A genetic risk score and diabetes predict development of alcohol-related cirrhosis in drinkers.

BACKGROUND & AIMS: Only a minority of excess alcohol drinkers develop cirrhosis. We developed and evaluated risk stratification scores to identify those at highest risk. METHODS: Three cohorts (GenomALC-1: n = 1,690, GenomALC-2: n = 3,037, UK Biobank: relevant n = 6,898) with a history of heavy alcohol consumption (≥80 g/day (men), ≥50 g/day (women), for ≥10 years) were included. Cases were participants with alcohol-related cirrhosis. Controls had a history of similar alcohol consumption but no evidence of liver disease. Risk scores were computed from up to 8 genetic loci identified previously as associated with alcohol-related cirrhosis and 3 clinical risk factors. Score performance for the stratification of alcohol-related cirrhosis risk was assessed and compared across the alcohol-related liver disease spectrum, including hepatocellular carcinoma (HCC). RESULTS: A combination of 3 single nucleotide polymorphisms (SNPs) (PNPLA3:rs738409, SUGP1-TM6SF2:rs10401969, HSD17B13:rs6834314) and diabetes status best discriminated cirrhosis risk. The odds ratios (ORs) and (95% CIs) between the lowest (Q1) and highest (Q5) score quintiles of the 3-SNP score, based on independent allelic effect size estimates, were 5.99 (4.18-8.60) (GenomALC-1), 2.81 (2.03-3.89) (GenomALC-2), and 3.10 (2.32-4.14) (UK Biobank). Patients with diabetes and high risk scores had ORs of 14.7 (7.69-28.1) (GenomALC-1) and 17.1 (11.3-25.7) (UK Biobank) compared to those without diabetes and with low risk scores. Patients with cirrhosis and HCC had significantly higher mean risk scores than patients with cirrhosis alone (0.76 ± 0.06 vs. 0.61 ± 0.02, p = 0.007). Score performance was not significantly enhanced by information on additional genetic risk variants, body mass index or coffee consumption. CONCLUSIONS: A risk score based on 3 genetic risk variants and diabetes status enables the stratification of heavy drinkers based on their risk of cirrhosis, allowing for the provision of earlier preventative interventions. LAY SUMMARY: Excessive chronic drinking leads to cirrhosis in some people, but so far there is no way to identify those at high risk of developing this debilitating disease. We developed a genetic risk score that can identify patients at high risk. The risk of cirrhosis is increased >10-fold with just two risk factors - diabetes and a high genetic risk score. Risk assessment using this test could enable the early and personalised management of this disease in high-risk patients.

Macrophage scavenger receptor 1 mediates lipid-induced inflammation in non-alcoholic fatty liver disease.

BACKGROUND & AIMS: Obesity-associated inflammation is a key player in the pathogenesis of non-alcoholic fatty liver disease (NAFLD). However, the role of macrophage scavenger receptor 1 (MSR1, CD204) remains incompletely understood. METHODS: A total of 170 NAFLD liver biopsies were processed for transcriptomic analysis and correlated with clinicopathological features. Msr1-/- and wild-type mice were subjected to a 16-week high-fat and high-cholesterol diet. Mice and ex vivo human liver slices were treated with a monoclonal antibody against MSR1. Genetic susceptibility was assessed using genome-wide association study data from 1,483 patients with NAFLD and 430,101 participants of the UK Biobank. RESULTS: MSR1 expression was associated with the occurrence of hepatic lipid-laden foamy macrophages and correlated with the degree of steatosis and steatohepatitis in patients with NAFLD. Mice lacking Msr1 were protected against diet-induced metabolic disorder, showing fewer hepatic foamy macrophages, less hepatic inflammation, improved dyslipidaemia and glucose tolerance, and altered hepatic lipid metabolism. Upon induction by saturated fatty acids, MSR1 induced a pro-inflammatory response via the JNK signalling pathway. In vitro blockade of the receptor prevented the accumulation of lipids in primary macrophages which inhibited the switch towards a pro-inflammatory phenotype and the release of cytokines such as TNF-ɑ. Targeting MSR1 using monoclonal antibody therapy in an obesity-associated NAFLD mouse model and human liver slices resulted in the prevention of foamy macrophage formation and inflammation. Moreover, we identified that rs41505344, a polymorphism in the upstream transcriptional region of MSR1, was associated with altered serum triglycerides and aspartate aminotransferase levels in a cohort of over 400,000 patients. CONCLUSIONS: Taken together, our data suggest that MSR1 plays a critical role in lipid-induced inflammation and could thus be a potential therapeutic target for the treatment of NAFLD. LAY SUMMARY: Non-alcoholic fatty liver disease (NAFLD) is a chronic disease primarily caused by excessive consumption of fat and sugar combined with a lack of exercise or a sedentary lifestyle. Herein, we show that the macrophage scavenger receptor MSR1, an innate immune receptor, mediates lipid uptake and accumulation in Kupffer cells, resulting in liver inflammation and thereby promoting the progression of NAFLD in humans and mice.

