Moving up a dimension: 3D in vitro models as effective alternatives to live fish studies

Lead Research Organisation: University of Plymouth
Department Name: Sch of Biological and Marine Sciences

Abstract

Whilst traditionally fundamental life processes are studied at whole organism level (i.e. 'in vivo'), in recent years for economic, ethical and legal reasons, there has been much emphasis on the use of cells, tissues or organs which are grown outside the body in plastic dishes or flasks, the so called 'in vitro' system. In this context, in recent decades due to population growth, industrialisation and increasing consumer demands, there has been an increase in the environmental stress or contaminants on the living systems. Many of these contaminants resulting from consumers or industrial activities end-up in the aquatic environment. Exposures to such stresses lead to an increase in many diseases and can impact wild species. In order to assess the potential detrimental impact of such contaminants, international law requires chemicals or consumer products to be tested on animals. While advances have been made to reduce the number of animals used in biomedical research, the use of fish is still required for assessing the potential toxic effects of contaminants. The use of cell or tissue-based models has been proposed for fish-based studies, with the benefit of requiring only a few donor animals. It is however not clear how well these models could predict the responses that are seen when using whole animals. There is therefore reluctance to accept these as a real alternative. The conventional suspension or single layer in vitro culture models also suffer from drawbacks, (e.g. they live for a short period in the laboratory, and some functions just do not work outside of the animal) making them unsuitable for long term studies.

Based on widely used mammalian models, including by our own team, we have developed a specialised culture from liver cells of fish that 'mimic' the responses of tissues in whole fish. These balls like structures called 'spheroids' live ten times longer than the existing single layer (monolayer) cells. We have demonstrated that they are able to take-up and metabolise chemicals in a way that is directly comparable to whole fish, suggesting that they could be used as a real alternative to whole animal usage. As these spheroid cells are prepared from only a handful of animals, as opposed to hundreds of animals typically needed to study biomedical procedures and to assess toxicity. Their small sizes mean that less material and smaller facilitates are required, making them less expensive to conduct research. Since billions of cells from several different organs can be harvested from a single fish means that far fewer fish will be used in research, and those that are will not be used directly in experiments, only their tissues harvested after sacrifice.

We aim to progress this model, beyond-the state-of art technologies to test the hypothesis that a combination of co-cultures (i.e. different cell types grown together) containing different cell or tissue types of fish (viz. gill, gut and liver) represents an effective alternative model to live fish studies, thus creating a 'virtual fish' alternative. This requires establishment of key biological responses at cellular or molecular levels called 'biomarkers', which are already well established in diagnosis, prognosis and treatments of a range of human diseases. Using these biomarkers, we aim to elucidate the mechanisms and physiological functions by which fish respond to such stresses including following chronic exposures to environmentally relevant chemicals. We then aim to make robust comparisons to assess whether the model is sensitive enough to be considered as an appropriate replacement to whole fish studies.

The proposal also aims to build strong links between industry, academia and stake holders, both nationally and internationally to share the knowledge base as well as to potentially gain wider regulatory approval as 'real' alternatives to animal tests to continue our commitment to supporting the 3R's (Reduction, Refinement & Replacement) initiative.

Technical Summary

The proposal aims to probe the primary hypothesis that a co-culture of 3D in vitro cultures (viz. liver-spheroids and gill / gut epithelia) from rainbow trout (Oncorhynchus mykiss) can represent an effective alternative model to live fish studies. Upon establishing standard conditions for the growth and maintenance of co-cultures, we aim to:

1.Fully characterise the system and further define the culture conditions to ensure basic functionality is maintained and viability is prolonged.

2.Establish baseline values and compare key proteomic and recognised molecular biomarkers in in vitro preparations and compare the values with available in vivo data.

3.Conduct a limited set of in vivo studies to provide material directly relevant to and from the same genetic stock as the cells used in vitro in order to confirm the comparability from in vivo to in vitro.

