Glycated haemoglobin is a major predictor of disease severity in patients with NAFLD.

OBJECTIVES: Currently, non-invasive scoring systems to stage the severity of non-alcoholic fatty liver disease (NAFLD) do not consider markers of glucose control (glycated haemoglobin, HbA1c); this study aimed to define the relationship between HbA1c and NAFLD severity in patients with and without type 2 diabetes. RESEARCH DESIGN AND METHODS: Data were obtained from 857 patients with liver biopsy staged NAFLD. Generalized-linear models and binomial regression analysis were used to define the relationships between histological NAFLD severity, age, HbA1c, and BMI. Paired biopsies from interventional studies (n = 421) were used to assess the impact of change in weight, HbA1c and active vs. placebo treatment on improvements in steatosis, non-alcoholic steatohepatitis (NASH), and fibrosis. RESULTS: In the discovery cohort (n = 687), risk of severe steatosis, NASH and advanced fibrosis correlated positively with HbA1c, after adjustment for obesity and age. These data were endorsed in a separate validation cohort (n = 170). Predictive modelling using HbA1c and age was non-inferior to the established non-invasive biomarker, Fib-4, and allowed the generation of HbA1c, age, and BMI adjusted risk charts to predict NAFLD severity. Following intervention, reduction in HbA1c was associated with improvements in steatosis and NASH after adjustment for weight change and treatment, whilst fibrosis change was only associated with weight change and treatment. CONCLUSIONS: HbA1c is highly informative in predicting NAFLD severity and contributes more than BMI. Assessments of HbA1c must be a fundamental part of the holistic assessment of patients with NAFLD and, alongside age, can be used to identify patients with highest risk of advanced disease.

Translating the potential of the urine steroid metabolome to stage NAFLD (TrUSt-NAFLD): study protocol for a multicentre, prospective validation study.

INTRODUCTION: Non-alcoholic fatty liver disease (NAFLD) affects approximately one in four individuals and its prevalence continues to rise. The advanced stages of NAFLD with significant liver fibrosis are associated with adverse morbidity and mortality outcomes. Currently, liver biopsy remains the 'gold-standard' approach to stage NAFLD severity. Although generally well tolerated, liver biopsies are associated with significant complications, are resource intensive, costly, and sample only a very small area of the liver as well as requiring day case admission to a secondary care setting. As a result, there is a significant unmet need to develop non-invasive biomarkers that can accurately stage NAFLD and limit the need for liver biopsy. The aim of this study is to validate the use of the urine steroid metabolome as a strategy to stage NAFLD severity and to compare its performance against other non-invasive NAFLD biomarkers. METHODS AND ANALYSIS: The TrUSt-NAFLD study is a multicentre prospective test validation study aiming to recruit 310 patients with biopsy-proven and staged NAFLD across eight centres within the UK. 150 appropriately matched control patients without liver disease will be recruited through the Oxford Biobank. Blood and urine samples, alongside clinical data, will be collected from all participants. Urine samples will be analysed by liquid chromatography-tandem mass spectroscopy to quantify a panel of predefined steroid metabolites. A machine learning-based classifier, for example, Generalized Matrix Relevance Learning Vector Quantization that was trained on retrospective samples, will be applied to the prospective steroid metabolite data to determine its ability to identify those patients with advanced, as opposed to mild-moderate, liver fibrosis as a consequence of NAFLD. ETHICS AND DISSEMINATION: Research ethical approval was granted by West Midlands, Black Country Research Ethics Committee (REC reference: 21/WM/0177). A substantial amendment (TrUSt-NAFLD-SA1) was approved on 26 November 2021. TRIAL REGISTRATION NUMBER: ISRCTN19370855.

Noninvasive assessment of liver disease severity in patients with nonalcoholic fatty liver disease (NAFLD) and type 2 diabetes.