Genome-wide Association Study and Meta-analysis on Alcohol-Associated Liver Cirrhosis Identifies Genetic Risk Factors.

BACKGROUND AND AIMS: Only a minority of heavy drinkers progress to alcohol-associated cirrhosis (ALC). The aim of this study was to identify common genetic variants that underlie risk for ALC. APPROACH AND RESULTS: We analyzed data from 1,128 subjects of European ancestry with ALC and 614 heavy-drinking subjects without known liver disease from Australia, the United States, the United Kingdom, and three countries in Europe. A genome-wide association study (GWAS) was performed, adjusting for principal components and clinical covariates (alcohol use, age, sex, body mass index, and diabetes). We validated our GWAS findings using UK Biobank. We then performed a meta-analysis combining data from our study, the UK Biobank, and a previously published GWAS. Our GWAS found genome-wide significant risk association of rs738409 in patatin-like phospholipase domain containing 3 (PNPLA3) (odds ratio [OR] = 2.19 [G allele], P = 4.93 × 10-17 ) and rs4607179 near HSD17B13 (OR = 0.57 [C allele], P = 1.09 × 10-10 ) with ALC. Conditional analysis accounting for the PNPLA3 and HSD17B13 loci identified a protective association at rs374702773 in Fas-associated factor family member 2 (FAF2) (OR = 0.61 [del(T) allele], P = 2.56 × 10-8 ) for ALC. This association was replicated in the UK Biobank using conditional analysis (OR = 0.79, P = 0.001). Meta-analysis (without conditioning) confirmed genome-wide significance for the identified FAF2 locus as well as PNPLA3 and HSD17B13. Two other previously known loci (SERPINA1 and SUGP1/TM6SF2) were also genome-wide significant in the meta-analysis. GeneOntology pathway analysis identified lipid droplets as the target for several identified genes. In conclusion, our GWAS identified a locus at FAF2 associated with reduced risk of ALC among heavy drinkers. Like the PNPLA3 and HSD17B13 gene products, the FAF2 product has been localized to fat droplets in hepatocytes. CONCLUSIONS: Our genetic findings implicate lipid droplets in the biological pathway(s) underlying ALC.

Obesity, Diabetes, Coffee, Tea, and Cannabis Use Alter Risk for Alcohol-Related Cirrhosis in 2 Large Cohorts of High-Risk Drinkers.

INTRODUCTION: Sustained high alcohol intake is necessary but not sufficient to produce alcohol-related cirrhosis. Identification of risk factors, apart from lifetime alcohol exposure, would assist in discovery of mechanisms and prediction of risk. METHODS: We conducted a multicenter case-control study (GenomALC) comparing 1,293 cases (with alcohol-related cirrhosis, 75.6% male) and 754 controls (with equivalent alcohol exposure but no evidence of liver disease, 73.6% male). Information confirming or excluding cirrhosis, and on alcohol intake and other potential risk factors, was obtained from clinical records and by interview. Case-control differences in risk factors discovered in the GenomALC participants were validated using similar data from 407 cases and 6,573 controls from UK Biobank. RESULTS: The GenomALC case and control groups reported similar lifetime alcohol intake (1,374 vs 1,412 kg). Cases had a higher prevalence of diabetes (20.5% (262/1,288) vs 6.5% (48/734), P = 2.27 × 10-18) and higher premorbid body mass index (26.37 ± 0.16 kg/m2) than controls (24.44 ± 0.18 kg/m2, P = 5.77 × 10-15). Controls were significantly more likely to have been wine drinkers, coffee drinkers, smokers, and cannabis users than cases. Cases reported a higher proportion of parents who died of liver disease than controls (odds ratio 2.25 95% confidence interval 1.55-3.26). Data from UK Biobank confirmed these findings for diabetes, body mass index, proportion of alcohol as wine, and coffee consumption. DISCUSSION: If these relationships are causal, measures such as weight loss, intensive treatment of diabetes or prediabetic states, and coffee consumption should reduce the risk of alcohol-related cirrhosis.

Genome-wide association study of non-alcoholic fatty liver and steatohepatitis in a histologically characterised cohort☆.