4.Determine the functionality of spheroids alone or as co-cultures with respect to basic bio-transformation pathways and metabolite formation following exposures to different chemical agents, (e.g. reference genotoxic chemicals: polycyclic aromatic hydrocarbons; environmentally relevant pharmaceuticals: non-steroidal anti-inflammatory, beta-blocker and selective serotonin re-uptake inhibitors; and environmentally relevant metals: copper).

5.Compare the comparative physiology (oxygen uptake, bio-energetics, metabolism and excretion rates) of the systems in response to 'normal' control conditions and exposure to environmentally relevant chemicals.

6.Evaluate 'mixture' toxicity from a 'toxicogenomic' perspective in both isolated tissue preparations and co-cultures.

7.Measure the bioconcentration of selected chemicals in all tissue types following chronic exposure to test chemicals using cutting edge analytical techniques.

8.Communicate the methodologies and findings to key industrial, academic and stakeholder groups.

Planned Impact

The outcome of this interdisciplinary proposal will make a significant difference to diverse groups of people in the wider scientific community in a variety of ways:

In terms of the applied science, one particular example that has great relevance to industry is the advent of the European chemical registration and approval (REACH ) that has lead to the pre-registration of >140,000 compounds, with >30,000 of these requiring significant in vivo testing. A large proportion of these will require the assessment of the bioaccumulation potential of these compounds via standard regulatory OECD tests such as the fish bioaccumulation test 305. In 2011 (according to the UK Government) 58,908 fish were reported as used in toxicology studies in the UK alone. Testing in other European countries, have similar rates. The majority of these fish are likely to be rainbow trout.

The requirement of extensive facilities, large amounts of test compound, often radio-labelled and significant numbers of fish clearly demonstrates the ethical and financial reasons that are preventing the rapid assessment of these compounds, which pose an unknown risk to the environment until they can be effectively tested. This information is essential for Environmental Risk Assessment and therefore the need for reliable, alternative testing methods are essential if these thousands of compounds are to be assessed without requiring millions of fish. As this field grows, and we help other groups to work with our model, we believe the studies will be fundamental and seminal work that will receive much attention and through open access publication and freely available protocols that will deliver well cited influential work.

We envisage that the developed model will serve as a spring board to enable many groups to conduct effective, small scale, low cost, high content, and higher throughput alternatives to study biological processes at whole organism level under in vitro conditions. Our model will offer a new, novel research tool for answering key scientific questions regarding an animal's physiology, that may have been previously too difficult to carry out with whole fish. The fact that these micro-tissues reform the organs outside of the fish means that researchers can see how cells communicate and interact using microscopic and molecular tools. This model could serve as spring board to stimulate design and developments of new investigations in the uptake and elimination of nutrients; cellular transport; immunity; disease progression; organ culture; signalling; cell biology such as cycling, division, replication, apoptosis and protein expression.

This project will provide unique understanding of the fundamental mechanisms, biochemical characteristics and extent of functionality of fish gill, gut and liver tissues both in life and in culture. These data will be of value to a range of fish biologists, and we anticipate that detailed information and understanding of this level of detail will quickly enter the teaching curricular of both fish and general biology.

The fully developed and validated model will be promoted for potential regulatory acceptance as true alternative to live fish bioaccumulation tests. Once accepted this would be necessarily widely taken up by many commercial laboratories in order to meet European and indeed worldwide legislation.

The research team is extensive expertise in disseminating research findings to School children, general public through outreach activities and popular articles in local media and in BBSRC and NERC journals. The PI has also experience in being invited by local BBC (radio and TV) and BBC radio five live as expert commentator. This would enhance not only the public understanding of the BBSRC funded science but will also encourage younger generation for a scientific career.
 
Title Virtual Fish 
Description Poster for Public Engagement Event 
Type Of Art Artwork 
Year Produced 2014 
Impact Public understanding of use of animals in research. 
 
