Trabajos científicos de Latinoamérica en la AASLD 2015

En San Francisco, EEUU, entre el 13 y 17 de noviembre, se realizó  el “Meeting” anual  66º, de la AASLD.

Se presentaron 2267 trabajos, tanto en modalidad oral como poster, de los cuales 36 correspondieron a países latinoamericanos. Brasil presentó 19 trabajos, México 6, Chile 6 y Argentina 5. A continuación les mostramos los abstracts.

 

Use of non-selective beta blockers in cirrhotic patients with ascites is associated with increased risk of acute kidney injury at hospital admission

Category: Portal Hypertension and Other Complications of Cirrhosis
Descriptor: PO2. Ascites, Renal Dysfunction, and Hepatorenal Syndrome
J.R. de Carvalho, R. Luz, H.M. Coelho, C. Villela-Nogueira, R.M. Perez, Hepatology Service, Federal University of Rio de Janeiro, Rio de Janeiro, Rio de Janeiro, BRAZIL

Introduction Use of non-selective beta blockers (NSBB) in patients with advanced cirrhosis has recently become an issue of debate, with concerns about their safety and possible association with complications such as acute kidney injury (AKI). Objective The aim of this study was to evaluate the association between use of NSBB and the presence of AKI in the first 48 hours of hospital admission in cirrhotic patients with ascites. Methods Hospitalizations of cirrhotic patients with ascites in an university hospital were retrospectively reviewed. AKI at hospital admission was defined as a difference of at least 0.3 mg/dL in two serum creatinine values obtained in the first 48 hours of admission. Association between use of NSBB and AKI was evaluated by logistic regression analysis. Results This study included 328 hospitalizations of cirrhotic patients with ascites, with a mean age of 59 (SD: ±12.7) years. Men represented 59% of cases. At admission, 133 patients (41%) were classified as Child B and 195 (59%) as Child C. History of previous gastrointestinal bleeding episode was present in 144 patients (44%) and 200 (61%) were on NSBB. AKI at hospital admission occurred in 196 patients (60%), being present in 66% of patients on use of NSBB and 50% of those not on use of the drug (p=0.004). In Child B patients, AKI occurred in 61% of those on NSBB and 51% of those without the drug (p=0.24). In Child C patients, a significant association between NSBB use and AKI was observed (69% vs 49%; p = 0.006). The variables age, gender, previous episode of gastrointestinal bleeding, previous episode of spontaneous bacterial peritonitis, history of diabetes mellitus, history of arterial hypertension, use of diuretics, use of NSBB, Child-Pugh score at admission and serum sodium at admission were included in a logistic regression model. In the final model, variables independently associated to AKI in the first 48 hours of admission were: age (OR: 1.03; 95% CI: 1.00 – 1.05; p=0.004), serum sodium at admission (OR: 0.95; 95% CI: 0.92 – 0.99; p=0.014) and use of NSBB (OR: 1.91; 95% CI: 1.20 – 3.06; p=0.007). Conclusion In Chid C cirrhotic patients with ascites, use of NSBB is associated with increased risk of AKI at hospital admission. Use of these drugs in this population should be reassessed.

 

Acute kidney injury and short-term mortality after single 5 liter paracentesis without albumin infusion in cirrhotic patients with diuretic-resistant ascites.

Category: Portal Hypertension and Other Complications of Cirrhosis
Descriptor: PO2. Ascites, Renal Dysfunction, and Hepatorenal Syndrome
R. Luz, J.R. de Carvalho, H.M. Coelho, R.M. Perez, C. Villela-Nogueira, Rio de Janeiro Federal University, Niteroi, BRAZIL
Backgroud and Aims: Currently, a single 5 liter paracentesis without albumin infusion is considered as a safe and an adequate short-term option for the management of patients with cirrhosis and diuretic-resistant ascites. The aims of this study was to assess the prevalence of acute kidney injury (AKI) and short-term survival after a single 5 liter paracentesis without albumin infusion in cirrhotic patients with diuretic-resistant ascites.

