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Background: Kinesin family member 12 (KIF12) mutation-related cholestatic disorder represents a rare subtype of progressive familial intrahepatic cholestasis (PFIC), referred to as PFIC Type 8, with only 21 reported cases globally to date., Methods: Here, we present a unique case of a 6-month-old boy diagnosed with homozygous KIF12 gene mutation, who successfully underwent a living donor liver transplant at our center for end-stage liver disease., Results: This case marks the youngest patient of KIF12-related cholestatic disorder necessitating a liver transplant to date. The child initially presented with neonatal cholestasis and then developed infantile hepatic decompensation. Our report discusses the diagnostic process and management strategies employed. It underscores the importance of prompt diagnosis through clinical suspicion, biochemical parameters, and genetic testing, as well as the adoption of suitable management strategies, including the early contemplation of liver transplant in such exceptional and rare cases of genetic intrahepatic cholestasis., Conclusion: KIF12-related genetic disease should be considered in neonatal cholestasis cases with high gamma glutamyl transpeptidase to differentiate from conditions like biliary atresia. Favorable outcomes post liver transplant stress the importance of early genetic testing and referral to liver transplant centers for unresponsive patients, potentially saving lives., (© 2024 Wiley Periodicals LLC.)

Aim: Hepatic fibrosis in injured liver is characterized by the activation of hepatic stellate cells (HSCs) from their quiescent state. Survivin (BIRC5) is one of the key genes that are upregulated during activation of HSCs but their role in HSC activation and fibrosis progression is unknown. Here, we have investigated the role of survivin protein in early fibrogenic activation of HSCs and fibrosis progression in chronic liver injury., Materials & Methods: Primary quiescent HSCs were isolated from healthy mice liver through perfusion and cultured for fibrogenic activation. Survivin expression was suppressed by its pharmacological suppressant, YM155. We developed chronic liver injury induced fibrotic mice model through administrating repeated dose of CCl 4 for 2 weeks and 4 weeks. Mice were pre-treated with YM155 a week before CCl 4 administration till 2nd week of dosing and then discontinued. Hepatic parameters were characterized and underlying mechanisms were investigated., Key Findings: Survivin expression gradually increased along with the expression of αSMA, collagen I activation maker in HSCs during their activation from quiescent state. Survivin suppression through YM155 downregulated αSMA, collagen I. Pre-treatment of YM155 in mice ceased the early activation of HSCs and onset of fibrosis in injured liver. However, discontinuation of YM155 initiated the activation of HSCs and fibrosis progression that shows survivin expression in HSCs is essential for their early activation and onset of liver fibrosis., Significance: Survivin expression induces with activation of HSCs and drives onset of liver fibrosis in injured liver. Targeting survivin protein in activated HSCs could be a potential anti-fibrotic therapeutic approach in chronic liver injury., (Copyright © 2021 Elsevier Inc. All rights reserved.)

The only definitive therapy for end-stage liver disease is whole-organ transplantation. The success of this intervention is severely limited by the complexity of the surgery, the cost of patient care, the need for long-term immunosuppression, and the shortage of donor organs. In rodents and humans, end-stage degeneration of hepatocyte function is associated with disruption of the liver-specific transcriptional network and a nearly complete loss of promoter P1-driven hepatocyte nuclear factor 4-alpha (P1-HNF4α) activity. Re-expression of HNF4α2, the predominant P1-HNF4α, reinstates the transcriptional network, normalizes the genes important for hepatocyte function, and reverses liver failure in rodents. In this study, we tested the effectiveness of supplementary expression of human HNF4α2 messenger RNA (mRNA) in primary human hepatocytes isolated from explanted livers of patients who underwent transplant for end-stage irreversibly decompensated liver failure (Child-Pugh B, C) resulting from alcohol-mediated cirrhosis and nonalcoholic steatohepatitis. Re-expression of HNF4α2 in decompensated cirrhotic human hepatocytes corrects the disrupted transcriptional network and normalizes the expression of genes important for hepatocyte function, improving liver-specific protein expression. End-stage liver disease in humans is associated with both loss of P1-HNF4α expression and failure of its localization to the nucleus. We found that while HNF4α2 re-expression increased the amount of P1-HNF4α protein in hepatocytes, it did not alter the ability of hepatocytes to localize P1-HNF4α to their nuclei. Conclusion: Re-expression of HNF4α2 mRNA in livers of patients with end-stage disease may be an effective therapy for terminal liver failure that would circumvent the need for organ transplantation. The efficacy of this strategy may be enhanced by discovering the cause for loss of nuclear P1-HNF4α localization in end-stage cirrhosis, a process not found in rodent studies., (© 2021 The Authors. Hepatology Communications published by Wiley Periodicals LLC on behalf of American Association for the Study of Liver Diseases.)

Pruritus is known to be a common complication in hepatitis patients, but the exact frequency and degree are not fully elucidated. Thus, we evaluated pruritus of 450 patients with chronic liver disease at our hospital. Pruritus was observed in 240 (53%) of the patients. Pruritus was significantly associated with males (OR = 1.51, P = 0.038) and patients with alkaline phosphatase (ALP) ≥ 200 U/L (OR = 1.56, P = 0.0495) and was significantly less in HBsAg-positive patients (OR = 0.449, P = 0.004). Seasonally, there was no difference in the frequency of pruritus between summer and winter. Of the 24 refractory pruritus patients treated with nalfurafine, 17 (71%) indicated improvement of itch, which is defined as a decrease in the visual analog scale score ≥ 30 mm. Pruritus was improved by nalfurafine both during daytime and nighttime in the Kawashima's scores evaluation. All patients who received nalfurafine exhibited improved Kawashima's scores ≥ 1 point during the daytime or nighttime. In conclusion, pruritus occurred in > 50% of patients with chronic liver disease, and predictors of pruritus were males and ALP ≥ 200 U/L. Nalfurafine may be useful for pruritus, regardless of whether daytime or nighttime.

Background and Aims: Portopulmonary hypertension (POPH) was previously associated with a single-nucleotide polymorphism (SNP) rs7175922 in aromatase (cytochrome P450 family 19 subfamily A member 1 [CYP19A1]). We sought to determine whether genetic variants and metabolites in the estrogen signaling pathway are associated with POPH., Approach and Results: We performed a multicenter case-control study. POPH patients had mean pulmonary artery pressure >25 mm Hg, pulmonary vascular resistance >240 dyn-sec/cm -5 , and pulmonary artery wedge pressure ≤15 mm Hg without another cause of pulmonary hypertension. Controls had advanced liver disease, right ventricular (RV) systolic pressure <40 mm Hg, and normal RV function by echocardiography. We genotyped three SNPs in CYP19A1 and CYP1B1 using TaqMan and imputed SNPs in estrogen receptor 1 using genome-wide markers. Estrogen metabolites were measured in blood and urine samples. There were 37 patients with POPH and 290 controls. Mean age was 57 years, and 36% were female. The risk allele A in rs7175922 (CYP19A1) was significantly associated with higher levels of estradiol (P = 0.02) and an increased risk of POPH (odds ratio [OR], 2.36; 95% confidence interval [CI], 1.12-4.91; P = 0.02) whereas other SNPs were not. Lower urinary 2-hydroxyestrogen/16-α-hydroxyestrone (OR per 1-ln decrease = 2.04; 95% CI, 1.16-3.57; P = 0.01), lower plasma levels of dehydroepiandrosterone-sulfate (OR per 1-ln decrease = 2.38; 95% CI, 1.56-3.85; P < 0.001), and higher plasma levels of 16-α-hydroxyestradiol (OR per 1-ln increase = 2.16; 95% CI, 1.61-2.98; P < 0.001) were associated with POPH., Conclusions: Genetic variation in aromatase and changes in estrogen metabolites were associated with POPH., (© 2020 by the American Association for the Study of Liver Diseases.)

