M. Romero-Gómez

Metabolomic Identification of Subtypes of Nonalcoholic Steatohepatitis

BACKGROUND & AIMS Nonalcoholic fatty liver disease (NAFLD) is a consequence of defects in diverse metabolic pathways that involve hepatic accumulation of triglycerides. Features of these aberrations might determine whether NAFLD progresses to nonalcoholic steatohepatitis (NASH). We investigated whether the diverse defects observed in patients with NAFLD are caused by different NAFLD subtypes with specific serum metabolomic profiles, and whether these can distinguish patients with NASH from patients with simple steatosis. METHODS We collected liver and serum from methionine adenosyltransferase 1a knockout (MAT1A-KO) mice, which have chronically low levels of hepatic S-adenosylmethionine (SAMe) and spontaneously develop steatohepatitis, as well as C57Bl/6 mice (controls); the metabolomes of all samples were determined. We also analyzed serum metabolomes of 535 patients with biopsy-proven NAFLD (353 with simple steatosis and 182 with NASH) and compared them with serum metabolomes of mice. MAT1A-KO mice were also given SAMe (30 mg/kg/day for 8 weeks); liver samples were collected and analyzed histologically for steatohepatitis. RESULTS Livers of MAT1A-KO mice were characterized by high levels of triglycerides, diglycerides, fatty acids, ceramides, and oxidized fatty acids, as well as low levels of SAMe and downstream metabolites. There was a correlation between liver and serum metabolomes. We identified a serum metabolomic signature associated with MAT1A-KO mice that also was present in 49% of the patients; based on this signature, we identified 2 NAFLD subtypes. We identified specific panels of markers that could distinguish patients with NASH from patients with simple steatosis for each subtype of NAFLD. Administration of SAMe reduced features of steatohepatitis in MAT1A-KO mice. CONCLUSIONS In an analysis of serum metabolomes of patients with NAFLD and MAT1A-KO mice with steatohepatitis, we identified 2 major subtypes of NAFLD and markers that differentiate steatosis from NASH in each subtype. These might be used to monitor disease progression and identify therapeutic targets for patients.

Nuevas perspectivas terapéuticas en la esteatohepatitis no alcohólica

Management of non-alcoholic steatohepatitis is focused on restitution of metabolic derangement, weight loss and drugs able to improve steatosis, ballooning and fibrosis. Life-style interventions based on Mediterranean diet and increasing physical activity are the first line therapy. In patients with unsuccessful life-style intervention several drugs are under development: agonist PPAR, agonist GLP-1R and agonist FXR together with drugs focussing on inflammation, ballooning, apoptosis and fibrosis. Bariatric surgery or advanced endoscopy are reserved for morbid obese without response to life-style intervention and weighting loss drugs.

Metabolomic‐based noninvasive serum test to diagnose nonalcoholic steatohepatitis: Results from discovery and validation cohorts

Nonalcoholic fatty liver disease (NAFLD) is the most common type of chronic liver disease worldwide and includes a broad spectrum of histologic phenotypes, ranging from simple hepatic steatosis or nonalcoholic fatty liver (NAFL) to nonalcoholic steatohepatitis (NASH). While liver biopsy is the reference gold standard for NAFLD diagnosis and staging, it has limitations due to its sampling variability, invasive nature, and high cost. Thus, there is a need for noninvasive biomarkers that are robust, reliable, and cost effective. In this study, we measured 540 lipids and amino acids in serum samples from biopsy‐proven subjects with normal liver (NL), NAFL, and NASH. Using logistic regression analysis, we identified two panels of triglycerides that could first discriminate between NAFLD and NL and second between NASH and NAFL. These noninvasive tests were compared to blinded histology as a reference standard. We performed these tests in an original cohort of 467 patients with NAFLD (90 NL, 246 NAFL, and 131 NASH) that was subsequently validated in a separate cohort of 192 patients (7 NL, 109 NAFL, 76 NASH). The diagnostic performances of the validated tests showed an area under the receiver operating characteristic curve, sensitivity, and specificity of 0.88 ± 0.05, 0.94, and 0.57, respectively, for the discrimination between NAFLD and NL and 0.79 ± 0.04, 0.70, and 0.81, respectively, for the discrimination between NASH and NAFL. When the analysis was performed excluding patients with glucose levels >136 mg/dL, the area under the receiver operating characteristic curve for the discrimination between NASH and NAFL increased to 0.81 ± 0.04 with sensitivity and specificity of 0.73 and 0.80, respectively. Conclusion: The assessed noninvasive lipidomic serum tests distinguish between NAFLD and NL and between NASH and NAFL with high accuracy. (Hepatology Communications 2018;2:807‐820)

