Unmasking the mechanisms of action of dietary bioactives in berries, cocoa, tea and wine
Lead Research Organisation:
University of East Anglia
Department Name: Pharmacy
Abstract
Despite nearly exponential growth in the past 20 years, flavonoid research has had limited success in realising the structure-function relationships and pathways behind the reported benefits of consumption. Initially, studies focused on antioxidant effects, without taking into account their limited bioavailability. New directions point towards the activity of flavonoid metabolites, which are much more abundant. The present project will identify mechanisms of action of human metabolites, ultimately informing the design of future interventions addressing dietary change for health benefit. Assessment will be focused on biomarkers and pathways involved in cardiovascular health. The project involves screening (protein, mRNA and transcription factor analysis, as well as functional trans-endothelial migration analysis), pathway inhibition (JNK/p38, ERK/Akt and other) and synergistic activity analysis using human cell culture models.
Studentship Projects
| Project Reference | Relationship | Related To | Start | End | Student Name |
|---|---|---|---|---|---|
| BB/M011216/1 | 01/10/2015 | 31/03/2024 | |||
| 1771116 | Studentship | BB/M011216/1 | 01/10/2016 | 31/03/2021 | Nursabah Atli |
| Description | A large number of most common flavonoid metabolites, related analogues and synthetic analogues of phenolic acids have been screened at physiological concentrations for their effect on expression of pro-inflammatory biomarker TNF-a in LPS-induced THP-1 cells. Our results, in line with our previous findings show that flavonoid metabolites did not poses a dose-response effect and more active at lower concentrations. The key finding from this screening was that the synthetic analogue of ferulic acid, ferulic-acid-hexyl-ester (FA-Hex-Es) and ferulic-acid-propyl-ester (FA-Pro-Es) showed a dose response on TNF-a protein secretion. We further investigated the effect of those compounds on TNF-a gene expression. Our results indicated that the effect was protein specific. Next, we studied their effect on the protein and the gene the expression of anti-inflammatory biomarkers: HO-1 and NQO1 as well as the activity of their transcription regulator, Nrf2. We found that both FA-Hex-Es and FA-Pro-Es elevated the gene and protein expression of HO-1 and a moderate but significant effect on NQO-1 protein expression. However, we found no significant effect on Nrf2 activity as well as no effect on NQO1 gene expression. In Conclusion, the data obtained from this study, suggest that hexyl ester combined with FA alkyl chain enhance bioactivity at physiologically achievable concentration and this effect is likely due to lipophilicity. This finding implies that metabolites most likely are transported into the cells and the site of action is intracellular rather than receptors. Furthermore, our protein versus gene expression data indicates that the mechanism of action of metabolites are likely to be post-translational modification. We have also established and trans-endothelial-migration (TEM) assay to screen and study the effect of flavonoid metabolites and phenolic acid analogues on migration of primary human monocytes and neutrophils through human dermal microvascular endothelium in response to chemotactic fMLP stimulation. In this study we found that few phenolic acids were able to significantly prevent trans-endothelial migration of primary human monocytes but no significant effect of neutrophils migration. To follow up TEM, we further investigated the effect of most potent compounds on MCP-1 secretion from neutrophil like cells (differentiated HL-60 cell line) and migration of THP-1 monocytes towards recombinant MCP-1. We find non effect on MCP-1 secretion and non-significant but as strong trend in inhibition of THP-1 migration. Furthermore, we investigated the effect of naturally occurring phenolic acids and their synthetic esters on pro-inflammatory biomarker IL-6 secretion in BV2 microglial cell line. FA-Hex-E significantly attenuated IL-6 gene and protein expression. Then we carried forward FA-Hex-Es to in investigate its MOA on transcription factor NF-kB and AP-1 as well as on JNK phosphorylation. FA-Hex-Es has significantly attenuated NF-kB. Abbreviations: LPS: lipopolysaccharide TNF-a: Tumor necrosis factor FA: ferulic acid FA-Pro-Es: ferulic acid hexyl ester FA-Hex-Es: ferulic acid hexyl ester TEM: trans-endothelial-migration fMLP: N-Formyl methionine-leucyl-phenylalanine MOA: mechanism of action HO-1: Heme oxygenase-1 NQO1: NAD(P)H dehydrogenase [quinone] 1 Nrf2: nuclear factor erythroid 2-related factor 2, (aka nuclear factor erythroid-derived 2-like 2) NF-kB: nuclear factor kappa-light-chain-enhancer of activated B cells AP-1: Activator protein 1 MCP-1: monocyte chemoattractant protein-1 (aka CCL2) JNK: c-Jun N-terminal kinases IL-6: Interleukin 6 |
| Exploitation Route | The uptake and solubility of flavonoid metabolites are not well studied. Our findings provide some evidences that the side of action of flavonoid metabolites are likely to be intracellular and through post-translational mechanism. These findings might help other researchers to explore site of action and narrow down the target molecules. |