BACKGROUND & AIMS: Genetic factors associated with non-alcoholic fatty liver disease (NAFLD) remain incompletely understood. To date, most genome-wide association studies (GWASs) have adopted radiologically assessed hepatic triglyceride content as the reference phenotype and so cannot address steatohepatitis or fibrosis. We describe a GWAS encompassing the full spectrum of histologically characterised NAFLD. METHODS: The GWAS involved 1,483 European NAFLD cases and 17,781 genetically matched controls. A replication cohort of 559 NAFLD cases and 945 controls was genotyped to confirm signals showing genome-wide or close to genome-wide significance. RESULTS: Case-control analysis identified signals showing p values ≤5 × 10-8 at 4 locations (chromosome [chr] 2 GCKR/C2ORF16; chr4 HSD17B13; chr19 TM6SF2; chr22 PNPLA3) together with 2 other signals with p <1 × 10-7 (chr1 near LEPR and chr8 near IDO2/TC1). Case-only analysis of quantitative traits showed that the PNPLA3 signal (rs738409) had genome-wide significance for steatosis, fibrosis and NAFLD activity score and a new signal (PYGO1 rs62021874) had close to genome-wide significance for steatosis (p = 8.2 × 10-8). Subgroup case-control analysis for NASH confirmed the PNPLA3 signal. The chr1 LEPR single nucleotide polymorphism also showed genome-wide significance for this phenotype. Considering the subgroup with advanced fibrosis (≥F3), the signals on chr2, chr19 and chr22 maintained their genome-wide significance. Except for GCKR/C2ORF16, the genome-wide significance signals were replicated. CONCLUSIONS: This study confirms PNPLA3 as a risk factor for the full histological spectrum of NAFLD at genome-wide significance levels, with important contributions from TM6SF2 and HSD17B13. PYGO1 is a novel steatosis modifier, suggesting that Wnt signalling pathways may be relevant in NAFLD pathogenesis. LAY SUMMARY: Non-alcoholic fatty liver disease is a common disease where excessive fat accumulates in the liver and may result in cirrhosis. To understand who is at risk of developing this disease and suffering liver damage, we undertook a genetic study to compare the genetic profiles of people suffering from fatty liver disease with genetic profiles seen in the general population. We found that particular sequences in 4 different areas of the human genome were seen at different frequencies in the fatty liver disease cases. These sequences may help predict an individual's risk of developing advanced disease. Some genes where these sequences are located may also be good targets for future drug treatments.

The degree of hepatic steatosis associates with impaired cardiac and autonomic function.

BACKGROUND & AIMS: Cardiovascular disease is the principle cause of death in patients with elevated liver fat unrelated to alcohol consumption, more so than liver-related morbidity and mortality. The aim of this study was to evaluate the relationship between liver fat and cardiac and autonomic function, as well as to assess how impairment in cardiac and autonomic function is influenced by metabolic risk factors. METHODS: Cardiovascular and autonomic function were assessed in 96 sedentary individuals: i) non-alcoholic fatty liver disease (NAFLD) (n = 46, hepatic steatosis >5% by magnetic resonance spectroscopy), ii) Hepatic steatosis and alcohol (dual aetiology fatty liver disease [DAFLD]) (n = 16, hepatic steatosis >5%, consuming >20 g/day of alcohol) and iii) CONTROL (n = 34, no cardiac, liver or metabolic disorders, <20 g/day of alcohol). RESULTS: Patients with NAFLD and DAFLD had significantly impaired cardiac and autonomic function when compared with controls. Diastolic variability and systolic variability (LF/HF-sBP [n/1]; 2.3 (1.7) and 2.3 (1.5) vs. 3.4 (1.5), p <0.01) were impaired in patients with NAFLD and DAFLD when compared to controls, with DAFLD individuals showing a decrease in diastolic variability relative to NAFLD patients. Hepatic steatosis and fasting glucose were negatively correlated with stroke volume index. Fibrosis stage was significantly negatively associated with mean blood pressure (r = -0.47, p = 0.02), diastolic variability (r = -0.58, p ≤0.01) and systolic variability (r = -0.42, p = 0.04). Hepatic steatosis was independently associated with cardiac function (p ≤0.01); TNF-α (p ≤0.05) and CK-18 (p ≤0.05) were independently associated with autonomic function. CONCLUSION: Cardiac and autonomic impairments appear to be dependent on level of liver fat, metabolic dysfunction, inflammation and fibrosis staging, and to a lesser extent alcohol intake. Interventions should be sought to moderate the excess cardiovascular risk in patients with NAFLD or DAFLD. LAY SUMMARY: Increased levels of fat in the liver impair the ability of the cardiovascular system to work properly. The amount of fat in the liver, metabolic control, inflammation and alcohol are all linked to the degree that the cardiovascular system is affected.

Systematic review assessing the effectiveness of dietary intervention on gut microbiota in adults with type 2 diabetes.