BACKGROUND AND AIMS: We evaluated the diagnostic accuracy of simple, noninvasive tests (NITs) in NAFLD patients with type 2 diabetes (T2D). METHODS AND RESULTS: This was an individual patient data meta-analysis of 1780 patients with biopsy-proven NAFLD and T2D. The index tests of interest were FIB-4, NAFLD Fibrosis Score (NFS), aspartate aminotransferase-to-platelet ratio index, liver stiffness measurement (LSM) by vibration-controlled transient elastography, and AGILE 3+. The target conditions were advanced fibrosis, NASH, and fibrotic NASH(NASH plus F2-F4 fibrosis). The diagnostic performance of noninvasive tests. individually or in sequential combination, was assessed by area under the receiver operating characteristic curve and by decision curve analysis. Comparison with 2278 NAFLD patients without T2D was also made. In NAFLD with T2D LSM and AGILE 3+ outperformed, both NFS and FIB-4 for advanced fibrosis (area under the receiver operating characteristic curve:LSM 0.82, AGILE 3+ 0.82, NFS 0.72, FIB-4 0.75, aspartate aminotransferase-to-platelet ratio index 0.68; p < 0.001 of LSM-based versus simple serum tests), with an uncertainty area of 12%-20%. The combination of serum-based with LSM-based tests for advanced fibrosis led to a reduction of 40%-60% in necessary LSM tests. Decision curve analysis showed that all scores had a modest net benefit for ruling out advanced fibrosis at the risk threshold of 5%-10% of missing advanced fibrosis. LSM and AGILE 3+ outperformed both NFS and FIB-4 for fibrotic NASH (area under the receiver operating characteristic curve:LSM 0.79, AGILE 3+ 0.77, NFS 0.71, FIB-4 0.71; p < 0.001 of LSM-based versus simple serum tests). All noninvasive scores were suboptimal for diagnosing NASH. CONCLUSIONS: LSM and AGILE 3+ individually or in low availability settings in sequential combination after FIB-4 or NFS have a similar good diagnostic accuracy for advanced fibrosis and an acceptable diagnostic accuracy for fibrotic NASH in NAFLD patients with T2D.

Biomarkers for staging fibrosis and non-alcoholic steatohepatitis in non-alcoholic fatty liver disease (the LITMUS project): a comparative diagnostic accuracy study.

BACKGROUND: The reference standard for detecting non-alcoholic steatohepatitis (NASH) and staging fibrosis-liver biopsy-is invasive and resource intensive. Non-invasive biomarkers are urgently needed, but few studies have compared these biomarkers in a single cohort. As part of the Liver Investigation: Testing Marker Utility in Steatohepatitis (LITMUS) project, we aimed to evaluate the diagnostic accuracy of 17 biomarkers and multimarker scores in detecting NASH and clinically significant fibrosis in patients with non-alcoholic fatty liver disease (NAFLD) and identify their optimal cutoffs as screening tests in clinical trial recruitment. METHODS: This was a comparative diagnostic accuracy study in people with biopsy-confirmed NAFLD from 13 countries across Europe, recruited between Jan 6, 2010, and Dec 29, 2017, from the LITMUS metacohort of the prospective European NAFLD Registry. Adults (aged ≥18 years) with paired liver biopsy and serum samples were eligible; those with excessive alcohol consumption or evidence of other chronic liver diseases were excluded. The diagnostic accuracy of the biomarkers was expressed as the area under the receiver operating characteristic curve (AUC) with liver histology as the reference standard and compared with the Fibrosis-4 index for liver fibrosis (FIB-4) in the same subgroup. Target conditions were the presence of NASH with clinically significant fibrosis (ie, at-risk NASH; NAFLD Activity Score ≥4 and F≥2) or the presence of advanced fibrosis (F≥3), analysed in all participants with complete data. We identified thres holds for each biomarker for reducing the number of biopsy-based screen failures when recruiting people with both NASH and clinically significant fibrosis for future trials. FINDINGS: Of 1430 participants with NAFLD in the LITMUS metacohort with serum samples, 966 (403 women and 563 men) were included after all exclusion criteria had been applied. 335 (35%) of 966 participants had biopsy-confirmed NASH and clinically significant fibrosis and 271 (28%) had advanced fibrosis. For people with NASH and clinically significant fibrosis, no single biomarker or multimarker score significantly reached the predefined AUC 0·80 acceptability threshold (AUCs ranging from 0·61 [95% CI 0·54-0·67] for FibroScan controlled attenuation parameter to 0·81 [0·75-0·86] for SomaSignal), with accuracy mostly similar to FIB-4. Regarding detection of advanced fibrosis, SomaSignal (AUC 0·90 [95% CI 0·86-0·94]), ADAPT (0·85 [0·81-0·89]), and FibroScan liver stiffness measurement (0·83 [0·80-0·86]) reached acceptable accuracy. With 11 of 17 markers, histological screen failure rates could be reduced to 33% in trials if only people who were marker positive had a biopsy for evaluating eligibility. The best screening performance for NASH and clinically significant fibrosis was observed for SomaSignal (number needed to test [NNT] to find one true positive was four [95% CI 4-5]), then ADAPT (six [5-7]), MACK-3 (seven [6-8]), and PRO-C3 (nine [7-11]). INTERPRETATION: None of the single markers or multimarker scores achieved the predefined acceptable AUC for replacing biopsy in detecting people with both NASH and clinically significant fibrosis. However, several biomarkers could be applied in a prescreening strategy in clinical trial recruitment. The performance of promising markers will be further evaluated in the ongoing prospective LITMUS study cohort. FUNDING: The Innovative Medicines Initiative 2 Joint Undertaking.

Association of changes in histologic severity of nonalcoholic steatohepatitis and changes in patient-reported quality of life.

