Validation of NIAAAm-CRP criteria to predict alcohol-associated steatohepatitis on liver histology.

Background & Aims: In clinical practice, the diagnosis of alcohol-associated hepatitis (AH) is mostly based on non-invasive criteria, which were defined at a consensus conference by the National Institute on Alcohol Abuse and Alcoholism (NIAAA). These criteria were recently modified by adding C-reactive protein (CRP) and termed NIAAAm-CRP criteria, which showed superior diagnostic accuracy for presence of alcohol-associated steatohepatitis (ASH) on liver histology. The aim of our study was to validate the diagnostic accuracy of both original NIAAA criteria and NIAAAm-CRP criteria for presence of ASH on liver histology in an independent cohort. Methods: Data from a large multinational cohort of 445 patients with alcohol-associated liver disease (ALD) that served to establish a novel grading and staging system of alcohol-associated liver disease were analyzed retrospectively. Diagnosis of ASH was based on presence of hepatocyte ballooning plus lobular neutrophil infiltration and established in virtual consensus meetings of multiple expert liver pathologists. Results: Complete data including CRP values were available in 346 patients. Overall diagnostic accuracy for prediction of ASH was 73% for NIAAA criteria and 77% for NIAAAm-CRP criteria. In a subgroup with suspected severe AH (MELD >20, n = 123), overall diagnostic accuracy for prediction of ASH was 69% for NIAAA criteria and 74% for NIAAAm-CRP criteria. Conclusion: Our findings confirm recent data on suboptimal diagnostic accuracy of original NIAAA criteria and validate slightly better but still suboptimal performance of NIAAAm-CRP criteria for presence of ASH. Impact and Implications: Alcohol-associated steatohepatitis (ASH) is diagnosed on liver histology but liver biopsy is not always feasible. Non-invasive diagnosis based on clinical findings has been proposed using the National Institute on Alcohol Abuse and Alcoholism (NIAAA) criteria and recently improved using NIAAAm-CRP criteria. Our findings validate slightly better but still suboptimal performance of NIAAAm-CRP criteria for the presence of histological ASH. Clinical trials of novel drugs should focus on histologically proven ASH.

Effect of albumin infusion on oxidative albumin modification and albumin binding capacity in chronic liver failure.

Oxidative albumin modification and impaired albumin binding function have been described both in chronic liver failure and for therapeutic albumin solutions. The aim of the present study was to evaluate the effect of albumin infusion on redox state and binding function of circulating albumin. We studied 20 patients with cirrhosis who routinely received albumin infusions for prevention of post-paracentesis circulatory dysfunction or treatment of hepatorenal syndrome. We measured albumin fractions by redox state of cysteine-34 and albumin binding properties using dansylsarcosine as site II ligand. Therapeutic albumin solutions showed high contents of human nonmercaptalbumin-1 and human nonmercaptalbumin-2, exceeding the respective values in our patients with decompensated cirrhosis. An initial protocol for the first nine patients sampled at baseline, 24 h and 48 h after albumin infusion revealed no significant changes of oxidized albumin species or albumin binding properties. However, a modified protocol for the remaining 11 patients sampled at baseline, <1 h after and 24 h after albumin infusion revealed short-lived changes of oxidized albumin species while no changes in albumin binding properties were observed. In conclusion, therapeutic albumin infusion transiently changed albumin redox state but did not improve binding function of circulating albumin in chronic liver failure.

Performance of non-invasive tests and histology for the prediction of clinical outcomes in patients with non-alcoholic fatty liver disease: an individual participant data meta-analysis.

BACKGROUND: Histologically assessed liver fibrosis stage has prognostic significance in patients with non-alcoholic fatty liver disease (NAFLD) and is accepted as a surrogate endpoint in clinical trials for non-cirrhotic NAFLD. Our aim was to compare the prognostic performance of non-invasive tests with liver histology in patients with NAFLD. METHODS: This was an individual participant data meta-analysis of the prognostic performance of histologically assessed fibrosis stage (F0-4), liver stiffness measured by vibration-controlled transient elastography (LSM-VCTE), fibrosis-4 index (FIB-4), and NAFLD fibrosis score (NFS) in patients with NAFLD. The literature was searched for a previously published systematic review on the diagnostic accuracy of imaging and simple non-invasive tests and updated to Jan 12, 2022 for this study. Studies were identified through PubMed/MEDLINE, EMBASE, and CENTRAL, and authors were contacted for individual participant data, including outcome data, with a minimum of 12 months of follow-up. The primary outcome was a composite endpoint of all-cause mortality, hepatocellular carcinoma, liver transplantation, or cirrhosis complications (ie, ascites, variceal bleeding, hepatic encephalopathy, or progression to a MELD score ≥15). We calculated aggregated survival curves for trichotomised groups and compared them using stratified log-rank tests (histology: F0-2 vs F3 vs F4; LSM: <10 vs 10 to <20 vs ≥20 kPa; FIB-4: <1·3 vs 1·3 to ≤2·67 vs >2·67; NFS: <-1·455 vs -1·455 to ≤0·676 vs >0·676), calculated areas under the time-dependent receiver operating characteristic curves (tAUC), and performed Cox proportional-hazards regression to adjust for confounding. This study was registered with PROSPERO, CRD42022312226. FINDINGS: Of 65 eligible studies, we included data on 2518 patients with biopsy-proven NAFLD from 25 studies (1126 [44·7%] were female, median age was 54 years [IQR 44-63), and 1161 [46·1%] had type 2 diabetes). After a median follow-up of 57 months [IQR 33-91], the composite endpoint was observed in 145 (5·8%) patients. Stratified log-rank tests showed significant differences between the trichotomised patient groups (p<0·0001 for all comparisons). The tAUC at 5 years were 0·72 (95% CI 0·62-0·81) for histology, 0·76 (0·70-0·83) for LSM-VCTE, 0·74 (0·64-0·82) for FIB-4, and 0·70 (0·63-0·80) for NFS. All index tests were significant predictors of the primary outcome after adjustment for confounders in the Cox regression. INTERPRETATION: Simple non-invasive tests performed as well as histologically assessed fibrosis in predicting clinical outcomes in patients with NAFLD and could be considered as alternatives to liver biopsy in some cases. FUNDING: Innovative Medicines Initiative 2.