AIMS/HYPOTHESIS: Despite improved understanding of the pathophysiology of type 2 diabetes mellitus, explanations for individual variability in disease progression and response to treatment are incomplete. The gut microbiota has been linked to the pathophysiology of type 2 diabetes mellitus and may account for this variability. We conducted a systematic review to assess the effectiveness of dietary and physical activity/exercise interventions in modulating the gut microbiota and improving glucose control in adults with type 2 diabetes mellitus. METHODS: A systematic search was conducted to identify studies reporting on the effect of dietary and physical activity/exercise interventions on the gut microbiota and glucose control in individuals with a confirmed diagnosis of type 2 diabetes mellitus. Study characteristics, methodological quality and details relating to interventions were captured using a data-extraction form. Meta-analyses were conducted where sufficient data were available, and other results were reported narratively. RESULTS: Eight studies met the eligibility criteria of the systematic review. No studies were found that reported on the effects of physical activity/exercise on the gut microbiota and glucose control. However, studies reporting on dietary interventions showed that such interventions were associated with modifications to the composition and diversity of the gut microbiota. There was a statistically significant improvement in HbA1c (standardised mean difference [SMD] -2.31 mmol/mol [95% CI -2.76, -1.85] [0.21%; 95% CI -0.26, -0.16]; I2 = 0%, p < 0.01), but not in fasting blood glucose (SMD -0.25 mmol/l [95% CI -0.85, 0.35], I2 = 87%, p > 0.05), fasting insulin (SMD -1.82 pmol/l [95% CI -7.23, 3.60], I2 = 54%, p > 0.05) or HOMA-IR (SMD -0.15 [95% CI -0.63, 0.32], I2 = 69%, p > 0.05) when comparing dietary interventions with comparator groups. There were no significant changes in the relative abundance of bacteria in the genera Bifidobacterium (SMD 1.29% [95% CI -4.45, 7.03], I2 = 33%, p > 0.05), Roseburia (SMD -0.85% [95% CI -2.91, 1.21], I2 = 79%, p > 0.05) or Lactobacillus (SMD 0.04% [95% CI -0.01, 0.09], I2 = 0%, p > 0.05) when comparing dietary interventions with comparator groups. There were, however, other significant changes in the gut microbiota, including changes at various taxonomic levels, including phylum, family, genus and species, Firmicutes:Bacteroidetes ratios and changes in diversity matrices (α and ß). Dietary intervention had minimal or no effect on inflammation, short-chain fatty acids or anthropometrics. CONCLUSIONS/INTERPRETATION: Dietary intervention was found to modulate the gut microbiota and improve glucose control in individuals with type 2 diabetes. Although the results of the included studies are encouraging, this review highlights the need for further well-conducted interventional studies to inform the clinical use of dietary interventions targeting the gut microbiota.

Modeling NAFLD disease burden in China, France, Germany, Italy, Japan, Spain, United Kingdom, and United States for the period 2016-2030.

BACKGROUND & AIMS: Non-alcoholic fatty liver disease (NAFLD) and non-alcoholic steatohepatitis (NASH) are increasingly a cause of cirrhosis and hepatocellular carcinoma globally. This burden is expected to increase as epidemics of obesity, diabetes and metabolic syndrome continue to grow. The goal of this analysis was to use a Markov model to forecast NAFLD disease burden using currently available data. METHODS: A model was used to estimate NAFLD and NASH disease progression in eight countries based on data for adult prevalence of obesity and type 2 diabetes mellitus (DM). Published estimates and expert consensus were used to build and validate the model projections. RESULTS: If obesity and DM level off in the future, we project a modest growth in total NAFLD cases (0-30%), between 2016-2030, with the highest growth in China as a result of urbanization and the lowest growth in Japan as a result of a shrinking population. However, at the same time, NASH prevalence will increase 15-56%, while liver mortality and advanced liver disease will more than double as a result of an aging/increasing population. CONCLUSIONS: NAFLD and NASH represent a large and growing public health problem and efforts to understand this epidemic and to mitigate the disease burden are needed. If obesity and DM continue to increase at current and historical rates, both NAFLD and NASH prevalence are expected to increase. Since both are reversible, public health campaigns to increase awareness and diagnosis, and to promote diet and exercise can help manage the growth in future disease burden. LAY SUMMARY: Non-alcoholic fatty liver disease and non-alcoholic steatohepatitis can lead to advanced liver disease. Both conditions are becoming increasingly prevalent as the epidemics of obesity and diabetes continue to increase. A mathematical model was built to understand how the disease burden associated with non-alcoholic fatty liver disease and non-alcoholic steatohepatitis will change over time. Results suggest increasing cases of advanced liver disease and liver-related mortality in the coming years.

Prednisolone or pentoxifylline for alcoholic hepatitis.