Nonalcoholic steatohepatitis (NASH) is a prevalent chronic disease that is associated with a spectrum of liver fibrosis and can lead to cirrhosis. Patients with NASH report lower health-related quality of life (HRQoL) than the general population. It remains uncertain how changes in histologic severity are associated with changes in HRQoL. This is a secondary analysis of the Farnesoid X Receptor Ligand Obeticholic Acid in NASH Treatment (FLINT) and Pioglitazone, Vitamin E, or Placebo for Nonalcoholic Steatohepatitis (PIVENS) randomized controlled trials in patients with biopsy-proven NASH. HRQoL was assessed using short form-36 at baseline and at follow-up biopsy (at 72 and 96 weeks, respectively). Adjusted linear regression models were used to examine the association between changes in liver fibrosis (primary analysis), nonalcoholic fatty liver disease (NAFLD) activity score (secondary analysis), and changes in HRQoL scores. Compared with stable fibrosis, improvement of fibrosis by at least one stage was significantly associated with improvements only in the physical function component by 1.8 points (95% confidence interval, 0.1, 3.5). Worsening of fibrosis by at least one stage was not associated with statistically significant changes in any HRQoL domain compared with stable fibrosis. Associations between HRQoL and NAFLD disease activity score in the secondary analysis were of similar magnitude. Weight loss was associated with small improvements in physical function, general health, and energy levels. Conclusion: Improvements in fibrosis stage were associated with improvements in the physical component of HRQoL, but the clinical impact was modest. As improving fibrosis may not meaningfully improve well-being, treatment for NASH will be cost effective only if it prevents long-term hepatic and cardiovascular disease.

Infection with the hepatitis C virus causes viral genotype-specific differences in cholesterol metabolism and hepatic steatosis.

Lipids play essential roles in the hepatitis C virus (HCV) life cycle and patients with chronic HCV infection display disordered lipid metabolism which resolves following successful anti-viral therapy. It has been proposed that HCV genotype 3 (HCV-G3) infection is an independent risk factor for hepatocellular carcinoma and evidence suggests lipogenic proteins are involved in hepatocarcinogenesis. We aimed to characterise variation in host lipid metabolism between participants chronically infected with HCV genotype 1 (HCV-G1) and HCV-G3 to identify likely genotype-specific differences in lipid metabolism. We combined several lipidomic approaches: analysis was performed between participants infected with HCV-G1 and HCV-G3, both in the fasting and non-fasting states, and after sustained virological response (SVR) to treatment. Sera were obtained from 112 fasting patients (25% with cirrhosis). Serum lipids were measured using standard enzymatic methods. Lathosterol and desmosterol were measured by gas-chromatography mass spectrometry (MS). For further metabolic insight on lipid metabolism, ultra-performance liquid chromatography MS was performed on all samples. A subgroup of 13 participants had whole body fat distribution determined using in vivo magnetic resonance imaging and spectroscopy. A second cohort of (non-fasting) sera were obtained from HCV Research UK for comparative analyses: 150 treatment naïve patients and 100 non-viraemic patients post-SVR. HCV-G3 patients had significantly decreased serum apoB, non-HDL cholesterol concentrations, and more hepatic steatosis than those with HCV-G1. HCV-G3 patients also had significantly decreased serum levels of lathosterol, without significant reductions in desmosterol. Lipidomic analysis showed lipid species associated with reverse cholesterol transport pathway in HCV-G3. We demonstrated that compared to HCV-G1, HCV-G3 infection is characterised by low LDL cholesterol levels, with preferential suppression of cholesterol synthesis via lathosterol, associated with increasing hepatic steatosis. The genotype-specific lipid disturbances may shed light on genotypic variations in liver disease progression and promotion of hepatocellular cancer in HCV-G3.

Increased serum miR-193a-5p during non-alcoholic fatty liver disease progression: Diagnostic and mechanistic relevance.

