ABSTRACT
Chronic infection by hepatitis B virus (HBV) is the major contributor to liver disease worldwide. Though HBV replicates via a nuclear episomal DNA (covalently closed circular DNA [cccDNA]), integration of HBV DNA into the host cell genome is regularly observed in the liver in infected patients. While reported as a prooncogenic alteration, the mechanism(s) and timing of HBV DNA integration are not well understood, chiefly due to the lack of in vitro infection models that have detectable integration events. In this study, we have established an in vitro system in which integration can be reliably detected following HBV infection. We measured HBV DNA integration using inverse nested PCR in primary human hepatocytes, HepaRG-NTCP, HepG2-NTCP, and Huh7-NTCP cells after HBV infection. Integration was detected in all cell types at a rate of >1 per 10,000 cells, with the most consistent detection in Huh7-NTCP cells. The integration rate remained stable between 3 and 9 days postinfection. HBV DNA integration was efficiently blocked by treatment with a 200 nM concentration of the HBV entry inhibitor Myrcludex B, but not with 10 µM tenofovir, 100 U of interferon alpha, or a 1 µM concentration of the capsid assembly inhibitor GLS4. This suggests that integration of HBV DNA occurs immediately after infection of hepatocytes and is likely independent of de novo HBV genome replication in this model. Site analysis revealed that HBV DNA integrations were distributed over the entire human genome. Further, integrated HBV DNA sequences were consistent with double-stranded linear HBV DNA being the major precursor. Thus, we have established an in vitro system to interrogate the mechanisms of HBV DNA integration.IMPORTANCE Hepatitis B virus (HBV) is a common blood-borne pathogen and, following a chronic infection, can cause liver cancer and liver cirrhosis. Integration of HBV DNA into the host genome occurs in all known members of the Hepadnaviridae family, despite this form not being necessary for viral replication. HBV DNA integration has been reported to drive liver cancer formation and persistence of virus infection. However, when and the mechanism(s) by which HBV DNA integration occurs are not clear. In this study, we have developed and characterized an in vitro system to reliably detect HBV DNA integrations that result from a true HBV infection event and that closely resemble those found in patient tissues. Using this model, we showed that integration occurs when the infection is first established. Importantly, we provide here a system to analyze molecular factors involved in HBV integration, which can be used to develop strategies to halt its formation.
Subject(s)
DNA, Viral/metabolism , Hepatitis B virus/physiology , Hepatocytes/virology , Organic Anion Transporters, Sodium-Dependent/metabolism , Symporters/metabolism , Cell Line , Hep G2 Cells , Hepatocytes/cytology , Hepatocytes/microbiology , Humans , Models, Biological , Virus Integration , Virus Internalization , Virus ReplicationABSTRACT
Despite the growing data supporting the role of microcirculation in regulating liver function, little of this knowledge has been translated into clinical practice. The aim of this study is to quantify hepatic microcirculation in vivo using sidestream dark field (SDF) imaging and correlate these findings with hepatic blood flow, hemodynamic parameters, and soluble mediators. Postreperfusion hepatic microcirculation was assessed using SDF imaging. Hepatic microcirculation measurements included functional sinusoidal density (cm/cm2 ), sinusoidal diameter (µm), red blood cell velocity (µm/second), volumetric blood flow (pl/second), and flow heterogeneity (FH) index. The serum concentrations of endothelin 1 (ET-1) and other inflammatory markers were analyzed with Luminex technology. Portal venous and hepatic artery flows were measured using a flowmeter. Twenty-eight patients undergoing cadaveric liver transplantations have been included in this study. Early allograft dysfunction (EAD) occurred in 7 (25%) patients and was associated with microcirculatory dysfunction. Low arterial and portal flow, high dose of inotropes, cold ischemia time, steatosis, and high ET-1 levels were all associated with impaired microcirculation. The time interval between portal venous and hepatic arterial reperfusion significantly correlated with the changes of the liver grafts' microcirculation. EAD patients tended to have higher serum levels of ET-1 on postoperative days 1, 2, 5, and 7 (all P < 0.01). Serum levels of ET-1 correlated significantly with microcirculation parameters. In conclusion, postreperfusion hepatic microcirculation is a determinant of organ dysfunction after liver reperfusion and could be used to identify very early patients at risk of EAD. Liver Transplantation 23 527-536 2017 AASLD.
