Arjuna Singanayagam

Update on acute liver failure.

Purpose of reviewAlthough advances in critical care management and liver transplantation have improved survival in acute liver failure (ALF), mortality remains significant. An evidence base to support management has been lacking, due to the conditions rarity, severity and heterogeneity. The purpose of this review is to critically appraise the latest evidence, updating clinicians on the current understanding of the best management. Recent findingsTransplant-free survival in acetaminophen-related ALF has improved considerably, such that reconsidering thresholds for transplant is required, perhaps utilizing biomarkers of liver regeneration. Autoimmune hepatitis-related ALF may be too advanced to permit rescue with corticosteroids, which could be deleterious in the sickest patients. Acute kidney injury is commoner in ALF than previously suspected. Intracranial pressure monitoring does not appear to alter mortality. Despite altered traditional indices of coagulation, new thrombin generation assays suggest a rebalanced coagulation in liver failure. Antimicrobial prophylaxis may not be required in all patients. Liver support systems remain controversial and require further evaluation. SummaryTraditional dogma in ALF management is questioned: transplant thresholds for acetaminophen overdose, steroid use in autoimmune ALF, routine antimicrobial prophylaxis, the coagulopathy of liver disease, the value of intracranial pressure monitoring and extracorporeal liver support.

Review article: the gut microbiome as a therapeutic target in the pathogenesis and treatment of chronic liver disease

Mortality from chronic liver disease is rising exponentially. The liver is intimately linked to the gut via the portal vein, and exposure to gut microbiota and their metabolites translocating across the gut lumen may impact upon both the healthy and diseased liver. Modulation of gut microbiota could prove to be a potential therapeutic target.

MerTK expressing hepatic macrophages promote the resolution of inflammation in acute liver failure

Objective Acute liver failure (ALF) is characterised by overwhelming hepatocyte death and liver inflammation with massive infiltration of myeloid cells in necrotic areas. The mechanisms underlying resolution of acute hepatic inflammation are largely unknown. Here, we aimed to investigate the impact of Mer tyrosine kinase (MerTK) during ALF and also examine how the microenvironmental mediator, secretory leucocyte protease inhibitor (SLPI), governs this response. Design Flow cytometry, immunohistochemistry, confocal imaging and gene expression analyses determined the phenotype, functional/transcriptomic profile and tissue topography of MerTK+ monocytes/macrophages in ALF, healthy and disease controls. The temporal evolution of macrophage MerTK expression and its impact on resolution was examined in APAP-induced acute liver injury using wild-type (WT) and Mer-deficient (Mer−/−) mice. SLPI effects on hepatic myeloid cells were determined in vitro and in vivo using APAP-treated WT mice. Results We demonstrate a significant expansion of resolution-like MerTK+HLA-DRhigh cells in circulatory and tissue compartments of patients with ALF. Compared with WT mice which show an increase of MerTK+MHCIIhigh macrophages during the resolution phase in ALF, APAP-treated Mer−/− mice exhibit persistent liver injury and inflammation, characterised by a decreased proportion of resident Kupffer cells and increased number of neutrophils. Both in vitro and in APAP-treated mice, SLPI reprogrammes myeloid cells towards resolution responses through induction of a MerTK+HLA-DRhigh phenotype which promotes neutrophil apoptosis and their subsequent clearance. Conclusions We identify a hepatoprotective, MerTK+, macrophage phenotype that evolves during the resolution phase following ALF and represents a novel immunotherapeutic target to promote resolution responses following acute liver injury.

CD14+CD15-HLA-DR- myeloid-derived suppressor cells impair antimicrobial responses in patients with acute-on-chronic liver failure