Documento de consenso. Manejo de la enfermedad hepática grasa no alcohólica (EHGNA). Guía de práctica clínica

Non-alcoholic fatty liver disease (NAFLD) is the main cause of liver diseases in Spain and the incidence is raising due to the outbreak of type 2 diabetes and obesity. This CPG suggests recommendation about diagnosis, mainly non-invasive biomarkers, and clinical management of this entity. Life-style modifications to achieve weight loss is the main target in the management of NAFLD. Low caloric Mediterranean diet and 200 minutes/week of aerobic exercise are encouraged. In non-responders patients with morbid obesity, bariatric surgery or metabolic endoscopy could be indicated. Pharmacological therapy is indicated in patients with NASH and fibrosis and non-responders to weight loss measures. NAFLD could influence liver transplantation, as a growing indication, the impact of steatosis in the graft viability, de novo NAFLD rate after OLT and a raised cardiovascular risk that modify the management of this entity. The current CPG was the result of the First Spanish NAFLD meeting in Seville.

Natural Extracts Abolished Lipid Accumulation in Cells Harbouring non-favourable PNPLA3 genotype

BACKGROUND & AIMS G-allele of PNPLA3 (rs738409) favours triglycerides accumulation and steatosis. In this study, we examined the effect of quercetin and natural extracts from mushroom and artichoke on reducing lipid accumulation in hepatic cells. MATERIAL AND METHODS Huh7.5 cells were exposed to oleic acid (OA) and treated with quercetin and extracts to observe the lipid accumulation, the intracellular-TG concentration and the LD size. Sterol regulatory element binding proteins-1 (SREBP-1), peroxisome proliferator-activated receptor (PPARα-γ) and cholesterol acyltransferase (ACAT) gene expression levels were analysed. RESULTS Quercetin decreased the intracellular lipids, LD size and the levels of intracellular-TG through the down-regulation of SREBP-1c, PPARγand ACAT1 increasing PPARα. The natural-extracts suppressed OA-induced lipid accumulation and the intracellular-TG. They down-regulate the hepatic lipogenesis through SREBP-1c, besides the activation of lipolysis through the increasing of PPARα expression. CONCLUSIONS Quercetin and the aqueous extracts decrease intracellular lipid accumulation by down-regulation of lipogenesis and up-regulation of lipolysis.Background & aims. G-allele of PNPLA3 (rs738409) favours triglycerides accumulation and steatosis. In this study, we examined the effect of quercetin and natural extracts from mushroom and artichoke on reducing lipid accumulation in hepatic cells. MATERIAL AND METHODS Huh7.5 cells were exposed to oleic acid (OA) and treated with quercetin and extracts to observe the lipid accumulation, the intracellular-TG concentration and the LD size. Sterol regulatory element binding proteins-1 (SREBP-1), peroxisome proliferator-activated receptor (PPARα-γ) and cholesterol acyltransferase (ACAT) gene expression levels were analysed. RESULTS Quercetin decreased the intracellular lipids, LD size and the levels of intracellular-TG through the down-regulation of SREBP-1c, PPARγ and ACAT1 increasing PPARα. The natural-extracts suppressed OA-induced lipid accumulation and the intracellular-TG. They down-regulate the hepatic lipogenesis through SREBP-1c, besides the activation of lipolysis through the increasing of PPARα expression. CONCLUSIONS Quercetin and the aqueous extracts decrease intracellular lipid accumulation by down-regulation of lipogenesis and up-regulation of lipolysis.