Genetic variation in TERT modifies the risk of hepatocellular carcinoma in alcohol-related cirrhosis: results from a genome-wide case-control study.

OBJECTIVE: Hepatocellular carcinoma (HCC) often develops in patients with alcohol-related cirrhosis at an annual risk of up to 2.5%. Some host genetic risk factors have been identified but do not account for the majority of the variance in occurrence. This study aimed to identify novel susceptibility loci for the development of HCC in people with alcohol related cirrhosis. DESIGN: Patients with alcohol-related cirrhosis and HCC (cases: n=1214) and controls without HCC (n=1866), recruited from Germany, Austria, Switzerland, Italy and the UK, were included in a two-stage genome-wide association study using a case-control design. A validation cohort of 1520 people misusing alcohol but with no evidence of liver disease was included to control for possible association effects with alcohol misuse. Genotyping was performed using the InfiniumGlobal Screening Array (V.24v2, Illumina) and the OmniExpress Array (V.24v1-0a, Illumina). RESULTS: Associations with variants rs738409 in PNPLA3 and rs58542926 in TM6SF2 previously associated with an increased risk of HCC in patients with alcohol-related cirrhosis were confirmed at genome-wide significance. A novel locus rs2242652(A) in TERT (telomerase reverse transcriptase) was also associated with a decreased risk of HCC, in the combined meta-analysis, at genome-wide significance (p=6.41×10-9, OR=0.61 (95% CI 0.52 to 0.70). This protective association remained significant after correction for sex, age, body mass index and type 2 diabetes (p=7.94×10-5, OR=0.63 (95% CI 0.50 to 0.79). Carriage of rs2242652(A) in TERT was associated with an increased leucocyte telomere length (p=2.12×10-44). CONCLUSION: This study identifies rs2242652 in TERT as a novel protective factor for HCC in patients with alcohol-related cirrhosis.

Diagnostic accuracy of non-invasive tests for advanced fibrosis in patients with NAFLD: an individual patient data meta-analysis.

OBJECTIVE: Liver biopsy is still needed for fibrosis staging in many patients with non-alcoholic fatty liver disease. The aims of this study were to evaluate the individual diagnostic performance of liver stiffness measurement by vibration controlled transient elastography (LSM-VCTE), Fibrosis-4 Index (FIB-4) and NAFLD (non-alcoholic fatty liver disease) Fibrosis Score (NFS) and to derive diagnostic strategies that could reduce the need for liver biopsies. DESIGN: Individual patient data meta-analysis of studies evaluating LSM-VCTE against liver histology was conducted. FIB-4 and NFS were computed where possible. Sensitivity, specificity and area under the receiver operating curve (AUROC) were calculated. Biomarkers were assessed individually and in sequential combinations. RESULTS: Data were included from 37 primary studies (n=5735; 45% women; median age: 54 years; median body mass index: 30 kg/m2; 33% had type 2 diabetes; 30% had advanced fibrosis). AUROCs of individual LSM-VCTE, FIB-4 and NFS for advanced fibrosis were 0.85, 0.76 and 0.73. Sequential combination of FIB-4 cut-offs (<1.3; ≥2.67) followed by LSM-VCTE cut-offs (<8.0; ≥10.0 kPa) to rule-in or rule-out advanced fibrosis had sensitivity and specificity (95% CI) of 66% (63-68) and 86% (84-87) with 33% needing a biopsy to establish a final diagnosis. FIB-4 cut-offs (<1.3; ≥3.48) followed by LSM cut-offs (<8.0; ≥20.0 kPa) to rule out advanced fibrosis or rule in cirrhosis had a sensitivity of 38% (37-39) and specificity of 90% (89-91) with 19% needing biopsy. CONCLUSION: Sequential combinations of markers with a lower cut-off to rule-out advanced fibrosis and a higher cut-off to rule-in cirrhosis can reduce the need for liver biopsies.

Hepatitis D virus (HDV) prevalence in Austria is low but causes considerable morbidity due to fast progression to cirrhosis.

