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.
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.
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

Lucendo-Villarin B
(2017)
Erratum to: Modelling foetal exposure to maternal smoking using hepatoblasts from pluripotent stem cells
in Archives of Toxicology

Lucendo-Villarin B
(2017)
Erratum to: Modelling foetal exposure to maternal smoking using hepatoblasts from pluripotent stem cells
in Archives of Toxicology

Lynch RW
(2018)
An efficient method to isolate Kupffer cells eliminating endothelial cell contamination and selective bias.
in Journal of leukocyte biology

Macfarlane D
(2011)
Metabolic pathways promoting intrahepatic fatty acid accumulation in methionine and choline deficiency: implications for the pathogenesis of steatohepatitis
in American Journal of Physiology-Endocrinology and Metabolism

McBride A
(2017)
In search of a small molecule agonist of the relaxin receptor RXFP1 for the treatment of liver fibrosis.
in Scientific reports

Mole D
(2009)
Alcohol and global health: focus on acute pancreatitis needed.
in Lancet (London, England)

Mole DJ
(2016)
Kynurenine-3-monooxygenase inhibition prevents multiple organ failure in rodent models of acute pancreatitis.
in Nature medicine

Murray IR
(2017)
av integrins on mesenchymal cells regulate skeletal and cardiac muscle fibrosis.
in Nature communications

Payne CM
(2011)
Persistence of functional hepatocyte-like cells in immune-compromised mice.
in Liver international : official journal of the International Association for the Study of the Liver

Pellicoro A
(2012)
Reversibility of liver fibrosis.
in Fibrogenesis & tissue repair
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 | Public |
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 | Public |
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 | Public |
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 | Public |
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 | Public |
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 | Public |
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 | Public |
Country | United Kingdom |
Start |
Description | Medical Research Council Programme Renewal |
Amount | £1,900,000 (GBP) |
Organisation | Medical Research Council (MRC) |
Sector | Public |
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 | Charity/Non Profit |
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 | 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 |
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 |