The role of elastin degradation in the pathogenesis of liver fibrosis

Lead Research Organisation: University of Edinburgh
Department Name: MRC Centre for Inflammation Research

Abstract

Long term or chronic liver disease, regardless of cause, results in scarring of the liver (termed fibrosis and cirrhosis). This scarring causes the liver to fail and is associated with significant illness and death. In the UK, the burden of cirrhosis has been identified by the Chief Medical Officer as a major challenge of the next decade. Currently there are no effective treatments for liver scarring. We have a programme of research investigating the factors which regulate recovery from scarring to inform and develop anti-scarring (so called anti-fibrotic) therapies.

We have recently demonstrated that if it is allowed to, the liver has a significant ability to break the scar tissue with a return towards more normal structure and function. In advanced liver disease though, the scar tissue persists and appears to be resistant to break-down. The presence of a protein called elastin seems to be critical to preventing effective breakdown of the scar tissue. In addition, we have preliminary evidence to suggest that a specialised form of inflammatory cell, the macrophage, produces a specific protein (enzyme) which breaks down elastin. Taken together these results suggest that elastin characterises irreversible scarring (cirrhosis) and that it accumulates as a result of a failure of the normal processes of breakdown.

Work we are undertaking will determine the cell source of elastin and the elastin degrading proteins and their relative levels at different stages of liver scarring (reversible and irreversible).
In addition, we will use experimental models to determine the mechanisms that make scarring resistant to breakdown and use infusions of purified inflammatory cells and specific population of stem cells to enhance scar breakdown.

Ultimately, by understanding the mechanisms regulating the turnover of elastin in liver scarring we will be better able to design effective anti-scarring therapies applicable in the clinic.

Technical Summary

Currently there are no effective treatments for liver fibrosis and cirrhosis which represent major health challenges worldwide. Therefore, there is a pressing need to develop antifibrotic therapies. We have recently demonstrated that experimental liver fibrosis is reversible and that reversal is characterised by matrix degradation and apoptosis of activated (myofibroblast-like) hepatic stellate cells (the major cell mediator of fibrosis). However, following prolonged experimental injury, fibrosis develops which is not degraded even after one year of recovery. Characterisation of the septa which fail to undergo degradation indicates that they contain elastin, in contrast to those which rapidly become remodelled, which do not. These septa are also crosslinked by tissue transglutaminase (tTG), for which elastin is a substrate.
In pilot studies, we have shown elastase (MMP-12) is expressed in the liver by tissue macrophages and after conditional depletion of macrophages, hepatic elastin degradation is impaired; indicating that the macrophages are critical to spontaneous recovery from fibrosis and are an important source of MMP-12.
We propose to investigate the hypotheses that: A failure of elastin degradation characterises mature liver fibrosis. The presence of elastin facilitates matrix cross-linking which becomes resistant to MMP mediated degradation and persists even during spontaneous resolution. Initially, we propose to quantitate the relative expression, distribution and cellular source of elastin and MMP-12 in rodent models of reversible and irreversible fibrosis. These will be completed by studies of cirrhotic human liver tissue. MMP-12 knockout mice and wild type mice will be examined after induction of liver fibrosis by CCl4 injection and during spontaneous recovery to examine whether elastin persistence results from a failure of elastin degradation. We will go on to ?rescue? the phenotype of MMP-12 knockout mice by restoring a wild type macrophage genotype via either haemopoetic stem cell infusion or direct macrophage injection from wild type animals. We will use DTR mice to examine in detail the effect of selective macrophage depletion on the degradation of elastin and fibrotic tissue in livers undergoing spontaneous recovery from experimentally induced liver fibrosis. Finally we will use a novel model to determine the relative role of stellate cell/myofibroblast derived MMP-12 vs macrophage derived MMP-12 in matrix turnover by seeding the hepatic scar with MMP-12 knockout stem cell derived stellate cells or macrophages. By completion of these studies we will have defined the role of elastin and regulation of its degradation by MMP-12 in the spontaneous resolution of liver fibrosis.