BACKGROUND: Hepatitis D virus (HDV) coinfection aggravates the course of hepatitis B virus (HBV). The prevalence of HDV in Austria is unknown. OBJECTIVE: This national study aimed at (i) recording the prevalence of HDV-infection in Austria and (ii) characterizing the "active" HDV cohort in Austria. METHODS: A total of 10 hepatitis treatment centers in Austria participated in this multicenter study and retrospectively collected their HDV patients between Q1/2010 and Q4/2020. Positive anti-HDV and/or HDV-RNA-polymerase chain reaction (PCR) results were retrieved from local database queries. Disease severity was assessed by individual chart review. Viremic HDV patients with clinical visits in/after Q1/2019 were considered as the "active" HDV cohort. RESULTS: A total of 347 anti-HDV positive patients were identified. In 202 (58.2%) patients, HDV-RNA-PCR test was performed, and 126/202 (62.4%) had confirmed viremia. Hepatocellular carcinoma was diagnosed in 7 (5.6%) patients, 7 (5.6%) patients underwent liver transplantation, and 11 (8.7%) patients died during follow-up. The "active" Austrian HDV cohort included 74 (58.7%) patients: Evidence for advanced chronic liver disease (ACLD, i.e., histological F3/F4 fibrosis, liver stiffness ≥10 kPa, varices, or hepatic venous pressure gradient ≥6 mmHg) was detected in 38 (51.4%) patients, including 2 (5.3%) with decompensation (ascites/hepatic encephalopathy). About 37 (50.0%) patients of the "active" HDV cohort had previously received interferon treatment. Treatment with the sodium-taurocholate cotransporting polypeptide inhibitor bulevirtide was initiated in 20 (27.0%) patients. CONCLUSION: The number of confirmed HDV viremic cases in Austria is low (<1% of HBV patients) but potentially underestimated. Testing all HBV patients will increase the diagnostic yield. More than half of viremic HDV patients had ACLD. Improved HDV testing and workup strategies will facilitate access to novel antiviral therapies.

Development and prognostic relevance of a histologic grading and staging system for alcohol-related liver disease.

BACKGROUND & AIMS: The SALVE Histopathology Group (SHG) developed and validated a grading and staging system for the clinical and full histological spectrum of alcohol-related liver disease (ALD) and evaluated its prognostic utility in a multinational cohort of 445 patients. METHODS: SALVE grade was described by semiquantitative scores for steatosis, activity (hepatocellular injury and lobular neutrophils) and cholestasis. The histological diagnosis of steatohepatitis due to ALD (histological ASH, hASH) was based on the presence of hepatocellular ballooning and lobular neutrophils. Fibrosis staging was adapted from the Clinical Research Network staging system for non-alcoholic fatty liver disease and the Laennec staging system and reflects the pattern and extent of ALD fibrosis. There are 7 SALVE fibrosis stages (SFS) ranging from no fibrosis to severe cirrhosis. RESULTS: Interobserver κ-value for each grading and staging parameter was >0.6. In the whole study cohort, long-term outcome was associated with activity grade and cholestasis, as well as cirrhosis with very broad septa (severe cirrhosis) (p <0.001 for all parameters). In decompensated ALD, adverse short-term outcome was associated with activity grade, hASH and cholestasis (p = 0.038, 0.012 and 0.001, respectively), whereas in compensated ALD, hASH and severe fibrosis/cirrhosis were associated with decompensation-free survival (p = 0.011 and 0.001, respectively). On multivariable analysis, severe cirrhosis emerged as an independent histological predictor of long-term survival in the whole study cohort. Severe cirrhosis and hASH were identified as independent predictors of short-term survival in decompensated ALD, and also as independent predictors of decompensation-free survival in compensated ALD. CONCLUSION: The SALVE grading and staging system is a reproducible and prognostically relevant method for the histological assessment of disease activity and fibrosis in ALD. LAY SUMMARY: Patients with alcohol-related liver disease (ALD) may undergo liver biopsy to assess disease severity. We developed a system to classify ALD under the microscope by grading ALD activity and staging the extent of liver scarring. We validated the prognostic performance of this system in 445 patients from 4 European centers.

Growing Human Hepatocellular Tumors Undergo a Global Metabolic Reprogramming.

Hepatocellular carcinoma (HCC) is a common malignancy with poor prognosis, high morbidity and mortality concerning with lack of effective diagnosis and high postoperative recurrence. Similar with other cancers, HCC cancer cells have to alter their metabolism to adapt to the changing requirements imposed by the environment of the growing tumor. In less vascularized regions of tumor, cancer cells experience hypoxia and nutrient starvation. Here, we show that HCC undergoes a global metabolic reprogramming during tumor growth. A combined proteomics and metabolomics analysis of paired peritumoral and tumor tissues from 200 HCC patients revealed liver-specific metabolic reprogramming and metabolic alterations with increasing tumor sizes. Several proteins and metabolites associated with glycolysis, the tricarboxylic acid cycle and pyrimidine synthesis were found to be differentially regulated in serum, tumor and peritumoral tissue with increased tumor sizes. Several prognostic metabolite biomarkers involved in HCC metabolic reprogramming were identified and integrated with clinical and pathological data. We built and validated this combined model to discriminate against patients with different recurrence risks. An integrated and comprehensive metabolomic analysis of HCC is provided by our present work. Metabolomic alterations associated with the advanced stage of the disease and poor clinical outcomes, were revealed. Targeting cancer metabolism may deliver effective therapies for HCC.

Long-term outcomes in patients with decompensated alcohol-related liver disease, steatohepatitis and Maddrey's discriminant function <32.