BACKGROUND & AIMS: Serum microRNA (miRNA) levels are known to change in non-alcoholic fatty liver disease (NAFLD) and may serve as useful biomarkers. This study aimed to profile miRNAs comprehensively at all NAFLD stages. METHODS: We profiled 2,083 serum miRNAs in a discovery cohort (183 cases with NAFLD representing the complete NAFLD spectrum and 10 population controls). miRNA libraries generated by HTG EdgeSeq were sequenced by Illumina NextSeq. Selected serum miRNAs were profiled in 372 additional cases with NAFLD and 15 population controls by quantitative reverse transcriptase PCR. RESULTS: Levels of 275 miRNAs differed between cases and population controls. Fewer differences were seen within individual NAFLD stages, but miR-193a-5p consistently showed increased levels in all comparisons. Relative to NAFL/non-alcoholic steatohepatitis (NASH) with mild fibrosis (stage 0/1), 3 miRNAs (miR-193a-5p, miR-378d, and miR378d) were increased in cases with NASH and clinically significant fibrosis (stages 2-4), 7 (miR193a-5p, miR-378d, miR-378e, miR-320b, miR-320c, miR-320d, and miR-320e) increased in cases with NAFLD activity score (NAS) 5-8 compared with lower NAS, and 3 (miR-193a-5p, miR-378d, and miR-378e) increased but 1 (miR-19b-3p) decreased in steatosis, activity, and fibrosis (SAF) activity score 2-4 compared with lower SAF activity. The significant findings for miR-193a-5p were replicated in the additional cohort with NAFLD. Studies in Hep G2 cells showed that following palmitic acid treatment, miR-193a-5p expression decreased significantly. Gene targets for miR-193a-5p were investigated in liver RNAseq data for a case subgroup (n = 80); liver GPX8 levels correlated positively with serum miR-193a-5p. CONCLUSIONS: Serum miR-193a-5p levels correlate strongly with NAFLD activity grade and fibrosis stage. MiR-193a-5p may have a role in the hepatic response to oxidative stress and is a potential clinically tractable circulating biomarker for progressive NAFLD. LAY SUMMARY: MicroRNAs (miRNAs) are small pieces of nucleic acid that may turn expression of genes on or off. These molecules can be detected in the blood circulation, and their levels in blood may change in liver disease including non-alcoholic fatty liver disease (NAFLD). To see if we could detect specific miRNA associated with advanced stages of NAFLD, we carried out miRNA sequencing in a group of 183 patients with NAFLD of varying severity together with 10 population controls. We found that a number of miRNAs showed changes, mainly increases, in serum levels but that 1 particular miRNA miR-193a-5p consistently increased. We confirmed this increase in a second group of cases with NAFLD. Measuring this miRNA in a blood sample may be a useful way to determine whether a patient has advanced NAFLD without an invasive liver biopsy.

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.

Association of Weight Changes With Changes in Histological Features and Blood Markers in Nonalcoholic Steatohepatitis.

BACKGROUND AND AIMS: Weight loss is recommended for patients with non-alcoholic steatohepatitis (NASH) but the impact of weight change on disease activity remains unclear. We examined the association between weight change (gain/loss) and changes in biochemical and histological features of NASH. METHODS: This was an analysis of the PIVENS and FLINT trials in adults with NASH who had liver biopsies at baseline and at either 1.5 years or 2 years. Multivariable regression models examined how weight change was associated with changes in (a) blood liver markers, (b) NASH resolution with no fibrosis worsening, (c) fibrosis improving with no NASH worsening, and (d) individual histological features. RESULTS: The BMI of the 421 participants was 34.3 kg/m2 (SD:6.5) and their mean weight change was +0.5 kg (SD:6.5). Weight change was independently and positively associated with changes in liver enzymes and the Fibrosis-4 score (all P < .001). Each kg of weight loss was associated with 7% (95% CI, 3%-10%; P < .001) increase in odds of achieving NASH resolution with no fibrosis worsening and with 5% (95% CI, 1%-8%; P = .01) increase in odds of achieving fibrosis improvement with no NASH worsening. Weight gain was associated with worsening of disease activity. For every kg of weight lost, the odds of fibrosis improving were 5% (95% CI, 2%-8%; P = .001). There was no evidence that the association between weight change and outcome depended upon pharmacological treatment, trial, body mass index, and baseline fibrosis. CONCLUSIONS: Weight change was independently and monotonically associated with changes in biochemical and histological features of NASH. Guidelines for NASH management should incorporate recommendations for both avoidance of weight gain and support to lose weight.

Assessment of hepatic steatosis by controlled attenuation parameter using the M and XL probes: an individual patient data meta-analysis.

