Adipose Tissue Insulin Resistance is Associated with Macrophage Activation in Non-Diabetic Patients with Non-Alcoholic Fatty Liver Disease

POSTER PRESENTATIONS the human transcriptome between a) NAFLD, b) AIH, and c) pathologically or clinically combined AIH/NAFLD. Methods: The training cohort (A) was comprised of pathologically and clinically proven 11 AIH and 9 NAFLD cases, including 8 nonalcoholic steatohepatitis (NASH). The test cohort (B) included four atypical cases, that is, 3 AIH with steatosis (grade 1 by NASH CRN score) and one NASH that obtained both biochemical and pathological remission by prednisolone, but had relapse after its cessation (case R). Pathological diagnosed was performed by a boardcertified pathologist. Hepatic RNA were extracted from biopsy samples and microarray hybridization was performed on Human Oligo chip 25 k array (TORAY, Tokyo, Japan), using Ambion Amino Allyl aRNA kit (Life Technologies, Carlsbad, U.S.A.) for RNA amplification and hybridization. With cohort A, we first extracted genes whose expression differed 1.5 fold< between NAFLD and AIH (FDR corrected p < 0.05, Genes X). Hierarchical clustering analysis was subsequently conducted with Genes X among all samples, including cohort B. Gene ontology (GO) analysis was performed using DAVID Bioinformatics Resources (Huang DW et al., Nature Protoc, 2009). Results: 2924 genes were identified as Genes X. GO analysis demonstrated that Genes X were significantly enriched in those associated with biological functions, such as immune response (2.3 × e−27), defense response (1.5 × e−26), inflammatory response (4.1 × e−18), and response to wounding (1.9 × e−15), in descending order. Using those genes, all samples were separated into two clusters: AIH and NAFLD in cohort A were exclusively included in Y and Z, respectively. Remarkably, two AIH with steatosis and longitudinal biopsies from case R (at initial diagnosis, in remission, and at relapse) were categorized into Z. Conclusions: The NAFLD transcriptome might produce a range of phenotypes, some of which cross over to the pathological and the clinical AIH. FRI-284 ROLE OF 13C-OCTANOATE BREATH TEST FOR NON-INVASIVE DIAGNOSIS OF NON-ALCOHOLIC STEATOHEPATITIS C. Fierbinteanu-Braticevici1, A. Moldoveanu1, L. Tribus1, A. Petrisor1, A. Necula1, O. Viasu1. 1Gastroenterology, Univeristy of Medicine and Pharmacy “Carol Davila” Bucharest, University Hospital Bucharest, Bucharest, Romania E-mail: [email protected] Background and Aims: Nonalcoholic fatty liver disease (NAFLD) represents a group of conditions ranging from simple steatosis to nonalcoholic steatohepatitis (NASH), which is characterized by a progressive course, evolving to cryptogenic cirrhosis. Hepatic mitochondrial dysfunction has been involved in progression of Nonalcoholic Fatty Liver to NASH esspecialy by oxidative stress. The 13 C- Octanoate breath test (OBT), enables mitochondrial activity evaluation, as a source of oxidative stress, implicated in the development of nonalcoholic steatohepatitis. Aim: to evaluate the efficacy of OBT in differentiating healthy subjects and patients with simple steatosis from NASH patients. Methods: C- Octanoate breath test was performed in 40 patients with histological proven NAFLD (ranging from simple steatosis to severe steatohepatitis) and in 20 healthy controls. The correlation between the 13C- Octanoate breath test and liver biopsy was tested using Spearman’s coefficient. The overall validity was measured using the area under receiver operating characteristic curve (AUROC) with 95% CI. Results: Delta over baseline values (DOB) of NASH patients at 15 min were significantly higher from controls and patients with simple steatosis (23.95; vs. 15.31 p < 0.001). The cumulative recovery of 13CO2 after 60 minutes (CUM 60) was significantly higher in NASH patients compared with controls and patients with simple steatosis (20.36 vs. 16.11 p < 0.001). The Spearman’s correlation coefficient between the OBT values and the histological fibrosis stages was significant, with values of 0.782 for D0B at 15 min and 0.706 for CUM at 60 min S476