BACKGROUND & AIMS: Patients with alcoholic hepatitis and a modified Maddrey's discriminant function (mDF) <32 have a low risk of short-term mortality. However, few data exist concerning long-term outcomes. The aims of this study were to evaluate 5-year survival rates and to identify predictive factors for long-term prognosis in this patient population. METHODS: We studied patients from 2 centers who were admitted for hepatic decompensation (ascites, hepatic encephalopathy, or jaundice) and who had histological findings of steatohepatitis and an mDF <32. Clinical and biological parameters were recorded at the time of liver biopsy and alcohol consumption was recorded during follow-up. We performed Cox proportional hazard survival analysis to identify factors associated with 5-year survival. RESULTS: One hundred and twenty-one patients were included (male: 64%, mean age: 51.5 ± 10.3 years, presence of cirrhosis: 84%). The median model for end-stage liver disease and mDF scores were 14 (IQR 11.7-16.1) and 19 (IQR 11.1-24), respectively. During follow-up, 30% of the patients remained abstinent. Survival rates at 1, 6, 12, 24, and 60 months were 96.7 ± 1.6%, 90.1 ± 2.7%, 80.8 ± 3.6%, 69.9 ± 4.3%, and 50.7 ± 4.9%, respectively. The majority of deaths (80%) were liver related. In multivariable analysis, encephalopathy at baseline and alcohol abstinence were predictive of 5-year survival. The 5-year survival rates of patients without and with encephalopathy at baseline were 60.5 ± 5.8% and 29.7 ± 8.0%, respectively, and the 5-year survival rates of abstinent and non-abstinent patients were 74.0 ± 8.0% and 40.9 ± 5.8%, respectively. CONCLUSIONS: The mortality rate of patients with alcoholic hepatitis and an mDF <32 is around 50% at 5 years. Hepatic encephalopathy at baseline and lack of alcohol abstinence impair long-term prognosis. New treatment strategies, including measures to ensure abstinence, are required. LAY SUMMARY: Patients with alcoholic hepatitis that is of intermediate severity have a low risk of short-term mortality but not much is known regarding long-term outcomes for these patients. This study clearly indicates that patients with intermediate disease characteristics have poor long-term outcomes. The presence of hepatic encephalopathy at the time of diagnosis and the absence of alcohol abstinence during follow-up are factors that predict poor long-term mortality.

Non-invasive diagnosis of cirrhosis and long-term disease monitoring by transient elastography in patients with Wilson disease.

BACKGROUND & AIMS: The value of liver stiffness measurement (LSM) by transient elastography (TE) for non-invasive fibrosis staging and disease monitoring has not been established in patients with Wilson disease (WD). METHODS: Liver stiffness measurement by TE and non-invasive fibrosis scores (APRI, FIB-4) were analysed from 188 WD patients with liver biopsy (LBX). Longitudinal LSM was performed in 128 (68.1%) patients. RESULTS: One hundred and eighty-eight patients (mean age: 35 ± 14 years, 54.8% women; 27.1% with histological cirrhosis) were studied. Forty-four[23.4%] patients were recently diagnosed with WD, while 144[76.6%] were previously diagnosed (>1 year between LBX and LSM). Overall, LSM (11.3 vs 6.1 kPa, P < .001), APRI (0.72 vs 0.38, P < .001) and FIB-4 (1.54 vs 0.89, P < .001) were higher in cirrhotic than in non-cirrhotic patients. This was even more pronounced in recently diagnosed patients (35.2 kPa vs 6.4 kPa, P < .001). Accuracy for diagnosing cirrhosis at an LSM cut-off ≥9.9 kPa was better in recently diagnosed (PPV: 74%, NPV: 100%) vs previously diagnosed (PPV: 53%, NPV: 82%) patients. Recently diagnosed patients had higher Area Under the Curve (AUC) for APRI (0.79 vs 0.61) and FIB-4 (0.84 vs 0.65) than previously diagnosed patients. At APRI <1.5 and FIB-4 <3.25 cirrhosis was ruled out with a specificity of 93% and 95% respectively. During a median follow-up of 46 (24-66) months, only 5.9% (5/85) of non-cirrhotic WD patients showed progression to cirrhotic LSM values, while 30.8% (4/13) of cirrhotic WD patients showed LSM suggestive of cirrhosis regression. CONCLUSION: TE-based LSM ≥9.9 kPa accurately identifies cirrhosis in WD patients. Next to TE-LSM <9.9 kPa, APRI <1.5 and FIB-4 <3.25 values assist to non-invasively rule out cirrhosis. LSM remains stable in most non-cirrhotic patients on WD therapy, while one-third of cirrhotic patients present clinically relevant decreases in LSM.

Genetic Variation in HSD17B13 Reduces the Risk of Developing Cirrhosis and Hepatocellular Carcinoma in Alcohol Misusers.

