Hepatic steatosis after liver transplantation: a systematic review and meta-analysis.

NAFLD can occur after liver transplantation (LT), as recurrence or de novo hepatic steatosis (HS). We aimed to evaluate the literature on prevalence, risk factors, and prognosis of post-LT HS. Systematic review with meta-analysis through a search on: PUBMED, Scopus, and Web-of-Science, from inception until the September 30, 2021. Forty studies were included, representing 6979 patients. The post-LT HS prevalence was 39.76% (95% CI, 34.06-45.46), with a rising kinetics (11.06% increase per decade, p =0.04), and a geographical distribution (15.10% more prevalent in American continent compared with Europe and Asia). Recurrent HS was up to 5-fold more likely than de novo HS [OR: 5.38 (2.69-10.76)]. Metabolic disturbances were stronger risk factors in the post-LT recipient [obesity: OR: 4.62 (3.07-6.96); metabolic syndrome: OR: 3.26 (2.03-5.25)] as compared with pre-LT recipients, with the exception of diabetes mellitus, which doubled the risk at any set [pre-LT diabetes mellitus: OR: 2.06 (1.58-2.68); post-LT diabetes mellitus: OR: 2.12 (1.73-2.59)]. Donor factors were not the relevant risk factors for post-LT HS and the only immunosuppressive drug associated with increased risk was sirolimus [OR: 1.68 (1.07-2.64)]. The prevalence of post-LT steatohepatitis was 28.82% (19.62-38.03) and the strongest risk factor was pre-LT NAFLD. Limited outcomes data suggest that post-LT HS did not increase the risk for liver cirrhosis or mortality in these studies. Two out of 5 patients submitted to LT will develop post-LT HS, being recurrent HS more common than de novo HS. Diabetes mellitus and post-LT metabolic syndrome are the strongest risk factors for HS and baseline NAFLD for steatohepatitis. All transplanted patients should be enrolled in lifestyle interventions to prevent post-LT metabolic syndrome, and sirolimus should be avoided in high-risk patients.

NAFLD Nutritional Management: Results from a Multidisciplinary Approach.

Introduction: Lifestyle changes are the mainstay treatment of nonalcoholic fatty liver disease (NAFLD). We aimed to assess the magnitude of weight loss in a group of NAFLD patients followed on a combined lifestyle intervention by a multidisciplinary team. Methods: Patients were assessed before and after a 12-month dietary intervention (Mediterranean diet aiming at weight loss). Patients who received a structured dietary plan along with general lifestyle recommendations were designated as the multidisciplinary treatment (MdT) group. Patients who declined follow-up still received general lifestyle recommendations and were designated as the conventional treatment group, being used as a control group. Results: From the 77 patients with documented NAFLD, 31.2% of patients were overweight and 55.8% obese; 66 patients constituted the MdT group and 11 the conventional treatment group. After 3 months, 89% of patients lost weight; at 6 months, 75.4% maintained the weight lost. At 12 months, 65% of patients still decreased their weight, with 92.2% of patients in the MdT group still maintaining a lower weight than baseline versus just 50% in the conventional group (p = 0.008). Only patients in the MdT group presented a weight loss higher than 10% (9.6%; n = 6). At 12 months patients in the MdT group presented an average reduction of 4.2 kg versus a reduction of just 0.6 kg in the conventional treatment group (p = 0.016). The MdT group, but not the conventional group, presented significant differences in liver enzymes at 12 months compared to baseline. Conclusion: Adherence to a multidisciplinary approach, compared to management solely by a hepatologist, in NAFLD patients, is effective with greater weight loss after a 12-month follow-up and a lower rate of weight gain recurrence.

