Southampton Imaging: 3D imaging at millimetre to nanometre scales for regenerative medicine using multiple complimentary modalities

Lead Research Organisation: University of Southampton
Department Name: Faculty of Medicine

Abstract

Regenerative medicine aims to make tissues and organs to repair damaged and diseased tissues and restore the body to its original health. It uses stems cells and precursor cells that, under exactly the right conditions, will change into the specialized cells needed to repair the tissue. Scaffolds upon which the cells can grow and be guided are used to help organize the cells into the right structures. We are now in a unique position to create new soft and hard tissues (e.g. liver, neural, cartilage, bone) to help aid treatment for all. Important in this goal of moving to clinical application will be to ensure the new formed tissue is completely safe.

One of the most important factors that determines the effectiveness and normal functioning of a tissue is the way it is structured or arranged (the architecture of the tissue- like a bridge). This tissue architecture is important at many different scales, from individual cell components to the large scale organization of structures such as bones or blood vessels and nerves supplying the organs. Therefore to understand the appropriate structures required and check that the tissue constructs made are performing correctly we need to be able to 'see' these by using microscopic imaging at different magnifications. Because the cells and tissues are three-dimensional (3D) structures, we need to see how they fit together in 3D to understand their architecture - in the same way we can understand how the elements of a building fit together and function effectively in 3D by walking around it.

This application is for three imaging systems specially designed to create 3D images of tissues and scaffolds at 3 different scales. The highest magnification is provided by a scanning electron microscope combined with a microtome- this gradually removes very thin (<50nm) slices from the sample imaging the surface that is revealed after each slice. This creates a stack of images representing the structure of the cells down to the components making up the cells and the fine detail of scaffolds- with this technique the membranes within the cells can clearly be seen. The next level of magnification uses a light microscope that shines a very thin sheet of light (4-10 micro meters). This allows us to look at much larger blocks of tissue (up to a cm cube - a sugar cube) without cutting it and again create a 3D image stack to represent this. Individual cells are easily seen and they can be labelled so we can identify cell types and track them over time. However there are some samples that light will not penetrate or that are too large - for these samples the third instrument, a high resolution microCT (computed tomography) imaging device is used. This uses X-rays to image through large samples without damaging them and the design of this new instrument can allows us to distinguish individual cells and some of their features in a way that is not currently possible.

These systems will add to the wide range of existing imaging facilities in Southampton that are supported by 12 expert imaging staff. The existing expertise in sample preparation and biological image interpretation is essential for these cutting edge imaging techniques to be used effectively. An additional problem in 3D imaging at all scales is the very large digital image sets that are produced- each taking large amounts of storage; 50-1000Gb (equivalent of 10-200 DVDs each!). Southampton University is a world leader in advanced computing and image processing - we already have state of the art computing hardware and software. In this project we will work with our collaborators in the University to further develop these specifically for the processing and analysis of images of regenerative medicine samples.

We expect regenerative medicine to transform human health over the next 10-30 years and in order to fulfill this promise as quickly and safely as possible it is essential we can image the generated structures and tissues.

Technical Summary

Increased aging populations pose new challenges and emphasize the need for approaches to augment and repair tissue lost through aging, trauma or disease. Regenerative medicine promises to deliver specifiable replacement tissues. As living tissue is inherently 3D, comprehensive quantitation of cell interaction, differentiation and tissue architecture necessitates the use of 3D analytical platforms with appropriate resolution, structural differentiation and volumetric analysis capabilities, along with time resolved, ex vivo and in vivo capabilities. Southampton Imaging offers an open access resource to imaging these tissue constructs pre- and post-implantation, vital to understand and validate the tissue structure and function.

To deliver appropriate tissue resolution we request a Gatan 3View system to produce nanometer scale 3D image stacks by electron microscopy, able to visualise cell organelles and local cell-scaffold interactions. This will be complemented by a LaVision light sheet microscope, capable of imaging blocks of up to 1 cm cube substantially extending the depth and volume that can be imaged. This would for example include representative vascular networks that can be specifically labelled by immuno fluorescence or affinity methods. The platform allows non-destructive imaging of smaller live samples and critically reduces the light flux permitting long-term imaging of live tissues and cell migration in 3D tissues. Together with an Xradia ultra-high resolution microCT with phase contrast imaging, we will be able to image large volumes of tissue constructs including bio-compatible matrices of all types as well as hard and soft tissue elements. The requested equipment will integrate with extensive imaging infrastructure (including 12 dedicated supported staff), computational and image processing capacity (including UK's most powerful supercomputer) providing a unique Image resource for the Regenerative Medicine community.

