Modelling therapies for surgical adhesions

Lead Research Organisation: University of Manchester
Department Name: School of Biological Sciences

Abstract

In this project we will discover new medical treatments to prevent the formation of internal scars, or 'adhesions'.

People often have to undergo operations which involve opening up their abdomens e.g. to cut out a tumour or an inflamed appendix. A common complication after these operations is the development of adhesions which can glue our guts together. These adhesions are painful and can even cause guts to become blocked and stop working. This then requires a major operation to put things right, yet these second operations may themselves lead to more adhesions.

We will work out how cells inside the abdomen form scars after operations and then use this knowledge to block the process. We will also work out ways to grow scars outside of the body. This will make it easier for us to monitor how well our new therapies are working.

By the end of the project, we aim to have a list of new medicines which can then be tested to help people who suffer from recurrent adhesions.

Technical Summary

Surgical adhesions can cause severe and chronic abdominal pain and life-threatening intestinal obstruction. Although they constitute an enormous socioeconomic burden, there are no effective preventative strategies. Reduced lysis of a post-surgical fibrin-rich clot is an important trigger in the initiation of adhesion development. Critically, a lack of understanding of the aberrant molecular and cellular mechanisms driving mature adhesion formation beyond this first step precludes the rational design of novel biological therapies.

Therefore, we will address the following related hypotheses. 1. Aberrant biological signalling pathways mediate key steps in adhesion formation. 2. Peritoneal mesothelial cells contribute to adhesions via epithelial-mesenchymal transition and 3. Inhibiting these key aberrant biological events will prevent adhesion formation.

We will combine surgical and transgenic strategies in mice with human culture models to successfully address these questions. First, we will replicate the human clinical scenario in a robust mouse model of surgical adhesion formation. Using Wt1CreERT2;Rosa26mTmG mice will allow us, for the first time, to lineage trace mesothelial cells during adhesion formation. We will define aberrant signalling pathways using biochemical techniques, and localise them to cell populations within the forming scar by immunohistology. Consequently, we will use specific reagents to block adhesion formation in the same in vivo model.

In parallel, we will establish monolayer and 3D human cell culture models in which mesothelial cells are triggered to form an adhesion by adding fibrin clot. With these systems, we will establish whether the same biological therapies tested in mice can block human adhesion formation ex vivo.

Our experimental strategy will not only illuminate the pathobiology of adhesion formation, but also pave the way for Phase 1 trials of novel biological therapies to prevent adhesions forming in humans.

Planned Impact

Scientific research impact. The basic science behind surgical adhesion formation has been a neglected research field, especially bearing in mind the scale of human morbidity and economic burden associated with the condition. Excitingly, however, links between tissue repair and fibrosis, and their relation to normal embryonic development and cancer progression, are increasingly appreciated. In particular, the ability of epithelial cells to change phenotype and become myofibroblasts through a process of epithelial-mesenchymal transition. Our project is timely and combines the clinical discipline of surgery with the basic sciences of mouse genetics and cell/molecular biology. Due to its broad scope, the results will be of direct relevance to the clinical and science research communities, including general surgeons, gynaecologists, and cell and developmental biologists.

Novel technology and translational impact. One aim is to generate a novel human 3D peritoneal call assay system which would have a huge impact on the academic and commercial community. The use of pure well characterised populations of primary human mesothelial cells derived by brushing the peritoneal surface (similar to bronchial brushings for airway epithelial cells) will be a first and banking these cells so they are available to the wider scientific community will be of immense and immediate value. In addition, a physiologically relevant in vitro model of the peritoneum that may be used in combination with diseased cells will be of great interest to a number of stakeholders as it will reduce and replace the requirement for animal studies (3Rs) in particular in the fields of tumour metastasis, peritonitis and endometriosis. Furthermore, the safety assessment committee urgently requires more human based research models to assure safety of their new compounds and so this model would be of great significance.

Societal and health impact. Surgical adhesions and their associated complications such as severe chronic pain, bowel obstruction and infertility in women, represent a huge health problem world-wide. Surgery is the only way to detect and treat already-formed adhesions but surgery itself also causes adhesions. The cost of adhesion-related complications for the NHS are an estimated £67 million each year. At present there are no pharmacological ways of preventing adhesion formation, and current physical therapies (e.g. degradable membranes and gel barriers) show limited efficacy. Any advance in developing efficient anti-adhesion agents would have enormous consequences for the range of surgical specialities. Furthermore, because surgical adhesions are associated or resemble other pathologies such as encapsulating peritoneal sclerosis, endometriosis, and cancer-, radiation-, infection- as well as dialysis-induced peritoneal sclerosis, the results of our project could potentially have widespread benefits in a number of clinically important areas.

Publications

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Mutsaers SE (2016) Mesothelial cells and peritoneal homeostasis. in Fertility and sterility

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Mutsaers SE (2015) Mesothelial cells in tissue repair and fibrosis. in Frontiers in pharmacology

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Pengelly S (2018) Outcome of open abdominal management following military trauma. in The British journal of surgery

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Siddique I (2015) Genetic Polymorphisms and Peritoneal Membrane Function. in Peritoneal dialysis international : journal of the International Society for Peritoneal Dialysis

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Wilm B (2016) Tools and Techniques for Wt1-Based Lineage Tracing. in Methods in molecular biology (Clifton, N.J.)