BACKGROUND: Alcoholic hepatitis is a clinical syndrome characterized by jaundice and liver impairment that occurs in patients with a history of heavy and prolonged alcohol use. The short-term mortality among patients with severe disease exceeds 30%. Prednisolone and pentoxifylline are both recommended for the treatment of severe alcoholic hepatitis, but uncertainty about their benefit persists. METHODS: We conducted a multicenter, double-blind, randomized trial with a 2-by-2 factorial design to evaluate the effect of treatment with prednisolone or pentoxifylline. The primary end point was mortality at 28 days. Secondary end points included death or liver transplantation at 90 days and at 1 year. Patients with a clinical diagnosis of alcoholic hepatitis and severe disease were randomly assigned to one of four groups: a group that received a pentoxifylline-matched placebo and a prednisolone-matched placebo, a group that received prednisolone and a pentoxifylline-matched placebo, a group that received pentoxifylline and a prednisolone-matched placebo, or a group that received both prednisolone and pentoxifylline. RESULTS: A total of 1103 patients underwent randomization, and data from 1053 were available for the primary end-point analysis. Mortality at 28 days was 17% (45 of 269 patients) in the placebo-placebo group, 14% (38 of 266 patients) in the prednisolone-placebo group, 19% (50 of 258 patients) in the pentoxifylline-placebo group, and 13% (35 of 260 patients) in the prednisolone-pentoxifylline group. The odds ratio for 28-day mortality with pentoxifylline was 1.07 (95% confidence interval [CI], 0.77 to 1.49; P=0.69), and that with prednisolone was 0.72 (95% CI, 0.52 to 1.01; P=0.06). At 90 days and at 1 year, there were no significant between-group differences. Serious infections occurred in 13% of the patients treated with prednisolone versus 7% of those who did not receive prednisolone (P=0.002). CONCLUSIONS: Pentoxifylline did not improve survival in patients with alcoholic hepatitis. Prednisolone was associated with a reduction in 28-day mortality that did not reach significance and with no improvement in outcomes at 90 days or 1 year. (Funded by the National Institute for Health Research Health Technology Assessment program; STOPAH EudraCT number, 2009-013897-42 , and Current Controlled Trials number, ISRCTN88782125 ).

Association of Liver Injury From Specific Drugs, or Groups of Drugs, With Polymorphisms in HLA and Other Genes in a Genome-Wide Association Study.

BACKGROUND & AIMS: We performed a genome-wide association study (GWAS) to identify genetic risk factors for drug-induced liver injury (DILI) from licensed drugs without previously reported genetic risk factors. METHODS: We performed a GWAS of 862 persons with DILI and 10,588 population-matched controls. The first set of cases was recruited before May 2009 in Europe (n = 137) and the United States (n = 274). The second set of cases were identified from May 2009 through May 2013 from international collaborative studies performed in Europe, the United States, and South America. For the GWAS, we included only cases with patients of European ancestry associated with a particular drug (but not flucloxacillin or amoxicillin-clavulanate). We used DNA samples from all subjects to analyze HLA genes and single nucleotide polymorphisms. After the discovery analysis was concluded, we validated our findings using data from 283 European patients with diagnosis of DILI associated with various drugs. RESULTS: We associated DILI with rs114577328 (a proxy for A*33:01 a HLA class I allele; odds ratio [OR], 2.7; 95% confidence interval [CI], 1.9-3.8; P = 2.4 × 10-8) and with rs72631567 on chromosome 2 (OR, 2.0; 95% CI, 1.6-2.5; P = 9.7 × 10-9). The association with A*33:01 was mediated by large effects for terbinafine-, fenofibrate-, and ticlopidine-related DILI. The variant on chromosome 2 was associated with DILI from a variety of drugs. Further phenotypic analysis indicated that the association between DILI and A*33:01 was significant genome wide for cholestatic and mixed DILI, but not for hepatocellular DILI; the polymorphism on chromosome 2 was associated with cholestatic and mixed DILI as well as hepatocellular DILI. We identified an association between rs28521457 (within the lipopolysaccharide-responsive vesicle trafficking, beach and anchor containing gene) and only hepatocellular DILI (OR, 2.1; 95% CI, 1.6-2.7; P = 4.8 × 10-9). We did not associate any specific drug classes with genetic polymorphisms, except for statin-associated DILI, which was associated with rs116561224 on chromosome 18 (OR, 5.4; 95% CI, 3.0-9.5; P = 7.1 × 10-9). We validated the association between A*33:01 terbinafine- and sertraline-induced DILI. We could not validate the association between DILI and rs72631567, rs28521457, or rs116561224. CONCLUSIONS: In a GWAS of persons of European descent with DILI, we associated HLA-A*33:01 with DILI due to terbinafine and possibly fenofibrate and ticlopidine. We identified polymorphisms that appear to be associated with DILI from statins, as well as 2 non-drug-specific risk factors.