Dubin-Johnson syndrome (DJS), an autosomal recessive disorder presenting with conjugated hyperbilirubinemia, is not associated with progression to chronic liver disease (CLD). Next-generation sequencing, application of bioinformatics pipeline, and segregation analysis were performed on 8 members of a consanguineous family with DJS and CLD. A novel variant, c.4406&#95;4407delTA (p.Leu1469fs), in the ABCC2-gene in a homozygous state was found to be associated with DJS and CLD in proband and afflicted family members. DJS may not be a benign entity and novel genetic variants may be associated with progressive liver disease.

Although, liver transplantation serves as the only curative treatment for patients with end-stage liver diseases, it is burdened with complications, which affect survival rates. In addition to clinical risk factors, contribution of recipient and donor genetic prognostic markers has been extensively studied in order to reduce the burden and improve the outcomes. Determination of single nucleotide polymorphisms (SNPs) is one of the most important tools in development of personalized transplant approach. To provide a better insight in recent developments, we review the studies published in the last three years that investigated an association of recipient or donor SNPs with most common issues in liver transplantation: Acute cellular rejection, development of new-onset diabetes mellitus and non-alcoholic fatty liver disease, hepatocellular carcinoma recurrence, and tacrolimus concentration variability. Reviewed studies confirmed previously established SNP prognostic factors, such as PNPLA3 rs738409 for non-alcoholic fatty liver disease development, or the role of CYP3A5 rs776746 in tacrolimus concentration variability. They also identified several novel SNPs, with a reasonably strong association, which have the potential to become useful predictors of post-transplant complications. However, as the studies were typically conducted in one center on relatively low-to-moderate number of patients, verification of the results in other centers is warranted to resolve these limitations. Furthermore, of 29 reviewed studies, 28 used gene candidate approach and only one implemented a genome wide association approach. Genome wide association multicentric studies are needed to facilitate the development of personalized transplant medicine., Competing Interests: Conflict-of-interest statement: No potential conflicts of interest., (©The Author(s) 2020. Published by Baishideng Publishing Group Inc. All rights reserved.)

Objective: To evaluate the efficacy of living-donor liver transplantation (LDLT) in children with tyrosinemia type I., Methods: Altogether 10 patients diagnosed with tyrosinemia type I underwent LDLT between June 2013 and April 2019. Cirrhosis was the indication for LDLT in all 10 patients, and hepatocellular carcinoma (HCC) was suspected in nine. Patients' outcomes, including liver function, restoration of metabolism, quality of life and physical development, were analyzed after LDLT., Results: All recipients were alive with a normal liver function after a median follow-up period of 49 months. Pathological examinations detected HCC in one patient, dysplasia in five and cirrhosis in all. Nine patients were found to have elevated alpha-fetoprotein level, and their median alpha-fetoprotein level dropped from 2520 ng/mL to a normal level after LDLT, with no recurrence of HCC detected during the follow-up. Tyrosine metabolism was restored to its normal level with normalized plasma tyrosine and succinylacetone concentrations. Moreover, urinary succinylacetone excretion decreased significantly during the follow up. LDLT improved patients' renal tubular function, as evidenced by the normalized plasma phosphate concentration and improved glomerular filtration rate. Severe rickets symptoms, including spontaneous fractures and bone pain, were also ameliorated. Improved motor function was reported by all patients' parents during the follow-up. Dietary restriction was no longer required, which was associated with a favorable catch-up in growth and improved quality of life. Complete resolution of hypertrophic cardiomyopathy was observed one year after LDLT in one patient., Conclusion: LDLT is an effective treatment for patients with end-stage liver disease resulting from tyrosinemia type I., (© 2020 Chinese Medical Association Shanghai Branch, Chinese Society of Gastroenterology, Renji Hospital Affiliated to Shanghai Jiaotong University School of Medicine and John Wiley & Sons Australia, Ltd.)

The presence of advanced fibrosis is an important measure of the severity of chronic liver disease. Prior works that have examined the gut microbiome as a novel biomarker for advanced fibrosis have only examined patients with nonalcoholic fatty liver disease. Therefore, our goal was to examine the gut microbiome across varying etiologies of liver disease to create a predictive model for liver fibrosis based upon a microbial signature. Stool samples were obtained from patients with chronic liver disease (n = 50) undergoing FibroScan (ultrasound elastography) at the VA Greater Los Angeles Healthcare System. Healthy control patients (n = 25) were also recruited as a reference population. Fecal samples underwent 16S ribosomal RNA sequencing. Using differentially abundant microbes, a random forest classifier model was created to distinguish advanced fibrosis from mild/moderate fibrosis. The findings were then validated in a separate cohort of chronic liver disease patients (n = 37). Etiologies for liver disease included non-alcoholic liver disease (58.0%), hepatitis C (26.0%), hepatitis B (10.0%), and alcohol (6.0%). Microbiome composition was distinct in liver patients with advanced fibrosis compared to those with minimal fibrosis and healthy controls (p = 0.003). In multivariate negative binomial modeling, 26 bacterial taxa were differentially abundant in patients with advanced fibrosis as compared to those with minimal/moderate fibrosis (q-value < 0.05). A random forests classifier based on these taxa had an AUROC of 0.90 to predict advanced fibrosis. Prevotella copri, which was enriched in patients with advanced fibrosis, was the most strongly predictive microbe in the classifier. The classifier had an AUROC of 0.82 for advanced fibrosis in the validation cohort and Prevotella copri remained the strongest predictive microbe for advanced fibrosis. There is a distinct microbial signature for patients with advanced fibrosis independent of liver disease etiology and other comorbidities. These results suggest that microbial profiles can be used as a non-invasive marker for advanced fibrosis and support the hypothesis that microbes and their metabolites contribute to hepatic fibrosis.

Alcoholic liver disease (ALD) is a complication that is a burden on global health and economy. Interleukin-33 (IL-33) is a newly identified member of the IL-1 cytokine family and is released as an "alarmin" during inflammation. Soluble suppression of tumourigenicity 2 (sST2), an IL-33 decoy receptor, has been reported as a new biomarker for the severity of systemic and highly inflammatory diseases. Here, we found the levels of plasma sST2, increased with the disease severity from mild to severe ALD. Importantly, the plasma sST2 levels in ALD patients not only correlated with scores for prognostic models (Maddrey's discriminant function, model for end-stage liver disease and Child-Pugh scores) and indexes for liver function (total bilirubin, international normalized ratio, albumin, and cholinesterase) but also correlated with neutrophil-associated factors as well as some proinflammatory cytokines. In vitro, lipopolysaccharide-activated monocytes down-regulated transmembrane ST2 receptor but up-regulated sST2 mRNA and protein expression and produced higher levels of tumour necrosis factor-α (TNF-α). By contrast, monocytes pretreated with recombinant sST2 showed decreased TNF-α production. In addition, although plasma IL-33 levels were comparable between healthy controls and ALD patients, we found the IL-33 expression in liver tissues from ALD patients was down-regulated at both RNA and protein levels. Immunohistochemical staining further showed that the decreased of IL-33-positive cells were mainly located in liver lobule area. These results suggested that sST2, but not IL-33, is closely related to the severity of ALD. Consequently, sST2 could be used as a potential biomarker for predicting the prognosis of ALD., (© 2018 The Authors. Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and Molecular Medicine.)

Objectives: Bile salt export pump (BSEP) deficiency is an important reason for chronic cholestasis leading to liver transplantation (LT) in early childhood. The underlying pathology is a dysfunction of BSEP due to various mutations in the ABCB11 gene. Cases of clinical recurrence after LT due to alloantibodies directed against BSEP (antibody-induced BSEP deficiency [AIBD]) have been reported. Most of these patients could be controlled by intensified immunosuppression., Methods: We here report on 3 children with BSEP-deficiency and end-stage liver disease, which developed AIBD after LT refractory to extensive immunosuppressive and immunomodulatory treatments; retransplantation was necessary in all 3 patients. In 1 patient, a stem cell transplantation was performed successfully., Results: AIBD seems to be induced by triggering factors such as initial impaired graft function or infections after LT., Conclusions: The underlying mutation may play a role in this process. Intensifying immunosuppression may be able to control AIBD, but some cases seem to be refractory to treatment and require retransplantation. Stem cell transplantation may provide a new therapeutic option for cases refractory to conservative treatment.

