Non-Technical Summary
Project duration
5 years 0 months
Project purpose
- Basic research
- Translational or applied research with one of the following aims:
- Avoidance, prevention, diagnosis or treatment of disease, ill-health or abnormality, or their effects, in man, animals or plants
Key words
Cardiovascular Disease, Non-alcoholic steatohepatitis, Heart Failure, Biomarkers, Therapy
Animal types & Life stages
- Mice - adult, embryo, neonate, juvenile, pregnant
- Rats - adult, embryo, neonate, juvenile, pregnant
Retrospective assessment
The Secretary of State has determined that a retrospective assessment of this licence is not required.
Objectives and benefits
Description of the projects objectives, for example the scientific unknowns or clinical or scientific needs it's addressing.
What's the aim of this project?
The project has several objectives that fall under one or both of two over-arching aims:
Identification of novel biomarkers to determine the risk of cardiovascular disease or non-alcoholic steatohepatitis (NASH) in the setting of obesity/metabolic syndrome or micronutrient dysregulation.
Identification of novel drug targets, and assessment of novel pharmacological agents directed at them, for the prevention of cardiovascular disease.
Potential benefits likely to derive from the project, for example how science might be advanced or how humans, animals or the environment might benefit - these could be short-term benefits within the duration of the project or long-term benefits that accrue after the project has finished.
Why is it important to undertake this work?
Heart and circulatory disease (Cardiovascular Disease; CVD) manifests in various ways, including high blood pressure (hypertension) and narrowing of the arteries (arteriosclerosis), which can lead to heart attack (acute myocardial infarction; AMI), heart failure and stroke. Despite a global reduction in the incidence of CVD, death rates from CVD in the UK remain amongst the highest in Western Europe and is the UK's biggest killer; statistics for 2020 show that 7.4 million people in the UK are living with CVD, and around 27% (170,000 people) of all deaths in the UK result from CVD. Although there are therapies that can help to control blood pressure and the development of arteriosclerosis, there are few (if any) therapies that can reduce the tissue injury that results from AMI or stroke. Consequently, for those living CVD there is decreased physical fitness and increased morbidity.
Approximately 80% of individuals with CVD have at least one other health condition linked to the development (such as metabolic syndrome or hypertension) or as a consequence (e.g. heart failure) of CVD. Metabolic syndrome (MetS) refers to a cluster of metabolic disorders associated with visceral obesity, insulin resistance, dyslipidaemia, hyperglycaemia, and hypertension, predisposing individuals to increased risk of arteriosclerosis, CVD, Type 2 diabetes (T2D) and non-alcoholic steatohepatitis (NASH). MetS is an even greater global public health problem, affecting 25% of the world’s adult population, with a similar prevalence reported for the UK. As no single treatment to control MetS exists, novel therapeutic strategies are required to improve population health. Therefore collectively, there is a continued need to find novel ways of preventing (e.g. through dietary intervention), detecting risk of developing (e.g. through detection of biomarkers) and treating (through novel therapies) CVD and its associated risk factors.
What outputs do you think you will see at the end of this project?
This project will provide new information about the cellular events by which dietary risk factors such as obesity, metabolic syndrome and micronutrient deficiency lead to the development of coronary heart disease, heart failure and fatty liver disease (comorbidities).This information will give an indication of where an intervention (for example a drug or nutritional supplement) targeting these events could prevent the development or halt the progression of these co-morbidities. Furthermore, it will identify novel biomarkers that can be used to either predict the likelihood of developing a co-morbidity or to determine the effectiveness of treatment. This information will be shared with the wider scientific community through conference presentations and published as scientific articles in peer-reviewed, high quality, open-access journals. Identification of drug targets, and novel compounds acting at them, or of novel biomarkers, may lead to the generation of intellectual property that could be licensed to the pharmaceutical industry for commercial development. In addition, identification of new dietary interventions to promote health and wellbeing could lead to the production of new nutritional supplements or nutraceuticals.
Who or what will benefit from these outputs, and how?
The beneficiaries of this information will be:
Researchers within the field of cardiovascular and metabolic science will be informed of ongoing work through conference presentations(2-3 per year) and eventual publication of the final data arising from different studies within the overall programme (within 6-9 months of completion). All scientific outputs will be in green open-access journals, and will be uploaded into the Institutional Repository in accordance with the University Data sharing policy and any journal- imposed restrictions on access.
The pharmaceutical industry undertaking R&D programmes to develop novel ligands targeting the receptor systems under study; where intellectual property with potential for commercialisation is generated, we will work with the Establishment IP officer to ensure that this is protected (up to 1 year). We will then explore the opportunities for licensing out this IP through discussions with companies working in the same field (~2-3 years from completion of the project).