Subject(s)
Diagnostic Imaging/methods , End Stage Liver Disease/surgery , Hemodynamics , Liver Circulation , Liver Transplantation/adverse effects , Microcirculation , Primary Graft Dysfunction/physiopathology , Adult , Aged , Allografts/blood supply , Biomarkers/blood , Blood Flow Velocity , Endothelin-1/blood , Female , Hepatic Artery/physiopathology , Humans , Liver/blood supply , Liver/diagnostic imaging , Male , Middle Aged , Portal Vein/physiopathology , Primary Graft Dysfunction/blood , Primary Graft Dysfunction/epidemiology , Primary Graft Dysfunction/etiology , Reperfusion/adverse effects , Severity of Illness IndexABSTRACT
Complications of end-stage chronic liver disease signify a major cause of mortality worldwide. Irrespective of the underlying cause, most chronic liver diseases are characterized by hepatocellular necrosis, inflammation, fibrosis, and proliferation of liver progenitor cells or ductular reactions. Vast differences exist between experimental models that mimic these processes, and their identification is fundamental for translational research. We compared two common murine models of chronic liver disease: the choline-deficient, ethionine-supplemented (CDE) diet versus thioacetamide (TAA) supplementation. Markers of liver injury, including serum alanine transaminase levels, apoptosis, hepatic fat loading, and oxidative stress, as well as inflammatory, fibrogenic and liver progenitor cell responses, were assessed at days 3, 7, 14, 21, and 42. This study revealed remarkable differences between the models. It identified periportal injury and fibrosis with an early peak and slow normalization of all parameters in the CDE regimen, whereas TAA-treated mice had pericentral patterns of progressive injury and fibrosis, resulting in a more severe hepatic injury phenotype. This study is the first to resolve two different patterns of injury and fibrosis in the CDE and TAA model and to indisputably identify the fibrosis pattern in the TAA model as driven from the pericentral vein region. Our data provide a valuable foundation for future work using the CDE and TAA regimens to model a variety of human chronic liver diseases.
Subject(s)
Disease Models, Animal , Hepatocytes/physiology , Liver Diseases/physiopathology , Stem Cells/physiology , Animals , Chronic Disease , Hepatocytes/pathology , Immunohistochemistry , Male , Mice , Mice, Inbred C57BL , Real-Time Polymerase Chain Reaction , Stem Cells/pathologyABSTRACT
BACKGROUND AND AIM: The glycoprotein CD147 has a role in tumor progression, is readily detectable in the circulation, and is abundantly expressed in hepatocellular carcinoma (HCC). Advanced HCC patients are a heterogeneous group with some individuals having dismal survival. The aim of this study was to examine circulating soluble CD147 levels as a prognostic marker in HCC patients. METHODS: CD147 was measured in 277 patients (110 HCC, 115 chronic liver disease, and 52 non-liver disease). Clinical data included etiology, tumor progression, Barcelona Clinic Liver Cancer (BCLC) stage, and treatment response. Patients with HCC were stratified into two groups based upon the 75th percentile of CD147 levels (24 ng/mL). RESULTS: CD147 in HCC correlated inversely with poor survival (P = 0.031). Increased CD147 predicted poor survival in BCLC stages C and D (P = 0.045), and CD147 levels >24 ng/mL predicted a significantly diminished 90-day and 180-day survival time (hazard ratio [HR] = 6.1; 95% confidence interval [CI]: 2.1-63.2; P = 0.0045 and HR = 2.8; 95% CI: 1.2-12.6; P = 0.028, respectively). In BCLC stage C, CD147 predicted prognosis; levels >24 ng/mL were associated with a median survival of 1.5 months compared with 6.5 months with CD147 levels ≤24 ng/mL (P = 0.03). CD147 also identified patients with a poor prognosis independent from treatment frequency, modality, and tumor size. CONCLUSIONS: Circulating CD147 is an independent marker of survival in advanced HCC. CD147 requires further evaluation as a potential new prognostic measure in HCC to identify patients with advanced disease who have a poor prognosis.
Subject(s)
Basigin/blood , Biomarkers, Tumor/blood , Carcinoma, Hepatocellular/diagnosis , Liver Neoplasms/diagnosis , Aged , Carcinoma, Hepatocellular/mortality , Disease Progression , Female , Humans , Liver Neoplasms/mortality , Male , Middle Aged , Predictive Value of Tests , Prognosis , Survival Rate , Time FactorsABSTRACT
In recent years, the global burden of obesity and diabetes has seen a parallel rise in other metabolic complications, such as non-alcoholic fatty liver disease (NAFLD). This condition, once thought to be a benign accumulation of hepatic fat, is now recognized as a serious and prevalent disorder that is conducive to inflammation and fibrosis. Despite the rising incidence of NAFLD, there is currently no reliable method for its diagnosis or staging besides the highly invasive tissue biopsy. This limitation has resulted in the study of novel circulating markers as potential candidates, one of the most popular being extracellular vesicles (EVs). These submicron membrane-bound structures are secreted from stressed and activated cells, or are formed during apoptosis, and are known to be involved in intercellular communication. The cargo of EVs depends upon the parent cell and has been shown to be changed in disease, as is their abundance in the circulation. The role of EVs in immunity and epigenetic regulation is widely attested, and studies showing a correlation with disease severity have made these structures a favorable target for diagnostic as well as therapeutic purposes. This review will highlight the research that is available on EVs in the context of NAFLD, the current limitations, and projections for their future utility in a clinical setting.