Objective Immune paresis in patients with acute-on-chronic liver failure (ACLF) accounts for infection susceptibility and increased mortality. Immunosuppressive mononuclear CD14+HLA-DR− myeloid-derived suppressor cells (M-MDSCs) have recently been identified to quell antimicrobial responses in immune-mediated diseases. We sought to delineate the function and derivation of M-MDSC in patients with ACLF, and explore potential targets to augment antimicrobial responses. Design Patients with ACLF (n=41) were compared with healthy subjects (n=25) and patients with cirrhosis (n=22) or acute liver failure (n=30). CD14+CD15−CD11b+HLA-DR− cells were identified as per definition of M-MDSC and detailed immunophenotypic analyses were performed. Suppression of T cell activation was assessed by mixed lymphocyte reaction. Assessment of innate immune function included cytokine expression in response to Toll-like receptor (TLR-2, TLR-4 and TLR-9) stimulation and phagocytosis assays using flow cytometry and live cell imaging-based techniques. Results Circulating CD14+CD15−CD11b+HLA-DR− M-MDSCs were markedly expanded in patients with ACLF (55% of CD14+ cells). M-MDSC displayed immunosuppressive properties, significantly decreasing T cell proliferation (p=0.01), producing less tumour necrosis factor-alpha/interleukin-6 in response to TLR stimulation (all p<0.01), and reduced bacterial uptake of Escherichia coli (p<0.001). Persistently low expression of HLA-DR during disease evolution was linked to secondary infection and 28-day mortality. Recurrent TLR-2 and TLR-4 stimulation expanded M-MDSC in vitro. By contrast, TLR-3 agonism reconstituted HLA-DR expression and innate immune function ex vivo. Conclusion Immunosuppressive CD14+HLA-DR− M-MDSCs are expanded in patients with ACLF. They were depicted by suppressing T cell function, attenuated antimicrobial innate immune responses, linked to secondary infection, disease severity and prognosis. TLR-3 agonism reversed M-MDSC expansion and innate immune function and merits further evaluation as potential immunotherapeutic agent.

Understanding infection susceptibility in patients with acute-on-chronic liver failure.

Infection accounts for over 50 % of admissions of cirrhotic patients to hospital and is the main precipitant for the development of multiple organ dysfunction syndrome (MODS), including hepatic encephalopathy, renal, respiratory and circulatory failure, a syndrome referred to as acute-on-chronic liver failure (AoCLF) [1]. Once established, AoCLF carries a prohibitively high 30-day mortality rate in excess of 25 % [2]. In recent census studies from Europe and USA [1, 2], 35 % of patients admitted with AoCLF require organ support, posing a significant burden on critical care services and resources. Despite advances in organ support, little progress has been made in understanding the pathogenesis and treatment of infections in patients with liver disease.

Balanced haemostasis with both hypo- and hyper-coagulable features in critically ill patients with acute-on-chronic-liver failure

Background: Cirrhotic patients have complex haemostatic abnormalities. Current evidence suggests stable cirrhotic (SC) patients have a “re‐balanced” haemostatic state. However, limited data exists in acute decompensated (AD) or acute on chronic liver failure (ACLF) patients. Methods: We utilised thrombin generation analysis, fibrinolysis assessment, and evaluation of haemostatic parameters to assess haemostasis in liver disease of progressive severity. Results: The study cohorts were comprised of: SC, n = 8; AD n = 44; ACLF, n = 17; and Healthy Control (HC), n = 35. There was a progressive increase across the cohorts in INR (p = 0.0001), Factor VIII (p = 0.0001) and VWF levels (p = 0.0001) and a correspondingly decrease in anti‐thrombin (p = 0.0001), ADAMTS‐13 (p = 0.01) and fibrinogen levels (p = 0.0001). In the presence of thrombomodulin, thrombin generation was equivalent or significantly higher in all the cohorts compared to HC (p = 0.0001). Compared to AD, ACLF had a lower ETP (p = 0.002) and thrombin peak (p = 0.0001). There was no significant difference across the cohorts in clot lysis time (p = 0.07), although compared to HC, AD had a significantly shorter lysis time (p = 0.001). Conclusions: Our cohorts, despite significant differences in haemostatic parameters, displayed intact thrombin generation but progressive hypo‐functional clot stability and potentially but not universal hyper‐functional haemostasis. Highlights:Haemostatic parameters change progressively from stable cirrhosis to acute‐on‐chronic liver failure.Thrombin generation remains intact in acute decompensated and acute‐on‐chronic liver failure.Clot stability varied significantly between the cohorts.Acute‐on‐chronic liver failure appears to have a separate haemostatic phenotype.

Increased Expression of Cytotoxic T-Lymphocyte−Associated Protein 4 by T Cells, Induced by B7 in Sera, Reduces Adaptive Immunity in Patients With Acute Liver Failure