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Wilm B (2016) The Role of WT1 in Embryonic Development and Normal Organ Homeostasis. in Methods in molecular biology (Clifton, N.J.)

 
Description Defining functional impacts of macrophages in the formation of surgical adhesions
Amount £575,759 (GBP)
Funding ID MR/S02560X/1 
Organisation Medical Research Council (MRC) 
Sector Public
Country United Kingdom
Start 08/2019 
End 10/2022
 
Description IKC POC Industrial partnership
Amount £70,000 (GBP)
Organisation Engineering and Physical Sciences Research Council (EPSRC) 
Sector Public
Country United Kingdom
Start 05/2018 
End 02/2019
 
Description RCS/BAPS Surgical Research Fellowship
Amount £59,013 (GBP)
Organisation Royal College of Surgeons of England 
Sector Charity/Non Profit
Country United Kingdom
Start 09/2016 
End 09/2017
 
Description grant applications 2016
Amount £13,900 (GBP)
Organisation Kidneys for Life 
Sector Charity/Non Profit
Country United Kingdom
Start 11/2016 
End 11/2017
 
Title Tamoxifen inducible lineage tracing of Wt1-expressing mesothelial cells 
Description We have established a genetic lineage tracing approach to follow the fate of adult mesothelial cells using Tamoxifen-driven Cre-ERT2 recombinase under control of the Wilms' tumour 1 gene. This system can be used to follow the fate of mesothelial cells in the embryo and adult. Using several different reporter lines, we have established a good understanding of this lineage tracing system for normal homeostasis and disease or injury conditions. 
Type Of Material Model of mechanisms or symptoms - mammalian in vivo 
Provided To Others? No  
Impact This tool is very useful for in vivo applications where the fate of mesothelial cells is followed after an intervention with mimicks a disease or injury situation. 
 
Title Using genetic lineage tracing to analyse the contribution of mesothelial cells to surgical adhesions 
Description We have used the genetic lineage tracing system to set up an in vivo model to follow the contribution of mesothelial cells to surgically-induced peritoneal adhesions. Our genetic system allows detailed analysis of the mesothelial cell contribution. 
Type Of Material Model of mechanisms or symptoms - mammalian in vivo 
Provided To Others? No  
Impact This method will allow us to demonstrate the contribution of mesothelial cells to adhesion formation, as well as to analyse the molecular mechanisms that regulate the process. 
 
Title purification of mesothelial cells 
Description Novel method developed to isolate mesothelial cells using magnetic bead sorting. 
Type Of Material Technology assay or reagent 
Year Produced 2017 
Provided To Others? Yes  
Impact Mesothelial cells are only available commercially from a limited number of suppliers so we have developed an in-house method to purify them from human omentum. 
 
Title RNA sequencing data for mesothelial transition 
Description We have performed RNA sequencing on mesothelial cells treated with TGF-B and discovered a number of novel up and down regulated genes. We are now understanding the role of the genes in mesothelial-mesenchymal transition. 
Type Of Material Database/Collection of data 
Provided To Others? No  
Impact We will publish the data generated which can then be used by other labs. New strategies to inhibit mesothelial mesenchymal transition will be tested leading to reduced peritoneal fibrosis and adhesion formation. 
 
Description Manchester Biogel 
Organisation Manchester BIOGEL
Country United Kingdom 
Sector Private 
PI Contribution Myself and Adrian Woolf were recently awarded an IKC proof of concept award (EPSRC funded through University of Leeds) to work on a project with our established collaborator, Peptigeldesign, an SME with specialist material fabrication technology. The project develops a unique medical device with regenerative properties for a new therapeutic application which shows clear commercial opportunities and has strong industry involvement.
Collaborator Contribution Peptigeldesign will provide the medical device and maximum input into study design, deliverables and future applications for funding, commercialisation and clinical trials.
Impact The collaboration is multi-disciplinary involving a SME, biomaterials and surgeons to complement our basic peritoneal biology expertise.
Start Year 2016
 
Description Community Science 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 General public engaged in hands-on activities around surgery and adhesions. Lots of interest and questions from children and parents/carers.
Year(s) Of Engagement Activity 2019
 
Description Community 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 I organised a community event involving a team of researchers including an Anatomy professor and a surgeon and myself and a post-doc. We had interactive games and stalls to show the general public about our research ie. the inside of your body is covered by a layer of special protective cells and when they are injured, such as after surgery, they can form scar tissue which may cause problems if not prevented.
Year(s) Of Engagement Activity 2018
 
Description Meet the mesothelium workshop 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Other audiences
Results and Impact We organised a symposium: Meet the Mesothelium. Aim of the symposium therefore was to get as many research groups, with interest in basic and translational biology of the mesothelium, as possible together to maintain and extend the existing contacts and collaborations and discuss the latest, unpublished developments in the mesothelium field.

Statement on diversity of speakers: Our symposium speakers represent a well-balanced mix of genders. While we have 4 speakers from the UK, we have excellent European coverage, with speakers from Sweden, Belgium, Spain, Italy and Poland. We are lucky to have Steve Mutsaers, a world expert on mesothelial biology who is from Australia, as speaker. Furthermore, our symposium speakers are at different stages of their careers, from senior (Prof Mutsaers, Prof Munoz-Chapuli) to junior (Christine-Maria Horejs).

We have been discussing larger grants between European groups.
Year(s) Of Engagement Activity 2016