Introdução: Mudanças no estilo de vida são a base do tratamento do fígado gordo não-alcoólico (FGNA). O nosso objetivo foi avaliar a magnitude da perda de peso num grupo de doentes com FGNA sujeitos a intervenção nutricional e acompanhados por uma equipa multidisciplinar. Métodos: Os doentes foram avaliados antes e após uma intervenção nutricional de 12 meses (dieta mediterrânica com objetivo de perda ponderal). Os doentes que recusaram follow-up multidisciplinar, receberam recomendações gerais de estilo de vida e foram designados como grupo de tratamento convencional e usados como comparação com o grupo de tratamento multidisciplinar (MD), que em adição às recomendações gerais, recebeu plano nutricional personalizado. Resultados: Dos 77 pacientes com FGNA documentado, 31.2% dos pacientes estavam em pré-obesidade e 55.8% eram obesos; 66 pacientes constituíram o grupo MD e 11 o grupo de tratamento convencional. Após 3 meses, 89% dos pacientes perderam peso, aos 6 meses, 75.4% mantiveram a perda de peso. Aos 12 meses, 65% dos pacientes ainda diminuíram o seu peso, com 92.2% dos pacientes no grupo MD mantendo um peso inferior ao inicial vs. 50% no grupo convencional (p = 0.008). Apenas os pacientes do grupo MD, conseguiram uma perda de peso superior a 10% (9.6%; n = 6). Aos 12 meses, os pacientes do grupo MD apresentaram redução média de 4.2 kg vs. redução de 0.6 kg no grupo de tratamento convencional (p = 0.016). O grupo MD apresentou diferenças significativas nas enzimas hepáticas aos 12 meses em comparação com valores iniciais. Conclusão: A abordagem multidisciplinar em pacientes com FGNA é efetiva, com maior perda de peso durante o acompanhamento de 12 meses e maior taxa de perda de peso mantida, em comparação com acompanhamento exclusivamente pelo hepatologista.

Adiponectin, Leptin, and IGF-1 Are Useful Diagnostic and Stratification Biomarkers of NAFLD.

Background: Nonalcoholic fatty liver disease (NAFLD) is the most common chronic liver disease where liver biopsy remains the gold standard for diagnosis. Here we aimed to evaluate the role of circulating adiponectin, leptin, and insulin-like growth factor 1 (IGF-1) levels as non-invasive NAFLD biomarkers and assess their correlation with the metabolome. Materials and Methods: Leptin, adiponectin, and IGF-1 serum levels were measured by ELISA in two independent cohorts of biopsy-proven obese NAFLD patients and healthy-liver controls (discovery: 38 NAFLD, 13 controls; validation: 194 NAFLD, 31 controls) and correlated with clinical data, histology, genetic parameters, and serum metabolomics. Results: In both cohorts, leptin increased in NAFLD vs. controls (discovery: AUROC 0.88; validation: AUROC 0.83; p < 0.0001). The leptin levels were similar between obese and non-obese healthy controls, suggesting that obesity is not a confounding factor. In the discovery cohort, adiponectin was lower in non-alcoholic steatohepatitis (NASH) vs. non-alcoholic fatty liver (AUROC 0.87; p < 0.0001). For the validation cohort, significance was attained for homozygous for PNPLA3 allele c.444C (AUROC 0.63; p < 0.05). Combining adiponectin with specific serum lipids improved the assay performance (AUROC 0.80; p < 0.0001). For the validation cohort, IGF-1 was lower with advanced fibrosis (AUROC 0.67, p < 0.05), but combination with international normalized ratio (INR) and ferritin increased the assay performance (AUROC 0.81; p < 0.01). Conclusion: Serum leptin discriminates NAFLD, and adiponectin combined with specific lipids stratifies NASH. IGF-1, INR, and ferritin distinguish advanced fibrosis.

Predictors for incomplete response to ursodeoxycholic acid in primary biliary cholangitis. Data from a national registry of liver disease.