Description • Developed and improved the use of a rainbow trout gill model to assess the uptake of environmental contaminants in trout.
• Developed a rainbow trout liver model to assess the metabolism of the compounds.
• Optimising spheroid models from different parts of the gut cells of the fish.
• Experiments are ongoing where we aim to combine these two models into the same experimental unit (i.e. liver and gill; liver and gut). This will allow assessment of both the uptake and metabolism of compounds in tissue specific manner at the same time.
• Using this combined model instead of using live fish will enable us to answer questions about the environmental safety of the contaminants without using the large numbers of fish currently required by regulations.Updated since last (2017) submission on 9.3.18: • Used a rainbow trout liver model to assess the accumulation and metabolism of a range of pharmaceuticals, demonstrating the benefits of in vitro alternatives to animal testing
• Improved a cell culture model of the rainbow trout gill so that it remains viable for a longer duration, and therefore allows improved testing of the toxicity of environmental contaminants.
• Optimised the culture of cells from four regions of the rainbow trout gut and characterised the functionality of the subsequent cultures.
• Developed a procedure to co-culture cells from two different rainbow trout organs (gill and liver) into a single experimental unit.
• Using the described models instead of using live fish will enable us to answer questions about the environmental safety of contaminants without using the large numbers of fish currently required by regulations. Further information accessible here: •Project leaflet summarising work
https://www.plymouth.ac.uk/uploads/production/document/path/9/9207/Virtual_Fish_leaflet_18_April_2017.pdf

• Symposium timetable (held during SETAC Europe meeting)
https://www.plymouth.ac.uk/uploads/production/document/path/9/9206/SETAC_Workshop_on_Fish_In_Vitro_Toxicology__Draft_Programme_18_April_17.pdf

• Conference Presentations Websites:
https://orlando.setac.org/wp-content/uploads/2016/10/SETAC-meeting-program-web-v4.pdf
http://www.sebiology.org/docs/default-source/Event-documents/seb_brighton_programme_web.pdf?sfvrsn=2

We have further progressed the work co-culturing the liver spheroids and gill organoids of fish to validate the 'vitual fish' model.
Exploitation Route Updated from last submission 2017 on 9.3.18: Our findings provide the foundation for future work developing in vitro techniques to reduce, refine and replace in vivo ecotoxicological testing. It is an emerging area of research and our findings justify further investment. There is scope to use the gill models in, for example longer term studies investigating chronic exposure of contaminants that has not previously been possible. Similar developments of novel gut cell cultures opens up the possibility of studying fish dietary uptake dynamics in vitro. The same cultures can also be used to tackle new questions in physiology and environmental monitoring. The continued development of protocols for tissue co-cultures will also enhance the next generation of in vitro models to improve future environmental risk assessments while reducing the number of live fish used.
We have collaborated with colleagues at Kings College London who specialised in gill cultures, shared our liver techniques that have been used in preliminary co-culture experiments and delivered a chapter in a successfully defended PhD thesis at Kings. We have collaborated with colleagues at Exeter University to quantify metabolism in spheroid models which is currently awaiting peer review, and colleagues at the Earlham institute to use RNA-seq to characterise genes present in the native gill, liver and intestinal tissue which will allow for quantification of differences between in vitro and in vivo predictions. Finally, we have successfully established for the first time a co-culture of gill and liver organoids and characterised response following exposure to pyrene, a ubiquitous environmental pollutant. These manuscripts are currently in preparation or in the process of submission for peer review.
We have collaborated with colleagues at Kings College London who specialised in gill cultures, shared our liver techniques that have been used in preliminary co-culture experiments and delivered a chapter in a successfully defended PhD thesis at Kings. We hope to see the manuscript submitted this year. Our findings provide the foundation for future work developing in vitro techniques to reduce, refine and replace in vivo ecotoxicological testing. It is an emerging area of research and our findings justify further investment. There is scope to use the gill models in, for example longer term studies investigating chronic exposure of contaminants that has not previously been possible. Similar developments of novel gut cell cultures opens up the possibility of studying fish dietary uptake dynamics in vitro. The same cultures can also be used to tackle new questions in physiology and environmental monitoring. The continued development of protocols for tissue co-cultures will also enhance the next generation of in vitro models to improve future environmental risk assessments while reducing the number of live fish used.
We have collaborated with colleagues at Kings College London who specialised in gill cultures, shared our liver techniques that have been used in preliminary co-culture experiments and delivered a chapter in a successfully defended PhD thesis at Kings. We have collaborated with colleagues at Exeter University to quantify metabolism in spheroid models which is currently awaiting peer review, and colleagues at the Earlham institute to use RNA-seq to characterise genes present in the native gill, liver and intestinal tissue which will allow for quantification of differences between in vitro and in vivo predictions. Finally, we have successfully established for the first time a co-culture of gill and liver organoids and characterised response following exposure to pyrene, a ubiquitous environmental pollutant. These manuscripts are currently in preparation or in the process of submission for peer review.

