect thiol couples. Foodstuffs containing the polyphenols and metabolites effective in the cell model will be studied in 2 day intervention studies in healthy elderly (80+ years) subjects. Urinary metabolites of polyphenols will be measured and lymphocytes analysed for thiol couples, redox-sensitive transcription factors and cytokine production. Subsequently the most effective foodstuff, or combination of foodstuffs, will be studied in a 10 week intervention examining chronic effects on circulating pro-inflammatory cytokines in subjects of different ages.

Who will benefit from this research? Industry will benefit from optimisation of existing foodstuffs and development of novel foodstuffs. The UK and international populations will benefit from development of novel formulations of polyphenol-enriched foods targeted at healthy ageing. Staff working directly on the project will also benefit from the research. How will they benefit from this research? The potential impact of this research is extensive. The proportion of elderly people in the UK population is increasing dramatically, particularly the frail elderly. Inflammation is a recognised risk factor underpinning many chronic age-related disorders including atherosclerosis, cancer and diabetes. Research has confirmed that diets rich in fruits and vegetables help maintain health and protect against age-related disorders, including some cancers and cardiovascular diseases. Data from this study will aid optimisation, targeted marketing and exploitation of existing polyphenol-rich foods such as those in specific beverages or cocoa/chocolate-based foodstuffs. Identification of novel polyphenols or polyphenol mixtures may lead to improvements in health of the elderly population. In the longer term the project may lead to a diet-based improvement in prevention of frailty in our ageing population with decreased susceptibility to infections, an overall reduction in the need for assisted care of the elderly and hence enhanced quality of life in older people. We anticipate that the design of the current study is such that information provided at the end of the study will have an immediate and direct application to the UK population. Staff working directly on the project will benefit from the development of skills related to industrial implementation of research outcomes, skills which are applicable to a number of employment sectors. What will be done to ensure that they benefit from this research? The applicants regularly give lay presentations to the public and schools in association with relevant charitable organisations such as Help the Aged (now AgeUK) and provide material for relevant local and national press, television and radio programmes. Professors Jackson and McArdle have worked with the Research into Ageing arm of Help the Aged for dissemination of research findings to the general public at public lectures and with the local and national press. Specific objectives by which the applicants will engage with beneficiaries: 1. The applicants will be fully engaged with the DRINC panel, working to disseminate information to industry, scientists and the general public by attendance at workshops, press releases and public presentations where appropriate. 2. The applicants have considerable experience in organising specialist short courses, conferences and workshops (eg Practical Workshop on the detection of ROS in cells and tissues; British Society for Research on Ageing, 2007-2009) attended by clinical and non-clinical scientists and industry. Media are informed of the event. The topic of this study will be incorporated into the programme of such activities. The outcome of such activities will include an academic review of the research area which details proposed ways forward. 3. The applicants will seek assistance from organisations such as AgeUK on dissemination of information to the general population by the most appropriate routes including web-based information, flyers and training for health professionals. 4. The applicants have been approached to produce text books for undergraduate education in this area of research and will examine the possibility of producing an equivalent text for the lay public. 5. The School of Clinical Sciences at the University of Liverpool is currently developing a strategy for research staff to engage more actively in local schools and community centres and we would foresee dissemination via those routes, particularly to target audiences.