Methods: This prospective study was held in an university hospital which is a reference in treatment of liver diseases. Cirrhotic patients with diuretic-resistant ascites and who required large volume paracentesis were enrolled. All patients underwent a paracentesis of a maximum of 5 liter without albumin replacement. Serum sodium, serum creatinine and plasmatic renin activity (PRA) were measured at baseline and repeated at day 6 after LVP. Acute kidney injury was defined as a difference of at least 0.3 mg/dL between baseline serum creatinine and serum creatinine at day 6. Patients were followed for 3 months after the procedure.

Results: Thirty-three patients were enrolled, of which 26 (78%) were men, mean age of 59±7 years. A mean of 4.8±0.5L of ascitic fluid were removed in the paracentesis. Seven patients (21%) fulfilled the diagnostic criteria for AKI. Patients who developed AKI had lower values of serum sodium (129 vs 133; p=0.02) and a higher frequency of use of non-selective beta blockers (61% vs 0%; p=0.009). PRA was obtained in 25 patients. PRA increase after paracentesis was higher in patients with AKI when compared to those who did not develop AKI (330% vs. 36.5%, p=0.07). The 90-day mortality was 90% in patients with AKI and 9% in patients without AKI (p=0.02).

Conclusions: A single 5 liter paracentesis without albumin replacement might impact on systemic and renal hemodynamics in a significanty proportion of patients with diuretic-resistant ascites. Hyponatremia and use of non-selective beta blockers are associated with a greater risk of AKI after the procedure and mortality significantly increases after the development of AKI. Albumin infusion might be considered in hiponatremic patients in use of beta-blockers.

 

Four years real-life experience with antiviral adherence in chronic hepatitis B infection in a tertiary university hospital.

Category: Hepatitis B
Descriptor: EO2. Diagnostics, Epidemiology, Prevention, and Natural History
R.M. Abreu, D. Paranaguá-Vezozzo, F.J. Carrilho, S.K. Ono, Gastroenterology, University of São Paulo School of Medicine, São Paulo, São Paulo, BRAZIL; R.M. Abreu, Division of Pharmacy, Clinical Hospital, University of São Paulo School of Medicine, São Paulo, BRAZIL; L.C. Bassit, S. Tao, Y. Jiang, R.F. Schinazi, Pediatrics, Emory University School of Medicine, and Veterans Affairs Medical Center, São Paulo, BRAZIL
Background: Evidence shows that antiviral treatment for chronic hepatitis B (CHB) infection suppress viral load, but does not eradicate hepatitis B virus (HBV). Among the factors directly linked to therapeutic success is adherence to treatment. The purpose of this study was to evaluate the causes of non-response to antiviral treatment in chronic HBV infection.

Methods: A prospective cohort study with CHB treated with adefovir, entecavir, lamivudine and / or tenofovir was performed in a Brazilian reference tertiary hospital. Treatment adherence was evaluated by a validate questionnaire named CEAT-HBV within December 2010 and August 2011. HBV drug resistance mutation and single-dose pharmacokinetics were determined by PCR sequencing and LC-MS respectively. The IRB approved the research study.

Results: Of the 183 individuals enrolled, CEAT-HBV identified 104 (57%) on HBV treatment adherence. Among the 79/183 (43%) individuals with non-adherence to antiviral treatment, 53/79 (67.1%) were more frequently viral load positive. The frequencies of HBV genotypes were A (14%), A1 (34%), A2 (3%), C (18%) and D (14%). However, 38% (70/183) had positive viral loads suggesting non-response to antiviral treatment. In the adherence group (n = 14) with positive viral load, 9 (64%) individuals had HBV with drug resistance mutations. In the positive viral load and non-adherence group (n = 40), 33 (83%) individuals had drug resistance HBV mutations. Sixteen out of 70 HBV positive plasma samples could not be amplified. The main factors involved with non-response to antiviral treatment were drug resistance mutations in HBV (51%), non-adherence without drug resistance mutations in HBV (37%), short treatment duration (8%) and non-determined (4%). The single-dose pharmacokinetics was used to confirm antiviral non-adherence. Two years after the first evaluation, the CEAT-HBV showed that 106/143 (74.1%) of individuals were on treatment adherent. However, 21.2% (40/183) individuals could not be evaluated and excluded. The main reasons for exclusion were death (20/183), 10 out 20 deaths due to hepatocellular carcinoma, loss to follow up (11/183) and others (9/183). It is worth noting that these 10 dead persons were adherent to antiviral treatment and had undetectable HBV DNA.