Further details of findings have been covered by the media, links are given below:

• Non-destructive technique measures oxygen levels in 3D cells used for toxicity testing. Plymouth University Press Release; 22 Feb 2016
https://www.plymouth.ac.uk/news/non-destructive-technique-measures-oxygen-levels-in-3d-cells-used-for-toxicity-testing
• Non-destructive technique measures oxygen levels in 3D cells used for toxicity testing Eurekalert, AAAS, 22 FEB 2016
http://www.eurekalert.org/pub_releases/2016-02/uop-ntm022216.php
• Direct Measurements of Oxygen Gradients in Spheroid Culture System Using Electron Parametric Resonance Oximetry; Paperity, Feb 2016
http://paperity.org/p/75344410/direct-measurements-of-oxygen-gradients-in-spheroid-culture-system-using-electron
• Innovative Method Measures O2 levels in 3D Cells Used for Tox Testing; Genetic Engineering and Biotechnology News (GEN); Feb 2016
http://www.genengnews.com/gen-news-highlights/innovative-method-measures-o2-levels-in-3-d-cells-used-for-tox-testing/81252396/

• Technique measures oxygen levels in 3D cells used for toxicity testing; Plymouth Daily, 24 Feb 2016
http://www.theplymouthdaily.co.uk/news/local-news/technique-measures-oxygen-levels-3d-cells-used-toxicity-testing
• Non-destructive technique measures oxygen levels in 3-D cells used for toxicity testing; Feb 2016
https://www.sciencedaily.com/releases/2016/02/160222151630.htm?utm_source=feedburner&utm_medium=feed&utm_campaign=Feed%3A+sciencedaily%2Fplants_animals%2Fbiology+%28Biology+News+--ScienceDaily%29&utm_content=Google+UK
Sectors Chemicals,Environment,Government, Democracy and Justice,Pharmaceuticals and Medical Biotechnology,Other