BACKGROUND: The current standard of treatment in primary biliary cholangitis (PBC) is ursodeoxycholic acid (UDCA), although a considerable proportion of patients show incomplete response resulting in disease progression. OBJECTIVE: This study aimed to assess the prevalence of incomplete response to UDCA and determine associated patients' characteristics. METHODS: Patients with PBC as main diagnosis were included from a national multicentric patient registry-Liver.pt. Main endpoints included incomplete response to UDCA treatment according to Barcelona, Paris I and Paris II criteria, Globe and UK PBC scores and the association between baseline characteristics and incomplete response according to Paris II criteria. RESULTS: A total of 434 PBC patients were identified, with a mean age of 55 years and 89.2% females. Nearly half of patients were asymptomatic at diagnosis and 93.2% had positive anti-mitochondrial antibodies. Almost all patients (95.6%) had been prescribed at least one drug for PBC treatment. At the last follow-up visit, 93.3% were under treatment of which 99.8% received UDCA. Incomplete response to UDCA was observed in 30.7%, 35.3%, 53.7% and 36.4% of patients according to Barcelona, Paris I, Paris II criteria and Globe score, respectively. After adjusting for age and sex, and accordingly to Paris II criteria, the risk for incomplete biochemical response was 25% higher for patients with cirrhosis at diagnosis (odds ratio [OR] = 1.25; 95% confidence interval [95%CI]: 1.02-1.54; p = 0.033) and 35% (95%CI:1.06-1.72; p = 0.016) and 5% (OR = 1.05; 95%CI:1.01-1.10; p = 0.013) for those with elevated gamma-glutamyl transferase (GGT) and alkaline phosphatase (ALP). CONCLUSION: A considerable proportion of patients showed incomplete biochemical response to UDCA treatment according to Paris II criteria. Cirrhosis, elevated GGT and ALP at diagnosis were identified as associated risk factors for incomplete response. Early identification of patients at risk of incomplete response could improve treatment care and guide clinical decision to a more careful patient monitorization.

Telemedicine as a tool for dietary intervention in NAFLD-HIV patients during the COVID-19 lockdown: A randomized controlled trial.

BACKGROUND & AIMS: Given reports of changes in dietary habits during covid-19 lockdown, our aim was to assess weight changes, over a 3-month Covid-19 national lockdown in a cohort of NAFLD-HIV patients on a dietary intervention trial. METHODS: After NAFLD screening in an outpatient Infectious Diseases Clinic, NAFLD patients were randomly allocated to general dietary recommendations (SC group) or to a structured dietary intervention based on the Mediterranean diet (intervention group). During lockdown, follow-up consultations in the intervention group were done by video and/or phone. After 3 months of lockdown, all patients (intervention and SC group) consented to a telephone interview which aimed to characterize eating habits and lifestyle changes and evaluate stress and depression. Biochemical data when available, was compared between the peri-period of confinement. RESULTS: One hundred and twelve patients were screened. From the 55 NAFDL identified, 27 were allocated to dietary intervention and 28 to SC and were followed before lockdown for a mean period of 5.0 ± 1.5 months in which SC group gained a median of 0.65 kg vs. a median loss of 1.5 kg in the intervention group (p < 0.001). During lockdown, 93.3% of patients in the SC group referred that "diet got worse" vs. 6.7% in the intervention group p < 0.01), and 35.3% vs. 15.7% (p = 0.014) reported increase in appetite, respectively. Both groups gained weight, SC group vs. 0.7 ± 1.7 kg in the intervention group, p < 0.001). Higher weight gain was associated with changes in the dietary pattern (3.8 ± 2.1 kg vs. 2.0 ± 1.3 kg in "no change in dietary pattern"; p = 0.002). Glucose blood levels increased after lockdown in the SC group, with a mean increase of 15 mg/dl (p = 0.023). The remaining metabolic parameters remained unchanged. CONCLUSION: The maintenance of dietary intervention, using telemedicine, can mitigate the adverse change in dietary habits and physical activity pattern, preventing a substantial increase in body weight.

What Is the Role of the New Index Relative Fat Mass (RFM) in the Assessment of Nonalcoholic Fatty Liver Disease (NAFLD)?