Plasma DNA methylation: a potential biomarker for stratification of liver fibrosis in non-alcoholic fatty liver disease.

OBJECTIVE: Liver biopsy is currently the most reliable way of evaluating liver fibrosis in patients with non-alcoholic fatty liver disease (NAFLD). Its inherent risks limit its widespread use. Differential liver DNA methylation of peroxisome proliferator-activated receptor gamma (PPARγ) gene promoter has recently been shown to stratify patients in terms of fibrosis severity but requires access to liver tissue. The aim of this study was to assess whether DNA methylation of circulating DNA could be detected in human plasma and potentially used to stratify liver fibrosis severity in patients with NAFLD. DESIGN: Patients with biopsy-proven NAFLD and age-matched controls were recruited from the liver and gastroenterology clinics at the Newcastle upon Tyne Hospitals NHS Foundation Trust. Plasma cell-free circulating DNA methylation of PPARγ was quantitatively assessed by pyrosequencing. Liver DNA methylation was quantitatively assessed by pyrosequencing NAFLD explant tissue, subjected to laser capture microdissection (LCM). Patients with alcoholic liver disease (ALD) were also subjected to plasma DNA and LCM pyrosequencing. RESULTS: 26 patients with biopsy-proven NAFLD were included. Quantitative plasma DNA methylation of PPARγ stratified patients into mild (Kleiner 1-2) and severe (Kleiner 3-4) fibrosis (CpG1: 63% vs 86%, p<0.05; CpG2: 51% vs 65% p>0.05). Hypermethylation at the PPARγ promoter of plasma DNA correlated with changes in hepatocellular rather than myofibroblast DNA methylation. Similar results were demonstrated in patients with ALD cirrhosis. CONCLUSIONS: Differential DNA methylation at the PPARγ promoter can be detected within the pool of cell-free DNA of human plasma. With further validation, plasma DNA methylation of PPARγ could potentially be used to non-invasively stratify liver fibrosis severity in patients with NAFLD. Plasma DNA methylation signatures reflect the molecular pathology associated with fibrotic liver disease.

Genetic Factors That Affect Risk of Alcoholic and Nonalcoholic Fatty Liver Disease.

Genome-wide association studies and candidate gene studies have informed our understanding of factors contributing to the well-recognized interindividual variation in the progression and outcomes of alcoholic liver disease and nonalcoholic fatty liver disease. We discuss the mounting evidence for shared modifiers and common pathophysiological processes that contribute to development of both diseases. We discuss the functions of proteins encoded by risk variants of genes including patatin-like phospholipase domain-containing 3 and transmembrane 6 superfamily member 2, as well as epigenetic factors that contribute to the pathogenesis of alcoholic liver disease and nonalcoholic fatty liver disease. We also discuss important areas of future genetic research and their potential to affect clinical management of patients.

Effects of Exercise on Liver Fat and Metabolism in Alcohol Drinkers.

BACKGROUND & AIMS: Exercise is an important component of obesity-associated disorders and has been shown to reduce markers of nonalcoholic fatty liver disease (NAFLD). However, little is known about how these effects are influenced by alcohol intake. The authors performed a randomized controlled trial to investigate the effects of exercise on hepatic triglyceride content (HTGC) and metabolism in overweight or obese patients who consume alcohol. METHODS: The authors performed a prospective study of 27 patients (mean 54 ± 11 years of age, body mass index [BMI] 31 ± 4 kg/m2) with >5% HTGC in the United Kingdom, consuming alcohol (mean 221 ± 75 g/week). Anthropometry, body composition, HTGC, and abdominal fat were measured using plethysmography and magnetic resonance imaging. Subjects were assigned to groups that exercised (3 times/week on nonconsecutive days) for 12 weeks (n = 14) or continued standard care (control group, n = 13), maintaining baseline weight and alcohol consumption. The exercise program consisted of aerobic exercise (static cycling) and a circuit of resistance exercise (free weights and machines). Patients were examined at baseline and at 12 weeks; data collected on HTGC, body composition, metabolic control, circulating inflammatory, and fibrosis markers were assessed at baseline and at 12 weeks. Between-group differences were evaluated using an unpaired t test and within-group differences using a paired t test. The primary outcomes for this study were changes in HTGC between baseline and 12 weeks. RESULTS: After 12 weeks, there was no significant difference between the exercise and control groups in HTGC (reduction of 0.1% ± 2.1% in exercisers vs increase of 0.5 ± 2.1% in control group; P > .05). At week 12, the exercise group had significant reductions in subcutaneous fat (loss of 23 ± 28 cm2 in the exercisers vs increase of 12 ± 19 cm2 in the control group; P < .01), and whole body fat (loss of 2.1 ± 1.1 kg in the exercisers vs increase of 0.2 ± 2.1 kg; P < .01). The exercise group also had a significantly greater increase in lean body mass (increase of 1.9 ± 1.4 kg for the exercisers vs increase of 0.7 ± 1.5 kg for the control group; P < .01) and a significantly greater reduction in level of cytokeratin 18 (reduction of 49 ± 82 U/L in exercisers vs increase of 17 ± 38 U/L in control group; P < .05). There were no differences between groups in changes in metabolic factors or markers of inflammation. CONCLUSIONS: In a randomized controlled trial of obese individuals who consume alcohol, exercise significantly improved body composition and reduced hepatocyte apoptosis (cytokeratin 18), but did not reduce HTGC. This finding could indicate that alcohol consumption reduces the effects of exercise on NAFLD observed in previous studies. Clinical care teams should look to use exercise as part of the management strategy for people consuming alcohol, but optimal benefit may be as an adjunct to alcohol reduction and weight management strategies. (ISRCTN.com, Number: ISRCTN90597099).