Publications

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Argyle DJ (2013) ECAT-V: where clinical and research training meet. in The Veterinary record

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Bird TG (2013) Bone marrow injection stimulates hepatic ductular reactions in the absence of injury via macrophage-mediated TWEAK signaling. in Proceedings of the National Academy of Sciences of the United States of America

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Campana L (2017) Regression of Liver Fibrosis. in Seminars in liver disease

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Conti JA (2008) The desmoplastic reaction surrounding hepatic colorectal adenocarcinoma metastases aids tumor growth and survival via alphav integrin ligation. in Clinical cancer research : an official journal of the American Association for Cancer Research

 
Description Academy of Medical Sciences Intermediate Fellowship (Dr D Mole)
Amount £538,239 (GBP)
Organisation Academy of Medical Sciences (AMS) 
Sector Charity/Non Profit
Country United Kingdom
Start 09/2008 
End 08/2013
 
Description Academy of Medical Sciences Intermediate Fellowship (Dr J Fallowfield)
Amount £743,397 (GBP)
Funding ID AMS-CSF3-Fallowfield 
Organisation Academy of Medical Sciences (AMS) 
Sector Charity/Non Profit
Country United Kingdom
Start 09/2009 
End 08/2014
 
Description Fellowship (Fallowfield)
Amount £60,000 (GBP)
Organisation Novartis 
Sector Private
Country Global
Start 02/2013 
End 01/2014
 
Description MRC CIR Centre Core Support Grant - Renewal
Amount £1,480,000 (GBP)
Organisation Medical Research Council (MRC) 
Sector Academic/University
Country United Kingdom
Start 09/2011 
End 09/2016
 
Description MRC Clinical Training Fellowship (Dr A Robson)
Amount £236,467 (GBP)
Organisation Medical Research Council (MRC) 
Sector Academic/University
Country United Kingdom
Start 04/2007 
End 03/2010
 
Description MRC Clinical Training Fellowship (Dr M Gibbons)
Amount £239,000 (GBP)
Organisation Medical Research Council (MRC) 
Sector Academic/University
Country United Kingdom
Start 03/2007 
End 02/2010
 
Description MRC Clinical Training Fellowship (Dr T Gordon-Walker)
Amount £223,799 (GBP)
Organisation Medical Research Council (MRC) 
Sector Academic/University
Country United Kingdom
Start 04/2008 
End 03/2011
 
Description MRC Collaborative Pogramme Grant (Prof CP Day)
Amount £2,400,000 (GBP)
Organisation Medical Research Council (MRC) 
Sector Academic/University
Country United Kingdom
Start 04/2007 
End 03/2012
 
Description MRC Programme Grant (with S Forbes)
Amount £1,200,000 (GBP)
Organisation Medical Research Council (MRC) 
Sector Academic/University
Country United Kingdom
Start 05/2007 
End 04/2012
 
Description MRC Strategic Grant (Forbes/Knight)
Amount £268,000 (GBP)
Organisation Medical Research Council (MRC) 
Sector Academic/University
Country United Kingdom
Start  
 
Description Medical Research Council Programme Renewal
Amount £1,900,000 (GBP)
Organisation Medical Research Council (MRC) 
Sector Academic/University
Country United Kingdom
Start 05/2012 
End 04/2017
 
Description Project grant (Mole)
Amount £241,552 (GBP)
Organisation GlaxoSmithKline (GSK) 
Sector Private
Country Global
Start 08/2013 
End 01/2014
 
Description Royal College of Surgeons Vacation Bursary (Ms Aysha Ali)
Amount £1,350 (GBP)
Organisation Royal College of Surgeons of England 
Sector Learned Society
Country United Kingdom
Start 06/2010 
End 07/2010
 
Description Stem Cells for Safer Medicine (Hay & Brickman)
Amount £180,000 (GBP)
Organisation Stem Cells for Safer Medicines (SC4SM) 
Sector Charity/Non Profit
Country United Kingdom
Start  
 
Description Technology Strategy Board Grant Roslin Cellab
Amount £140,937 (GBP)
Organisation Innovate UK 
Sector Public
Country United Kingdom
Start  
 
Description UKSCF/UK Stem Cell Foundation
Amount £1,700,000 (GBP)
Organisation UK Stem Cell Foundation 
Sector Charity/Non Profit
Country United Kingdom
Start 10/2008 
End 09/2010
 
Description Wellcome Trust Equipment Grant (Mullins)
Amount £668,183 (GBP)
Funding ID 092155/Z/10/Z 
Organisation Wellcome Trust 
Sector Charity/Non Profit
Country United Kingdom
Start 09/2010 
End 08/2015
 
Description Wellcome Trust Intermediate Fellowship (Dr N Henderson)
Amount £877,220 (GBP)
Funding ID 085187/Z/08/Z 
Organisation Wellcome Trust 
Sector Charity/Non Profit
Country United Kingdom
Start 02/2009 
End 01/2013
 