Background: Patients with end-stage liver disease (ESLD) have a compromised nutritional status because of the liver crucial role in regulating metabolic homeostasis and energy balance., Data Sources: A systematic review of literature based on extensive relevant articles published from 2001 to 2017 in English in PubMed database was performed by searching keywords such as liver disease, non-alcoholic liver disease, alcoholic liver disease, malnutrition, epigenetics, gut microbiota, and probiotics., Results: Liver transplantation would be one eligible therapy for ESLD patients, even if, the clinical outcome is negatively influenced by malnutrition and/or infections. The malnutrition is a condition of nutrient imbalance with a high incidence in ESLD patients. An accurate evaluation of nutritional status could be fundamental for reducing complications and prolonging the survival of ESLD patients including those undergoing liver transplantation. In addition, the interaction among nutrients, diet and genes via epigenetics has emerged as a potential target to reduce the morbidity and mortality in ESLD patients. The malnutrition induces changes in gut microbiota causing dysbiosis with a probable translocation of bacteria and/or pathogen-derived factors from the intestine to the liver. Gut microbiota contribute to the progression of chronic liver diseases as well as hepatocellular carcinoma. The administration of probiotics modulating gut microbiota could improve all chronic liver diseases., Conclusions: This review provides an update on malnutrition status linked to epigenetics and the potential benefit of some probiotics on the management of ESLD patients. In support of this view and to reveal the constant and growing interest in this field, some clinical trials are reported., (Copyright © 2018 First Affiliated Hospital, Zhejiang University School of Medicine in China. Published by Elsevier B.V. All rights reserved.)

Liver diseases are a growing epidemic worldwide. If unresolved, liver fibrosis develops and can lead to cirrhosis and clinical decompensation. Around 5% of cirrhotic liver diseased patients develop hepatocellular carcinoma (HCC), which in its advanced stages has limited therapeutic options and negative survival outcomes. CEPBA is a master regulator of hepatic function where its expression is known to be suppressed in many forms of liver disease including HCC. Injection of MTL-CEBPA, a small activating RNA oligonucleotide therapy (CEBPA-51) formulated in liposomal nanoparticles (NOV340- SMARTICLES) upregulates hepatic CEBPA expression. Here we show how MTL-CEBPA therapy promotes disease reversal in rodent models of cirrhosis, fibrosis, hepatosteatosis, and significantly reduces tumor burden in cirrhotic HCC. Restoration of liver function markers were observed in a carbon-tetrachloride-induced rat model of fibrosis following 2 weeks of MTL-CEBPA therapy. At 14 weeks, animals showed reduction in ascites and enhanced survival rates. MTL-CEBPA reversed changes associated with hepatosteatosis in non-alcoholic methionine and cholic-deficient diet-induced steaotic liver disease. In diethylnitrosamine induced cirrhotic HCC rats, MTL-CEBPA treatment led to a significant reduction in tumor burden. The data included here and the rapid adoption of MTL-CEBPA into a Phase 1 study may lead to new therapeutic oligonucleotides for undruggable diseases.

The aim of this study is to elucidate the biogenetic modification of donor and recipient interleukin-28B (IL-28B) genotypes in liver graft biopsies after living donor liver transplantation (LDLT) for chronic hepatitis C virus- (HCV-) related, end-stage liver disease. Fifty liver graft biopsies were collected from recipients during LDLT treatment for HCV-related, end-stage liver disease. DNA was extracted from all 50 liver tissues, and the IL-28B single-nucleotide polymorphisms (SNPs) rs8099917 and rs12979860 were studied for allelic discrimination by real-time PCR analysis. Blood samples were obtained from donors and recipients on postoperative day 0 (POD0), POD7, and POD30. We randomly selected five liver biopsies and isolated the hepatocytes by laser capture microdissection (LCM) to evaluate genotype modifications resulting from LDLT. After LDLT, the IL-28B SNP rs8099917 was identified not only in the liver graft biopsies and donors' sera (TT = 41 : 43; GT = 9 : 5; GG = 0 : 2), but also in liver graft biopsies and recipients' sera on POD0 (TT = 41 : 44; GT = 9 : 4; GG = 0 : 2), POD7 (TT = 41 : 30; GT = 9 : 18; GG = 0 : 2), and POD30 (TT = 41 : 29; GT = 9 : 19; GG = 0 : 2). A significant difference was observed between the rs8099917 allele frequencies of liver graft biopsies and recipients' sera on POD30 ( p = 0.039). In addition, a significant difference was also noted between the rs12979860 allele frequencies of liver graft biopsies and donors' sera (CT = 49 : 39; TT = 1 : 10) ( p = 0.012) and of liver graft biopsies and recipients' sera on POD0 (CT = 49 : 39; TT = 1 : 11) ( p = 0.002), POD7 (CT = 49 : 42; TT = 1 : 8) ( p = 0.016), and POD30 (CT = 49 : 41; TT = 1 : 9) ( p = 0.008). This phenomenon was confirmed by pyrosequencing of hepatocytes isolated by LCM. Following LDLT, the TT-to-GT IL-28B genotype modification predominated in rs8099917, and the CC-to-CT modification predominated in rs12979860. In conclusion, these modified phenomena suggested that the selected donor with a predictable and favourable IL-28B genotype will not confer a benefit on the recipient in the living donor liver transplantation setting.

Objective: In the liver, a contributing factor in the pathogenesis of non-alcoholic fatty liver disease is oxidative stress leading to the accumulation of highly reactive electrophilic α/β unsaturated aldehydes. The objective of this study was to determine if significant differences were evident when evaluating carbonylation in human end-stage fatty nonalcoholic steatohepatitis (fNASH) compared to end-stage nonfatty NASH (nfNASH)., Methods: Using hepatic tissue obtained from healthy humans and patients diagnosed with end stage nfNASH or fNASH, overall carbonylation was assessed by immunohistochemistry (IHC) and LC-MS/MS followed by bioinformatics., Results: Picrosirius red staining revealed extensive fibrosis in both fNASH and nfNASH which corresponded with increased reactive aldehyde staining. Although significantly elevated when compared to normal hepatic tissue, no significant differences in overall carbonylation and fibrosis were evident when comparing fNASH with nfNASH. Examining proteins that are critical for anti-oxidant defense revealed elevated expression of thioredoxin, thioredoxin interacting protein, glutathione S-transferase p1 and mitochondrial superoxide dismutase in human NASH. As important, using immunohistochemistry, significant colocalization of the aforementioned proteins occurred in cytokeratin 7 positive cells indicating that they are part of the ductular reaction. Expression of catalase and Hsp70 decreased in both groups when compared to normal human liver. Mass spectrometric analysis revealed a total of 778 carbonylated proteins. Of these, 194 were common to all groups, 124 unique to tissue prepared from healthy individuals, 357 proteins exclusive to NASH, 124 proteins distinct to samples from patients with fNASH and 178 unique to nfNASH. Using functional enrichment analysis of hepatic carbonylated proteins revealed a propensity for increased carbonylation of proteins regulating cholesterol and Huntington's disease related pathways occurred in nfNASH. Examining fNASH, increased carbonylation was evident in proteins regulating Rho cytoskeletal pathways, nicotinic acetylcholine receptor signaling and chemokine/cytokine inflammatory pathways. Using LC-MS/MS analysis and trypsin digests, sites of carbonylation were identified on peptides isolated from vimentin, endoplasmin and serum albumin in nfNASH and fNASH respectively., Conclusions: These results indicate that cellular factors regulating mechanisms of protein carbonylation may be different depending on pathological diagnosis of NASH. Furthermore these studies are the first to use LC-MS/MS analysis of carbonylated proteins in human NAFLD and explore possible mechanistic links with end stage cirrhosis due to fatty liver disease and the generation of reactive aldehydes., (Copyright © 2017 Elsevier Inc. All rights reserved.)