The clinical community since, although there are limitations to attempting to translate experimental findings into the clinic, the new information could inform hypotheses for subsequent collaborative controlled clinical studies, for example validation of a biomarker to demonstrate the stage of disease progression; we already work with clinical colleagues to acquire patient blood samples from specific patient groups alongside some of our in vivo experimental studies.
The general public, who will be kept abreast of developments through press releases (where appropriate) on relevant research outcomes timed to coincide with the publication of a full article. We also encourage our PhD students to participate in public engagement activities.
How will you look to maximise the outputs of this work?
To avoid unnecessary replication of data by other groups, where appropriate, raw data arising from the studies will be made available to the wider scientific community in accordance with the Establishment's Data sharing policy and provided as supplementary information for published articles.
We believe that negative data is as important as positive data, and should be published to inform the scientific community. Many peer-reviewed journals now agree with this philosophy, and publish negative studies as long as the underlying science is robust. Therefore, in the event that we fail to generate data to support our original hypothesis we will publish these findings to prevent other scientists from travelling down the wrong route.
Collaboration is key to the success of our research programmes, and we already collaborate widely with colleagues in academia both nationally and internationally (e.g. Australia, France, Italy). We also collaborate closely with industries involved in pharmaceutical R&D and those involved in isolation/production of health-promoting bioactives from by-products of raw material processing, such as marine algae and agricultural crops.
Species and numbers of animals expected to be used
- Mice: 1500
- Rats: 250
Predicted harms
Typical procedures done to animals, for example injections or surgical procedures, including duration of the experiment and number of procedures.
Explain why you are using these types of animals and your choice of life stages.
Rats and mice are well-established models to study the impact of diets aimed at inducing obesity and metabolic syndrome, and also to determine the impact of micronutrient deficiencies, on cardiovascular physiology and tissue structure and function. Since diet-related problems in humans arise as adults, the most appropriate life stage to mimic these events in animals is at the adult stage. Genetically modified animals that express alterations in genes that code for proteins linked to energy metabolism are important models to use to be able to understand the importance of these proteins in the development of diet-induced cardiac and liver disorders. Although breeding of these animals involves animals at all life stages, but only adult animals will be used for any experimental procedures.
Typically, what will be done to an animal used in your project?
A typical experiment will involve feeding wild type and GM mice a high cholesterol diet for 12 weeks, with weekly measurement of body weight and non-invasive assessment of body composition (lean vs fat content) at the start and end of the diet period. When testing for the effects of a drug intervention, this would be given in the drinking water for the last 2-4 weeks of dietary intervention. 2-3 days before the end of diet intervention, animals will be tested for their glucose status; this involves withdrawal of food (for up to 8 hours) prior to an injection of either glucose or insulin. After the injection blood samples are taken approximately every 10 minutes, over a 1-hour period, via tail prick and blood glucose measured using glucometer strips. At the end of the diet intervention animals are either humanely killed using an authorised method of euthanasia, or terminally anaesthetised for measurement of cardiovascular variables prior to euthanasia.
What are the expected impacts and/or adverse effects for the animals during your project?
The animals will not experience adverse effects that are more than mild and transient.
Expected severity categories and the proportion of animals in each category, per species.
What are the expected severities and the proportion of animals in each category (per animal type)?
With the exception of surgical pump implantation, all of the procedures are of mild severity, that is they are of low impact, non-painful, have no lasting effect and exhibit a very rapid return to normal. Around 5% of animals may undergo pump implantation, which is of moderate severity as they undergo surgery under general anaesthesia. However, measures such as pre- and post-operative pain relief reduce the impact of surgery and the animals return to normal within one or two days.
What will happen to animals at the end of this project?
- Killed
Replacement
State what non-animal alternatives are available in this field, which alternatives you have considered and why they cannot be used for this purpose.
Why do you need to use animals to achieve the aim of your project?
Cardiovascular and liver disease arises from metabolic syndrome or dietary deficiency as a result of complex interactions between tissues and organs within the body. While we can study some of the direct influences of dietary modification in isolated cells, tissues or organs, to gain a full understanding of how changes in one tissue (e.g. fat tissue) influence the normal functioning of another organ (e.g. the heart or liver) means that this can only be assessed in a whole animal.
Which non-animal alternatives did you consider for use in this project?
We perform a broad range of experiments in cultured cells and tissues to understand the key changes, for example alterations in anti-oxidant status or release of biochemical markers, that we would expect to see in response to alterations in the conditions aimed at mimicking dietary insufficiency or simulating metabolic syndrome. Where suitable genetic strains of the roundworm, C. elegans, exist we perform experiments to assess certain events, such as fat accumulation in response to metabolic stimuli. Only once we have gathered sufficient information to justify the need for in vivo studies will we use animals.
Why were they not suitable?