Description Wellcome Trust Intermediate Fellowship (T Kendall)
Amount £867,024 (GBP)
Funding ID 095898 
Organisation Wellcome Trust 
Sector Charity/Non Profit
Country United Kingdom
Start 10/2011 
End 09/2015
 
Description Wellcome Trust PhD Portfolio for Clinicians (ECAT)
Amount £5,000,000 (GBP)
Organisation Wellcome Trust 
Sector Charity/Non Profit
Country United Kingdom
Start 10/2008 
End 09/2013
 
Description Wellcome Trust Research Training Fellowship (Dr D McFarlane)
Amount £135,404 (GBP)
Organisation Wellcome Trust 
Sector Charity/Non Profit
Country United Kingdom
Start 03/2007 
End 02/2009
 
Description Wellcome Trust Research Training Fellowship (Dr P Ramachandran)
Amount £242,492 (GBP)
Funding ID 083869/Z/07/Z 
Organisation Wellcome Trust 
Sector Charity/Non Profit
Country United Kingdom
Start 09/2008 
End 08/2011
 
Description Wellcome Trust Research Training Fellowship (Dr T Bird)
Amount £240,000 (GBP)
Funding ID 081604/Z/06/Z 
Organisation Wellcome Trust 
Sector Charity/Non Profit
Country United Kingdom
Start 05/2007 
End 04/2010
 
Description Wellcome Trust STMTI Fellowship (V Snowdon)
Amount £244,706 (GBP)
Funding ID 096528 
Organisation Wellcome Trust 
Sector Charity/Non Profit
Country United Kingdom
Start 04/2011 
End 03/2014
 
Description Wellcome Trust Vacation Scholarship (M Rygier)
Amount £1,440 (GBP)
Organisation Wellcome Trust 
Sector Charity/Non Profit
Country United Kingdom
Start 06/2009 
End 07/2009
 
Description Wellcome Trust Vacation Scholarship (Pei Pei Lee)
Amount £1,400 (GBP)
Organisation Wellcome Trust 
Sector Charity/Non Profit
Country United Kingdom
Start 06/2009 
End 07/2009
 
Title iPSC & HuESC derived hepatocytes 
Description We have employed ActivinA and Wnt3a signalling to produce efficient levels of hepatocyte like cells (approximately 90%) which display hepatic function, including: ureagenesis, gluconeogenesis, albumin secretion and CYP1A2 activity from hESCs and iPSCs (Hay etal.,2008 ; Sullivan etal.,2010). Although hepatic endoderm (HE) generated from hESCs and iPSCs is a genotypically diverse, cheap and stable source of "hepatocytes", HE routine application is limited due to phenotypic instability in culture. Adopting an unbiased approach we screened polymer microarrays and identified a polyurethane matrix which promoted hepatic endoderm viability, hepatocellular gene expression, drug inducible metabolism and function. We have identified a manufacturable sub-cellular matrix capable of supporting long-term differentiated cell function which represents a step forward in developing scalable and phenotypicaly stable hESC derived hepatocytes. 
Type Of Material Technology assay or reagent 
Year Produced 2009 
Provided To Others? Yes  
Impact We have employed ActivinA and Wnt3a signalling to produce efficient levels of hepatocyte like cells (approximately 90%) which display hepatic function, including: ureagenesis, gluconeogenesis, albumin secretion and CYP1A2 activity from hESCs and iPSCs (Hay etal.,2008 ; Sullivan etal.,2010). Although hepatic endoderm (HE) generated from hESCs and iPSCs is a genotypically diverse, cheap and stable source of "hepatocytes", HE routine application is limited due to phenotypic instability in culture. Adopting an unbiased approach we screened polymer microarrays and identified a polyurethane matrix which promoted hepatic endoderm viability, hepatocellular gene expression, drug inducible metabolism and function. We have identified a manufacturable sub-cellular matrix capable of supporting long-term differentiated cell function which represents a step forward in developing scalable and phenotypicaly stable hESC derived hepatocytes. 
 