Exercise Reduces Liver Lipids and Visceral Adiposity in Patients With Nonalcoholic Steatohepatitis in a Randomized Controlled Trial.

BACKGROUND & AIMS: Pharmacologic treatments for nonalcoholic steatohepatitis (NASH) are limited. Lifestyle interventions are believed to be effective in reducing features of NASH, although the effect of regular exercise, independent of dietary change, is unclear. We performed a randomized controlled trial to study the effect of exercise on hepatic triglyceride content (HTGC) and biomarkers of fibrosis in patients with NASH. METHODS: Twenty-four patients (mean age, 52 ± 14 y; body mass index, 33 ± 6 kg/m2) with sedentary lifestyles (<60 min/wk of moderate-vigorous activity) and biopsy-proven NASH were assigned randomly to groups that exercised (n = 12) or continued standard care (controls, n = 12) for 12 weeks while maintaining their weight. The exercise (cycling and resistance training) was supervised at an accredited sports center and supervised by a certified exercise specialist and recorded 3 times per week on nonconsecutive days. We measured HTGC, body composition, circulating markers of inflammation, fibrosis, and glucose tolerance at baseline and at 12 weeks. RESULTS: Compared with baseline, exercise significantly reduced HTGC (reduction of 16% ± 24% vs an increase of 9% ± 15% for controls; P < .05), visceral fat (reduction of 22 ± 33 cm2 vs an increase of 14 ± 48 cm2 for controls; P < .05), plasma triglycerides (reduction of 0.5 ± 1.0 mmol/L vs an increase of 0.3 ± 0.4 mmol/L for controls; P < .05), and γ-glutamyltransferase (reduction of 10 ± 28 U/L-1 vs a reduction of 17 ± 38 U/L-1 for controls; P < .05). There were no effects of exercise on liver enzyme levels, metabolic parameters, circulatory markers of inflammation (levels of interleukin 6, tumor necrosis factor-α, or C-reactive protein) and fibrosis. CONCLUSIONS: In a randomized controlled trial, 12 weeks of exercise significantly reduced HTGC, visceral fat, and plasma triglyceride levels in patients with NASH, but did not affect circulating markers of inflammation or fibrosis. Exercise without weight loss therefore affects some but not all factors associated with NASH. Clinical care teams should consider exercise as part of a management strategy of NASH, but weight management strategies should be included. Larger and longer-term studies are required to determine the effects of exercise in patients with NASH. ISRCTN registry.com: ISRCTN16070927.

Age as a Confounding Factor for the Accurate Non-Invasive Diagnosis of Advanced NAFLD Fibrosis.

OBJECTIVES: Non-invasive fibrosis scores are widely used to identify/exclude advanced fibrosis in patients with non-alcoholic fatty liver disease (NAFLD). However, these scores were principally developed and validated in patients aged between 35 and 65 years of age. The objective of this study was to assess the effect of age on the performance of non-invasive fibrosis tests in NAFLD. METHODS: Patients were recruited from European specialist hepatology clinics. The cohort was divided into five age-based groups: ≤35 (n=74), 36-45 (n=96), 46-55 (n=197), 56-64 (n=191), and ≥65 years (n=76), and the performance of the aspartate aminotransferase (AST)/alanine transaminase (ALT) ratio, fibrosis 4 (FIB-4), and NAFLD fibrosis score (NFS) for advanced fibrosis (stage F3-F4) for each group was assessed using liver biopsy as the standard. RESULTS: Six hundred and thirty-four patients were included. The diagnostic accuracy of the AST/ALT ratio was lower than NFS and FIB-4 in all the age groups. The AST/ALT ratio, NFS, and FIB-4 score performed poorly for a diagnosis of advanced fibrosis in those aged ≤35 years (area under the receiver operating characteristic curves (AUROCs 0.52, 0.52, and 0.60, respectively). For all groups >35 years, AUROCs for advanced fibrosis were similar for the NFS and FIB-4 score (range 0.77-0.84). However, the specificity for advanced fibrosis using the FIB-4 and NFS declined with age, becoming unacceptably low in those aged ≥65 years (35% for FIB-4 and 20% for NFS). New cutoffs were derived (and validated) for those aged ≥65 years, which improved specificity to 70% without adversely affecting sensitivity (FIB-4 2.0, sensitivity 77%; NFS 0.12, sensitivity 80%). CONCLUSIONS: The NFS and FIB-4 scores have similar accuracy for advanced fibrosis in patients aged >35 years. However, the specificity for advanced fibrosis is unacceptably low in patients aged ≥65 years, resulting in a high false positive rate. New thresholds for use in patients aged ≥65 years are proposed to address this issue.