BACKGROUND AND AIMS: Carriage of rs738409:G in patatin-like phospholipase domain containing 3 (PNPLA3) is associated with an increased risk for developing alcohol-related cirrhosis and hepatocellular carcinoma (HCC). Recently, rs72613567:TA in hydroxysteroid 17-beta dehydrogenase 13 (HSD17B13) was shown to be associated with a reduced risk for developing alcohol-related liver disease and to attenuate the risk associated with carriage of PNPLA3 rs738409:G. This study explores the risk associations between these two genetic variants and the development of alcohol-related cirrhosis and HCC. APPROACH AND RESULTS: Variants in HSD17B13 and PNPLA3 were genotyped in 6,171 participants, including 1,031 with alcohol-related cirrhosis and HCC, 1,653 with alcohol-related cirrhosis without HCC, 2,588 alcohol misusers with no liver disease, and 899 healthy controls. Genetic associations with the risks for developing alcohol-related cirrhosis and HCC were determined using logistic regression analysis. Carriage of HSD17B13 rs72613567:TA was associated with a lower risk for developing both cirrhosis (odds ratio [OR], 0.79; 95% confidence interval [CI], 0.72-0.88; P = 8.13 × 10-6 ) and HCC (OR, 0.77; 95% CI, 0.68-0.89; P = 2.27 × 10-4 ), whereas carriage of PNPLA3 rs738409:G was associated with an increased risk for developing cirrhosis (OR, 1.70; 95% CI, 1.54-1.88; P = 1.52 × 10-26 ) and HCC (OR, 1.77; 95% CI, 1.58-1.98; P = 2.31 × 10-23 ). These associations remained significant after adjusting for age, sex, body mass index, type 2 diabetes, and country. Carriage of HSD17B13 rs72613567:TA attenuated the risk for developing cirrhosis associated with PNPLA3 rs738409:G in both men and women, but the protective effect against the subsequent development of HCC was only observed in men (ORallelic , 0.75; 95% CI, 0.64-0.87; P = 1.72 × 10-4 ). CONCLUSIONS: Carriage of variants in PNPLA3 and HSD17B13 differentially affect the risk for developing advanced alcohol-related liver disease. A genotypic/phenotypic risk score might facilitate earlier diagnosis of HCC in this population.

Blood metabolomics uncovers inflammation-associated mitochondrial dysfunction as a potential mechanism underlying ACLF.

BACKGROUND & AIMS: Acute-on-chronic liver failure (ACLF), which develops in patients with cirrhosis, is characterized by intense systemic inflammation and organ failure(s). Because systemic inflammation is energetically expensive, its metabolic costs may result in organ dysfunction/failure. Therefore, we aimed to analyze the blood metabolome in patients with cirrhosis, with and without ACLF. METHODS: We performed untargeted metabolomics using liquid chromatography coupled to high-resolution mass spectrometry in serum from 650 patients with AD (acute decompensation of cirrhosis, without ACLF), 181 with ACLF, 43 with compensated cirrhosis, and 29 healthy individuals. RESULTS: Of the 137 annotated metabolites identified, 100 were increased in patients with ACLF of any grade, relative to those with AD, and 38 comprised a distinctive blood metabolite fingerprint for ACLF. Among patients with ACLF, the intensity of the fingerprint increased across ACLF grades, and was similar in patients with kidney failure and in those without, indicating that the fingerprint reflected not only decreased kidney excretion but also altered cell metabolism. The higher the ACLF-associated fingerprint intensity, the higher the plasma levels of inflammatory markers, tumor necrosis factor α, soluble CD206, and soluble CD163. ACLF was characterized by intense proteolysis and lipolysis; amino acid catabolism; extra-mitochondrial glucose metabolism through glycolysis, pentose phosphate, and D-glucuronate pathways; depressed mitochondrial ATP-producing fatty acid ß-oxidation; and extra-mitochondrial amino acid metabolism giving rise to metabotoxins. CONCLUSIONS: In ACLF, intense systemic inflammation is associated with blood metabolite accumulation and profound alterations in major metabolic pathways, in particular inhibition of mitochondrial energy production, which may contribute to the development of organ failures. LAY SUMMARY: Acute-on-chronic liver failure (ACLF), which develops in patients with cirrhosis, is characterized by intense systemic inflammation and organ failure(s). Because systemic inflammation is energetically expensive, its metabolic costs may result in organ dysfunction/failure. We identified a 38-metabolite blood fingerprint specific for ACLF that revealed mitochondrial dysfunction in peripheral organs. This may contribute to organ failures.

Evaluation and comparison of six noninvasive tests for prediction of significant or advanced fibrosis in nonalcoholic fatty liver disease.

Background: In nonalcoholic fatty liver disease (NAFLD), advanced fibrosis has been identified as an important prognostic factor with increased liver-related mortality and treatment need. Due to the high prevalence of NAFLD, noninvasive risk stratification is needed to select patients for liver biopsy and treatment. Objective: To compare the diagnostic accuracy of several widely available noninvasive tests for assessment of fibrosis among patients with NAFLD with or without nonalcoholic steatohepatitis (NASH). Methods: We enrolled consecutive patients with NAFLD admitted to two Austrian referral centers who underwent liver biopsy. Liver stiffness measurement (LSM) was obtained by vibration-controlled transient elastography (VCTE, FibroScan) and blood samples were collected for determination of enhanced liver fibrosis (ELF) test, FibroMeterV2G, FibroMeterV3G, NAFLD fibrosis score (NFS), and fibrosis-4 index (FIB-4). Results: Our study cohort contained 186 patients with histologically confirmed NAFLD. On liver histology, NASH was present in 92 patients (50%), significant fibrosis (F ≥ 2) in 71 patients (38%), advanced fibrosis (F ≥ 3) in 49 patients (26%), and F ≥ 3 plus NASH in 35 patients (19%). For diagnosis of F ≥ 2, F ≥ 3, and F ≥ 3 plus NASH, respectively, receiver operating characteristic (ROC) analysis revealed superior diagnostic accuracy of ELF score (area under ROC curve (AUROC) 0.85, 0.90, 0.90), FibroMeterV2G (AUROC 0.86, 0.88, 0.89), FibroMeterV3G (AUROC 0.84, 0.88, 0.88), and LSM per protocol (AUROC 0.87, 0.95, 0.91) versus FIB-4 (AUROC 0.80, 0.82, 0.81) or NFS (AUROC 0.78, 0.80, 0.79). Conclusion: Proprietary fibrosis panels and VCTE show superior diagnostic accuracy for noninvasive diagnosis of fibrosis stage in NAFLD as compared to FIB-4 and NFS.