Planned Impact

A multidisciplinary open-access 3D imaging service for regenerative medicine will substantially impact on the ability to provide and apply best cutting-edge practice in 3D imaging. We anticipate that project outputs will have significant bearing on the quality of life through improved regenerative medicine capability. By imaging the whole 3D structure of substantive tissue or implant volumes, it will enhance the development and implementation of effective biomaterial and implants. Quantitation of cell interactions, differentiation and tissue architecture necessitates the use of these 3D analytical platforms (X-ray, light and electron) with appropriate resolution including time resolved, ex vivo and in vivo analysis.

The extent and degree of detailed analysis possible with these technologies will also be improved, deepening understanding and reducing the time required to develop clinically effective tissue regenerative solutions. Critically, by building on extensive existing imaging facilities and expertise in the biological, imaging, engineering and computing domains the time from identifying a regenerative medicine imaging requirement to its application will be reduced. Shared experience and solutions in Southampton will allow data handling and analysis problems to be effectively provided, increasing the value of the data and reducing the time taken to appropriately analyse the images.

Our strategy is to share our data and the approaches developed and used for analysis with the wider scientific community to provide immediate access, enhancing collaboration and research.

We will also disseminate results at various stem cell, regenerative medicine, tissue engineering and microfluidics meetings. The applicants engage with the regenerative research community and have collectively presented keynote and plenary lectures at over 30 meetings around the world in the last 2 years. The enhanced research capability proposed holds significant translational therapeutic potential. As part of our established Excellence with Impact programme, we will build on links with Business Fellows to promote innovative collaborations such as that already sought by Xradia in anticipation of the project (see letter attached).

The research team will work as part of an active multidisciplinary team of biologists, tissue engineers, bio-engineers and, where appropriate, with clinicians and industry. This equipment grant will contribute substantially to the training and development of the Regenerative Medicine community through accredited postgraduate courses and the ability to train researchers at the interdisciplinary boundaries of regenerative medicine and imaging.

In order to maximize broader impact to the general public, we will organize a number of educational outreach programmes (Oreffo as part of the Acellular Hub will bid for outreach funds available in the hub) building on our links to a new school education programme, the LifeLab Southampton. This invites local school pupils, their parents and teachers to experience laboratory-based experiments learn how lifestyle affects lifelong health. We will continue our active involvement with BBSRC & EPSRC public outreach. All applicants have enjoyed significant media coverage for their work with presentations on Sky News, BBC TV stations, Radio Stations and the national and international press and science press (The Engineer, Materials Today and Nature Middle East amongst others). We will continue to actively engage with the press to showcase Southampton Imaging.

Publications

10 25 50
 
Company Name Renovos Biologics Limited 
Description Business Summary Renovos is a new company based in Southampton capitalising on decades of orthopaedic research. The company aims to be a leader in bone repair technologies. Business Description Renovos will offer: • products (stem cells for research) • services (low-cost, high-value in vivo screening assay) • therapeutics (patent-pending nanoclay preparations for drug delivery and tissue repair) 
Year Established 2017 
Impact Formed only 3 months ago- yet to reach potential
Website http://renovos.co.uk/
 
Description BBSRC GB Bioscience 2014 Festival 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? Yes
Geographic Reach International
Primary Audience Public/other audiences
Results and Impact Large science festival over four days - reaching politicians and policy makers on day 1, school children on day 2 and public on days 3 and 4

Event held November 13-16th - over 6000 attended our stand over four days form school children to aged individuals, research councils and various education stakeholders
Year(s) Of Engagement Activity 2014
URL http://www.bbsrc.ac.uk/society/exhibitions/gb-bioscience-festival/scaffolds-cells-making-replacement...
 