BACKGROUND In patients with human immunodeficiency virus (HIV) and hepatitis C virus (HCV) co-infection, HIV can modulate HCV replication and immune response as well as accelerate liver fibrosis. The role of miRNA in HIV/HCV co-infection is not fully elucidated. The aim of this study was to examine the differential expression of miRNAs in the liver. MATERIAL AND METHODS Thirteen patients who had undergone a liver transplant (7 HCV-infected and 6 HIV/HCV-co-infected patients) were examined using a miRNA array containing 1347 human miRNAs. To confirm the microarray results, data for 20 patients (10 HCV-infected and 10 HIV/HCV-co-infected) were validated using real-time polymerase chain reaction probing for miR101b, miR149, and miR200c. This miRNA was selected based on microarray results and its biological significance in liver fibrosis. RESULTS Microarray analysis revealed 22 miRNAs that were differentially expressed in the HIV/HCV-co-infected group compared to the HCV-infected group (p<0.05). The expression of miR-101b and miR149 was significantly decreased in the HIV/HCV-co-infected group compared to that in the HCV-infected group (miR101b, 0.103±0.09 vs. 0.0157±0.0093, p=0.007; miR149, 0.152±0.159 vs. 0.0192±0.015, p=0.025). CONCLUSIONS HIV/HCV co-infection may promote liver fibrosis by modulating miRNA expression.

Objectives: Hepatitis B viral infection is among the most common causes of cirrhosis and hepatocellular carcinoma and a frequent viral indication for liver transplant. Cytokine-mediated immunity plays a critical role in introducing and promoting hepatitis B virus outcomes and in graft microenvironment. Interleukin 27 is a heterodimeric cytokine and a member of interleukin-6/interleukin-12 family. Interleukin-27 shows a broad range of pro- and antiinflammatory properties and plays a determining role during immune responses in combating hepatitis B virus. Therefore, in this study, the possible association between expressions of interleukin-27 gene with hepatitis B virus infection was evaluated in liver transplant patients., Materials and Methods: In a cross-sectional study from liver transplant patients with the risk of hepatitis B virus infection who admitted to Namazi Hospital affiliated to Shiraz University of Medical Sciences, 50 patients were selected and subgrouped to 25 hepatitis B virus-infected and 25 noninfected ones between years 2011 and 2013. The 25 healthy controls also were enrolled in this study. The presence of hepatitis B virus infection was assessed using polymerase chain reaction and enzyme-linked immunosorbent assay protocols in liver transplant patients. In addition, the interleukin-27 gene expression level was analyzed using an in-house-SYBER Green real time polymerase chain reaction method. The rate of interleukin-27 gene expression level was statistically analyzed in studied patient groups and controls using the Livak (2-▵▵CT) method., Results: The expression level of interleukin-27 gene was increased 10.27- and 2.36-fold in hepatitis B virus-infected and uninfected liver transplanted patients compared with healthy controls., Conclusion: Hepatitis B virus infection can lead to overexpression of interleukin-27 gene in liver transplant patients compared with uninfected ones and controls. However, further studies are needed to characterize the effective antihepatitis B virus effects of interleukin-27 in liver transplant patients.

Orthotopic liver transplant (OLT) remains the standard of care for end stage liver disease. To circumvent allo-rejection, OLT subjects receive gluococorticoids (GC). We investigated the effects of GC on endogenous mesenchymal stem (stromal) cells (MSCs) in OLT. This question is relevant because MSCs have regenerative potential and immune suppressor function. Phenotypic analyses of blood samples from 12 OLT recipients, at pre-anhepatic, anhepatic and post-transplant (2 h, Days 1 and 5) indicated a significant decrease in MSCs after GC injection. The MSCs showed better recovery in the blood from subjects who started with relatively low MSCs as compared to those with high levels at the prehepatic phase. This drop in MSCs appeared to be linked to GC since similar change was not observed in liver resection subjects. In order to understand the effects of GC on decrease MSC migration, in vitro studies were performed in transwell cultures. Untreated MSCs could not migrate towards the GC-exposed liver tissue, despite CXCR4 expression and the production of inflammatory cytokines from the liver cells. GC-treated MSCs were inefficient with respect to migration towards CXCL12, and this correlated with retracted cytoskeleton and motility. These dysfunctions were partly explained by decreases in the CXCL12/receptor axis. GC-associated decrease in MSCs in OLT recipients recovered post-transplant, despite poor migratory ability towards GC-exposed liver. In total, the study indicated that GC usage in transplant needs to be examined to determine if this could be reduced or avoided with adjuvant cell therapy.

Introduction: Nonobstructive cholestasis after pediatric liver transplantation is a common diagnostic and therapeutic dilemma. We describe a girl with neonatal cholestasis because of progressive familial intrahepatic cholestasis 2 (PFIC-2) and presence of a homozygous splice site mutation in the ABCB11 gene. Liver transplantation was performed because of end-stage liver disease at the age of 6. Cholestasis with normal gamma-glutamyl transferase (GGT) developed 8 years after liver transplantation. A liver biopsy showed canalicular cholestasis and giant cell hepatitis without evidence of rejection, mimicking PFIC-2. Immunofluorescence staining of normal human liver sections with patient's serum revealed reactivity toward a canalicular epitope, which could be identified as bile salt export pump (BSEP) using BSEP-yellow fluorescent protein (YFP) transfected cells. Our patient developed a recurrence of a PFIC-2 phenotype due to production of antibodies against BSEP (alloimmune BSEP disease [AIBD]). Intensification of immunosuppressive therapy as well as antibody treatment with plasmapheresis and Rituximab were initiated, leading to stabilization of the clinical condition and depletion of anti-BSEP antibodies in serum. However, after 1 year liver transplantation was necessary again because of end-stage liver insufficiency. Afterward, immunomodulatory treatment consisted of tacrolimus, mycophenolate mofetil, prednisone, immunoadsorption, and high-dose immunoglobulin therapy (1 g/kg/d)., Conclusion: Cholestasis after liver transplantation may indicate an AIBD with a PFIC-2 phenotype. Besides enhancement of immunosuppressive therapy, an antibody depletion with plasmapheresis, immunoadsorption, immunoglobulins, and B-cell depletion represents a therapeutic option., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Background: Hepatitis B virus (HBV) occult infection (OBI) is a risk factor to be taken into account in transfusion, hemodialysis and organ transplantation. The aim of this study was to identify and characterize at the molecular level OBI cases in patients with end-stage liver disease., Methods: Sixty-six liver samples were obtained from patients with diagnosis of end-stage liver disease submitted to liver transplantation in Medellin (North West, Colombia). Samples obtained from patients who were negative for the surface antigen of HBV (n = 50) were tested for viral DNA detection by nested PCR for ORFs S, C, and X and confirmed by Southern-Blot. OBI cases were analyzed by sequencing the viral genome to determine the genotype and mutations; additionally, viral genome integration events were examined by the Alu-PCR technique., Results: In five cases out of 50 patients (10%) the criteria for OBI was confirmed. HBV genotype F (subgenotypes F1 and F3), genotype A and genotype D were characterized in liver samples. Three integration events in chromosomes 5q14.1, 16p13 and 20q12 affecting Receptor-type tyrosine-protein phosphatase T, Ras Protein Specific Guanine Nucleotide Releasing Factor 2, and the zinc finger 263 genes were identified in two OBI cases. Sequence analysis of the viral genome of the 5 OBI cases showed several punctual missense and nonsense mutations affecting ORFs S, P, Core and X., Conclusions: This is the first characterization of OBI in patients with end-stage liver disease in Colombia. The OBI cases were identified in patients with HCV infection or cryptogenic cirrhosis. The integration events (5q14.1, 16p13 and 20q12) described in this study have not been previously reported. Further studies are required to validate the role of mutations and integration events in OBI pathogenesis.