INTRODUCTION: Nonalcoholic fatty liver disease (NAFLD) is the liver manifestation of adiposopathy. Recently, a new score was developed to estimate body fat percentage (relative fat mass, RFM). We aimed to evaluate the value of RFM in predicting the presence and severity of NAFLD, compared with other anthropometric measurements. METHODS: RFM, body mass index (BMI), and other anthropometric measurements were evaluated in two cohorts of subjects: a cohort from a Portuguese prospective epidemiological study (e_Cor) and morbidly obese patients with biopsy-proven NAFLD. We evaluated if RFM and BMI were related with the presence and severity of liver disease, which was assessed by noninvasive tools in the first cohort and by liver histology in the morbidly obese cohort. The independence of relations found in univariate analysis was assessed with multivariable logistic regression analysis. RESULTS: In the general population cohort, 744 subjects (48% male) were enrolled. BMI-defined obesity was present in 23% and RFM-defined obesity in 86%. Insulin resistance (IR) related with BMI-defined obesity (OR 4.37 [2.16-8.84]) and weight (OR 1.05 [1.02-1.08]) in men, and waist circumference (WC) (OR 1.07 [1.03-1.11]) in women. Dyslipidemia and hypertension related with RFM-defined obesity in men (OR 2.96 [1.36-6.47] and OR 5.37 [1.31-22.06], respectively). Ultrasound-diagnosed NAFLD in 33% related with weight in men (OR 1.03 [1.003-1.06] and WC in women (OR 1.06 [1.02-1.10]). In men, ALT elevation related with weight (OR 1.04 [1.02-1.07]). In women, advanced fibrosis (estimated by NAFLD Fibrosis Score) associated with BMI-defined obesity (OR 42.43 [3.61-498.13]). In the morbidly obese cohort, 152 subjects were enrolled, of whom 84% were female, 37% had steatohepatitis, and 9.4% had advanced fibrosis. Adiponectin associated inversely and leptin positively with RFM in men. The severity of steatosis increased linearly with BMI and WC in women. Higher BMI associated with steatohepatitis in women and advanced fibrosis in men. CONCLUSION: RFM-defined obesity better predicted dyslipidemia and hypertension (though not IR) and adipokine imbalance; however, it did not add value to BMI-defined obesity in predicting NAFLD or liver injury.

Skeletal muscle miR-34a/SIRT1:AMPK axis is activated in experimental and human non-alcoholic steatohepatitis.

Non-alcoholic fatty liver disease (NAFLD) pathogenesis associates with intramyocellular lipid deposition and mitochondrial dysfunction. microRNAs (miRs), including pro-apoptotic miR-34a, are modulated during disease progression in liver tissue and plasma. We aimed to investigate the functional role of the miR-34a/SIRT1:AMP-activated protein kinase (AMPK) pathway in modulating local mitochondrial dysfunction in the skeletal muscle of human and experimental non-alcoholic steatohepatitis. Muscle biopsies were obtained from morbid obese NAFLD patients undergoing bariatric surgery. C57BL/6N mice were fed different NAFLD-inducing diets and C2C12 muscle cells incubated with palmitic acid (PA) in the presence or absence of an AMPK activator, or upon miR-34a functional modulation. Several muscle miRNAs, including miR-34a, were found increased with human NAFLD progression. Activation of the miR-34a/SIRT1:AMPK pathway, concomitant with impairment in insulin signalling mediators and deregulation of mitochondrial-shaping proteins, was evident in C2C12 cells incubated with PA, as well as in the skeletal muscle of all three diet-induced NAFLD mice models. Functional studies established the association between miR-34a- and PA-induced muscle cell deregulation. Of note, activation of AMPK almost completely prevented miR-34a- and PA-induced cellular stress. In addition, the miR-34a/SIRT1:AMPK pathway and mitochondrial dynamics dysfunction were also found amplified in muscle of human NAFLD. Finally, muscle miR-34a expression and mitofusin 2 (Mfn2) protein levels correlated with hallmarks of NAFLD and disease progression. Our results indicate that activation of the miR-34a/SIRT1:AMPK pathway leads to mitochondrial dynamics dysfunction in skeletal muscle of human and experimental NAFLD, representing an appealing prospective target in metabolic syndrome. KEY MESSAGES: Skeletal muscle microRNAs are modulated during NAFLD progression. Palmitic acid-induced muscle cell dysfunction occurs, at least in part, through activation of the miR-34a/SIRT1:AMPK pathway. miR-34a/SIRT1:AMPK activation associates with mitochondria dynamics dysfunction in human NAFLD.

miR-21 ablation and obeticholic acid ameliorate nonalcoholic steatohepatitis in mice.