( p < 0.001). DOB at 15 min with a cutoff value of 19 ‰, was the best parameter for identifying the patients with histological proven NASH vs. simple steatosis with an AUROC of 0.921, 95% CI – 0.886–1.000, a sensitivity of 100% and a specificity of 83%, positive predictive value 85%, negative predictive value 100%. Conclusions: Evaluation of liver function by using 13C-Octanoate Breath Test provides a novel non-invasive method for distinguishing NASH patients from patients with simple liver steatosis, as an alternative of liver biopsy. FRI-285 ADIPOSE TISSUE INSULIN RESISTANCE IS ASSOCIATED WITH MACROPHAGE ACTIVATION IN NON-DIABETIC PATIENTS WITH NON-ALCOHOLIC FATTY LIVER DISEASE C. Rosso1, K. Kazankov2, M. Gaggini3, C. Saponaro3, M. Marietti1, H.J. Møller4, G.P. Caviglia1, E. Buzzigoli3, M.L. Abate1, A. Smedile1, G.M. Saracco5, H. Vilstrup2, J. George6, A. Gastaldelli3, H. Grønbæk2, E. Bugianesi1. 1Department of Medical Sciences, University of Turin, Turin, Italy; 2Department of Hepatology and Gastroenterology, Aarhus University Hospital, Aarhus, Denmark; 3Cardiometabolic Risk Unit, Institute of Clinical Physiology, CNR, Pisa, Italy; 4Department of Clinical Biochemistry, Aarhus University Hospital, Aarhus, Denmark; 5 Department of Oncology, University of Turin, Turin; 6Storr Liver Centre, University of Sydney, Westmead, Italy E-mail: [email protected] Background and Aims: Non-alcoholic fatty liver disease (NAFLD) has a bidirectional relationship with insulin resistance (IR): the liver is the target of an increased flux of Free Fatty Acids (FFAs) and adipokines stemming from a dysfunctional adipose tissue (AT) but a fatty liver actively contributes to the dyslipidemic profile and to the chronic low grade inflammation. Soluble CD163 (sCD163), a marker of hepatic macrophages activation, has been associated with fibrosis in NAFLD. Our aim is to elucidate the link between IR in the liver and in the AT, hepatic macrophages activation and liver damage in 40 non-diabetic patients with NAFLD. Methods: All study subjects underwent tracers studies with [2H5] glycerol and [2H2]glucose in fasting conditions. AT-IR was calculated as FFAs × insulin (AT-IR1) and as Glycerol Ra x insulin (AT-IR2). Hepatic-IR was derived from endogenous glucose production × insulin. sCD163 levels were measured by an enzyme-linked immunosorbent assay. Hepatic fat was assessed by liver biopsy while visceral fat (VF) and subcutaneous fat (SF) were measured with standard magnetic resonance imaging (MRI). Histology was scored according to Kleiner. Results: AT-IR showed a significant association with hepatic fat (ATIR1: r = 0.50, p = 0.001; AT-IR2: r = 0.44, p = 0.004), with NAS score ( p = 0.006 and 0.05 respectively) and with fibrosis ( p = 0.001 for both) at liver biopsy. Plasma levels of sCD163 were significantly associated with fasting plasma levels of FFAs and with lipolysis (r = 0.35, p = 0.026; r = 0.35, p = 0.028, respectively). sCD163 levels were also directly related to AT-IR (AT-IR1 r = 0.38, p = 0.016 and AT-IR2 r = 0.31, p = 0.005) and with liver fat (r = 0.53; p = 0.005), while no correlation was found with Hepatic-IR (r = 0.22, p = 0.170), VF (r = 0.15, p = 0.407) or SF (r = 0.08, p = 0.655). Among histological features, sCD163 plasma levels increased in proportion to the NAS score (r = 0.54; p = 0.003) and to the degree of fibrosis ( p < 0.001). At logistic regression analysis, sCD163 plasma levels better predicted moderate/severe (≥F2) fibrosis than AT-IR (OR 5.2, CI:1.1–24.6). Conclusions: We hypothesize that in NAFLD AT-IR can stimulate hepatic macrophage activation via an increased flux of FFAs thus concurring to liver damage. Funded by FP7/2007–2013under grant agreement no.HEALTH-F22009-241762 for the project FLIP;PRIN2009ARYX4 T. Horizon2020 under grant agreement no.634413 for the project EPoS.

Journal of Hepatology 2016 vol. 64 | S425–S630