Description Identifying Antifibrotic Targets 
Organisation Novartis
Country Global 
Sector Private 
PI Contribution I have acted as a consultant to the above facilitating the development of the above programme.
Collaborator Contribution Novartis and their predecessor Corthera have supplied recombinant relaxin and worked with us on the deployment of relaxin as an antifibrotic and portal hypotensive agent. We are currently in discussions over the feasibility of an experimental medicine study in human subjects
Impact Currently discusing clinical application of relaxin as antifibrotic and portal hypotensive agent
Start Year 2008
 
Description Imaging Liver Fibrosis 
Organisation Bayer
Department Bayer HealthCare
Country Germany 
Sector Private 
PI Contribution I have acted a s a consultant to the above facilitating the development of the above programme.
Impact None at present
Start Year 2006
 
Description polymers for growing cells 
Organisation University of Edinburgh
Department MRC Centre for Reproductive Health
Country United Kingdom 
Sector Public 
PI Contribution Providing liver biology expertise for extended culture of functional human hepatocyte like cells
Collaborator Contribution Added a multidisciplinary component to the MRC CIRAs before
Impact Patent : (WO/2010/106345) POLYMERS FOR GROWING CELLS
Start Year 2010
 
Description polymers for growing cells 
Organisation University of Edinburgh
Department School of Chemistry
Country United Kingdom 
Sector Academic/University 
PI Contribution Providing liver biology expertise for extended culture of functional human hepatocyte like cells
Collaborator Contribution Added a multidisciplinary component to the MRC CIRAs before
Impact Patent : (WO/2010/106345) POLYMERS FOR GROWING CELLS
Start Year 2010
 
Title Polymers for growing cells 
Description The present invention provides a polymer substrate for use in the attachment and functioning of hepatocyte and hepatocyte like cells. In particular, the polymer substrate is a polyurethane polymer. 
IP Reference WO2010106345 
Protection Patent application published
Year Protection Granted 2010
Licensed Commercial In Confidence
Impact The ability to culture functional human heptocyte like cells over long periods of time in vitro
 
Title Polymers for growing cells 
Description The present invention provides a polymer substrate for use in the attachment and functioning of hepatocyte and hepatocyte like cells. In particular, the polymer substrate is a polyurethane polymer. 
Type Support Tool - For Fundamental Research
Current Stage Of Development Refinement. Non-clinical
Year Development Stage Completed 2009
Development Status Under active development/distribution
Impact Ability to maintain functional human hepatocyte like cells for extended periods of culture in vitro 
 
Description BASL and AASLD 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Type Of Presentation Keynote/Invited Speaker
Geographic Reach International
Primary Audience Health professionals
Results and Impact Presentations have been made by our group at the major UK and North American liver disease meetings: BASL and AASLD.

Presentations at the AASLD have recieved the President's commendation (top 10% of scientific presentations)
Year(s) Of Engagement Activity 2008,2009,2010
 
Description Inaugural Lecture 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Type Of Presentation Keynote/Invited Speaker
Geographic Reach National
Primary Audience Public/other audiences
Results and Impact 1 hour public lecture with reception afterwards.

Wide ranging audience from high school students
Year(s) Of Engagement Activity 2007
 
Description Medical Detectives Lecture 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Type Of Presentation Keynote/Invited Speaker
Geographic Reach National
Primary Audience Public/other audiences
Results and Impact a public lecture as part of a series celebrating Conan Doyle's medical studentship in Edinburgh 45 mins with discusion, wide ranging audience from high school students to pensioners

it was posted on youtube and I have received feedback from around the world
Year(s) Of Engagement Activity 2009
 
Description Primary Futures: Who's in Health? campaign launch 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach Regional
Primary Audience Schools
Results and Impact Iredale launched "Who's in Health" in Scotland with the Chief Medical Officer for Scotland, Dr Catherine Calderwood at Sciennes Primary School in Edinburgh on the 1st of December. This event launched the Primary Futures Campaign to help young children understand how people in the health sector use literacy, maths and science in their jobs.
Year(s) Of Engagement Activity 2015
URL http://news.scotland.gov.uk/News/Primary-Futures-Who-s-In-Health-1fd9.aspx
 
Description Public Presentation 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? Yes
Type Of Presentation Keynote/Invited Speaker
Geographic Reach National
Primary Audience Public/other audiences
Results and Impact A public presentation with colleagues from Oxford and KCL exploring the potential of regenerative medicine. This was followed by an informal public breakout session. The presentation comprised MRC's contribution to the Times Cheltenham Science Festival June 2012.

Wide ranging audience comprising members of the public, patients and other interested parties. Uniformly positive feedback received.
Year(s) Of Engagement Activity 2012