Subject(s)
Biomarkers/blood , Extracellular Vesicles/metabolism , Non-alcoholic Fatty Liver Disease/diagnosis , Animals , Early Diagnosis , HumansABSTRACT
Hepatic artery stenosis (HAS) is thought to predispose patients to biliary complications secondary to ischemic injury. Despite this, the clinical significance of HAS remains poorly defined. The aims of this study were to determine the prevalence and outcomes of HAS and to define which patients might benefit from endovascular treatment. From a prospective database of 662 adult patients undergoing liver transplantation between 2000 and 2011, we identified 54 patients who developed HAS. HAS was defined as any stenosis > 70% that was seen during multidetector computed tomographic angiography (MDCTA) or digital subtraction angiography. The benefit of endovascular therapy was evaluated with propensity score matching. New biliary complications occurred in 17 patients (31.4%), and 23 of the 54 study patients with HAS received endovascular treatment. Among the propensity score-matched patients, the biliary stricture-free survival time was significantly longer for those who received endovascular therapy (P = 0.03). An incidental diagnosis (P = 0.07) and a time from transplantation > 6 months (P = 0.021) were associated with a reduced risk of developing biliary stricture. Patients with symptomatic HAS who received treatment had better biliary stricture-free survival than patients who were treated conservatively, although no significant difference was recorded (P = 0.11). No patient with asymptomatic HAS and normal liver function tests developed biliary strictures. In conclusion, HAS intervention was associated with improved biliary stricture-free survival. In patients with late-onset HAS (≥6 months) and asymptomatic patients, endovascular treatment is not warranted.
Subject(s)
Constriction, Pathologic/therapy , Endovascular Procedures , Hepatic Artery/physiopathology , Liver/physiology , Adult , Angiography, Digital Subtraction , Bile Ducts/physiopathology , Constriction, Pathologic/etiology , Databases, Factual , Female , Humans , Ischemia/pathology , Liver Transplantation/adverse effects , Male , Middle Aged , Prospective Studies , Time Factors , Tomography, X-Ray ComputedABSTRACT
Although chronic hepatitis B virus (HBV) infection is a known risk factor for the development of hepatocellular carcinoma (HCC), the steps involved in the progression from normal liver to HCC are poorly understood. In this review, we apply five conceptual models, previously proposed by Vineis et al. to explain carcinogenesis in general, to explore the possible steps involved in the initiation and evolution of HBV-associated HCC. Available data suggest that the most suitable and inclusive model is based on evolution of hepatocyte subpopulations. In this evolutionary model, HCC-associated changes are driven by selection and subsequent clonal expansion of phenotypically altered hepatocyte subpopulations in the microenvironment of the HBV-infected liver. This model can incorporate the wide range of mechanisms proposed to play a role in the initiation of HCC including oncogenic HBV proteins, integration of HBV DNA and chronic inflammation of the liver. The model may assist in the early prevention, detection and treatment of HCC and may guide future studies of the initiation of HBV-associated HCC.
Subject(s)
Carcinoma, Hepatocellular/virology , Cell Transformation, Viral , Hepatitis B virus/pathogenicity , Hepatitis B, Chronic/virology , Liver Neoplasms/virology , Models, Biological , Animals , Biological Evolution , Biomarkers, Tumor/genetics , Biomarkers, Tumor/metabolism , Carcinoma, Hepatocellular/genetics , Carcinoma, Hepatocellular/metabolism , Carcinoma, Hepatocellular/pathology , Epigenesis, Genetic , Gene Expression Regulation, Neoplastic , Gene Expression Regulation, Viral , Genetic Predisposition to Disease , Hepatitis B virus/genetics , Hepatitis B virus/growth & development , Hepatitis B, Chronic/complications , Hepatitis B, Chronic/diagnosis , Humans , Liver Neoplasms/genetics , Liver Neoplasms/metabolism , Liver Neoplasms/pathology , Mutation , Tumor Microenvironment , Virus ReplicationABSTRACT
Chronic liver disease causes significant morbidity and mortality through progressive fibrosis, cirrhosis, and liver cancer. The classical theory of fibrogenesis has hepatic stellate cells (HSCs) as the principal and only significant source of abnormal extracellular matrix (ECM). Further, HSCs have the major role in abnormal ECM turnover. It is the death of hepatocytes, as the initial target of injury, that initiates a sequence of events including the recruitment of inflammatory cells and activation of HSCs. Following this initial response, the ongoing insult to hepatocytes is regarded as perpetuating injury, but otherwise, hepatocytes are regarded as "victims" and "bystanders" in progressive fibrosis. Recent developments, however, challenge this view and suggest the concept of the hepatocyte being an active participant in liver injury. It is clear now that hepatocytes undergo phenotypic changes, adapt to injury, and react to the altered microenvironment. In this review, we describe studies showing that hepatocytes contribute to progressive fibrosis by direct manipulation of the surrounding ECM and through signaling to effector cells, particularly HSCs and intrahepatic immune cells. Together, these findings suggest an active "accomplice" role for the hepatocyte in progressive liver fibrosis and highlight novel pathways that could be targeted for development of future anti-fibrotic therapies.