BACKGROUND: Diagnostic tools for liver disease can now include estimation of the grade of hepatic steatosis (S0 to S3). Controlled attenuation parameter (CAP) is a non-invasive method for assessing hepatic steatosis that has become available for patients who are obese (FibroScan XL probe), but a consensus has not yet been reached regarding cutoffs and its diagnostic performance. We aimed to assess diagnostic properties and identify relevant covariates with use of an individual patient data meta-analysis. METHODS: We did an individual patient data meta-analysis, in which we searched PubMed and Web of Science for studies published from database inception until April 30, 2019. Studies reporting original biopsy-controlled data of CAP for non-invasive grading of steatosis were eligible. Probe recommendation was based on automated selection, manual assessment of skin-to-liver-capsule distance, and a body-mass index (BMI) criterion. Receiver operating characteristic methods and mixed models were used to assess diagnostic properties and covariates. Patients with non-alcoholic fatty liver disease (NAFLD) were analysed separately because they are the predominant patient group when using the XL probe. This study is registered with PROSPERO, CRD42018099284. FINDINGS: 16 studies reported histology-controlled CAP including the XL probe, and individual data from 13 papers and 2346 patients were included. Patients with a mean age of 46·5 years (SD 14·5) were recruited from 20 centres in nine countries. 2283 patients had data for BMI; 673 (29%) were normal weight (BMI <25 kg/m2), 530 (23%) were overweight (BMI ≥25 to <30 kg/m2), and 1080 (47%) were obese (BMI ≥30 kg/m2). 1277 (54%) patients had NAFLD, 474 (20%) had viral hepatitis, 285 (12%) had alcohol-associated liver disease, and 310 (13%) had other liver disease aetiologies. The XL probe was recommended in 1050 patients, 930 (89%) of whom had NAFLD; among the patients with NAFLD, the areas under the curve were 0·819 (95% CI 0·769-0·869) for S0 versus S1 to S3 and 0·754 (0·720-0·787) for S0 to S1 versus S2 to S3. CAP values were independently affected by aetiology, diabetes, BMI, aspartate aminotransferase, and sex. Optimal cutoffs differed substantially across aetiologies. Risk of bias according to QUADAS-2 was low. INTERPRETATION: CAP cutoffs varied according to cause, and can effectively recognise significant steatosis in patients with viral hepatitis. CAP cannot grade steatosis in patients with NAFLD adequately, but its value in a NAFLD screening setting needs to be studied, ideally with methods beyond the traditional histological reference standard. FUNDING: The German Federal Ministry of Education and Research and Echosens.

Systemic iron reduction by venesection alters the gut microbiome in patients with haemochromatosis.

BACKGROUND & AIMS: Iron reduction by venesection has been the cornerstone of treatment for haemochromatosis for decades, and its reported health benefits are many. Repeated phlebotomy can lead to a compensatory increase in intestinal iron absorption, reducing intestinal iron availability. Given that most gut bacteria are highly dependent on iron for survival, we postulated that, by reducing gut iron levels, venesection could alter the gut microbiota. METHODS: Clinical parameters, faecal bacterial composition and metabolomes were assessed before and during treatment in a group of patients with haemochromatosis undergoing iron reduction therapy. RESULTS: Systemic iron reduction was associated with an alteration of the gut microbiome, with changes evident in those who experienced reduced faecal iron availability with venesection. For example, levels of Faecalibacterium prausnitzii, a bacterium associated with improved colonic health, were increased in response to faecal iron reduction. Similarly, metabolomic changes were seen in association with reduced faecal iron levels. CONCLUSION: These findings highlight a significant shift in the gut microbiome of patients who experience reduced colonic iron during venesection. Targeted depletion of faecal iron could represent a novel therapy for metabolic and inflammatory diseases, meriting further investigation. LAY SUMMARY: Iron depletion by repeated venesection is the mainstay of treatment for haemochromatosis, an iron-overload disorder. Venesection has been associated with several health benefits, including improvements in liver function tests, reversal of liver scarring, and reduced risk of liver cancer. During iron depletion, iron absorption from the gastrointestinal (GI) tract increases to compensate for iron lost with treatment. Iron availability is limited in the GI tract and is crucial to the growth and function of many gut bacteria. In this study we show that reduced iron availability in the colon following venesection treatment leads to a change in the composition of the gut bacteria, a finding that, to date, has not been studied in patients with haemochromatosis.

Accurate non-invasive diagnosis and staging of non-alcoholic fatty liver disease using the urinary steroid metabolome.

BACKGROUND: The development of accurate, non-invasive markers to diagnose and stage non-alcoholic fatty liver disease (NAFLD) is critical to reduce the need for an invasive liver biopsy and to identify patients who are at the highest risk of hepatic and cardio-metabolic complications. Disruption of steroid hormone metabolic pathways has been described in patients with NAFLD. AIM(S): To assess the hypothesis that assessment of the urinary steroid metabolome may provide a novel, non-invasive biomarker strategy to stage NAFLD. METHODS: We analysed the urinary steroid metabolome in 275 subjects (121 with biopsy-proven NAFLD, 48 with alcohol-related cirrhosis and 106 controls), using gas chromatography-mass spectrometry (GC-MS) coupled with machine learning-based Generalised Matrix Learning Vector Quantisation (GMLVQ) analysis. RESULTS: Generalised Matrix Learning Vector Quantisation analysis achieved excellent separation of early (F0-F2) from advanced (F3-F4) fibrosis (AUC receiver operating characteristics [ROC]: 0.92 [0.91-0.94]). Furthermore, there was near perfect separation of controls from patients with advanced fibrotic NAFLD (AUC ROC = 0.99 [0.98-0.99]) and from those with NAFLD cirrhosis (AUC ROC = 1.0 [1.0-1.0]). This approach was also able to distinguish patients with NAFLD cirrhosis from those with alcohol-related cirrhosis (AUC ROC = 0.83 [0.81-0.85]). CONCLUSIONS: Unbiased GMLVQ analysis of the urinary steroid metabolome offers excellent potential as a non-invasive biomarker approach to stage NAFLD fibrosis as well as to screen for NAFLD. A highly sensitive and specific urinary biomarker is likely to have clinical utility both in secondary care and in the broader general population within primary care and could significantly decrease the need for liver biopsy.