Conclusion: Non-adherence was high among CHB individuals receiving antiviral treatment. Drug resistance mutations and non-adherence to anti-HBV therapy were the main cause of treatment failure in CHB individuals. This study demonstrates the urgency to implement more effective procedures to improve adherence to antiviral treatment for chronic hepatitis B.

 

High prevalence of PNPLA3 rs738409 (I148M) polymorphism in Chilean latino population and its association to non-alcoholic fatty liver disease risk and histological disease severity.

Category: Steatosis and Steatohepatitis
Descriptor: QO2. Steatohepatitis: Clinical and Therapeutic
M. Arrese, J.P. Arab, A. Riquelme, C.E. Benítez, F. Barrera, A. Soza, J.F. Miquel, C. Hernandez-Rocha, C. Robles, D. Sandoval, Departamento de Gastroenterología , Facultad de Medicina, Pontificia Universidad Católica de Chile , Santiago, CHILE; J. Suazo, Nutrición, Escuela de Medicina, Universidad de Chile, Santiago, CHILE; I. Duarte, Patología, Facultad de Medicina Pontificia Universidad Católica de Chile , Santiago, CHILE; J. Santos, Departamento de Nutrición, Facultad de Medicina Pontificia Universidad Católica de Chile , Santiago, CHILE
Background: A frequent non-synonymous variant of PNPLA3 gene (rs738409) has been associated with higher frequency of nonalcoholic-fatty-liver-disease (NAFLD) and its most severe phenotype, nonalcoholic-steatohepatitis (NASH). The aim of the present study was to assess the association between this PNPLA3 polymorphism and both the risk and the histological severity of NAFLD in Chilean latinos.

Methods: For this purpose two sub-studies were carried out: first sub-study was derived from a large population-based study with or without ultrasound-probed NAFLD. In the second sub-study, patients with biopsy-proven NAFLD for which NASH-activity score (NAS) and fibrosis stage were evaluated. Both groups were genotyped for the PNPLA3 rs738409 variant. Clinical and biochemical variables were evaluated.

Results: In the first sub-study, 166 individuals with NAFLD and 460 individuals without NAFLD were evaluated. Allelic frequencies of high-risk allele G were 0.63 and 0.52, respectively. Both G-allele and GG genotype frequencies were significantly higher in NAFLD cases than controls (p= 0.0003 and p= 0.0004, respectively). In the multivariate analysis, three variables were significantly associated with NAFLD: Body mass index (OR 1.11; p< 0.0001), type 2 diabetes mellitus (OR 2.53; p= 0.02) and GG genotype of PNPLA3 (OR 1.98; p= 0.001). In the second sub-study, 56 patients were included. Patients with the GG genotype showed a significantly higher degree of lobular inflammation (p= 0.027) and NAS (p= 0.015) compared to CC/CG genotype. There were no differences in grade of fibrosis.

 

High prevalence of undiagnosed liver cirrhosis and advanced fibrosis in type 2 diabetic outpatients.

Category: Steatosis and Steatohepatitis
Descriptor: QO2. Steatohepatitis: Clinical and Therapeutic
F. Barrera, J.P. Arab, C.E. Benítez, A. Riquelme, M. Arrese, Department of Gastroenterology, Escuela de Medicina, Pontificia Universidad Catolica de Chile, Santiago, CHILE
Background: Patients with Type 2 Diabetes Mellitus (T2DM) are at risk for developing end-stage liver disease due to nonalcoholic steatohepatitis (NASH). Non-invasive methods have been validated for assessing the severity of nonalcoholic fatty liver disease (NAFLD). Data on prevalence of advanced fibrosis among T2DM patients evaluated by these methods is scarce.