Subject(s)
Hepatocytes/physiology , Liver Diseases/etiology , Cell Death , Disease Progression , Extracellular Matrix , Hepatocytes/immunology , Hepatocytes/pathology , Humans , Liver Cirrhosis/etiology , Liver Cirrhosis/immunology , Liver Cirrhosis/pathology , Liver Diseases/immunology , Liver Diseases/pathology , Liver RegenerationABSTRACT
Fibrosis in livers with hepatitis C virus (HCV) recurrence after liver transplantation (LT) can be rapidly progressive, and the mechanisms underlying this process are poorly understood. In livers with HCV infections in the non-LT setting, there is a significant relationship between the development of structures known as the ductular reaction (DR), hepatic progenitor cells (HPCs), and fibrosis. This study characterizes the DR, HPCs, and fibrosis associated with HCV recurrence after LT. Immunohistochemistry and confocal microscopy were used to characterize the DR, HPC, and fibrosis in liver biopsy specimens. Key findings were confirmed in a separate, independent cohort. The initial characterization cohort had 194 biopsy samples from 105 individuals with HCV recurrence after LT. The immunophenotype, morphology, and location of the DR were consistent with an HPC origin. The DR correlated with intrahepatic fibrosis (rs = 0.529, P < 0.001) and the number of activated hepatic stellate cells (HSCs; rs = 0.446, P < 0.001). There was an early occurrence of hepatocyte replicative arrest as well as increased hepatocyte proliferation that correlated with the DR (rs = 0.295, P < 0.001). Replicative arrest preceded hepatocyte proliferation in early-stage injury. Hepatocyte proliferation decreased with advanced fibrosis; in contrast, the extent of the DR and the number of activated HSCs continued to increase. In the second cohort of 37 individuals, the DR and the number of HPCs similarly correlated with fibrosis and inflammation after LT. In conclusion, this is the first characterization of the DR in HCV-associated liver injury after LT. There was a significant correlation between the DR and the development of progressive fibrosis in HCV recurrence. These results suggest a pivotal role for both the DR and the HPC responses in the aggressive fibrosis seen with HCV recurrence after LT.
Subject(s)
Hepatic Duct, Common/virology , Hepatitis C/surgery , Hepatitis C/virology , Liver Failure/virology , Liver Transplantation , Aged , Cell Proliferation , Disease Progression , Female , Fibrosis/complications , Hepacivirus , Hepatic Duct, Common/pathology , Hepatocytes/cytology , Humans , Immunohistochemistry , Immunophenotyping , Inflammation , Liver Failure/therapy , Liver Transplantation/adverse effects , Male , Microscopy, Confocal , Microscopy, Fluorescence , Middle Aged , Recurrence , Retrospective Studies , Stem Cells/cytology , Treatment OutcomeABSTRACT
PURPOSE OF REVIEW: Improved understanding of the pathophysiology of fibrosis and recent technological advances have resulted in the development of several serum biomarkers and imaging tools as noninvasive alternatives to biopsy. Most of these markers have been developed in chronic hepatitis C (CHC) and nonalcoholic fatty liver disease (NAFLD) patients. This review highlights some of the recent advances and potential clinical application of these modalities. RECENT FINDINGS: Many noninvasive approaches initially developed for binary disease staging in CHC continue to be refined for diagnostic use in other chronic liver disease such as NAFLD. A combination of serum markers and imaging tools appears useful in reducing the need for biopsy for the diagnosis of cirrhosis, and providing functional assessment in advanced stage disease. Cytokeratin-18 fragments, controlled attenuation parameter (CAP), real-time elastography, and magnetic resonance imaging approaches appear promising for NAFLD, but require further validation. SUMMARY: Current noninvasive tests of fibrosis provide good diagnostic and prognostic utility for advanced stage liver disease, and have been adapted into clinical practice for CHC. Reliable biomarkers for steatosis, nonalcoholic steatohepatitis, and assessment of fibrosis progression in chronic liver disease are still required. Continued advances in bioimaging and functional genomics will be important for quantitative assessment of fibrosis and future biomarker development.