End-stage liver disease is a common cause of non-AIDS-related mortality in HIV + patients, despite effective anti-retroviral therapies (ARTs). HIV-1 infection causes gut CD4 depletion and is thought to contribute to increased gut permeability, bacterial translocation, and immune activation. Microbial products drain from the gut into the liver via the portal vein where Kupffer cells (KCs), the resident liver macrophage, clear translocated microbial products. As bacterial translocation is implicated in fibrogenesis in HIV patients through unclear mechanisms, we tested the hypothesis that HIV infection of KCs alters their response to LPS in a TLR4-dependent manner. We showed that HIV-1 productively infected KCs, enhanced cell-surface TLR4 and CD14 expression, and increased IL-6 and TNF-α expression, which was blocked by a small molecule TLR4 inhibitor. Our study demonstrated that HIV infection sensitizes KCs to the proinflammatory effects of LPS in a TLR4-dependent manner. These findings suggest that HIV-1-infected KCs and their dysregulated innate immune response to LPS may play a role in hepatic inflammation and fibrosis and represent a novel target for therapy., (© Society for Leukocyte Biology.)

Hypoxia-inducible factor (HIF)-1α and HIF-2α play an important role in liver fibrosis. von Hippel-Lindau protein (VHL), a key mediator of HIF-α, regulates fibrosis in an organ- and cell-specific way. In this study, human liver samples were collected from hepatitis C-, alcoholic-, and cholestatic-associated fibrotic and healthy individuals. Two mouse models of liver fibrosis were established: bile duct ligation and carbon tetrachloride injection. We constructed adenovirus vectors to overexpress VHL, normoxia-active HIF-α, and lentiviral vectors to silence HIF-α. The results showed that liver sections from fibrosis patients had a lower level of VHL and higher levels of HIF-1α and HIF-2α compared with healthy sections, a finding which was confirmed in mice. Overexpression of VHL attenuated liver fibrosis, downregulated fibrogenic genes, and inhibited liver inflammation, apoptosis, and angiogenesis. Overexpression of VHL was more successful at inhibiting fibrosis compared with silencing HIF-1α plus HIF-2α. Normoxia-active HIF-1α or HIF-2α prevented the inhibitory effect of VHL on liver fibrosis, indicating that attenuating fibrosis via VHL is HIF-1α- and HIF-2α-dependent to some extent. In addition, overexpression of VHL inhibited mouse hepatic stellate cells activation and proliferation and promoted apoptosis. Taken together, VHL may be considered a new target to inhibit liver fibrosis.

We describe a case of fulminant intraoperative thrombosis during deceased donor liver transplantation. Despite significant medical bleeding, the patient suddenly developed diffuse thrombosis in all chambers of the heart and pulmonary vasculature resulting in intraoperative death. The patient's postmortem genetic analysis demonstrated a deleterious missense mutation in a coagulation pathway gene, SERPINC1, which codes for antithrombin III. The level of antithrombin III was not available to directly prove the causality of thrombosis, but our findings suggest that this mutation, in combination with antifibrinolytic administration in a hypercoagulable cirrhotic patient, might have contributed to the development of this catastrophic thrombotic event.

Hepatocellular adenoma is considered to occur exclusively in non-fibrotic livers. It is a heterogeneous entity and a molecular classification is now widely accepted. The most frequent hepatocellular adenoma subtype, namely inflammatory adenoma, harbor somatic activating mutations of genes involved in the interleukin-6 pathway that lead to high C-reactive protein and serum amyloid A expression. The aim of our study was to investigate a series of benign hepatocellular neoplasms developed on cirrhotic livers and characterized by an unequivocal histological diagnosis. We performed a clinical, pathological, and molecular study of 10 benign hepatocellular neoplasms developed in three patients with cirrhosis. Markers allowing hepatocellular adenoma classification were assessed by quantitative real-time PCR and immunohistochemistry. Samples were sequenced for CTNNB1, HNF1A, IL6ST, GNAS, STAT3, and TERT (promoter) mutations. A control series of 32 classical macronodules developed in cirrhosis related to various etiologies was screened by immunohistochemistry and gene sequencing. The three patients had cirrhosis related to metabolic syndrome and/or alcohol intake; two had a single tumor, while the third developed more than 30 lesions. Microscopic examination showed well-differentiated neoplasms sharing features with inflammatory adenoma including inflammatory infiltrates, sinusoidal dilatation, and dystrophic vessels. Sequencing revealed classical hotspot somatic mutations (IL6ST, n=8; STAT3, n=1; and GNAS, n=1) known to be responsible for IL-6/JAK/STAT pathway activation. Two classical high-grade macronodules demonstrated high serum amyloid A and/or C-reactive protein expression, without gene mutations. Altogether, our findings support the existence of rare inflammatory adenoma developed in cirrhosis.

Background: The KEAP1-Nrf2 antioxidant signaling pathway is important in protecting liver from various insults. However, little is known about the expression of Nrf2-related genes in human liver in different diseases., Methods: This study utilized normal donor liver tissues (n=35), samples from patients with hepatocellular carcinoma (HCC, n=24), HBV-related cirrhosis (n=27), alcoholic cirrhosis (n=5) and end-stage liver disease (n=13). All of the liver tissues were from the Oriental Liver Transplant Center, Beijing, China. The expressions of Nrf2 and Nrf2-related genes, including its negative regulator Kelch-like ECH-associated protein 1 (KEAP1), its targeted gene NAD(P)H-quinone oxidoreductase 1 (NQO1), glutamate-cysteine ligase catalytic subunit (GCLC) and modified subunit (GCLM), heme oxygenase 1 (HO-1) and peroxiredoxin-1 (PRDX1) were evaluated., Results: The expression of Nrf2 was decreased in HCC, increased in alcoholic cirrhosis and end-stage liver disease. The expression of KEAP1 was increased in all of the liver samples. The most notable finding was the increased expression of NQO1 in HCC (18-fold), alcoholic cirrhosis (6-fold), end-stage liver disease (5-fold) and HBV-related cirrhosis (3-fold). Peri-HCC also had 4-fold higher NQO1 mRNA as compared to the normal livers. GCLC mRNA levels were lower only in HCC, as compared to the normal livers and peri-HCC tissues. GCLM mRNA levels were higher in HBV-related cirrhosis and end-stage liver disease. HO-1 mRNA levels were increased in all liver tissues except for HCC. Peri-HCC had higher PRDX1 mRNA levels compared with HCC and normal livers., Conclusion: Nrf2 and Nrf2-related genes are aberrantly expressed in the liver in different diseases and the increase of NQO1 was the most notable finding, especially in HCC.

Background: Until recently, liver transplantation (Ltx) was the only available treatment for hereditary transthyretin (TTR) amyloidosis; today, however, several pharmacotherapies are tested. Herein, we present survival data from the largest available database on transplanted hereditary TTR patients to serve as a base for comparison., Methods: Liver transplantation was evaluated in a 20-year retrospective analysis of the Familial Amyloidosis Polyneuropathy World Transplant Registry., Results: From April 1990 until December 2010, data were accumulated from 77 liver transplant centers. The Registry contains 1940 patients, and 1379 are alive. Eighty-eight Ltx were performed in combination with a heart and/or kidney transplantation. Overall, 20-year survival after Ltx was 55.3%. Multivariate analysis revealed modified body mass index, early onset of disease (<50 years of age), disease duration before Ltx, and TTR Val30Met versus non-TTR Val30Met mutations as independent significant survival factors. Early-onset patients had an expected mortality rate of 38% that of the late-onset group (P < 0.001). Furthermore, Val30Met patients had an expected mortality rate of 61% that of non-TTR Val30Met patients (P < 0.001). With each year of duration of disease before Ltx, expected mortality increased by 11% (P < 0.001). With each 100-unit increase in modified body mass index at Ltx, the expected mortality decreased to 89% of the expected mortality (P < 0.001). Cardiovascular death was markedly more common than that observed in patients undergoing Ltx for end-stage liver disease., Conclusions: Long-term survival after Ltx, especially for early-onset TTR Val30Met patients, is excellent. The risk of delaying Ltx by testing alternative treatments, especially in early-onset TTR Val30Met patients, requires consideration.