FibroScan-AST (FAST) score for the non-invasive identification of patients with non-alcoholic steatohepatitis with significant activity and fibrosis: a prospective derivation and global validation study.

BACKGROUND: The burden of non-alcoholic fatty liver disease (NAFLD) is increasing globally, and a major priority is to identify patients with non-alcoholic steatohepatitis (NASH) who are at greater risk of progression to cirrhosis, and who will be candidates for clinical trials and emerging new pharmacotherapies. We aimed to develop a score to identify patients with NASH, elevated NAFLD activity score (NAS≥4), and advanced fibrosis (stage 2 or higher [F≥2]). METHODS: This prospective study included a derivation cohort before validation in multiple international cohorts. The derivation cohort was a cross-sectional, multicentre study of patients aged 18 years or older, scheduled to have a liver biopsy for suspicion of NAFLD at seven tertiary care liver centres in England. This was a prespecified secondary outcome of a study for which the primary endpoints have already been reported. Liver stiffness measurement (LSM) by vibration-controlled transient elastography and controlled attenuation parameter (CAP) measured by FibroScan device were combined with aspartate aminotransferase (AST), alanine aminotransferase (ALT), or AST:ALT ratio. To identify those patients with NASH, an elevated NAS, and significant fibrosis, the best fitting multivariable logistic regression model was identified and internally validated using boot-strapping. Score calibration and discrimination performance were determined in both the derivation dataset in England, and seven independent international (France, USA, China, Malaysia, Turkey) histologically confirmed cohorts of patients with NAFLD (external validation cohorts). This study is registered with ClinicalTrials.gov, number NCT01985009. FINDINGS: Between March 20, 2014, and Jan 17, 2017, 350 patients with suspected NAFLD attending liver clinics in England were prospectively enrolled in the derivation cohort. The most predictive model combined LSM, CAP, and AST, and was designated FAST (FibroScan-AST). Performance was satisfactory in the derivation dataset (C-statistic 0·80, 95% CI 0·76-0·85) and was well calibrated. In external validation cohorts, calibration of the score was satisfactory and discrimination was good across the full range of validation cohorts (C-statistic range 0·74-0·95, 0·85; 95% CI 0·83-0·87 in the pooled external validation patients' cohort; n=1026). Cutoff was 0·35 for sensitivity of 0·90 or greater and 0·67 for specificity of 0·90 or greater in the derivation cohort, leading to a positive predictive value (PPV) of 0·83 (84/101) and a negative predictive value (NPV) of 0·85 (93/110). In the external validation cohorts, PPV ranged from 0·33 to 0·81 and NPV from 0·73 to 1·0. INTERPRETATION: The FAST score provides an efficient way to non-invasively identify patients at risk of progressive NASH for clinical trials or treatments when they become available, and thereby reduce unnecessary liver biopsy in patients unlikely to have significant disease. FUNDING: Echosens and UK National Institute for Health Research.

Nonalcoholic Fatty Liver Disease in Adults: Current Concepts in Etiology, Outcomes, and Management.

Nonalcoholic fatty liver disease (NAFLD) is a spectrum of disease, extending from simple steatosis to inflammation and fibrosis with a significant risk for the development of cirrhosis. It is highly prevalent and is associated with significant adverse outcomes both through liver-specific morbidity and mortality but, perhaps more important, through adverse cardiovascular and metabolic outcomes. It is closely associated with type 2 diabetes and obesity, and both of these conditions drive progressive disease toward the more advanced stages. The mechanisms that govern hepatic lipid accumulation and the predisposition to inflammation and fibrosis are still not fully understood but reflect a complex interplay between metabolic target tissues including adipose and skeletal muscle, and immune and inflammatory cells. The ability to make an accurate assessment of disease stage (that relates to clinical outcome) can also be challenging. While liver biopsy is still regarded as the gold-standard investigative tool, there is an extensive literature on the search for novel noninvasive biomarkers and imaging modalities that aim to accurately reflect the stage of underlying disease. Finally, although no therapies are currently licensed for the treatment of NAFLD, there are interventions that appear to have proven efficacy in randomized controlled trials as well as an extensive emerging therapeutic landscape of new agents that target many of the fundamental pathophysiological processes that drive NAFLD. It is highly likely that over the next few years, new treatments with a specific license for the treatment of NAFLD will become available.