HSD17B13 truncated variant is associated with a mild hepatic phenotype in Wilson's Disease.

HSD17B13 encodes hydroxysteroid 17-ß dehydrogenase 13, a novel liver lipid-droplet associated protein that is involved in the regulation of lipid biosynthetic processes. A protein-truncating HSD17B13 variant (rs72613567) was shown to protect individuals from alcoholic and non-alcoholic liver disease. Since steatosis is a common feature in Wilson's disease (WD), we aimed to assess whether the HSD17B13 variant modulates the phenotypic presentation and progression of WD. METHODS: The HSD17B13:TA (rs72613567) variant was determined by allelic discrimination real-time PCR in 586 patients. The HSD17B13 genotype was correlated with the phenotypic presentation. The age of onset and the type of symptoms at presentation were used as markers of the WD phenotype. RESULTS: The overall HSD17B13:TA allele frequency in patients with WD was 23.3% (273/1,172), not significantly different from the reported minor allele frequency. There was a significantly lower HSD17B13:TA allele frequency in patients with fulminant WD compared to all other phenotypic WD groups (11.0% vs. 24.0%, p < 0.01). Among the patients with fulminant WD there was a trend for a gender effect; none of the male patients carried the HSD17B13:TA allele. HSD17B13:TA allele frequency was more common in patients with minimal or no fibrosis (49 [31.1%] had simple steatosis and 20 minimal changes at biopsy) than in patients with cirrhosis or advanced fibrosis (22.3%, p = 0.025). CONCLUSIONS: The HSD17B13:TA allele modulates the phenotype and outcome of WD. This allele likely ameliorates hepatic fibrosis and reduces the transition from copper induced hemolysis to fulminant disease in patients with WD. LAY SUMMARY: Wilson's disease is a hereditary disease caused by accumulation of copper in the liver and other tissues. It presents with a variety of clinical symptoms. In this study we explored the role of a recently described gene mutation (HSD17B13:TA) which apparently protects the liver against toxins like alcohol. The results indicate that this mutation plays a role in the evolution of liver disease. Patients with Wilson's disease who carry this mutation are more likely to have mild disease, while the absence of the mutation is associated with the most severe form - fulminant Wilson's disease.

Oxidized Albumin Triggers a Cytokine Storm in Leukocytes Through P38 Mitogen-Activated Protein Kinase: Role in Systemic Inflammation in Decompensated Cirrhosis.

Decompensated cirrhosis is characterized by exuberant systemic inflammation. Although the inducers of this feature remain unknown, the presence of circulating forms of oxidized albumin, namely human nonmercaptalbumin 1 (HNA1) and HNA2, is a common finding in cirrhosis. The aim of this study was to explore the ability of these oxidized albumin forms to induce systemic inflammation by triggering the activation of peripheral leukocytes. We observed significantly higher plasma levels of HNA1 and HNA2 in patients with cirrhosis (n = 256) compared to healthy volunteers (n = 48), which gradually increased during the course from compensated to decompensated to acute-on-chronic liver failure. Plasma HNA1 and HNA2 levels significantly correlated with inflammatory markers (i.e., interleukin-6 [IL-6], IL-1ß, tumor necrosis factor-alpha [TNF-α] and IL-8) in patients with cirrhosis. To directly test the inflammatory effects of HNA1 and HNA2 on leukocytes, these oxidized albumin forms were prepared ex vivo and their posttranslational modifications monitored by liquid chromatography (LC)-quadrupole time-of-flight/mass spectrometry (MS). HNA1, but not HNA2, increased IL-1ß, IL-6, and TNF-α mRNA and protein expression in leukocytes from both healthy volunteers and patients with cirrhosis. Moreover, HNA1 up-regulated the expression of eicosanoid-generating enzymes (i.e., cyclooxygenase-2 [COX-2] and microsomal prostaglandin E [PGE] synthase 1) and the production of inflammatory eicosanoids (PGE2 , PGF2α , thromboxane B2 , and leukotriene B4 ), as determined by LC-electrospray ionization-MS/MS. The inflammatory response to HNA1 was more pronounced in peripheral blood mononuclear cells (PBMCs) and marginal in polymorphonuclear neutrophils. Kinome analysis of PBMCs revealed that HNA1 induced the phosphorylation of p38 mitogen-activated protein kinase, the inhibition of which blocked HNA1-induced cytokine and COX-2 induction. Conclusion: HNA1 triggers an inflammatory response in PBMCs, providing a rationale for its removal and replacement by reduced albumin in the prevention of systemic inflammation in patients with advanced liver disease.