Pyrrole-imidazole (PI) polyamide is a novel gene regulating agent that competitively inhibits transcription factor binding to the promoter of the specific target gene. Liver fibrosis is an integral stage in the development of chronic liver disease, and transforming growth factor β (TGFβ) is known to play a central role in the progression of this entity. The aim of this study was to evaluate the effect of PI polyamide targeting TGFβ1 on rat liver fibrosis. PI polyamide was designed to inhibit activator protein 1 (AP-1) transcription factor binding to the TGFβ1 gene promoter. The effect of PI polyamide on hepatic stellate cells was evaluated by real time polymerase chain reaction (PCR) in RI-T cells. To determine the effect of PI polyamide in vivo, PI polyamide was intravenously administered at a dose of 3 mg/kg/week in dimethylnitrosamine (DMN)-induced rat model of liver fibrosis. Treatment of RI-T cells with 1.0 µM PI polyamide targeting TGFβ1 significantly inhibited TGFβ1 mRNA expression. Azan staining showed that DMN treatment significantly increased areas of fibrous materials compared with controls. PI polyamide targeting TGFβ1 significantly decreased the fibrous area compared with DMN group. mRNA expression levels of α-smooth muscle actin and matrix metalloproteinase-2 were significantly increased in DMN-treated group compared with control. Treatment with TGFβ1 PI polyamide significantly decreased mRNA expression of these genes compared with DMN group. The novel gene regulator PI polyamide targeting TGFβ1 may be a feasible therapeutic agent for the treatment of chronic liver disease.

Background & Aims: In children with liver failure, as many as half remain of indeterminate aetiology. This hinders timely consideration of optimal treatment options. We posit that a significant subset of these children harbour known inherited metabolic liver diseases with atypical presentation or novel inborn errors of metabolism. We investigated the utility of whole-exome sequencing in three children with advanced liver disease of indeterminate aetiology., Methods: Patient 1 was a 10 year-old female diagnosed with Wilson disease but no detectable ATP7B mutations, and decompensated liver cirrhosis who underwent liver transplant and subsequently developed onset of neurodegenerative disease. Patient 2 was a full-term 2 day-old male with fatal acute liver failure of indeterminate aetiology. Patient 3 was an 8 year-old female with progressive syndromic cholestasis of unknown aetiology since age 3 months., Results: Unbiased whole-exome sequencing of germline DNA revealed homozygous mutations in MPV17 and SERAC1 as the disease causing genes in patient 1 and 2, respectively. This is the first demonstration of SERAC1 loss-of-function associated fatal acute liver failure. Patient 1 expands the phenotypic spectrum of the MPV17-related hepatocerebral mitochondrial DNA depletion syndrome. Patient 3 was found to have syndromic cholestasis due to bi-allelic NOTCH2 mutations., Conclusions: Our findings validate the application of whole-exome sequencing in the diagnosis and management of children with advanced liver disease of indeterminate aetiology, with the potential to enhance optimal selection of treatment options and adequate counselling of families. Moreover, whole-exome sequencing revealed a hitherto unrecognized phenotypic spectrum of inherited metabolic liver diseases., (Copyright © 2014 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.)

Aim: To investigate the expression of P450 enzyme genes by using end-stage liver disease samples and trimmed normal Chinese donor livers., Methods: The end-stage liver disease samples [n = 93, including hepatocellular carcinoma (HCC), peri-HCC tissue, hepatitis B virus cirrhosis, alcoholic cirrhosis, and severe cirrhosis] and trimmed normal Chinese donor livers (n = 35) from The Institute of Organ Transplantation in Beijing, China. Total RNA was extracted, purified, and subjected to real-time RT-PCR analysis., Results: For cytochrome P450 enzymes 1 (CYP1) family, the expression of CYP1A2 was decreased 90% in HCC, 80% in alcoholic cirrhosis, and 65% in severe cirrhosis. For CYP2 family, the expression of CAR was decreased 50% in HCC, but increased 50% in peri-HCC tissues. Similar decreases (about 50%) of CYP2B6, CYP2C9, CYP2C19, CYP2D6 and CYP2E1 were observed in HCC, as compared to peri-HCC tissues and normal livers. CYP2C19 were decreased in all end-stage liver diseases and CYP2E1 also decreased in alcoholic cirrhosis and severe cirrhosis. For CYP3 family, the expression of PXR was decreased 60% in HCC, together with decreases in CYP3A4, CYP3A5, and CYP3A7. In contrast, the expression of CYP3A7 was slightly increased in HBV cirrhosis. The expression of CYP4A11 was decreased 85% in HCC, 7% in alcoholic cirrhosis and severe liver cirrhosis, along with decreases in PPARα. The 93 end-stage livers had much higher inter-individual variations in gene expression than 35 normal livers., Conclusion: The expression of CYP enzyme genes and corresponding nuclear receptors was generally decreased in end-stage liver diseases, and significant differences in gene expression were evident between peri-HCC and HCC.

Unlabelled: Proliferating ducts, termed "oval cells," have long been thought to be bipotential, that is, produce both biliary ducts and hepatocytes during chronic liver injury. The precursor to oval cells is considered to be a facultative liver stem cell (LSC). Recent lineage tracing experiments indicated that the LSC is SRY-related HMG box transcription factor 9 positive (Sox9(+) ) and can replace the bulk of hepatocyte mass in several settings. However, no clonal relationship between Sox9(+) cells and the two epithelial liver lineages was established. We labeled Sox9(+) mouse liver cells at low density with a multicolor fluorescent confetti reporter. Organoid formation validated the progenitor activity of the labeled population. Sox9(+) cells were traced in multiple oval cell injury models using both histology and fluorescence-activated cell sorting. Surprisingly, only rare clones containing both hepatocytes and oval cells were found in any experiment. Quantitative analysis showed that Sox9(+) cells contributed only minimally (<1%) to the hepatocyte pool, even in classic oval cell injury models. In contrast, clonally marked mature hepatocytes demonstrated the ability to self-renew in all classic mouse oval cell activation injuries. A hepatocyte chimera model to trace hepatocytes and nonparenchymal cells also demonstrated the prevalence of hepatocyte-driven regeneration in mouse oval cell injury models., Conclusion: Sox9(+) ductal progenitor cells give rise to clonal oval cell proliferation and bipotential organoids, but rarely produce hepatocytes in vivo. Hepatocytes themselves are the predominant source of new parenchyma cells in prototypical mouse models of oval cell activation., (© 2014 by the American Association for the Study of Liver Diseases.)

Recently, suppressor of cytokine signaling-3 (SOCS3) has been shown to be an inducible endogenous negative regulator of Janus kinase/signal transducers and activators of transcription (JAK/STAT) pathway which is relevant in inflammatory response, while its functions in acute liver failure and HBV-induced acute-on-chronic liver failure (HBV-ACLF) have not been fully elucidated. In this study, we explored the role of SOCS3 in the development of mouse hepatitis virus strain 3 (MHV-3)-induced acute liver failure and its expression in liver and peripheral blood mononuclear cells (PBMCs) of patients with HBV-ACLF. Inflammation-related gene expression was detected by real-time PCR, immunohistochemistry and Western blotting. The correlation between SOCS3 level and liver injury was studied. Our results showed that the SOCS3 expression was significantly elevated in both the liver tissue and PBMCs from patients with HBV-ACLF compared to mild chronic hepatitis B (CHB). Moreover, a time course study showed that SOCS3 level was increased remarkably in the liver of BALB/cJ mice at 72 h post-infection. Pro-inflammatory cytokines, interleukin (IL)-1β, IL-6, and tumor necrosis factor (TNF)-α, were also increased significantly at 72 h post-infection. There was a close correlation between hepatic SOCS3 level and IL-6, and the severity of liver injury defined by alanine aminotransferase (ALT) and aspartate aminotransferase (AST) levels, respectively. These data suggested that SOCS3 may play a pivotal role in the pathogenesis of MHV-3-induced acute liver failure and HBV-ACLF.