URL https://www.plymouth.ac.uk/research/genetic-ecotoxicology/virtual-fish
 
Description Updated since last submission 2017, 9.3.18: AstraZeneca (the industrial partner) have been closely involved in this project. Their interest was principally to see the development of biologically relevant in vitro models that could be used in decision cascades and part of weight-of-evidence arguments to direct and justify appropriate in vivo tests. To this end, the work has indeed developed biologically relevant models. The gut model is the first to facilitate the potential to understand xenobiotic dietary uptake in vitro. It is harvested from the appropriate part of the donor animal and provides extensive replication and longevity measured in weeks that was significantly better than the anticipated model at the outset. As with the gill model, the intestine model is able to withstand water exposure making it a potential tool for environmental monitoring. The spheroids are proven robust and reliable and show metabolic capacity that appears similar to in vivo. The development of the gill model to withstand water exposure for several weeks is a significant and remarkable step forward. The toolbox offered by this project underpins a significant European Innovative Medicines Initiative and the data generated used to parameterise examples in that project. Through the collaboration with AstraZeneca, the project has therefore contributed to the Innovative Medicines Initiative (IMI) under grant agreement no.115735-iPiE: Intelligent led assessment of Pharmaceuticals in the Environment; resources of which are composed of financial contribution from the European Union's Seventh Framework Programme (FP7/2015-2018) and European Federation of Pharmaceutical Industries and Associations (EFPIA) companies' in kind contribution. This is a €10m project to help prioritise the environmental risks posed by legacy human pharmaceuticals. In vitro methods are essential to understand uptake and metabolism of these compounds in fish. The methods developed here will be used by a range of pharmaceutical companies in 2018 as part of this IMI project. Companies include both UK pharma (AstraZeneca and GSK) and 10 other global players. Therefore, this project has contributed to the overall reduction in future animal studies, saved significant financial costs and will help deliver a better understanding of the environmental risk of pharmaceuticals. AstraZeneca (the industrial partner) have been closely involved in this project. Their interest was principally to see the development of biologically relevant in vitro models that could be used in decision cascades and part of weight-of-evidence arguments to direct and justify appropriate in vivo tests. To this end, the work has indeed developed biologically relevant models. The gut model is the first to facilitate the potential to understand xenobiotic uptake in vitro. It is harvested from the appropriate part of the donor animal and provides extensive replication and longevity measured in weeks that was significantly better than the anticipated model at the outset. The spheroids are proven robust and reliable and show metabolic capacity that appears similar to in vivo. The development of the gill model to withstand water exposure for several weeks is a significant and remarkable step forward. The toolbox offered by this project underpins a significant European Innovative Medicines Initiative and the data generated used to parameterise examples in that project. Through the collaboration with AstraZeneca, the project has therefore contributed to the Innovative Medicines Initiative (IMI) under grant agreement no.115735-iPiE: Intelligent led assessment of Pharmaceuticals in the Environment; resources of which are composed of financial contribution from the European Union's Seventh Framework Programme (FP7/2015-2018) and European Federation of Pharmaceutical Industries and Associations (EFPIA) companies' in kind contribution. This is a €10m project to help prioritise the environmental risks posed by legacy human pharmaceuticals. In vitro methods are essential to understand uptake and metabolism of these compounds in fish. The methods developed here will be used by a range of pharmaceutical companies in 2018 as part of this IMI project. Companies include both UK pharma (AstraZeneca and GSK) and 10 other global players. Therefore, this project has contributed to the overall reduction in future animal studies, saved significant financial costs and will help deliver a better understanding of the environmental risk of pharmaceuticals.• The work has been presented in public meetings and international conferences which was attended by different stakeholders (e.g. regulators, NGOs, Govt. organisations and other industries). AstraZeneca (the industrial partner) have been closely involved in this project. Their interest was principally to see the development of biologically relevant in vitro models that could be used in decision cascades and part of weight-of-evidence arguments to direct and justify appropriate in vivo tests. To this end, the work has indeed developed biologically relevant models. The gut model is the first to facilitate the potential to understand xenobiotic uptake in vitro. It is harvested from the appropriate part of the donor animal and provides extensive replication and longevity measured in weeks that was significantly better than the anticipated model at the outset. The spheroids are proven robust and reliable and show metabolic capacity that appears similar to in vivo. The development of the gill model to withstand water exposure for several weeks is a significant and remarkable step forward. The toolbox offered by this project underpins a significant European Innovative Medicines Initiative and the data generated used to parameterise examples in that project. Through the collaboration with AstraZeneca, the project has therefore contributed to the Innovative Medicines Initiative (IMI) under grant agreement no.115735-iPiE: Intelligent led assessment of Pharmaceuticals in the Environment; resources of which are composed of financial contribution from the European Union's Seventh Framework Programme (FP7/2015-2018) and European Federation of Pharmaceutical Industries and Associations (EFPIA) companies' in kind contribution. This is a €10m project to help prioritise the environmental risks posed by legacy human pharmaceuticals. In vitro methods are essential to understand uptake and metabolism of these compounds in fish. The methods developed here will be used by a range of pharmaceutical companies in 2018 as part of this IMI project. Companies include both UK pharma (AstraZeneca and GSK) and 10 other global players. Therefore, this project has contributed to the overall reduction in future animal studies, saved significant financial costs and will help deliver a better understanding of the environmental risk of pharmaceuticals.
First Year Of Impact 2016
Sector Chemicals,Education,Energy,Environment,Healthcare,Manufacturing, including Industrial Biotechology,Pharmaceuticals and Medical Biotechnology
Impact Types Cultural,Societal,Economic