Subject(s)
Biomarkers/blood , Liver Cirrhosis/diagnosis , Biopsy , Disease Progression , Elasticity Imaging Techniques/methods , Humans , Liver/pathology , Non-alcoholic Fatty Liver Disease/diagnosis , PrognosisABSTRACT
BACKGROUND & AIMS: Obese Alms1 mutant (foz/foz) NOD.B10 mice develop diabetes and fibrotic NASH when fed high-fat(HF) diet. To establish whether diabetes or obesity is more closely associated with NASH fibrosis, we compared diabetic foz/foz C57BL6/J with non-diabetic foz/foz BALB/c mice. We also determined hepatic cytokines, growth factors and related profibrotic pathways. METHODS: Male and female foz/foz BALB/c and C57BL6/J mice were fed HF or chow for 24 weeks before determining metabolic indices, liver injury, cytokines, growth factors, pathology/fibrosis and matrix deposition pathways. RESULTS: All foz/foz mice were obese. Hepatomegaly, hyperinsulinemia, hyperglycaemia and hypoadiponectinaemia occurred only in foz/foz C57BL6/J mice, whereas foz/foz BALB/c formed more adipose. Serum ALT, steatosis, ballooning, liver inflammation and NAFLD activity score were worse in C57BL6/J mice. In HF-fed mice, fibrosis was severe in foz/foz C57BL6/J, appreciable in WT C57BL6/J, but absent in foz/foz BALB/c mice. Hepatic mRNA expression of TNF-α, IL-12, IL-4, IL-10 was increased (but not IFN-γ, IL-1ß, IL-17A), and IL-4:IFN-γ ratio (indicating Th-2 predominance) was higher in HF-fed foz/foz C57BL6/J than BALB/c mice. In livers of HF-fed foz/foz C57BL6/J mice, TGF-ß was unaltered but PDGFα and CTGF were increased in association with enhanced α-SMA, CD147and MMP activity. CONCLUSIONS: In mice with equivalent genetic/dietary obesity, NASH development is linked to strain differences in hyperinsulinaemia and hyperglycaemia inversely related to lipid partitioning between adipose and liver. Diabetes-mediated CTGF-regulation of MMPs as well as cytokines/growth factors (Th-2 cytokine predominant, PDGFα, not TGF-ß) mobilized in the resultant hepatic necroinflammatory change may contribute to strain differences in NASH fibrosis.
Subject(s)
DNA-Binding Proteins/genetics , Diabetes Mellitus, Type 2/etiology , Diet, High-Fat/adverse effects , Liver Cirrhosis/etiology , Non-alcoholic Fatty Liver Disease/etiology , Analysis of Variance , Animals , Cell Cycle Proteins , Cytokines/metabolism , Diabetes Mellitus, Type 2/metabolism , Female , Fluorescent Antibody Technique , Intercellular Signaling Peptides and Proteins/metabolism , Liver/metabolism , Liver/pathology , Liver Cirrhosis/pathology , Male , Matrix Metalloproteinases/metabolism , Mice , Mice, Inbred BALB C , Mice, Inbred C57BL , Mice, Inbred NOD , Non-alcoholic Fatty Liver Disease/pathology , Species SpecificityABSTRACT
Hepatocellular carcinoma (HCC) is a prevalent primary liver cancer that is derived from hepatocytes and is characterised by high mortality rate and poor prognosis. While HCC is driven by cumulative changes in the hepatocyte genome, it is increasingly recognised that the liver microenvironment plays a pivotal role in HCC propensity, progression and treatment response. The microenvironmental stimuli that have been recognised as being involved in HCC pathogenesis are diverse and include intrahepatic cell subpopulations, such as immune and stellate cells, pathogens, such as hepatitis viruses, and non-cellular factors, such as abnormal extracellular matrix (ECM) and tissue hypoxia. Recently, a number of novel environmental influences have been shown to have an equally dramatic, but previously unrecognized, role in HCC progression. Novel aspects, including diet, gastrointestinal tract (GIT) microflora and circulating microvesicles, are now being recognized as increasingly important in HCC pathogenesis. This review will outline aspects of the HCC microenvironment, including the potential role of GIT microflora and microvesicles, in providing new insights into tumourigenesis and identifying potential novel targets in the treatment of HCC.
