The role of monocytes and macrophages in the outcome of acute liver failure

Liver disease is the 5th commonest cause of death in the UK. The majority of it in Western communities is caused by the effects of excess alcohol consumption. Alcoholic liver disease is increasing in the UK and alcoholic hepatitis (AH) is it's most florid manifestation, affecting 5 in every 100,000 people. AH hospitalises relatively young people, the average age being 51, and so it impacts heavily on both quality and quantity of life in our communities.

In the UK, acute liver failure (ALF) occurs in up to one thousand people per year. It is a devastating condition that carries a high mortality rate (>40%) despite the use of liver transplantation. ALF accounts for 10-20% of all liver transplants performed in the UK. At present only 6-7 hundred donor organs are available each year for liver transplantation. This fails to meet the current demand resulting in deaths on the waiting list, some 30-50% of patients listed not proceeding to transplantation. Acute liver failure cases are listed under an emergency category and given priority on the waiting list over elective cases, exacerbating the risk of death on waiting list for patients with end-stage chronic liver disease. Drug induced liver injury, particularly paracetamol toxicity is the commonest cause of ALF.

Infection is very common in both AH and ALF and is associated with a rapid decline in liver function and premature death in over fifty percent of these patients. The cause for this marked susceptibility to infections is largely unknown, but our recent data indicate that monocytes, a subset of circulating white blood cells, are markedly defective and unable to combat infectious agents such as bacteria and viruses. A significant proportion of this research will study monocytes from the bloodstream of patients suffering from AH and ALF and we will assess their ability to fight infectious agents. I will consider whether patients with poorly functioning monocytes have an increased risk of infection by following their admission to hospital. In addition, I will look for factors in their blood which prevent these immune cells from working properly which, if removed, could restore immune function. This understanding will allow us to develop targeted, individualised treatments that restore the immune response to infection and save lives in patients with AH and ALF.

Currently, treatment strategies for ALF are limited with no specific targeted therapies in current clinical care used to ameliorate the severity of liver damage. The second component of this research will focus on the role of macrophages in the damaged and inflamed livers of patients with ALF and AH. Macrophages, which reside in tissue, represent more mature immune cell population, derived from circulating monocytes and are major determinants of the body's response to tissue injury. They possess both tissue destructive capacity, required to clear damaged cells, and also tissue repair properties. Their role during ALF and AH is not fully understood. We have recently shown that, within the liver of patients undergoing emergency liver transplantation for ALF, there is an abundance of macrophages attempting to promote tissue repair. We would like to investigate the role played by hepatic macrophages in during experimental and human models of acute liver injury. In particular, we would like to focus our work towards understanding whether these immune cells resolve liver damage and promote tissue repair. Better understanding of the mechanisms that control the function of this pivotal immune cell should provide novel insight into the progression of acute liver injury and identify key therapeutic targets to promote recovery following acute liver injury. Recovery of patients with ALF without need for transplantation would impact greatly on health resource utilization and further improve availability of organs for patients with chronic liver disease.

Sepsis is of major cause of mortality in both acute liver failure (ALF) and severe alcoholic hepatitis (AH), termed ALF syndromes. Our work indicates that circulating monocytes (Mo) and hepatic macrophages (H-mac) are responsible for the observed immunoparesis and play a major role in the pathogenesis and outcome of ALF syndromes.

Our objectives are to assess whether circulating Mo dysfunction accounts for the susceptibility to sepsis and poor outcome in ALF syndromes. We will investigate if changes in functional characteristics of H-Mac and their production of anti-inflammatory mediators are responsible for monocyte dysfunction in ALF syndromes.
Mo/H-Mac will be isolated and phenotypically characterised using a FACS-based lineage gating strategy. Functional analyses will consist of a number of FACS and ELISA based techniques that detect the secretion of inflammatory mediators, TLR intracellular signalling pathways, the degree of apoptosis, phagocytosis/oxidative burst activity and antigen presentation capabilities to recall antigen. In addition, in vitro analyses will assess the effects of ALF/AH sera and anti-inflammatory mediators on monocyte function.