Description Biomedical Imaging Unit and µ-VIS joint one day meeting on 3D Imaging fro Life Sciences 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach Regional
Primary Audience Postgraduate students
Results and Impact Biomedical Imaging Unit and µ-VIS joint one day meeting - 3D imaging for Life Sciences
The meeting set out to showcase the range and diversity of imaging available for life sciences research at these two University of Southampton core facilities
Year(s) Of Engagement Activity 2018
 
Description Cheltenham 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 6-11 June - The Stem Cell featured at the festival and the team interacted with an estimated 1800 visitors - led to student interest in visiting our group.
Year(s) Of Engagement Activity 2017
 
Description Cheltenham Science Festival - Central Exhibit in Discover Zone 
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 The Stem Cell Mountain featured in 'Doc Brown's Lab' (in keeping with the 2015 Back to the Future Theme) at the famous Cheltenham Science Festival Discover Zone. The team interacted with an estimated 1800 visitors over 6 days. Received feedback such as the following: "Great explanation from volunteers about the stem cell model. My Year 6 pupils could then explain the purpose of the model - great" and "That's how you teach science!"
Year(s) Of Engagement Activity 2015
URL http://www.cheltenhamfestivals.com/science/
 
Description LifeLab - School Interaction 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? Yes
Type Of Presentation Poster Presentation
Geographic Reach Regional
Primary Audience Schools
Results and Impact Several schools involved in learninig about health, nutrition and disease risk - up to 100 Children involved

Schools interested in continuing the programme
Year(s) Of Engagement Activity 2011,2013
 
Description MRC/BBSRC Southampton Imaging launch 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach Regional
Primary Audience Professional Practitioners
Results and Impact The event was to launch Southampton imaging- as part of the UKRMP £1.17M award - to showcase the new tools and equipment in imaging available to the Regenerative Medicine Community, locally and Regionally, The event led ot signifcant collaborative research activity

New grant collaborations - eg at least 2 new PhD studnetships based on the Imaging technology
Year(s) Of Engagement Activity 2014
URL http://www.southampton.ac.uk/southamptonimaging/
 
Description NHS OPEN DAY 
Form Of Engagement Activity Participation in an open day or visit at my research institution
Part Of Official Scheme? No
Geographic Reach Regional
Primary Audience Public/other audiences
Results and Impact A full day presentation on Scaffolds and cells - making replacement body parts in the lab at Southampton General Hospital- attended by over 1000 people

Strong interest from children as a result of a new "Stem Cell Pinball" display built specifically for the event
Year(s) Of Engagement Activity 2014
URL http://www.stemcells.org.uk/
 
Description National Science Week 
Form Of Engagement Activity Participation in an open day or visit at my research institution
Part Of Official Scheme? Yes
Geographic Reach Regional
Primary Audience Public/other audiences
Results and Impact Several thousand visitied University of Southampton Science event- the Bone and Joint Musculoskeletal Activity drew approx 200-300 individuals who activelty participated (made models, discussed bone repair, handeled samples) in the last science week exhibition.

Candidates asked specifically about Medicine Bachelor of Medicine programmes and how to apply. Patients asked if they could jopin clinical trials in the future as availalble for impaction bone grafting - so extremely positive feedback for the researchers on this project.
Year(s) Of Engagement Activity 2009,2010,2011,2012,2013
 
Description Open Conference "Stem cells for bone regeneration: state-of-the-art research and solutions" 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact Invited Speaker - Bonebank Conference, Odense, Denmark. "Skeletal stem cells for bone tissue regeneration.".
The audience were peer scientists, clinicians and industrialist to inform, educate and debate developments in translational bone research.
Year(s) Of Engagement Activity 2016
 
Description Postgraduate Day 
Form Of Engagement Activity Participation in an open day or visit at my research institution
Part Of Official Scheme? No
Geographic Reach Regional
Primary Audience Postgraduate students
Results and Impact Scaffolds and cells - making replacement body parts in the lab - an exhibition showcasing the ability to harness cells and scaffolds - including 3D print technology for orthopaedic regenerative medicine applications

Interest for future research projects and collaborations
Year(s) Of Engagement Activity 2014
URL http://www.stemcells.org.uk/
 