The pathogenesis of liver cirrhosis is not completely elucidated. Although in the majority of patients, the risk factors may be identified in B and C viral hepatitis, alcohol intake, drugs or fatty liver disease, there is a small percentage of patients with no apparent risk factors. In addition, the evolution of chronic liver disease is highly heterogeneous from one patient to another. Among patient with identical risk factors, some rapidly progress to cirrhosis and hepatocellular carcinoma (HCC) whereas others have a benign course. Therefore, a genetic predisposition may contribute to the development of cirrhosis and HCC. Evidence supporting the role of genetic factors as a risk for cirrhosis has been accumulating during the past years. In addition to the results from epidemiological studies, polymorphisms studies and data on twins, the concept of telomere shortening as a genetic risk factor for chronic liver disease and HCC has been proposed. Here we review the literature on telomerase mutations, telomere shortening and liver disease including hepatocellular carcinoma.

Background & Aims: Augmented susceptibility to infections increases mortality in patients with end-stage liver disease (ESLD). We sought to determine the contribution of selected genetic variants involved in inflammatory signalling downstream of the Toll-like receptor 4 (TLR4) to severe bacterial infections (SBIs) in patients with ESLD., Methods: We retrospectively assessed incidence of SBIs in 336 adult ESLD patients enlisted for orthotopic liver transplantation (OLT) and genotyped them for TLR4 c.+1196C/T, CD14 c.-159C/T, TNFA c.-238G/A, TNFA c.-863C/A, IL1B c.-31C/T and IL1RN variable number of tandem repeats allelic variants. Principal findings were validated in an independent cohort of 332 ESLD patients., Results: Thirty-four percent of patients from the identification cohort and 40% of patients from the validation cohort presented with SBI while enlisted for OLT. The presence of the variant allele TNFA c.-238A (rs361525) was associated with lower serum levels of TNF-α, and with significantly decreased risk of SBI in both cohorts. Multivariate analysis showed that the relative protection from SBI associated with this allele almost completely negated the increased susceptibility to SBI owed to advanced ESLD. Although not predictive of overall mortality, the presence of the TNFA c.-238A allele was associated with a complete prevention of SBI-related pre-transplant deaths., Conclusions: Our results suggest that genetic variability in inflammatory signalling is associated with the development of SBI in patients with ESLD. Specifically, we identified the importance of the TNFA c.-238A allele as a strong predictor of protection from SBI, and as a genetic marker associated with significantly improved pre-transplant survival in patients with SBI., (Copyright © 2013 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.)

Hepatitis B virus (HBV) infection is one of the major causes of chronic liver inflammation. Tim-3 acts as a negative regulatory molecule and plays a critical role in immune tolerance. In the current study, we investigated Tim-3 expression on peripheral monocytes and CD3+CD16/CD56+ natural killer like T (NKT-like) cells in chronic hepatitis B (CHB) patients. Peripheral blood mononuclear cells (PBMCs) were isolated from 52 CHB patients and 60 healthy controls. Tim-3+CD14+ cells and Tim-3+CD3+CD16/CD56+ cells were analyzed by flow cytometry. Results showed that expression of Tim-3 was significantly increased on both the monocytes and NKT-like cells in CHB patients than in controls (P = 0.002 and P < 0.001, respectively). Tim-3 levels on monocytes and NKT-like cells were further upregulated in patients with acute-on-chronic liver failure (ACLF). In addition, we assessed the correlation of Tim-3 expression with levels of alanine aminotransferase (ALT) and tumor necrosis factor alpha (TNF-α). Data revealed that Tim-3 expression on both monocytes and NKT-like cells was positively correlated with level of ALT (r = 0.59, P < 0.001, and r = 0.60, P < 0.001, respectively), whereas Tim-3 expression on NKT-like cells was negatively correlated with serum level of TNF-α (r = -0.54, P < 0.001) in CHB patients. Our results suggest that Tim-3 may play important roles in the pathogenesis of CHB., (© 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Hepatitis B virus (HBV) infection is one of the major causes of chronic liver inflammation. Toll-like receptor 3 (TLR3) plays a key role in innate immunity and is responsible for recognizing viral pathogens. It has been reported that the TLR3 C1234T polymorphism is associated with various diseases. The aim of this study was to investigate whether TLR3 polymorphisms were correlated with susceptibility to chronic HBV infection. Two polymorphisms in the TLR3 gene, A952T and C1234T, were tested by direct sequencing in 452 chronic hepatitis B (CHB) patients and 462 healthy controls. Data showed that subjects carrying 1234CT genotype and TT genotype had 1.42-fold and 2.31-fold increased risk of chronic HBV infection compared to those with CC genotype (95 % confidence interval [CI] = 1.08-1.86, p = 0.012; 95 % CI = 1.34-3.96, p = 0.002, respectively). Further analysis revealed that the prevalence of 1234CT genotype and T allele was significantly increased in CHB patients with acute-on-chronic liver failure (ACLF) than those without ACLF (odds ratio [OR] = 1.55, p = 0.030; OR = 1.43, p = 0.040, respectively). These results indicate that TLR3 C1234T polymorphism could be a risk factor for the development of chronic HBV infection, especially the CHB-related ACLF.

Liver transplantation represents the treatment of choice for acute hepatic failure or chronically induced end stage of liver disease. Molecular characterization of hepatic injury and recovery processes encloses the key for biomarker discovery in the liver transplantation field. Several pathological hepatic processes were demonstrated to be associated with deregulated miRNA profiles. Moreover, abnormal concentration levels of circulating cell-free miRNAs correlate with specific hepatic injury. The high molecular stability and emerging rapid assessment techniques invite further consideration of miRNAs as feasible monitoring and outcome predictive biomarkers for liver disease and liver transplantation. The present review aims to provide an overview of recent achievements in research on the potential applicability of miRNAs as biomarkers in liver disease and liver transplantation.

High mobility group box chromosomal protein 1 (HMGB1) is an important proinflammatory molecule in many inflammatory disorders, but little is known about its role in acute-on-chronic liver failure (ACLF). Here, we investigated the relationship between the expression of HMGB1 and the disease onset and severity of ACLF patients and mice with acute liver injury/failure induced by concanavalin A (ConA). Peripheral blood mononuclear cells (PBMCs) and serum from ACLF patients were collected, and a mouse model of acute liver injury/failure was induced by ConA. HMGB1 mRNA expression in patient PBMCs or in murine livers and serum HMGB1 protein in ACLF patients and mice were assayed by RT-PCR and Western blotting, respectively. HMGB1 translocation in hepatocytes of ConA-treated mice was assessed by immunohistochemical staining. Up-regulated HMGB1 mRNA levels in PBMCs and accumulated protein in serum were both correlated with disease severity in ACLF patients. In the animal model, HMGB1 levels increased at 4 h and reached its peak value at 8-12 h after challenge with ConA, which suggests that HMGB1 is a relatively late proinflammatory cytokine compared with TNF-α. Translocation of HMGB1 from the nucleus to the cytoplasm in hepatocytes was correlated with the severity of liver injury in mice. While specific anti-HMGB1 antibodies and nicotine protected mice from acute liver injury/failure by reducing mortality and improving liver tissue injury, treatment with recombinant HMGB1 led to an increased mortality due to ConA challenge. Thus, the data from the present study suggest that HMGB1 plays a critical role in the systemic inflammation of ACLF and could be a potential therapeutic target in the treatment of ACLF., (Copyright © 2012 Elsevier Inc. All rights reserved.)

Background: Expression of drug-metabolizing enzymes and drug transporters in liver is mainly regulated by a system of nuclear receptors. The aim of the current study was to investigate the expression of nuclear receptors, as well as these enzymes and transporters, in liver samples from patients suffering from end-stage liver disease of various etiologies (HCV infection, alcohol liver disease, and primary sclerosis cholangitis)., Methods: Gene expression was measured using quantitative real-time PCR with surgical specimens from livers of patients with end-stage liver disease, and non-tumoral liver tissue that served as control., Results: Our study confirmed that the expression of most phase I enzymes is suppressed in end-stage liver disease, and is correlated with a decrease in NR1I2 and NR1I3, the main regulators of xenobiotic metabolism. While mRNA levels of phase II enzymes were generally unchanged, some ABC transporters were up-regulated. The most spectacular increases in expression were observed with ABCC4 (MRP4) - at the mRNA level, and CYP1B1 - at both the mRNA and protein levels. We also demonstrated that IL-6 can induce CYP1B1 expression independently of CYP1A1, in a human hepatocellular liver carcinoma cell line., Conclusions: As CYP1B1 is an enzyme which converts various substrates into carcinogenous metabolites, its overexpression in liver may be one of the factors increasing the risk of hepatic cancers in patients with liver disease. CYP1A1 and CYP1B1 are often referred to as model AHR target genes, but CYP1A1 was down-regulated in diseased liver samples. This points to the existence of differences in regulation of these two genes.