Functional characterisation of H-macs will be performed at the different phases of experimental ALF and compared to that of explanted H-macs. Using co-culture of H-macs with parenchymal and non-parenchymal cells and knock-out in experimental ALF models, we will determine key mediators and mechanisms that induce a switch in function of h-macs to promote the resolution of inflammation, tissue repair/regenerative processes.

Liver disease is the 5th commonest cause of death in the UK and rising sharply in prevalence. This work will increase our understanding of mechanisms of hepatic injury and infection in ALF syndromes, generate targeted therapies to attenuate liver injury and reduce infection, thus improving the prohibitively high mortality rate encountered in these conditions.

This project will beneficially impact the following groups:
1. Patients suffering from acute hepatic syndromes who may require urgent liver transplantation
2. Reduce the requirement for transplantation for patients with acute hepatic syndromes thus improving organ availability for patients with chronic liver disease.
3. Patients with septic complications of liver syndromes where mortality is high and where transplantation is often contraindicated

Three key ways in which this research will benefit patient health are described in detail below:
Liver disease is the 5th most common cause of death in the UK and alcohol is the chief cause. Alcoholic hepatitis (AH) affects a third of heavy drinkers. Importantly, it may afflict patients who had not previously been advised to stop drinking, for whom prognosis may be bleak: short term mortality can be as high as 35%. A better understanding of the pathogenesis of AH is needed in order to open new avenues for therapy to the disease itself. There are extensive animal data highlighting monocytes/macrophages as key orchestrators of ethanol-mediated liver injury but a corresponding paucity of data from human studies regarding macrophage and circulating monocyte function in AH. We envisage gaining significant insight into the role of monocytes/macrophages in the pathogenesis of AH, reveal a number of potential therapeutic targets on which to base interventional strategies with novel agents.

ALF is a devastating condition that carries a mortality rate >40% despite the use of liver transplantation. ALF accounts for 10-20% of liver transplants performed in the UK. At present only 6-7 hundred donor organs are available each year for liver transplantation. This fails to meet the current demand resulting in deaths on the waiting list, some 30-50% of patients listed not proceeding to transplantation. Acute liver failure cases are listed under an emergency category and given priority on the waiting list over elective cases, exacerbating the risk of death on waiting list for patients with end-stage chronic liver disease. Drug induced liver injury; particularly paracetamol (acetaminophen) toxicity is the commonest cause of ALF. The role of the innate immune system, in particular macrophages, in the propagation of acute liver injury is not well understood in ALF. Better understanding of the mechanisms that control the function of this pivotal cell in ALF should provide insight into the progression of acute liver injury and identify key therapeutic targets to promote recovery. Recovery of patients with ALF without need for transplantation would impact greatly on health resource utilization and further improve availability of organs for patients with chronic liver disease. Currently, treatment strategies for ALF are limited with no commercially available, specific therapies in current clinical usage highlighting an unmet need for specific, targeted therapies in ALF.

Infection is the commonest cause of mortality in both ALF and AH. Insights gained from this project have the potential to reduce morbidity and mortality due to sepsis. If we can demonstrate that phenotypic or functional monocyte defects correlate with susceptibility to infection, we could accurately stratify their disease severity and rationalise their medical care, such as in the use of anti-microbial and immunosuppressive agents. For example, if impairment in monocyte oxidative burst predicts likelihood of sepsis, clinicians may elect to avoid immunosuppressive treatment for AH, such as corticosteroid therapy, in those patients demonstrated to have poor monocyte oxidative burst capacity. Alternatively, antibiotic prophylaxis or therapy may be focussed on patients with distinct immune defects, which may in turn be targeted at particular pathogens. We would anticipate the identifying and assessing the potential utility of such biomarkers over a 5 year period.