Subject(s)
Carcinoma, Hepatocellular/pathology , Liver Neoplasms/pathology , Liver/pathology , Tumor Microenvironment , Animals , Carcinoma, Hepatocellular/genetics , Carcinoma, Hepatocellular/immunology , Carcinoma, Hepatocellular/therapy , DNA Damage , Disease Progression , Gastrointestinal Tract/microbiology , Humans , Inflammation/complications , Inflammation/immunology , Liver/immunology , Liver/metabolism , Liver Neoplasms/genetics , Liver Neoplasms/immunology , Liver Neoplasms/therapy , Signal TransductionABSTRACT
BACKGROUND AND AIM: Metabolic dysfunction-associated steatohepatitis (MASH) is a metabolic disorder with limited treatment options. The thyroid hormone receptor (THR)-ß agonist resmetirom/MGL-3196 (MGL) increases liver fat oxidation and has been approved for treating adult MASH. However, over 60% of patients receiving MGL treatment do not achieve MASH resolution. Therefore, we investigated the potential for combination therapy of MGL with the mitochondrial uncoupler BAM15 to improve fatty liver disease outcomes in the GAN mouse model of MASH. METHODS: C57BL/6J male mice were fed GAN diet for 38 weeks before stratification and randomization to treatments including MGL, BAM15, MGL + BAM15, or no drug control for 8 weeks. Treatments were admixed in diet and mice were pair-fed to control for drug intake. Treatment effectiveness was assessed by body weight, body composition, energy expenditure, glucose tolerance, tissue lipid content, and histological analyses. RESULTS: MGL + BAM15 treatment resulted in better efficacy versus GAN control mice than either monotherapy in the context of energy expenditure, liver fat loss, glucose control, and fatty liver disease activity score. Improvements in ALT, liver mass, and plasma cholesterol were primarily driven by MGL, while improvements in body fat were primarily driven by BAM15. No treatments altered liver fibrosis. CONCLUSIONS: MGL + BAM15 treatment had overall better efficacy to improve metabolic outcomes in mice fed GAN diet than either monotherapy alone. These data warrant further investigation into combination therapies of THR-ß agonists and mitochondrial uncouplers for the potential treatment of disorders related to fatty liver, obesity, and insulin resistance.
ABSTRACT
Discoidin domain receptor 1 (DDR1) is a receptor tyrosine kinase that binds and is activated by collagens. Transcriptional profiling of cirrhosis in human liver using a DNA array and quantitative PCR detected elevated mRNA expression of DDR1 compared with that in nondiseased liver. The present study characterized DDR1 expression in cirrhotic and nondiseased human liver and examined the cellular effects of DDR1 expression. mRNA expression of all five isoforms of DDR1 was detected in human liver, whereas DDR1a demonstrated differential expression in liver with hepatitis C virus and primary biliary cirrhosis compared with nondiseased liver. In addition, immunoblot analysis detected shed fragments of DDR1 more readily in cirrhotic liver than in nondiseased liver. Inasmuch as DDR1 is subject to protease-mediated cleavage after prolonged interaction with collagen, this differential expression may indicate more intense activation of DDR1 protein in cirrhotic compared with nondiseased liver. In situ hybridization and immunofluorescence localized intense DDR1 mRNA and protein expression to epithelial cells including hepatocytes at the portal-parenchymal interface and the luminal aspect of the biliary epithelium. Overexpression of DDR1a altered hepatocyte behavior including increased adhesion and less migration on extracelular matrix substrates. DDR1a regulated extracellular expression of matrix metalloproteinases 1 and 2. These data elucidate DDR1 function pertinent to cirrhosis and indicate the importance of epithelial cell-collagen interactions in chronic liver injury.