Background & Aims Patients with acute liver failure (ALF) have defects in innate immune responses to microbes (immune paresis) and are susceptible to sepsis. Cytotoxic T-lymphocyte−associated protein 4 (CTLA4), which interacts with the membrane receptor B7 (also called CD80 and CD86), is a negative regulator of T-cell activation. We collected T cells from patients with ALF and investigated whether inhibitory signals down-regulate adaptive immune responses in patients with ALF. Methods We collected peripheral blood mononuclear cells from patients with ALF and controls from September 2013 through September 2015 (45 patients with ALF, 20 patients with acute-on-chronic liver failure, 15 patients with cirrhosis with no evidence of acute decompensation, 20 patients with septic shock but no cirrhosis or liver disease, and 20 healthy individuals). Circulating CD4+ T cells were isolated and analyzed by flow cytometry. CD4+ T cells were incubated with antigen, or agonist to CD3 and dendritic cells, with or without antibody against CTLA4; T-cell proliferation and protein expression were quantified. We measured levels of soluble B7 molecules in supernatants of isolated primary hepatocytes, hepatic sinusoidal endothelial cells, and biliary epithelial cells from healthy or diseased liver tissues. We also measured levels of soluble B7 serum samples from patients and controls, and mice with acetaminophen-induced liver injury using enzyme-linked immunosorbent assays. Results Peripheral blood samples from patients with ALF had a higher proportion of CD4+ CTLA4+ T cells than controls; patients with infections had the highest proportions. CD4+ T cells from patients with ALF had a reduced proliferative response to antigen or CD3 stimulation compared to cells from controls; incubation of CD4+ T cells from patients with ALF with an antibody against CTLA4 increased their proliferative response to antigen and to CD3 stimulation, to the same levels as cells from controls. CD4+ T cells from controls up-regulated expression of CTLA4 after 24−48 hours culture with sera from patients with ALF; these sera were found to have increased concentrations of soluble B7 compared to sera from controls. Necrotic human primary hepatocytes exposed to acetaminophen, but not hepatic sinusoidal endothelial cells and biliary epithelial cells from patients with ALF, secreted high levels of soluble B7. Sera from mice with acetaminophen-induced liver injury contained high levels of soluble B7 compared to sera from mice without liver injury. Plasma exchange reduced circulating levels of soluble B7 in patients with ALF and expression of CTLA4 on T cells. Conclusions Peripheral CD4+ T cells from patients with ALF have increased expression of CTLA4 compared to individuals without ALF; these cells have a reduced response to antigen and CD3 stimulation. We found sera of patients with ALF and from mice with liver injury to have high concentrations of soluble B7, which up-regulates CTLA4 expression by T cells and reduces their response to antigen. Plasma exchange reduces levels of B7 in sera from patients with ALF and might be used to restore antimicrobial responses to patients.

Transplantation for the Very Sick Patient—Donor and Recipient Factors

Purpose of ReviewPatients with end-stage chronic liver disease (CLD) frequently deteriorate with development of a ‘frail’ state and progress to development of extra-hepatic organ failures. Making the clinical judgement when a patient is ‘too sick’ to proceed with liver transplantation is challenging; we summarise recent studies that may inform decision-making.Recent FindingsReports quantify the impact of both recipient and donor factors related to early post-transplant survival, with consistent adverse association of advanced donor and recipient age, under-nutrition, sarcopenia and impaired functional status, and requirement for critical care unit admission and organ support. Survival for ‘very sick’ recipients has shown progressive improvement over time, but evidence is increasing of much greater post-transplant resource use and cost.SummaryFactors associated with adverse outcomes in very sick recipients are becoming more clearly delineated. The increasing success of transplantation and exploration of its extended use must be tempered by the recognition of the consequences upon constrained clinical resources.

P1294 : Proof-of-principle evaluation of immunomodulatory drugs in promoting phagocytosis capacity in patients with liver failure

6.6ng/ml respectively at 15 days post-LT. 20 (16%) on basiliximab had ACR vs 72/188 (38%) controls (p = 0.001). Of those not on basiliximab, 35% (35/97) with TAC levels >5ng/ml vs 41% (38/92) with TAC 50 years (p 5ng/ml at week 1 post-LT (p = 0.003, OR=0.4, 95%CI 0.3–0.7) and mean CNI levels >7ng/ml up to 15 days postLT (p = 0.019, OR=0.85, 95%CI 0.75–0.97). The same factors were associated with mild renal impairment (eGFR 7ng/ml at day 15 post-LT (p = 0.003, OR=1.9, 95%CI 1.2–2.9) and use of steroids >3 months post-LT (p = 0.006, OR=0.7, 95%CI 0.5–0.9). Conclusions: Basiliximab use allows reduced TAC trough levels with less episodes of ACR compared with the control group. However, TAC trough levels <7ng/ml at 15 days post-LT regardless of basiliximab use, were protective of renal function without predisposing to ACR in patients with renal impairment at baseline.