| Sectors | Agriculture, Food and Drink,Manufacturing, including Industrial Biotechology,Pharmaceuticals and Medical Biotechnology |
| Description | Collaboration with Dr Colin Kay, North Carolina State University |
| Organisation | North Carolina State University |
| Department | Plants for Human Health Institute |
| Country | United States |
| Sector | Academic/University |
| PI Contribution | We are providing synthetic metabolites and Dr Kay is using them as research tools to look for them in biological samples |
| Collaborator Contribution | Dr Kay uses mass spectrometry to investigate the presence of flavonoid metabolites in human biological samples. Based upon metabolite profiles, he has predicted structures that may be presented and we have synthesised the target compounds. He is now matching the fingerprint of the samples with the synthetic compounds. |
| Impact | No publications to date from this funded project |
| Start Year | 2016 |
| Description | Collaboration with Unilever |
| Organisation | Unilever |
| Department | Unilever Research and Development Vlaardingen B.V. |
| Country | Netherlands |
| Sector | Private |
| PI Contribution | Provision of compounds and expertise on flavonoid metabolites, analysis of results |
| Collaborator Contribution | Expertise in structure activity relationships, transendothelial migration assay and statistics. Technique transferred to O'Connell lab in the UK for further optimisation. |
| Impact | Manuscript in preparation, Transfer of compounds and transendothelial migration technique to O'Connell lab in the UK |
| Start Year | 2017 |
| Description | Event at the Norfolk Show |
| Form Of Engagement Activity | Participation in an activity, workshop or similar |
| Part Of Official Scheme? | No |
| Geographic Reach | International |
| Primary Audience | Public/other audiences |
| Results and Impact | We ran a range of information and hands on activities on the beneficial effect of flavonoids and health for the general public at the Norfolk Show, the largest country show in the UK. More than 350 people across a wide age range took part in the hands on activities, including games, quizzes, drawing superfruit heroes, making chemical models etc and more asked information about diet. People commented that this had raised their awareness of this research. |
| Year(s) Of Engagement Activity | 2019 |
| Description | Norwich Science Festival |
| Form Of Engagement Activity | Participation in an activity, workshop or similar |
| Part Of Official Scheme? | No |
| Geographic Reach | National |
| Primary Audience | Public/other audiences |
| Results and Impact | Participated in British Immunological Society allergy busters stand and involved in a range of information and hand on activities. More than 700 people from all ages participated in the quizzes, games, drawing the nose bugs, making snot, the ink lungs and mast cell and observing lung tissue and lung cells under microscope. More information asked about allergies. Many comments from the public that this rise their awareness of the science behind. |
| Year(s) Of Engagement Activity | 2019 |
| URL | http://www.uea.ac.uk/pharmacy/news-and-events/-/asset_publisher/w7O8j7rUDTtg/blog/id/31133551 |
| Description | Norwich Science Festival |
| Form Of Engagement Activity | Participation in an activity, workshop or similar |
| Part Of Official Scheme? | No |
| Geographic Reach | National |
| Primary Audience | Public/other audiences |
| Results and Impact | I engaged in different activities ranging from explaining molecular medicines to what a pharmacist does, from amino acids to proteins and drug targets as well as DNA and gene expression. The majority of public were kids with varying ages and backgrounds. To drag their attention and to engage them, I have invented a small play where kids could guess one shared common thing between five different molecules and make a hexagonal shape out of colourful sticks. At the end of the day, we have heard that our stand was one of the busiest stand of the festival. |
| Year(s) Of Engagement Activity | 2018 |
| Description | Question and answer session for final year Undergrad Pharmacy students |
| Form Of Engagement Activity | A talk or presentation |
| Part Of Official Scheme? | No |
| Geographic Reach | Local |
| Primary Audience | Undergraduate students |
| Results and Impact | I have participated in a 40 minutes question and answer session for final year undergrad pharmacy students who were preparing to conduct their final year research project in their chosen lab. Together with three other PhD students, we inform the students about the rules applying their final year project. Besides the rules, we gave the students some tips how they could achieve better scores by showing interest and initiative about their own projects and being curious about what other people doing in the lab as well as being prepared and organised and keeping good records about their lab activities. Finally, we inspire them with our personal reason for doing a PhD and a brief explanation of what our PhD projects are about. |
| Year(s) Of Engagement Activity | 2019 |