Lysyl oxidase-like protein 2 (LOXL2) modulates barrier function in cholangiocytes in cholestasis.

BACKGROUND & AIMS: The lysyl oxidase-like protein 2 (LOXL2) promotes stabilization of the extracellular matrix, chemotaxis, cell growth and cell mobility. We aimed to (i) identify stimuli of LOXL2 in cholangiopathies, (ii) characterize the effects of LOXL2 on biliary epithelial cells' (BECs) barrier function, (iii) compare LOXL2 expression in primary sclerosing cholangitis (PSC), primary biliary cholangitis, and disease controls, and (iv) to determine LOXL2 expression and its cellular sources in four mouse models of cholangiopathies. METHODS: Cultured murine BECs were challenged with well-known triggers of cellular senescence, hypoxia, phospholipid-deficient Abcb4-/- mouse bile and chenodeoxycholic acid and investigated for LOXL2, SNAIL1 and E-cadherin expression and transepithelial electrical resistance with and without LOX-inhibition. In vivo, LOXL2 expression was studied in PSC livers, and controls and mouse models. We compared LOXL2 serum levels in patients with PSC, secondary SC, primary biliary cholangitis, and controls. RESULTS: Cellular senescence, hypoxia, Abcb4-/- bile and chenodeoxycholic acid induced LOXL2 and SNAIL1 expression, repressed E-cadherin expression, and significantly reduced transepithelial electrical resistance in BECs. Notably, all of the pathological changes could be recovered via pharmacological LOX-inhibition. Mouse models showed induced LOXL2 expression in the portal region and in association with ductular reaction. LOXL2 serum levels were significantly elevated in patients with cholangiopathies. In PSC, LOXL2 expression was located to characteristic periductal onion skin-type fibrosis, ductular reaction, Kupffer cells, and fibrotic septa. Importantly, in PSC, LOXL2 overexpression was paralleled by E-cadherin loss in BECs from medium-sized bile ducts. CONCLUSIONS: Reactive BECs produce LOXL2, resulting in increased tight junction permeability, which can be ameliorated by pharmacological LOX-inhibition in vitro. Reactive BECs, portal myofibroblasts, and Kupffer cells are the main sources of LOXL2 in cholangiopathies. LAY SUMMARY: In this study, we investigate the role of lysyl oxidase-like protein 2 (LOXL2), an enzyme pivotal in the development of organ fibrosis, in the pathogenesis of cholangiopathies (diseases of bile ducts), such as primary sclerosing cholangitis. We found LOXL2 to be expressed in association with bile duct epithelial injury and uncovered mechanisms for its upregulation and the subsequent effects in vitro and in vivo. Our findings support testing of anti-LOXL2 treatment strategies for patients with primary sclerosing cholangitis.

Sorafenib in Patients with Hepatocellular Carcinoma-Results of the Observational INSIGHT Study.

Purpose: Sorafenib is the only currently approved systemic therapy for advanced hepatocellular carcinoma (HCC). We aimed to evaluate the safety and efficacy of sorafenib therapy in patients with HCC under real-life conditions regarding patient, tumor characteristics, and any adverse events at study entry and at follow-up visits every 2 to 4 months.Experimental Design: The current INSIGHT study is a noninterventional, prospective, multicenter, observational study performed in 124 sites across Austria and Germany between 2008 and 2014.Results: Median overall survival and time to progression (RECIST) were found to be dependent on baseline Barcelona Clinic Liver Cancer (BCLC) tumor stage (A: 29.2, B: 19.6, C: 13.6, D: 3.1 and A: 6.0, B: 5.5, C: 3.9, and D: 1.7 months, respectively), Child-Pugh liver function (A: 17.6, B: 8.1, C: 5.6 and A: 5.3, B: 3.3, C: 2.5 months, respectively), and performance status of the patient; however, age did not affect prognosis. Sorafenib-related adverse events at any grade occurred in 64.9% of patients, with diarrhea (35.4%), hand-foot-skin reaction (16.6%), nausea (10.3%), and fatigue (11.2%) occurring most frequently.Conclusions: Sorafenib treatment was shown to be effective in a real-life setting, in agreement with previously reported clinical trial data. The therapy was found to have an acceptable safety profile, with predominantly mild to moderate side effects. Clin Cancer Res; 23(19); 5720-8. ©2017 AACR.

Lack of ABCG2 Leads to Biventricular Dysfunction and Remodeling in Response to Hypoxia.