 
Description NC3Rs sponsored Session at Plymouth
Geographic Reach Multiple continents/international 
Policy Influence Type Influenced training of practitioners or researchers
Impact we aim to present our work in the forthcoming 38th Annual Conference of United Kingdom Environmental Mutagen Society (UKEMS) to be organised at Plymouth University for 12-15 July, 2015. A session sponsored by NC3Rs on 'Alternative approaches to genotoxicity and ecotoxicity' is planned for this conference and we aim to show case research work to be carried out in this conference for the benefit of wider scientific community.
 
Title Further characterisation and validation of 'Virtual Fish' model using different tissues 
Description Further characterisation of double-seeded, permeable insert intestinal model: • Morphological assessment • Trans-epithelial resistance (TER) measurements • EROD activity after exposure to positive control chemical (ß-napthoflavone) • Uptake and distribution of copper and nano-silver in different spheroid models 
Type Of Material Database/Collection of data 
Year Produced 2014 
Provided To Others? Yes  
Impact Once develop the model will reduce the number of fish being used in toxicological studies. This will also provide mechanistic understanding of environmental contaminants on fish cells. 
 
Description EURASMUS Student exchange with University of Ancona, Italy and University of Nottingham, UK 
Organisation Marche Polytechnic University
Country Italy 
Sector Academic/University 
PI Contribution We established the collaboration in order to validate the developed in vitro models collaboration where established with Marche Polytechnic University and the University of Nottingham, Nanotechnology Centre, Department of Chemistry. We carried out the experimental work, at Plymouth University, under our laboratory conditions.
Collaborator Contribution The researcher from Marche Polytechnic University brought some complimentary techniques to add to the project. The University of Nottingham provided well characterised manufactured nano particles. The developed spheriod methods where exposed to these manufactured nano particles. The University of Nottingham made further characterisations of nano particles uptake and interaction in the spheriod systems.
Impact Title: Uptake, distribution and toxicological effects of two different silver nanomaterials in 3-D in vitro fish cultures Published Date: November 2015 Primary Author: Vicari T Secondary Authors: Baron M; Maunder R; La Torre A; Rance G; Khlobystov A; Jha A Publication Type: Conference Proceeding Abstract Is Published: Yes Journal: Mutagenesis (2015) 30, 875 (Abstract)
Start Year 2014
 
Description EURASMUS Student exchange with University of Ancona, Italy and University of Nottingham, UK 
Organisation University of Nottingham
Country United Kingdom 
Sector Academic/University 
PI Contribution We established the collaboration in order to validate the developed in vitro models collaboration where established with Marche Polytechnic University and the University of Nottingham, Nanotechnology Centre, Department of Chemistry. We carried out the experimental work, at Plymouth University, under our laboratory conditions.
Collaborator Contribution The researcher from Marche Polytechnic University brought some complimentary techniques to add to the project. The University of Nottingham provided well characterised manufactured nano particles. The developed spheriod methods where exposed to these manufactured nano particles. The University of Nottingham made further characterisations of nano particles uptake and interaction in the spheriod systems.
Impact Title: Uptake, distribution and toxicological effects of two different silver nanomaterials in 3-D in vitro fish cultures Published Date: November 2015 Primary Author: Vicari T Secondary Authors: Baron M; Maunder R; La Torre A; Rance G; Khlobystov A; Jha A Publication Type: Conference Proceeding Abstract Is Published: Yes Journal: Mutagenesis (2015) 30, 875 (Abstract)
Start Year 2014
 