This corrects the article DOI: 10.1038/cddis.2017.172.

miR-21 ablation and obeticholic acid ameliorate nonalcoholic steatohepatitis in mice.

microRNAs were recently suggested to contribute to the pathogenesis of nonalcoholic fatty liver disease (NAFLD), a disease lacking specific pharmacological treatments. In that regard, nuclear receptors are arising as key molecular targets for the treatment of nonalcoholic steatohepatitis (NASH). Here we show that, in a typical model of NASH-associated liver damage, microRNA-21 (miR-21) ablation results in a progressive decrease in steatosis, inflammation and lipoapoptosis, with impairment of fibrosis. In a complementary fast food (FF) diet NASH model, mimicking features of the metabolic syndrome, miR-21 levels increase in both liver and muscle, concomitantly with decreased expression of peroxisome proliferator-activated receptor α (PPARα), a key miR-21 target. Strikingly, miR-21 knockout mice fed the FF diet supplemented with farnesoid X receptor (FXR) agonist obeticholic acid (OCA) display minimal steatosis, inflammation, oxidative stress and cholesterol accumulation. In addition, lipoprotein metabolism was restored, including decreased fatty acid uptake and polyunsaturation, and liver and muscle insulin sensitivity fully reinstated. Finally, the miR-21/PPARα axis was found amplified in liver and muscle biopsies, and in serum, of NAFLD patients, co-substantiating its role in the development of the metabolic syndrome. By unveiling that miR-21 abrogation, together with FXR activation by OCA, significantly improves whole body metabolic parameters in NASH, our results highlight the therapeutic potential of nuclear receptor multi-targeting therapies for NAFLD.

Epidemiological modifiers of non-alcoholic fatty liver disease: Focus on high-risk groups.

An improved understanding of non-alcoholic fatty liver disease epidemiology would lead to identification of individuals at high risk of developing chronic liver disease and extra-hepatic complications, thus contributing to more effective case finding of non-alcoholic fatty liver disease among selected groups. We aimed to illustrate the epidemiology of non-alcoholic fatty liver disease in high-risk groups, which were identified based on existing literature. To this end, PubMed was searched to retrieve original articles published until May 2015 using relevant and pertinent keywords "nonalcoholic fatty liver disease" and "diabetes", "obesity", "hyperlipidaemia", "familial heterozygous hypobetalipoproteinaemia", "hypertension", "metabolic syndrome", "ethnicity", "family history" or "genetic polymorphisms". We found that age, sex and ethnicity are major physiological modifiers of the risk of non-alcoholic fatty liver disease, along with belonging to "non-alcoholic fatty liver disease families" and carrying risk alleles for selected genetic polymorphisms. Metabolic syndrome, diabetes, obesity, mixed hyperlipidaemia and hypocholesterolaemia due to familial hypobetalipoproteinaemia are the major metabolic modifiers of non-alcoholic fatty liver disease risk. Compared with these metabolic conditions, however, arterial hypertension appears to carry a relatively more modest risk of non-alcoholic fatty liver disease. A better understanding of the epidemiology of non-alcoholic fatty liver disease may result in a more liberal policy of case finding among high-risk groups.

Proangiogenic factors in the development of HCC in alcoholic cirrhosis.

Alcoholic liver disease, the most common cause of liver cirrhosis, is associated with an increased risk for hepatocellular carcinoma. Angiogenic factors have been implicated in pathophysiology of cirrhosis, and of hepatocellular carcinoma, and in particular of alcoholic liver cirrhosis, due to alcohol induced hypoxia associated with increased hepatic oxygen consumption. In one study, it was found that among genetic polymorphisms in proangiogenic factors, KDR and VEFGA may confer an increased risk of HCC, in patients with ALD. There is need of further studies of the proangiogenic factors in HCC, in order to help us define their use as prognostic markers and also as markers of response to treatment.

Schistosome-induced cholangiocyte proliferation and osteopontin secretion correlate with fibrosis and portal hypertension in human and murine schistosomiasis mansoni.