Cell-base or non-mammalian in vivo models (i.e. C. elegans) can provide a wealth of information. However, because they do not allow the study of how different organs/tissues in the body respond collectively to changes in diet and metabolism (cells) or do not have the same physiology as mammals (including humans) they cannot provide the full picture.
Reduction
Explain how the numbers of animals for this project were determined. Describe steps that have been taken to reduce animal numbers, and principles used to design studies. Describe practices that are used throughout the project to minimise numbers consistent with scientific objectives, if any. These may include e.g. pilot studies, computer modelling, sharing of tissue and reuse.
How have you estimated the numbers of animals you will use?
We have many years of experience of undertaking dietary intervention studies and so we can use our previous data to determine how many animals will be required to obtain scientifically sound data upon which we can form solid conclusions; this will take account of the inherent biological variability for the desired end points that will be measured. In addition, we can gain further insight with regards to experimental group sizes from the wider scientific literature.
What steps did you take during the experimental design phase to reduce the number of animals being used in this project?
In addition to using previous data to inform our decisions around the number of animals to use in any particular study, we also make use of online tools, such as the NC3R's Experimental Design Assistant, and follow the ARRIVE guidelines on designing and reporting experiments using animals. We also seek advice from the Establishment statistician when required.
What measures, apart from good experimental design, will you use to optimise the number of animals you plan to use in your project?
For studies using GM animals we only breed the numbers required for a particular study; where possible we utilise both male and female animals unless there is a clear scientific reason to only perform studies in one sex. When we perform studies that involve drug intervention we perform pilot studies in a small number of animals to determine the most appropriate dose of drug to use for the larger study, thus minimising the need to study multiple doses, which would need more experimental groups.
All of our studies are designed to ensure that we obtain as much information as possible from each animal; this includes animal characteristics (phenotype), changes in physiology (such as blood pressure), alterations in blood biochemistry (such as blood cholesterol) and tissue structures. Where there may be tissues/organs that are not required for a particular study (e.g. blood vessels) we try to use these for isolated tissue experiments that will provide pilot data for future studies. Any other tissues are retained in a biobank, details of which will be available in the Institutional repository, in the event that they may be of use at some point in the future by ourselves of other academic groups.
Refinement
Give examples of the specific measures (e.g., increased monitoring, postoperative care, pain management, training of animals) to be taken, in relation to the procedures, to minimise welfare costs (harms) to the animals. Describe the mechanisms in place to take up emerging refinement techniques during the lifetime of the project.
Which animal models and methods will you use during this project? Explain why these models and methods cause the least pain, suffering, distress, or lasting harm to the animals.
Some animals will be given novel drugs for a period of time; in the majority of cases this will be achieved by adding the drugs to their food or drinking water, but in some cases (<5% of animals) where this is not possible (for example palatability problems that cannot be overcome by the use of treats) we may need to administer drugs by a small pump implanted under the skin. This procedure results in transient moderate discomfort that is managed by the use of appropriate pain relief. We will also determine the glucose status of some animals, by giving an injection of either insulin or glucose followed by blood sampling; the technique used for blood sampling from the tail is similar to that used by individuals with diabetes who measure their own blood glucose by finger prick.
Why can’t you use animals that are less sentient?
At the end of the diet period any further investigations are performed under terminal anaesthesia (i.e. the animals do not regain consciousness). For some studies, where relevant genetic strains exist, we employ the C. elegans roundworm to perform in vivo studies, but although they have a close genetic profile to humans, their physiology is very different from mammals.
How will you refine the procedures you're using to minimise the welfare costs (harms) for the animals?
We have already incorporated as many refinements as possible to minimise the mild impact of the dietary interventions in our animals; animals are group housed and provided with environmental enrichment. Any animals undergoing surgical implantation of pumps for drug delivery are given peri-operative pain relief. For animals that are tested for fasting glucose status we minimise the period of time required for fasting and use a method of blood sampling that causes only mild and very transient discomfort.
What published best practice guidance will you follow to ensure experiments are conducted in the most refined way?
We refer to the NC3R's guidance on experimental design and follow their best practice approaches for refining our experimental methods.
How will you stay informed about advances in the 3Rs, and implement these advances effectively, during the project?
The Project Licence holder attends local meetings with other Project Licensees to learn and present about 3R approaches in different research groups, and the Establishment senior animal technician is part of a regional group that meets to discuss refinements in animal husbandry etc; both of these networks are a valuable source of information and inform us at to what improvements to make within our own establishment. The Establishment also has a NC3Rs/ARRIVE committee that meets 2-3 times per annum to discuss recent advances in the 3R's and agree on the implementation of changes; this committee also scrutinises external grant proposals for projects involving animals to ensure that the 3R's and ARRIVE guidelines are adhered to in the experimental design.