The Functional Significance of the Fn14/TWEAK Receptor-Ligand System in Ductular Reactive Cell Differentiation and Bile Ductule Neogenesis.

Liver disease is now a significant health problem in the UK. It accounts for increasing ill-health in the community, hospital admissions and death rates. It also contributes to increasingly higher healthcare costs to the National Health Service. Diseases involving the bile ducts within the liver have also increased dramatically. These diseases include primary biliary cirrhosis (PBC), primary sclerosing cholangitis (PSC) and organ rejection after liver transplantation. It is also observed in livers donated for transplantation after cardiac death, a source of organs on which the NHS Transplant service is becoming more reliant.

Normally the bile ducts drain bile from the liver to the gut to help with digestion. Diseased bile ducts are progressively destroyed, termed ductopenia. The liver has a tremendous capacity to regenerate and make new bile ducts. However, eventually ductopenia from overwhelming destruction of the bile ducts causes an accumulation of bile in the liver and leads to end-stage liver disease. At that point the main treatment is a liver transplant which itself is associated with potentially life-threatening complications. Because the number of organs available for transplantation currently does not meet demand, there is an urgent need to understand the mechanisms which regulate ductopenia and bile duct regeneration.

Precursor cells near the bile ducts, termed ductular reactive cells, promote regeneration of the bile ducts. A molecule called Fn14 is found on these ductular reactive cells. The corresponding molecule to Fn14 called TWEAK is found on certain white blood cells near diseased bile ducts. The interaction of Fn14 and TWEAK might play a crucial role in bile duct regeneration. I propose to study the effects of Fn14 and TWEAK on the bile ducts and the ductular reactive cells in the Centre for Liver Research at the University of Birmingham. This will primarily use bile ducts and ductular reactive cells that have been isolated from diseased livers from the liver transplant programme at the Queen Elizabeth Hospital in Birmingham and obtained with patients' permission. I will initially investigate how much Fn14 is present on the bile ducts and ductular reactive cells in these livers. Additionally I will study what happens to the bile ducts and the ductular reactive cells when the Fn14 and TWEAK molecules interact. Further investigation will focus on if and how the Fn14 molecule influences the ductular reactive cells become regenerating and functioning bile ducts.

Overall this research will enable a better understanding of bile duct regeneration and potentially lead to the development of treatments for ductopenia to ultimately help prevent patients needing a liver transplant.

The novel TNF receptor Fn14 is highly expressed on cholangiocytes in areas of ductular regeneration in the inflammed liver. This project will characterise the role of Fn14 and its ligand TWEAK in remodelling and differentiating intrahepatic bile ducts by cholangiocytes and their ductular reactive cell precursors. This will determine the Fn14/TWEAK system as a potential therapeutic target for chronic ductopenic liver diseases.

Fundamental aims are to define:
1.Fn14/TWEAK expression in normal and ductopenic livers.
2.The functional role of Fn14/TWEAK in biliary remodelling by mature cholangiocytes.
3.The phenotype of ductular reactive cells allowing isolation and culturing of cholangiocyte precursors.
4.The role of Fn14/TWEAK in ductular reactive cell differentiation forming functional biliary architecture.

Project design:
(1)Human studies: cholangiocytes will be isolated from explanted livers. Detailed analysis of Fn14/TWEAK expression and functional outcome of Fn14/TWEAK interaction will utilise multicolour confocal microscopy, flow cytometry and cell IQ for cholangiocyte migration.
(2)Isolation of human ductular reactive cells: by antibody phenotyping before positive and negative fluorescent cell-sorting.
(3)Murine studies: to determine the effects of Fn14/TWEAK using wildtype and Fn14 knockout specimens in liver injury models including carbon tetrachloride fibrosis, bile duct ligation cholestasis and partial hepatectomy.

This project contains a number of novel scientific aspects as previously outlined. Therefore the immediate economic and societal impact is likely to be limited. Nevertheless it is important to plan for the longer term benefits of this research.

This research will firstly emphasise the importance of medical staff to investigating the Fn14/TWEAK receptor-ligand system in hepatic inflammation and repair and their significant contribution towards consent and acquisition of human tissue by surgeons as well as the processing of samples by pathologists. Not only does this have workforce planning implications if an increased number of human samples are processed from other centres, but with appropriate engagement with trainee doctors also encourage more of them to take time out-of-programme from their respective Deaneries to undertake research, ideally at the NIHR Biomedical Research Unit and Centre for Liver Research. This would have workspace implications within the laboratory due to the increasing demand of researchers actively wanting to join the laboratory. It could promote expanding our research facilities at the University of Birmingham.

If it can be demonstrated that the Fn14/TWEAK receptor-ligand system is an appropriate therapeutic target then the commercialisation then this would, in conjunction with our collaborators at Biogen Idec Inc., lead to the creation of spin-off companies in the UK. The University of Birmingham can specifically deal with intellectual property issues relating to commercialisation. Such companies could promote investment into the research and development facilities within or near to our research establishment, contributing to our local economy and the creation of jobs. These are longer term impacts.

Further long term impacts revolve around the need for rigorous clinical trials to ascertain the population effects of this novel therapy. The appropriate clinicians would need to be aware of these ongoing trials. If successful then this may influence provision of NHS services which currently can be time and cost demanding. It may obviate the need for invasive radiological procedures required to stent biliary strictures, prevent biliary reconstructive surgery or possibly orthotopic liver transplantation thus improving the morbidity and mortality of patients. The investment in a therapeutic target for Fn14/TWEAK would be offset by the significant reduction in costs from current treatment options.

Throughout this project there would need to be significant public engagement to ensure that they are aware of firstly the impact of cholangiopathies on morbidity and mortality from chronic liver disease and secondly that research that could have a significant impact on public health is being conducted.