Description Scaffolds and cells - making replacement body parts in the lab - Exhibit at the BBSRC Great British Bioscience 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 Engaged over 6500 visitors from across the UK as part of the BBSRC run GBBF. EPSRC grant generated the centre piece of our exhibit - the Stem cell mountain - a large interactive pinball machine built in collaboration with Winchester Science Centre to communicate the concept of Stem Cell Potential. In response to the feedback question 'Tell us something from your visit that you found particularly interesting' the answer 'Stem Cells' was one of the most frequently given. Through a conversation at this exhibition a talented international MSc student from Queen Mary's was inspired to commence a PhD project (self-funded) in my group.
Year(s) Of Engagement Activity 2014
 
Description Science Day at University of Southampton 
Form Of Engagement Activity Participation in an open day or visit at my research institution
Part Of Official Scheme? No
Geographic Reach Regional
Primary Audience Public/other audiences
Results and Impact Science Day ('Stem Cell Mountain' & Bone regenerative medicine display') at University of Southampton - 14/03/15. (Open to the general public - 1000+ visitors)
Year(s) Of Engagement Activity 2008,2011,2013,2015,2016
 
Description Southampton Science and Engineering Festival 
Form Of Engagement Activity Participation in an open day or visit at my research institution
Part Of Official Scheme? No
Geographic Reach Regional
Primary Audience Public/other audiences
Results and Impact 18 March 2017 - Stem Cell Mountain at the Southampton Science and Engineering Festival for 1 day. regional general public. predominantly families. Interest in students to visit our laboratories
Year(s) Of Engagement Activity 2017
 
Description Speaker at UKRMP Conference UK 
Form Of Engagement Activity A magazine, newsletter or online publication
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact Invited Speaker - Inaugural UK Regenerative Medicine Conference, London. "Size matters - Harnessing biomimetic materials for skeletal tissue engineering."
The audience were peer scientists, clinicians and industrialist to inform, educate and debate developments in translational bone research.
Year(s) Of Engagement Activity 2016
 
Description Stem Cell Mountain at the Glastonbury Music Festival Science Tent 
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 Stem Cell Mountain exhibit in shared Science Tent, mixed Festival audience (academics, post-docs, science specialists, general public, families and young people) Measure 1: visitors passing stand; 1100 per day = 5500 Measure 2: visitors interacting with researchers on stand; 500 per day = 2500 Engaged classically hard to reached audience. Quote: "I was considering doing a PhD at one point - I didn't think I'd reconsider it here"
Year(s) Of Engagement Activity 2015
 
Description Stem Cell Outreach Programme ( New Forest Country Show) 
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 25 - 27 July 2017 - Stem Cell Mountain outreach programme at the New Forest Country Show (general public, farmers, agricultural community) - led ot interest by students to visit our laboratories
Year(s) Of Engagement Activity 2017
 
Description Thomas Hardye School Science Day 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Schools
Results and Impact A Dorset secondary school open day engaging 300 students with translational stem cell medicine. First experience of public engagement for several colleagues who reported great satisfaction with the experience.
Year(s) Of Engagement Activity 2015
URL http://www.southampton.ac.uk/per/university/roadshow.page
 
Description Wessex Medical Charity Talk 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach Regional
Primary Audience Public/other audiences
Results and Impact Approx 80 attendees attended presentation - this resulted in extensive discussion and debate afterwards and over dinner

Request for a repeat presentation for Wessex Charity
Year(s) Of Engagement Activity 2009,2010,2015
 
Description Winchester Science Festival 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach Regional
Primary Audience Public/other audiences
Results and Impact 29 - 30 July 2017 - Stem Cell Mountain at a three day festival. Engaged largely with family audience and science festival talk attendees - led ot gernerla interest in our basic research programmes
Year(s) Of Engagement Activity 2017
 
Description Winchester Science Festival 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach Regional
Primary Audience Public/other audiences
Results and Impact Stem Cell Mountain at a three day up and coming science festival. Engaged largely family audience and science festival talk attendees Measure 1: visitors passing stand; 300 per day = 900 Measure 2: visitors interacting with researchers on stand; 150 per day = 450
Year(s) Of Engagement Activity 2015