A multidisciplinary approach to the management of NAFLD is associated with improvement in markers of liver and cardio-metabolic health.

Non-alcoholic fatty liver disease (NAFLD) is a globally prevalent health problem, associated in its more severe forms with increased liver-related and cardiovascular-related morbidity and mortality. We established a multidisciplinary metabolic hepatology clinic in 2014 and have analysed the clinical data to evaluate the effectiveness of this service. Patients with NAFLD (n=165) who had attended two or more appointments were included. Prespecified clinical data were collected prospectively at clinic appointments and analysed retrospectively. Interventions offered included lifestyle advice, signposting to weight loss services and pharmacological treatment of diabetes and cardiovascular risk factors. Median follow-up was 13 months (range: 2-34). 59% (n=97) of patients had type 2 diabetes mellitus (T2DM). 53% (n=87) underwent liver biopsy of whom 18% (n=16) had cirrhosis. Median alanine aminotransferase (ALT) reduced by 11 IU/L (p<0.0001), median weight reduced by 3.3 kg (p=0.0005). There were significant reductions in HbA1c, total cholesterol and liver stiffness. Specifically, in patients with T2DM, HbA1c decreased by 4 mmol/mol (p=0.01) with significant reductions in ALT, weight and total cholesterol. Relative cardiovascular risk assessed by the QRISK3 score reduced in the whole cohort and in those with T2DM. Health economic modelling suggested the clinic intervention among those patients with poorly controlled T2DM was cost-effective. In conclusion, a multidisciplinary approach to the management of patients with NAFLD in this observational cohort study was associated with improvements in liver-related and cardio-metabolic related health parameters and with evidence of cost-effectiveness in patients with poorly controlled T2DM.

Immunotherapy-related hepatitis: real-world experience from a tertiary centre.

OBJECTIVE: Immune checkpoint inhibitors like anti-programmed cell death protein 1 (PD-1) drugs Nivolumab and Pembrolizumab and anti-cytotoxic T-lymphocyte associated (CTLA-4) drug Ipilimumab have become standard of care in many metastatic cancers. Immunotherapy-related hepatitis and cholangitis present a diagnostic and management challenge, being rare and incompletely characterised. We aim to report the incidence, features and treatments used for this in a real-world setting and to identify useful biomarkers, which can be used to predict effective use of steroids. DESIGN: Retrospective review of 453 patients started on immunotherapy over 7 years. SETTING: Tertiary hepatology and oncology centre. PATIENTS: 21 patients identified with immunotherapy-related hepatotoxicity. RESULTS: Hepatitis was most common in those receiving dual therapy (incidence 20%), with 75% of Grade 4 hepatitis cases occurring with ipilimumab-containing regimens. Corticosteroid monotherapy is first line treatment, but doses above 60 mg OD prednisolone do not demonstrate any additional benefit in time to hepatitis resolution. The alanine transaminase (ALT) reduction in steroid-responsive hepatitis is typically rapid (with a halving of ALT within 11 days). The commencement of additional immunosuppression (typically mycophenolate) appears safe and prompts a more rapid fall in ALT than corticosteroid use alone. Infliximab was safely used twice as hepatitis treatment. We also describe one patient with rare immunotherapy-induced biliary disease. CONCLUSIONS: Vigilance is required for detection of immunotherapy-associated liver disease as, other than dual immunotherapy, we can identify no predictive factors for its development. Our data suggest that corticosteroid response is not dependent on the higher dosing regimens. Early escalation of immunosuppression may be of benefit in the absence of a rapid response to corticosteroids.

Non-invasive assessment of portal hypertension by multi-parametric magnetic resonance imaging of the spleen: A proof of concept study.

BACKGROUND AND AIMS: Non-invasive assessment of portal hypertension is an area of unmet need. This proof of concept study aimed to evaluate the diagnostic accuracy of a multi-parametric magnetic resonance technique in the assessment of portal hypertension. Comparison to other non-invasive technologies was a secondary aim. METHODS: T1 and T2* maps through the liver and spleen were acquired prior to trans-jugular liver biopsy and hepatic vein pressure gradient (HVPG) measurement. T1 measurements reflect changes in tissue water content, but this relationship is confounded by the presence of iron, which in turn can be quantified accurately from T2* maps. Data were analysed using LiverMultiScan (Perspectum Diagnostics, Oxford, UK) which applies an algorithm to remove the confounding effect of iron, yielding the "iron corrected T1" (cT1). Sensitivity, specificity, diagnostic values and area under the curve were derived for spleen cT1, liver cT1, transient elastography, and serum fibrosis scores. HVPG was the reference standard. RESULTS: Nineteen patients (15 men) with median age 57 years were included. Liver disease aetiologies included non-alcoholic fatty liver disease (n = 9; 47%) and viral hepatitis (n = 4; 21%). There was strong correlation between spleen cT1 and HVPG (r = 0.69; p = 0.001). Other non-invasive biomarkers did not correlate with HVPG. Spleen cT1 had excellent diagnostic accuracy for portal hypertension (HVPG >5 mmHg) and clinically significant portal hypertension (HVPG ≥10 mmHg) with an area under the receiver operating characteristic curve of 0.92 for both. CONCLUSION: Spleen cT1 is a promising biomarker of portal pressure that outperforms other non-invasive scores and should be explored further.