Aims: The ATP-binding cassette (ABC)G2 transporter protects the heart from pressure overload-induced ventricular dysfunction but also protects cancer cells from chemotherapeutic agents. It is upregulated in the myocardium of heart failure patients and clears hypoxia-induced intracellular metabolites. This study employs ABCG2 knockout (KO) mice to elucidate the relevance of ABCG2 for cardiac and pulmonary vascular structure and function in chronic hypoxia, and uses human primary cardiac fibroblasts to investigate the potential role of ABCG2 in cardiac fibrosis. Methods and results: ABCG2 KO and control mice (n = 10) were subjected to 4 weeks normoxia or hypoxia. This allowed for investigation of the interaction between genotype and hypoxia (GxH). In hypoxia, KO mice showed pronounced right (RV) and left (LV) ventricular diastolic dysfunction. Compared to normoxia, end-diastolic pressure (EDP) was increased in control vs. KO mice by +1.1 ± 0.3 mmHg vs. +4.8 ± 0.3 mmHg, p for GxH < 0.001 (RV) and +3.9 ± 0.5 mmHg vs. +11.5 ± 1.6 mmHg, p for GxH = 0.110 (LV). The same applied for myocardial fibrosis with +0.3 ± 0.1% vs. 1.3 ± 0.2%, p for GxH = 0.036 (RV) and +0.06 ± 0.03% vs. +0.36 ± 0.08%, p for GxH = 0.002 (LV), whereas systolic function and capillary density was unaffected. ABCG2 deficiency did not influence hypoxia-induced pulmonary hypertension or vascular remodeling. In line with these observations, human cardiac fibroblasts showed increased collagen production upon ABCG2 silencing in hypoxia (p for GxH = 0.04). Conclusion: Here we provide evidence for the first time that ABCG2 membrane transporter can play a crucial role in ventricular dysfunction and fibrosis in hypoxia-induced pulmonary hypertension.

Histological parameters and alcohol abstinence determine long-term prognosis in patients with alcoholic liver disease.

BACKGROUND & AIMS: Few data exist on predictors of long-term prognosis in patients with alcoholic liver disease (ALD). Most studies have only assessed short-term prognosis in patients with advanced ALD. We aimed to assess the prognostic impact of clinical, biochemical and histological parameters on long-term prognosis in patients with early/compensated and decompensated ALD. METHODS: Consecutive patients (n=192) with biopsy-proven liver disease due to alcohol abuse were analyzed retrospectively. Prognostic factors were evaluated in patients with early/compensated ALD (n=60) and in patients with decompensated ALD (clinical decompensation and/or bilirubin >3mg/dl at entry) (n=132). Factors that predict long-term survival were identified using Cox regression models. RESULTS: Liver-related mortality at 5years was 13% in early/compensated and 43% in decompensated ALD. In early/compensated ALD patients, long-term prognosis was determined by fibrosis stage, but not by clinical or biochemical variables. Severe fibrosis (F3/4) was present in 52% and had a major impact on 10-year mortality (F3/4: 45% vs. F0-2: 0%, p<0.001). In contrast, in decompensated patients, a combination of clinical features (sex), biochemical markers of liver failure (bilirubin, international normalized ratio [INR]), and histological features (pericellular fibrosis) predicted long-term survival. During follow-up, abstinence from alcohol was an important predictor of survival in both early/compensated and decompensated ALD. CONCLUSION: Fibrosis stage is the main predictor of long-term survival in patients with early/compensated ALD, while clinical, biochemical and histological parameters predict survival in patients with decompensated disease. Promoting abstinence may improve survival in patients with both early and advanced ALD. LAY SUMMARY: In this study, we evaluated long-term outcome in 192 patients with alcoholic liver disease who underwent liver biopsy: 60 patients with early disease (no symptoms) and 132 patients with advanced disease (jaundice, complications of cirrhosis). Importantly, half of the patients with 'early' disease already had severe fibrosis or cirrhosis on liver histology and dismal outcome (45% mortality at 10years). Abstinence from alcohol improved the prognosis in both early and advanced stages of the disease.

Daclatasvir combined with sofosbuvir or simeprevir in liver transplant recipients with severe recurrent hepatitis C infection.

Daclatasvir (DCV) is a potent, pangenotypic nonstructural protein 5A inhibitor with demonstrated antiviral efficacy when combined with sofosbuvir (SOF) or simeprevir (SMV) with or without ribavirin (RBV) in patients with chronic hepatitis C virus (HCV) infection. Herein, we report efficacy and safety data for DCV-based all-oral antiviral therapy in liver transplantation (LT) recipients with severe recurrent HCV. DCV at 60 mg/day was administered for up to 24 weeks as part of a compassionate use protocol. The study included 97 LT recipients with a mean age of 59.3 ± 8.2 years; 93% had genotype 1 HCV and 31% had biopsy-proven cirrhosis between the time of LT and the initiation of DCV. The mean Model for End-Stage Liver Disease (MELD) score was 13.0 ± 6.0, and the proportion with Child-Turcotte-Pugh (CTP) A/B/C was 51%/31%/12%, respectively. Mean HCV RNA at DCV initiation was 14.3 × 6 log10 IU/mL, and 37% had severe cholestatic HCV infection. Antiviral regimens were selected by the local investigator and included DCV+SOF (n = 77), DCV+SMV (n = 18), and DCV+SMV+SOF (n = 2); 35% overall received RBV. At the end of treatment (EOT) and 12 weeks after EOT, 88 (91%) and 84 (87%) patients, respectively, were HCV RNA negative or had levels <43 IU/mL. CTP and MELD scores significantly improved between DCV-based treatment initiation and last contact. Three virological breakthroughs and 2 relapses occurred in patients treated with DCV+SMV with or without RBV. None of the 8 patient deaths (6 during and 2 after therapy) were attributed to therapy. In conclusion, DCV-based all-oral antiviral therapy was well tolerated and resulted in a high sustained virological response in LT recipients with severe recurrent HCV infection. Most treated patients experienced stabilization or improvement in their clinical status.