Description 38th Annual Conference of UK Environmental Mutagen Society (UKEMS), July 2015, Plymouth 
Form Of Engagement Activity A formal working group, expert panel or dialogue
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Policymakers/politicians
Results and Impact Result Description The 38th Annual Conference of United Kingdom Environmental Mutagen Society (UKEMS) was hosted at Plymouth University with a dedicated session on 'Next steps towards the applications of alternative approaches to animal testing' , which was sponsored by NC3Rs UK. Posters and platform presentations were made from the members of the project team (including PDRF, Dr Baron) which stimulated discussion to move the science forward. These presentations have been published as abstracts in the journal Mutagenesis, Vol. 30, 2015. (http://mutage.oxfordjournals.org/content/30/6/851.full.pdf+html)

Most important impact? Increase in requests about (further) participation or involvement. Further suggestions to improve the quality of the work.
Year(s) Of Engagement Activity 2015
URL http://www.plymouthherald.co.uk/Plymouth-University-host-conference-damaging/story-26886749-detail/s...
 
Description Developing realistic fish in vitro models to assess impact of contaminants in the aquatic environment 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact Communication, awareness and interest in alternative animal alternative models. Raising interest in how they compare and contrast to existing work in mammalian and human research. Raising awareness with the in vitro toxicology community of other predictive models, which could be used for environmental regulatory submissions for general chemical (REACH), biocide and pharmaceutical registration. Discussions included how comparable and predictive the proposed models could be of in vivo response.
Year(s) Of Engagement Activity 2017
 
Description NC3Rs Workshop: Pathways-based approaches across the biosciences: Towards application in practice. 28 April 2016, London, UK. 
Form Of Engagement Activity A formal working group, expert panel or dialogue
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact Main purpose: Discussion of project applications to AOP monitoring and possible future collaborations in this field
Year(s) Of Engagement Activity 2016
URL https://www.nc3rs.org.uk/events/pathways-based-approaches-across-biosciences-towards-application-pra...
 
Description Organised a dedicated workshop on Current trends in vitro toxicology: Applications of 3Rs principles 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact Discussion with wider research community about the current progress in using in vitro models in toxicological assessments. Communicate work to wider audience with new ideas and alternative approaches and discuss future direction of work.
Wider knowledge of work within broader science community.
Increase in requests about (further) participation and collaborations to explore new avenues of research and applications.
Year(s) Of Engagement Activity 2017
 
Description Presentation at 27th SETAC Europe meeting, Brussels, Belgium 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact Discussion and progression of project ideas with key Ecotoxicological research community at international research conference. Communicate work to new audience and make new collaborations. Dissemination of project science to wider audience and feedback gained on new applications.
Year(s) Of Engagement Activity 2017
 
Description PubhD, Public Engagement platform explaining the theme of the project - 22nd November 2016, Plymouth, UK. 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Public/other audiences
Results and Impact Public Engagement platform: explaining the theme of the project with complementary PhD project to the public and graduate students (age 18-65) (co-sponsored by industrial partner AstraZeneca).
Main purpose: To share information and answer questions
Communicate work to wider public audience; explain purpose of work and potential benefits. Discuss societal worries and misconceptions about toxicity testing.

Most important impact: Increased understanding of animal testing in a toxicity context.
Year(s) Of Engagement Activity 2016
URL https://pubhd.wordpress.com
 
Description Regulatory Science Associates workshop - platform presentation of an overview of the project and the context of the developed tools. 
Form Of Engagement Activity A formal working group, expert panel or dialogue
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Industry/Business
Results and Impact Regulatory Science Associates workshop -
Endocrine Disruption: What Does the Future Hold for Safety Assessment? Workgroup 'Minimising the use of fish in tests'
January 31st - February 1st 2017 Cranage Hall, Cheshire, UK

Main purpose: Promote project work to regulatory audience and wider knowledge of work within relevant science community
Communication, awareness and interest in engaging appropriate studies to reduce in vivo animal exposure using a screening cascade as part of a weight of evidence in negotiation with regulatory authorities.