PYY plays a key role in the resolution of diabetes following bariatric surgery in humans.

BACKGROUND: Bariatric surgery leads to early and long-lasting remission of type 2 diabetes (T2D). However, the mechanisms behind this phenomenon remain unclear. Among several factors, gut hormones are thought to be crucial mediators of this effect. Unlike GLP-1, the role of the hormone peptide tyrosine tyrosine (PYY) in bariatric surgery in humans has been limited to appetite regulation and its impact on pancreatic islet secretory function and glucose metabolism remains under-studied. METHODS: Changes in PYY concentrations were examined in obese patients after bariatric surgery and compared to healthy controls. Human pancreatic islet function was tested upon treatment with sera from patients before and after the surgery, in presence or absence of PYY. Alterations in intra-islet PYY release and insulin secretion were analysed after stimulation with short chain fatty acids (SCFAs), bile acids and the cytokine IL-22. FINDINGS: We demonstrate that PYY is a key effector of the early recovery of impaired glucose-mediated insulin and glucagon secretion in bariatric surgery. We establish that the short chain fatty acid propionate and bile acids, which are elevated after surgery, can trigger PYY release not only from enteroendocrine cells but also from human pancreatic islets. In addition, we identify IL-22 as a new factor which is modulated by bariatric surgery in humans and which directly regulates PYY expression and release. INTERPRETATION: This study shows that some major metabolic benefits of bariatric surgery can be emulated ex vivo. Our findings are expected to have a direct impact on the development of new non-surgical therapy for T2D correction.

Accuracy of FibroScan Controlled Attenuation Parameter and Liver Stiffness Measurement in Assessing Steatosis and Fibrosis in Patients With Nonalcoholic Fatty Liver Disease.

BACKGROUND & AIMS: We estimated the accuracy of FibroScan vibration-controlled transient elastography controlled attenuation parameter (CAP) and liver stiffness measurement (LSMs) in assessing steatosis and fibrosis in patients with suspected nonalcoholic liver disease (NAFLD). METHODS: We collected data from 450 consecutive adults who underwent liver biopsy analysis for suspected NAFLD at 7 centers in the United Kingdom from March 2014 through January 2017. FibroScan examinations with M or XL probe were completed within the 2 weeks of the biopsy analysis (404 had a valid examination). The biopsies were scored by 2 blinded expert pathologists according to nonalcoholic steatohepatitis clinical research network criteria. Diagnostic accuracy was estimated using the area under the receiver operating characteristic curves (AUROCs) for the categories of steatosis and fibrosis. We assessed effects of disease prevalence on positive and negative predictive values. For LSM, the effects of histological parameters and probe type were appraised using multivariable analysis. RESULTS: Using biopsy analysis as the reference standard, we found that CAP identified patients with steatosis with an AUROC of 0.87 (95% confidence interval [CI] 0.82-0.92) for S≥S1, 0.77 (95% CI 0.71-0.82) for S≥S2, and 0.70 (95% CI 0.64-0.75) for S=S3. Youden cutoff values for S≥S1, S≥S2, and S≥S3 were 302 dB/m, 331 dB/m, and 337 dB/m, respectively. LSM identified patients with fibrosis with AUROCs of 0.77 (95% CI 0.72-0.82) for F≥F2, 0.80 (95% CI 0.75-0.84) for F≥F3, and 0.89 (95% CI 0.84-0.93) for F=F4. Youden cutoff values for F≥F2, F≥F3, and F=F4 were 8.2 kPa, 9.7 kPa, and 13.6 kPa, respectively. Applying the optimal cutoff values, determined from this cohort, to populations of lower fibrosis prevalence increased negative predictive values and reduced positive predictive values. Multivariable analysis found that the only parameter that significantly affected LSMs was fibrosis stage (P<10-16); we found no association with steatosis or probe type. CONCLUSIONS: In a prospective analysis of patients with NAFLD, we found CAP and LSM by FibroScan to assess liver steatosis and fibrosis, respectively, with AUROC values ranging from 0.70 to 0.89. Probe type and steatosis did not affect LSM. STUDY REGISTRATION: ClinicalTrials.gov Identifier: NCT01985009.