Variants in the LGALS9 Gene Are Associated With Development of Liver Disease in Heavy Consumers of Alcohol.

BACKGROUND & AIMS: Alcohol consumption is a major cause of chronic liver disease and contributes to a large proportion of cirrhosis-related deaths worldwide. However, only a fraction of heavy consumers of alcohol develop advanced alcoholic liver disease (ALD), so there are likely to be other risk factors. We investigated whether polymorphisms in the gene encoding galectin-9 (LGALS9), previously shown to mediate liver injury, were associated with the development of ALD. METHODS: We isolated DNA from peripheral blood mononuclear cells (PBMCs) of 575 individuals with at-risk alcohol consumption but no other risk factors for chronic liver disease; all subjects were white Europeans who had consumed more than 80 grams ethanol per day. Of the subjects, 388 had ALD (including, 268 with cirrhosis and 74 with alcoholic hepatitis; mean age, 49 y; 72% male) and 187 had normal liver function with no biochemical or clinical evidence of liver disease (controls; mean age, 42 y; 73% male). Select LGALS9 polymorphisms were genotyped using allelic discrimination. We also genotyped and measured expression of LGALS9 messenger RNA in PBMCs from individuals who were not heavy consumers of alcohol. RESULTS: We used data from the HapMap project to identify 5 single-nucleotide polymorphisms (SNPs) that tag all the common haplotypes. When we looked for these SNPs in individuals with vs without liver disease, 4 (rs3751093, rs4239242, rs732222, and rs4794976) were associated with an increased risk of developing ALD. We found that levels of LGALS9 messenger RNA and protein expressed were associated with an allele carried by PBMCs. Multivariate analysis confirmed that rs4239242 and rs4794976 were associated with an increased risk of ALD. CONCLUSIONS: In a genetic analysis of heavy consumers of alcohol, we associated 2 SNPS in LGALS9 with the development of ALD. Although larger studies are required, this information could be used to determine the risk of individuals developing ALD or to develop therapeutic agents.

How to Diagnose Nonalcoholic Fatty Liver Disease.

Patients with nonalcoholic fatty liver disease (NAFLD) are asymptomatic and present with either unexplained abnormal liver blood tests or a bright liver on ultrasonography. Some patients will have normal liver blood tests raising the issue of whether patients with risk factors for NAFLD (diabetes and/or metabolic syndrome [MS]) should be screened for its presence with biomarkers, such as the fatty liver index (FLI). The diagnosis of NAFLD requires the exclusion of other causes of chronic liver disease and steatosis, especially heavy alcohol consumption and viral hepatitis particularly HCV genotype 3. Diagnostic work-up should include evaluation of family and personal history of components of the MS and assessment of liver tests, fasting blood glucose, triglycerides and HDL levels. A drug history is important due to a number being associated with steatosis. To confirm the diagnosis of NAFLD and quantify steatosis, ultrasound (US) and MRI-based techniques are available but none are in routine use outside clinical trials. Standard US is no more accurate than biomarkers such as FLI. The accurate staging of NAFLD requires liver biopsy; however, this is clearly impractical for such a prevalent disease. Accordingly, a number of imaging and blood-based biomarker tests have been evaluated. While none have proved reliable for the diagnosis of nonalcoholic steatohepatitis, several have proved accurate in diagnosing the presence of stage 3 or 4 fibrosis, including the NAFLD fibrosis score, fibrosis-4 and the enhanced liver fibrosis test. Of the imaging techniques, elastography has received the most attention and is being used in routine clinical practice. US acoustic radiation force impulse imaging, and MR-based elastography have recently been described but none are sufficiently accurate to replace liver biopsy for clinical trials as yet or are cost effective for use in routine clinical settings.