Schistosomiasis is a major cause of portal hypertension worldwide. It associates with portal fibrosis that develops during chronic infection. The mechanisms by which the pathogen evokes these host responses remain unclear. We evaluated the hypothesis that schistosome eggs release factors that directly stimulate liver cells to produce osteopontin (OPN), a pro-fibrogenic protein that stimulates hepatic stellate cells to become myofibroblasts. We also investigated the utility of OPN as a biomarker of fibrosis and/or severity of portal hypertension. Cultured cholangiocytes, Kupffer cells and hepatic stellate cells were treated with soluble egg antigen (SEA); OPN production was quantified by quantitative reverse transcriptase polymerase chain reaction (qRTPCR) and ELISA; cell proliferation was assessed by BrdU (5-bromo-2'-deoxyuridine). Mice were infected with Schistosoma mansoni for 6 or 16 weeks to cause early or advanced fibrosis. Liver OPN was evaluated by qRTPCR and immunohistochemistry (IHC) and correlated with liver fibrosis and serum OPN. Livers from patients with schistosomiasis mansoni (early fibrosis n=15; advanced fibrosis n=72) or healthy adults (n=22) were immunostained for OPN and fibrosis markers. Results were correlated with plasma OPN levels and splenic vein pressures. SEA-induced cholangiocyte proliferation and OPN secretion (P<0.001 compared with controls). Cholangiocytes were OPN (+) in Schistosoma-infected mice and humans. Liver and serum OPN levels correlated with fibrosis stage (mice: r=0.861; human r=0.672, P=0.0001) and myofibroblast accumulation (mice: r=0.800; human: r=0.761, P=0.0001). Numbers of OPN (+) bile ductules strongly correlated with splenic vein pressure (r=0.778; P=0.001). S. mansoni egg antigens stimulate cholangiocyte proliferation and OPN secretion. OPN levels in liver and blood correlate with fibrosis stage and portal hypertension severity.

Genetic ancestry analysis in non-alcoholic fatty liver disease patients from Brazil and Portugal.

AIM: To study the association between genetic ancestry, non-alcoholic fatty liver disease (NAFLD) metabolic characteristics in two cohorts of patients, from Brazil and Portugal. METHODS: We included 131 subjects from Brazil [(n = 45 with simple steatosis (S. Steatosis) and n = 86 with nonalcoholic steatohepatitis (NASH)] and 90 patients from Portugal (n = 66, S. Steatosis; n = 24, NASH). All patients had biopsy-proven NAFLD. In histologic evaluation NAFLD activity score was used to assess histology and more than 5 points defined NASH in this study. Patients were divided into two groups according to histology diagnosis: simple steatosis or non-alcoholic statohepatitis. Genetic ancestry was assessed using real-time polymerase chain reaction. Seven ancestry informative markers (AT3-I/D, LPL, Sb19.3, APO, FY-Null, PV92, and CKMM) with the greatest ethnic-geographical differential frequencies (≥ 48%) were used to define genetic ancestry. Data were analyzed using R PROJECTS software. Ancestry allele frequencies between groups were analyzed by GENEPOP online and the estimation of genetic ancestry contribution was evaluated by ADMIX-95 software. The 5% alpha-error was considered as significant (P < 0.05). RESULTS: In the Brazilian sample, NASH was significantly more frequent among the elderly patients with diabetes (NASH 56 ± 1.1 years old vs S. Steatosis 51 ± 1.5 years old, P = 3.7 x 10(-9)), dyslipidemia (NASH 63% vs S. Steatosis 37%, P = 0.009), higher fasting glucose levels (NASH 124 ± 5.2 vs S. Steatosis 106 ± 5.3, P = 0.001) and Homeostatic Model of Assessment index > 2.5 [NASH 5.3 (70.8%) vs S. Steatosis 4.6 (29.2%) P = 0.04]. In the Portuguese study population, dyslipidemia was present in all patients with NASH (P = 0.03) and hypertension was present in a larger percentage of subjects in the S. Steatosis group (P = 0.003, respectively). The genetic ancestry contribution among Brazilian and Portuguese individuals with NASH was similar to those with S. Steatosis from each cohort (Brazilian cohort: P = 0.75; Portuguese cohort: P = 0.97). Nonetheless, the genetic ancestry contribution of the Brazilian and Portuguese population were different, and a greater European and Amerindian ancestry contribution was detected in the Portuguese population while a higher African genetic ancestry contribution was observed in Brazilian population of both NASH and S. Steatosis groups. CONCLUSION: There was no difference between the genetic ancestry contribution among Brazilian and Portuguese individuals with NASH and S. Steatosis from each cohort.