Background: The genetic diversity of chemokines and chemokine receptors has been associated with the outcome of hepatitis B virus infection. The aim of this study was to evaluate whether the copy number variation in the CCL3L1 gene and the polymorphisms of CCR5Δ32 and CCR5-2459A→G (rs1799987) are associated with recurrent hepatitis B in liver transplantation for hepatitis B virus infection-related end-stage liver disease., Methods: A total of 185 transplant recipients were enrolled in this study. The genomic DNA was extracted from whole blood, the copy number of the CCL3L1 gene was determined by a quantitative real-time PCR based assay, CCR5Δ32 was detected by a sizing PCR method, and a single-nucleotide polymorphism in CCR5-2459 was detected by restriction fragment length polymorphism PCR., Results: No CCR5Δ32 mutation was detected in any of the individuals from China. Neither copy number variation nor polymorphism in CCR5-2459 was associated with post-transplant re-infection with hepatitis B virus. However, patients with fewer copies (<4) of the CCL3L1 gene compared with the population median in combination with the CCR5G allele had a significantly higher risk for recurrent hepatitis B (odds ratio=1.93, 95% CI: 1.00-3.69; P=0.047)., Conclusion: Patients possessing the compound decreased functional genotype of both CCL3L1 and CCR5 genes might be more likely to have recurrence of hepatitis B after transplantation.

Objective: To study the clinical characteristics of hepatitis B virus-related acute-on-chronic liver failure patients with familial aggregation., Methods: 275 patients with hepatitis B virus--related acute-on-chronic liver failure were investigated. The patients were divided into familial aggregation and non-familial aggregation group basis on their epidemiological features. Clinical data and biochemical indicators between the two groups were analyzed statistically., Results: 93 of 275 patients (33.82%) case were family aggregation. There was no significant difference compared with chronic hepatitis B patients (38.3%). The mean age of the two groups was 45.98 and 43.61 years old, respectively (P > 0.05). The rates of liver cirrhosis in family aggregation group were significant higher than non-familial aggregation group (73.91% vs 58.24%, p < 0.05). Serum total (TBil) and prothrombin activities (PTA) were no significant difference between the two groups, but ALT level in familial aggregation group was much higher (407.80 U/L vs 256.45 U/L, P 0.05)., Conclusion: Familial aggregation were not related to acute-on-chronic liver failure in chronic HBV hepatitis patients. But the rate of liver cirrhosis were higher in patients with familial aggregation.

Promoter methylation of glutathione-S-transferase P1 (GSTP1) may be involved in liver damage caused by oxidative stress in acute-on-chronic hepatitis B-induced liver failure (ACHBLF). This study aimed to explore GSTP1 promoter methylation status and oxidative stress in such patients. DNA was extracted from peripheral blood mononuclear cells (PBMCs) of patients with acute-on-chronic liver hepatitis B-induced liver failure, chronic hepatitis B (CHB) and normal controls, followed by sodium-bisulfite treatment and methylation-specific PCR (MSP) analysis. Plasma malondialdehyde (MDA) adducts levels were detected by enzyme-linked immunosorbent assay as oxidative stress marker. Model for end-stage liver disease (MELD) score was employed to estimate the severity of the liver failure. Eleven of 35 patients with acute-on-chronic liver failure and 3 of 35 patients with stab le hepatitis B displayed GSTP1 promoter methylation, and the difference was significant (χ2)  = 5.71, P = 0.02). No differences in standard liver function tests were found in patients with acute-on-chronic liver failure with and without GSTP1 promoter methylation although the levels of total bilirubin were greater in those with methylation. The levels of MDA adducts were significantly higher in patients with liver failure when compared to those with CHB (12.44 ± 5.38 pmol/mg vs 8.42 ± 5.49 pmol/mg, P < 0.01), and in the patients with liver failure who had promoter methylation the levels were higher than in those who did not (15.2 ± 4.68 pmol/mg vs 11.17 ± 5.29 pmol/mg, P < 0.01). The MELD score was not significantly different between methylated and unmethylated patients with liver failure (P > 0.05), although MDA adducts were correlated with MELD scores in patients with acute-on-chronic liver failure (r = 0.579, P < 0.01). GSTP1 promoter methylation may facilitate oxidative stress-associated liver damage in ACHBLF, and oxidative stress is correlated with ACHBLF severity., (© 2011 Blackwell Publishing Ltd.)

Aim: TAP2 genes are involved in antigen presentation by MHC class I molecules, especially in the transport of endogenous peptides. As for most MHC genes, a polymorphism has been described and the possibility that it could influence the recipient immune response by modulating antigen presentation in renal disorders. The aim of our study was to analyze TAP2 gene polymorphism in between ESRD (end stage renal disease) patients and healthy controls in our North Indian population., Materials and Method: We analyzed 3 polymorphisms in the TAP2 gene viz. 565 (G→A, Ala→Thr), 379 (G→A, Val→Ile) and 665 (A→G, Thr→Ala), by amplification refractory mutation system-polymerase chain reaction, (ARMS-PCR) methodology using SPSS 11.0 (Chicago, IL, U.S.A.) in a hospital based case-control study in 148 ESRD cases and 230 healthy controls. They were age and sex matched and were of similar ethnicity., Results: Our results showed that only TAP2 G>A 379 Val/Ile was found to be significant for end stage renal disease patients. Furthermore, 2 haplotypes, Ile(379)-Thr(665) and Ile(379)-Ala(665) (p=0.001; OR=0.28 and p=0.030; OR=4.30 respectively) were also associated with the disease and on the other hand gene combination effect revealed protective pattern with ESRD (Val/Ile(379)-Thr/Thr(665) p=0.007; OR=0.30). These results suggested that the currently described polymorphism in the limited coding region of TAP2 genes individually does not influence end stage renal disease risk. It however, proposes a strong link when in combination with each other., Conclusion: The study suggests that the possibility exists that some alleles, in the TAP genes, might confer susceptibility or protection in patients with ESRD which might prove to be helpful in identifying individuals at a higher risk for progressive renal insufficiency., (Copyright © 2011 Elsevier B.V. All rights reserved.)

Background: Accumulating evidence indicates that multiple genetic factors are involved in the pathogenesis of primary biliary cirrhosis (PBC). The aim of this study was to investigate whether polymorphisms of the integrin αV subunit gene (ITGAV), a component of integrin αVβ6, which plays an important role in the process of fibrosis, are associated with susceptibility to the onset and/or progression of PBC., Methods: In the primary study, eight tag single nucleotide polymorphisms (SNPs) in ITGAV were analyzed by polymerase chain reaction (PCR)-restriction fragment length polymorphism, direct DNA sequencing, or high-resolution melting curve analysis in 309 Japanese patients with PBC who were registered in the National Hospital Organization Study Group for Liver Disease in Japan (PBC cohort I) and 293 gender-matched healthy Japanese volunteers (control subjects). For the replication study, 35 PBC patients who progressed to end-stage hepatic failure and underwent liver transplantation (PBC cohort II) were also analyzed., Results: Three tag SNPs (rs3911238, rs10174098, and rs1448427) in ITGAV were significantly associated with the severe progression of PBC, but not with susceptibility to the onset of PBC, in the primary study (PBC cohort I). Among these SNPs, rs1448427 was also significantly associated with the severe progression to end-stage hepatic failure in the replication study of PBC patients who underwent liver transplantation (PBC cohort II)., Conclusions: ITGAV is a genetic determinant for the severe progression of PBC in Japanese patients. Genetic polymorphisms of ITGAV may be useful for identifying high-risk Japanese PBC patients, including those who will require liver transplantation, at the time of initial diagnosis.