Subject(s)
Liver Cirrhosis/enzymology , Liver Cirrhosis/pathology , Liver/enzymology , Liver/pathology , Receptor Protein-Tyrosine Kinases/metabolism , Adolescent , Adult , Cell Adhesion , Cell Line , Cell Movement , Discoidin Domain Receptor 1 , Female , Gene Expression Regulation , Green Fluorescent Proteins/metabolism , Humans , Immunohistochemistry , Male , Matrix Metalloproteinases/metabolism , Middle Aged , Protein Isoforms/chemistry , Protein Isoforms/genetics , Protein Isoforms/metabolism , RNA, Messenger/genetics , RNA, Messenger/metabolism , Receptor Protein-Tyrosine Kinases/chemistry , Receptor Protein-Tyrosine Kinases/genetics , Recombinant Fusion Proteins/metabolism , Transfection , Young AdultABSTRACT
BACKGROUND: A CD antibody microarray has been previously developed allowing semi-quantitative identification of greater than 80 CD antigens on circulating leucocytes from peripheral blood samples. This assay, which uses a live cell-capture technique, enables an extensive leucocyte immunophenotype determination in a single analysis and to date this has been used successfully to characterise diseases including human leukaemias and HIV infection. AIMS: To determine CD antigen expression profiles for patients with various liver diseases and to look for preserved disease-specific signatures. METHODS: Three liver disease groups including hepatitis C (HCV) (n = 35), non-alcoholic steatohepatitis (NASH) (n = 21) and alcohol-related liver disease (n = 14) were compared with a normal group (n = 23). Hierarchal Clustering (HCL) and Principal Component Analysis (PCA) of the data revealed distinct binding patterns for patients with and without cirrhosis. RESULTS: Patients with cirrhosis and portal hypertension compared with those without cirrhosis had significantly reduced expression of several markers of T-cell function including CD45, CD8, CD28 and TCR α/ß. Disease prediction algorithms based on the expression data were able to discriminate cirrhotics from non-cirrhotics with 71% overall success, which improved to 77% when only patients with HCV were considered. CONCLUSIONS: These results demonstrate disease-specific consensus patterns of expression of CD antigens for patients with chronic liver disease, suggesting that the CD antibody array is a promising tool in the analysis of human liver disease, and with further refinement may have future research and clinical utility.
Subject(s)
Algorithms , Antibodies , Antigens, CD/metabolism , Immunophenotyping/methods , Liver Diseases/diagnosis , Protein Array Analysis/methods , Adult , Aged , Aged, 80 and over , Antibodies/metabolism , Cluster Analysis , Female , Humans , Leukocytes/metabolism , Male , Middle Aged , Principal Component AnalysisABSTRACT
BACKGROUND AND AIM: Donor liver steatosis can impact on liver allograft outcomes. The aim of the present study was to comprehensively report on the impact of type and grade of donor steatosis, as well as donor and recipient factors, including the reported Donor Risk Index (DRI), on liver allograft outcomes. METHODS: A review of unit data for all adult liver transplant procedures from 2001 to 2007, as well as donor offers. Donor liver biopsies were regraded for steatosis by an experienced histopathologist. RESULTS: Steatosis was detected in 184/255 (72%) of biopsies, of which 114 (62%) had microvesicular steatosis (MiS; 68 mild, 22 moderate, 24 severe) and 70 (38%) macrovesicular steatosis (MaS; 59 mild, 7 moderate, 4 severe). The majority (66/70, 94%) of biopsies with MaS also contained MiS. Allograft steatosis was associated with increasing donor body mass index (P = 0.000), plus donor male sex (P < 0.05). Primary non function (P = 0.002), early renal failure (P = 0.040), and requirement for retransplantation (P = 0.012) were associated only with severe MaS. Early biliary complications were associated with moderate MaS (P = 0.039). Only severe MaS was significantly associated with inferior allograft survival at 3 months (relative risk = 12.09 [8.75-19.05], P = 0.000) and 1 year (P = 0.000). CONCLUSIONS: MiS is a common finding and frequently coexists with MaS on liver allograft biopsy, while isolated MaS is uncommon. Only the presence of moderate to severe MaS is associated with inferior early allograft outcomes. The impact of severe MaS on allograft survival appears greater than other donor factors, including the calculated DRI.
Subject(s)
Fatty Liver/complications , Fatty Liver/pathology , Graft Survival , Liver Transplantation/physiology , Renal Insufficiency/etiology , Adolescent , Adult , Aged , Analysis of Variance , Body Mass Index , Fatty Liver/classification , Female , Graft Survival/physiology , Humans , Kaplan-Meier Estimate , Liver Transplantation/adverse effects , Male , Middle Aged , Proportional Hazards Models , Reoperation , Severity of Illness Index , Sex Factors , Young AdultABSTRACT
Liver cancer has variable incidence worldwide and high mortality. Histologically, the most common subtype of liver cancer is hepatocellular carcinoma (HCC). Approximately 30-40% of HCC patients are diagnosed at an advanced stage, and at present, there are limited treatment options for such patients. The current first-line therapy with tyrosine kinase inhibitors, sorafenib or lenvatinib, prolongs survival by a median of about 2.5-3 months after which the disease normally progresses. Additionally, many patients discontinue the use of tyrosine kinase inhibitors due to toxicity or may not be suitable candidates due to co-morbidity or frailty. It is, therefore, imperative to identify novel therapeutic targets for advanced HCC patients. Persistent injury to the liver as a result of insults such as hepatitis B or C viral (HBV or HCV) infections, alcohol abuse, and non-alcoholic fatty liver disease (NAFLD), results in chronic inflammation, which progresses to hepatic fibrosis and later, cirrhosis, provides the conditions for initiation of HCC. One of the key pathways studied for its role in inflammation and carcinogenesis is the eicosanoid pathway. In this review, we briefly outline the eicosanoid pathway, describe the mechanisms by which some pathway members either facilitate or counter the development of liver diseases, with the focus on NAFLD/hepatic fibrosis/cirrhosis, and HCC. We describe the link between the eicosanoid pathway, inflammation and these liver diseases, and identify components of the eicosanoid pathway that may be used as potential therapeutic targets in HCC.