Genetic polymorphisms of proangiogenic factors seem to favor hepatocellular carcinoma development in alcoholic cirrhosis.

OBJECTIVE: Angiogenesis has been associated with hepatic cirrhosis and hepatocellular carcinoma (HCC). Alcohol promotes liver hypoxia, a trigger of angiogenesis. We aimed to evaluate whether the frequency of three polymorphisms in hypoxia-induced factor-1α (HIF-1α), vascular endothelial growth factor A (VEGFA), and KDR (encoding vascular endothelial growth factor receptor 2) genes was higher in alcoholics presenting liver disease (ALD) and ALD patients who developed HCC. MATERIALS AND METHODS: Functional HIF-1α 1744C/T, VEGFA 2578C/A, and KDR 1416A/T single-nucleotide polymorphisms were studied in 125 ALD patients and 88 heavy drinkers without liver disease (NLD). ALD patients were followed up to 9 years or until they died; 26 patients developed HCC. RESULTS: ALD patients were older than NLD (56±11 vs. 50±13, P<0.001), but drank less (215±164 vs. 331±293 g/day, P<0.001). No differences were found between HIF-1α, VEGFA, or KDR allelic frequencies or genotypes, isolated or simultaneously, between ALD and NLD. In ALD patients, those who developed HCC had a higher KDR 1416T allele frequency (36 vs. 15%, P=0.004; odds ratio 2.72; 95% confidence interval 1.35-5.46). There was also a progressive increase in genotypes with one or two T alleles in patients who developed HCC: AA 50 vs. 73%, AT 35 vs. 23%, and TT 15 vs. 4% (P=0.009). The simultaneous presence of KDR 1416T and VEGFA 2578A was associated with an increased risk of HCC (odds ratio 3.088; 95% confidence interval 1.20-7.96). CONCLUSION: Genetic polymorphisms in proangiogenic factors did not associate with the risk of ALD in heavy drinkers. However, KDR and VEFGA polymorphisms may confer an increased risk of HCC in patients with ALD.

Alcohol activates the hedgehog pathway and induces related procarcinogenic processes in the alcohol-preferring rat model of hepatocarcinogenesis.

BACKGROUND: Alcohol consumption promotes hepatocellular carcinoma (HCC). The responsible mechanisms are not well understood. Hepatocarcinogenesis increases with age and is enhanced by factors that impose a demand for liver regeneration. Because alcohol is hepatotoxic, habitual alcohol ingestion evokes a recurrent demand for hepatic regeneration. The alcohol-preferring (P) rat model mimics the level of alcohol consumption by humans who habitually abuse alcohol. Previously, we showed that habitual heavy alcohol ingestion amplified age-related hepatocarcinogenesis in P rats, with over 80% of alcohol-consuming P rats developing HCCs after 18 months of alcohol exposure, compared with only 5% of water-drinking controls. METHODS: Herein, we used quantitative real-time PCR and quantitative immunocytochemistry to compare liver tissues from alcohol-consuming P rats and water-fed P rat controls after 6, 12, or 18 months of drinking. We aimed to identify potential mechanisms that might underlie the differences in liver cancer formation and hypothesized that chronic alcohol ingestion would activate Hedgehog (HH), a regenerative signaling pathway that is overactivated in HCC. RESULTS: Chronic alcohol ingestion amplified age-related degenerative changes in hepatocytes, but did not cause appreciable liver inflammation or fibrosis even after 18 months of heavy drinking. HH signaling was also enhanced by alcohol exposure, as evidenced by increased levels of mRNAs encoding HH ligands, HH-regulated transcription factors, and HH target genes. Immunocytochemistry confirmed increased alcohol-related accumulation of HH ligand-producing cells and HH-responsive target cells. HH-related regenerative responses were also induced in alcohol-exposed rats. Three of these processes (i.e., deregulated progenitor expansion, the reverse Warburg effect, and epithelial-to-mesenchymal transitions) are known to promote cancer growth in other tissues. CONCLUSIONS: Alcohol-related changes in Hedgehog signaling and resultant deregulation of liver cell replacement might promote hepatocarcinogenesis.

NAFLD, NASH and liver cancer.