Most important impact: Raising awareness with the regulatory community including a wide range of contract consultants responsible for a large proportion of all environmental regulatory submissions for general chemical (REACH), biocide and pharmaceutical registration. Discussions included when to use these tools and the science underpinning the techniques
Year(s) Of Engagement Activity 2017
URL http://www.plymouthherald.co.uk/Plymouth-University-host-conference-damaging/story-26886749-detail/s...
 
Description SEB 2016 Animal Symposium 'Improving experimental approaches in animal biology: Implementing the 3Rs. 29th June - 1 July 2016, London, UK. 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact Scientific meeting platform and poster presentations
Main purpose: To share information and wider knowledge of work within relevant science community
Active contribution to sessions and discussions at international research conference. Communicate work to wider audience, forge links, collaborations and share ideas through discussion.

Most important impact: Increase in requests about (further) participation and collaborations to explore new avenues of research and applications.
Year(s) Of Engagement Activity 2016
 
Description SEB Annual Meeting, 3-7 July 2016, Brighton, UK 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact Scientific meeting poster presentation
Main purpose: To share information and Wider knowledge of work within broader science community
Discussion with wider research community at international research conference. Communicate work to wider audience with new ideas and alternative approaches.

Most important impact: Increase in requests about (further) participation and collaborations to explore new avenues of research and applications
Year(s) Of Engagement Activity 2016
URL http://www.sebiology.org/events/meetings_archive/SEB_Brighton_2016
 
Description SETAC World Congress meeting, 6 - 10th November 2016, Orlando, Florida, USA 
Form Of Engagement Activity A formal working group, expert panel or dialogue
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact Scientific meeting platform presentations (150 in audience, 2500 at meeting)
Main purpose To share information and knowledge exchange and enhancement of project ideas
Discussion and progression of project ideas with key Ecotoxicological research community at international research conference. Communicate work to new audience and make new collaborations

Most important impact: Dissemination of project science to wider audience and feedback gained on new applications.
Year(s) Of Engagement Activity 2016
URL https://orlando.setac.org
 
Description STEM Ambassador session at local school (Devonport High School for Boys, Plymouth). 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Schools
Results and Impact Year 8 curriculum enrichment day - 5 × 50 min lessons
Main purpose: To communicate project objectives and information
Communicate project and broader importance of environmental awareness to students. Inspire next generation in scientific work. Forge links to local community.

Most important impact: Inspiration and promotion of scientific study to students, including wider knowledge of work within local community and inspiration for future career choices.
Year(s) Of Engagement Activity 2016
 
Description Society of Environmental Toxicology and Chemistry, Barcelona, 3rd -7th May 2015 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Policymakers/politicians
Results and Impact How many people? 2000+

Result Description Active contribution to session and discussions at international research conference. Communicate work to wider audience; forge links, collaborations and share ideas through discussion.

Impact Description Wider knowledge of work within relevant science community

Most important impact? Increase in requests about (further) participation and collaborations to explore new avenues of research and applications
Year(s) Of Engagement Activity 2015
URL http://barcelona.setac.eu/?contentid=767
 
Description The AstraZeneca Global Environment Symposium, Alderley Park Conference Centre, Manchester, 15 -16th December, 2015 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Policymakers/politicians
Results and Impact Result Description Active contribution to session and discussions at industry symposia. Communicate work to wider audience; forge links, collaborations and share ideas through discussion.

Impact Description Wider knowledge of work within relevant UK science community

Most important impact? Increase in requests about (further) participation and collaborations to explore new avenues of research and applications
Year(s) Of Engagement Activity 2014,2015