ABSTRACT
Although 1-year survival rates following liver transplantation over the last 20 years may have improved, there is doubt about improvement in long-term survival. We examined survival with and without initial 12-month mortality in adult liver transplant recipients over a 20-year period. Patient and allograft survival for 3 different time periods was compared: 1986-1994 (group 1, n = 547), 1995-2000 (group 2, n = 735), and 2000-2005 (group 3, n = 749). After this, all deaths in the first 12 months of each group were removed. Patient and allograft survival was then once again compared across the 3 groups. There was significant improvement in both patient and allograft survival across the 20-year period (P < 0.001). Overall patient and allograft survival improved in non-hepatitis C virus (HCV) patients but not in HCV patients. A similar comparison with deaths in the first year removed, however, showed no difference in patient survival (P = 0.07) and only a marginal improvement in allograft survival (P = 0.048) between the 3 time periods. When patients were divided into HCV-positive and HCV-negative groups with deaths in the first year removed, there was, however, improved patient and allograft survival in the HCV-negative group but not in the HCV-positive group. The causes of death between 1 and 5 years were then compared. There were 48 deaths in period 1, 63 in period 2, and 43 in period 3 (P = not significant). There were more deaths due to cardio/cerebrovascular disease and hepatitis B virus recurrence in the first time period, but there were more deaths due to recurrent HCV and de novo malignancy in later time periods. In conclusion, although overall survival following liver transplantation in adults seems to be improving over time, the long-term results are not, particularly in HCV patients.
Subject(s)
Hepatitis C, Chronic/mortality , Hepatitis C, Chronic/surgery , Liver Failure/mortality , Liver Failure/surgery , Liver Transplantation/mortality , Adolescent , Adult , Aged , Australia/epidemiology , Carcinoma, Hepatocellular/mortality , Carcinoma, Hepatocellular/surgery , Child , Cholangitis, Sclerosing/mortality , Cholangitis, Sclerosing/surgery , Hepatitis, Autoimmune/mortality , Hepatitis, Autoimmune/surgery , Humans , Kaplan-Meier Estimate , Liver Cirrhosis, Biliary/mortality , Liver Cirrhosis, Biliary/surgery , Liver Neoplasms/mortality , Liver Neoplasms/surgery , Middle Aged , New Zealand/epidemiology , Transplantation, Homologous , Young AdultABSTRACT
The outcomes of right lobe split (RLS) liver transplantation are variable in adult recipients. This report is an analysis of outcomes of our initial 5-year experience with the right lobe trisegment split graft. A retrospective analysis was performed of the recipient and graft outcomes from July 2002 to March 2007 of all adult recipients of RLS grafts versus recipients of whole grafts (WGs). All data were analyzed with Stata version 8 (Stata Corp., Texas). There were 43 (19.1%) RLS recipients and 182 (80.9%) WG recipients. The median Model for End-Stage Liver Disease score was 13 (7-23) in the RLS group and 18 (6-50) in the WG group (P < 0.001). Hepatocellular carcinoma and primary sclerosing cholangitis were more common in the RLS group (P < 0.05), whereas alcoholic cirrhosis and chronic hepatitis C were more common in the WG group. The median donor age was lower in the RLS group at 39 (13-61) years versus the WG group at 47 (12-79) years (P < 0.001). Primary nonfunction occurred in 1.6% of the WG patients only. Biliary complications occurred in 28% of the RLS patients versus 28% of the WG patients. Vascular complications occurred in 18% of the RLS patients versus 14% of the WG patients. The retransplantation rate was similar at 2.3% in the RLS group versus 4.9% in the WG group (P = not significant). Overall 3-year recipient survival was 92.7% in the RLS group versus 82.7% in the WG group (P = 0.284). Graft survival was 88.4% in the RLS group at 3 years versus 78.5% in the WG group (P = 0.304). In conclusion, good outcomes can be achieved with RLS liver transplantation in adult recipients without a detrimental effect on recipient or graft survival.