NAFLD affects a large proportion of the US population and its incidence and prevalence are increasing to epidemic proportions around the world. As with other liver diseases that cause cirrhosis, NAFLD increases the risk of liver cancer, a disease with poor outcomes and limited therapeutic options. The incidences of hepatocellular carcinoma (HCC) and intrahepatic cholangiocarcinoma are also rising, and HCC is now the leading cause of obesity-related cancer deaths in middle-aged men in the USA. In this Review, we summarize the correlations between liver cancer and NAFLD-related cirrhosis, and the role of the metabolic syndrome in the development of liver cancer from diverse aetiologies, including HCV-mediated cirrhosis. Recent advances in understanding the progression of NAFLD to HCC from preclinical models will also be discussed. Targeted genetic manipulation of certain metabolic or stress-response pathways, including one-carbon metabolism, NF-κB, PTEN and microRNAs, has been valuable in elucidating the pathways that regulate carcinogenesis in NAFLD. Although tremendous advances have occurred in the identification of diagnostic and therapeutic opportunities to reduce the progression of NAFLD, considerable gaps in our knowledge remain with regard to the mechanisms by which NAFLD and its risk factors promote liver cancer.

miR-34a/SIRT1/p53 is suppressed by ursodeoxycholic acid in the rat liver and activated by disease severity in human non-alcoholic fatty liver disease.

BACKGROUND & AIMS: Non-alcoholic fatty liver disease (NAFLD) comprises a spectrum of stages from simple steatosis to non-alcoholic steatohepatitis (NASH). However, disease pathogenesis remains largely unknown. microRNA (miRNA or miR) expression has recently been reported to be altered in human NASH, and modulated by ursodeoxycholic acid (UDCA) in the rat liver. Here, we aimed at evaluating the miR-34a/Sirtuin 1(SIRT1)/p53 pro-apoptotic pathway in human NAFLD, and to elucidate its function and modulation by UDCA in the rat liver and primary rat hepatocytes. METHODS: Liver biopsies were obtained from NAFLD morbid obese patients undergoing bariatric surgery. Rat livers were collected from animals fed a 0.4% UDCA diets. Primary rat hepatocytes were incubated with bile acids or free fatty acids (FFAs) and transfected with a specific miRNA-34a precursor and/or with a p53 overexpression plasmid. p53 transcriptional activity was assessed by ELISA and target reporter constructs. RESULTS: miR-34a, apoptosis and acetylated p53 increased with disease severity, while SIRT1 diminished in the NAFLD liver. UDCA inhibited the miR-34a/SIRT1/p53 pathway in the rat liver in vivo and in primary rat hepatocytes. miR-34a overexpression confirmed its targeting by UDCA, which prevented miR-34a-dependent repression of SIRT1, p53 acetylation, and apoptosis. Augmented apoptosis by FFAs in miR-34a overexpressing cells was also inhibited by UDCA. Finally, p53 overexpression activated miR-34a/SIRT1/p53, which in turn was inhibited by UDCA, via decreased p53 transcriptional activity. CONCLUSIONS: Our results support a link between liver cell apoptosis and miR-34a/SIRT1/p53 signaling, specifically modulated by UDCA, and NAFLD severity. Potential endogenous modulators of NAFLD pathogenesis may ultimately provide new tools for therapeutic intervention.

Non-invasive diagnosis of non-alcoholic fatty liver disease. A critical appraisal.

Non-alcoholic fatty liver disease (NAFLD) affects one in every three subjects in the occidental world. The vast majority will not progress, but a relevant minority will develop liver cirrhosis and its complications. The classical gold standard for diagnosing and staging NAFLD and assessing fibrosis is liver biopsy (LB). However, it has important sample error issues and subjectivity in the interpretation, apart from a small but real risk of complications. The decision to perform an LB is even harder in a condition so prevalent such as NAFLD, in which the probability of finding severe liver injury is low. In an attempt to overcome LB and to subcategorize patients with NAFLD in different prognoses allowing better management decisions, several non-invasive methods have been studied in the last decade. The literature is vast and confusing. This review will summarize which methods have been tested and how they perform, which tests are adequate for clinical practice and how they can change the management of these patients.