Inflammasome and extrahepatic organ failure in acute-on-chronic liver failure

Acute-on-chronic liver failure (ACLF) is a syndrome characterized by acute deterioration of liver function in patients with pre-existent chronic liver disease together with organ failure (OF). Six organs have been defined: brain, circulation, coagulation, kidneys, lungs and liver. Consequently, it is associated with high short-term mortality (1). The prevalence of ACLF is around 30% among patients with end-stage cirrhosis hospitalized with acute decompensation (AD). Patients are dynamically classified as having ACLF according to the number of OFs (corresponding to three grades of severity), with an increased risk of short-term death. Mortality correlates better with the early clinical course of ACLF than with the initial grade at presentation, especially between the third and seventh day after diagnosis (2).

The Oral Glutamine Challenge in Liver Cirrhosis

Oral glutamine challenge is a test which aims to measure increase in blood ammonia after glutamine intake, what is similar to the situation that occurs after a protein meal. Oral glutamine challenge induces an increase in blood ammonia in patients with cirrhosis but not in healthy control or transplanted subjects. A pathological response curve for glutamine tolerance is defined as an ammonia rise to 128 mg/dl at 60 min after the glutamine ingestion. No serious adverse effects have been described in the different published studies. Oral glutamine challenge is able to predict minimal hepatic encephalopathy, as well as is associated with a poor survival. On the other hand, ammonia is mainly produced by glutaminase. Therefore, glutaminase enzyme is an excellent target to develop new drugs that reduced its activity.

P0938 : Olive oil comsumption ameliorates non-alcoholic steatohepatitis induced by hypercaloric and hyperlipidic western diets

Background and Aims: Activation of hepatic stellate cells (HSC) and dysregulation of several mediators such as matrix metalloproteinases (MMPs) and their inhibitors (TIMPs) play a determinant role in the fibrogenesis during the progression of NAFLD to NASH. This study was aimed to establish the interplay between hepatocytes and HSC in an in vitro cell model of NASH. Methods: The effect of free fatty acids (FFA) (Oleic:Palmitic, 2:1) was analyzed at short (24h) and long (96h) exposure times in different experimental set-ups: (1) Monoculture of each cell type; (2) Transwell system (soluble mediators effects) and (3) simultaneous co-culture (SCC) by seeding both cell types together (cell-to-cell interaction). In each system was assessed the amount of steatosis; expression of HSC activation marker (a-SMA), ECM turnover regulators (MMP-2 and TIMP-2) as well as collagen biosynthesis in comparison to untreated cells (ctrl). Results: The amount of steatosis was comparable among all the experimental set-ups. However, HSC activation in terms of a-SMA gene (2.20±0.25-folds; p < 0.01) and protein (1.70±0.20folds; p < 0.01) expression was only increased in the SCC and was maximal after 24h of FFA exposure. Similarly, the close contact of the two cell types induced an up-regulation of TIMP2 protein (1.42±0.27-folds; p < 0.05) which was inversely correlated both with MMP-2 protein (0.58±0.10-folds; p < 0.01) and activity (0.70±0.13folds; p < 0.05). This dysregulation was accompanied by an increase of collagen biosynthesis at longer FFA exposure times (1.5±0.10folds, p < 0.01). Any of these effects was directly induced by FFA (monoculture) nor by the soluble mediators (transwell). Conclusions: Our data suggest that hepatocytes-to-HSC proximity/interaction is essential for fibrosis initiation in NASH.

606 METFORMIN PREVENTS OVERT HEPATIC ENCEPHALOPATHY (HE) IN CIRRHOTICS: A RETROSPECTIVE OBSERVATIONAL STUDY

605 METFORMIN TREATMENT REDUCES HEPATOCELLULAR CARCINOMA (HCC) RISK IN PATIENTS WITH CIRRHOSIS C: A RETROSPECTIVE OBSERVATIONAL STUDY OF TWO INDEPENDENT COHORTS J. Ampuero, M. Fernandez-Gil, D. Nunez, S. Alonso, M. Maraver, M.L. Gutierrez, C. Cano, A. Rojas, J.L. Lledo, M. Millan, R. Gallego-Duran, I. Camacho, B. Figueruela, E. Suarez, C. Fernandez-Rodriguez, M. Romero-Gomez. UCM Digestive Diseases and CIBERehd, Valme University Hospital, Sevilla, Digestive Diseases Unit, Alcorcon University Hospital, Madrid, Spain E-mail: javi.ampuero@gmail.com