Aim: To evaluate prevalence of steatosis, advanced fibrosis and cirrhosis by non-invasive methods in T2DM patients.

Methods: We conducted a cross-sectional study in 145 consecutive T2DM patients (>55 years-old). The presence of cirrhosis and advanced fibrosis was evaluated by liver morphology assessed by magnetic resonance imaging (MRI) and NAFLD fibrosis score (NFS) respectively. Exclusion criteria included significant alcohol consumption, viral hepatitis or other liver diseases and exposure to hepatotoxic agents.

Results: 52.6% were women, average age was 60 years (57-64), BMI was 29.6±4.7 kg/m2 and diabetes duration was 7.6±6.9 years. A high prevalence of liver steatosis (62.4%) and steatosis and abnormal ALT (37.6%) was found. The prevalence of advanced fibrosis using the NFS was 12.8 and evidence of liver cirrhosis on MRI was 5.3%. In a multivariate analysis GGT >82 IU/L (P=0.004) and no alcohol intake (P=0.032) were independently associated to advanced fibrosis.

Conclusion: A high frequency of undiagnosed advanced fibrosis and cirrhosis was observed in otherwise unselected T2DM patients older than 55 y/o. These patients are at high risk of developing liver-related complications such as portal hypertension and hepatocellular carcinoma. Routine screening for liver disease should be considered in this population (Grant Support: FONDECYT 1150327 to M.A.and 1150311 to F.B) .

 

Sublingual administration of tacrolimus is a feasible route and requires a lower dose to achieve similar drug exposition in adult liver transplant recipients.

Category: Liver Transplantation and Liver Surgery
Descriptor: LO3. Immunosuppression, Outcomes, and Complications
A. Cancino, B.M. Norero, R. Wolff, F. Barrera, A. Soza, M. Arrese, C.E. Benítez, Gastroenterology, Pontificia Universidad Católica de Chile, Santiago, RM, CHILE; S. Solari, Laboratorio Clínico, Pontificia Universidad Católica de Chile, Santiago, CHILE; J. Guerra, N. Jarufe, J.A. Martínez, Digestive Surgery, Pontificia Universidad Católica de Chile, Santiago, CHILE
Introduction: Oral administration of immunosuppressive drugs is not always feasible after liver transplantation and parenteral administration of tacrolimus (TAC) implies a significantly higher expense of resources. The aim of this study is to evaluate the feasibility of sublingual (sl) administration of tacrolimus in adult liver transplant recipients.

Methods: In patients on per oral administration (po) serum TAC levels were determined at different time points (0, 0.5, 1, 2, 4, 6 and 12 hours), then patients were switched to sl administration and TAC dose was tapered (starting with a 20-30% reduction of the po dose) to obtain a similar trough level that in po administration. Then TAC levels were determined at the same time points. The exposition to TAC (AUC) was estimated and compared for SL and po administration.

Results: Seventeen recipients were enrolled. The mean time required to taper de sl dose to obtain a proper trough level was 16.41±10.5 days (range 6-40 days). There were no differences on trough levels on po and sl administrations (7.06±2.2 vs 7.19±2.0 ng/ml respectively, p=ns). Remarkably, when AUC was evaluated, no differences were found in both groups (111.4±32.23 ng/ml/h vs 111.4±35.65 ng/ml/h, p=ns). However, the dose required to achieve a similar trough level was significantly higher when TAC was administered on po vs sl route (4.68±2.32 vs 2.94±1.7 mg/d, p=0.018), meaning a 38% reduction. The maximum concentration after TAC administration (Cmax) was similar in sl and po groups (19.65±9.97 vs 15.88±6.91 ng/ml, p=ns). No severe adverse effects were registered on sl group and was well tolerated. In one patient very few and small blisters were transiently observed for a few days and in 4 patients a spicy taste was transiently observed with the meals on sl group.

Conclusion: Sublingual administration of tacrolimus is a feasible alternative to po administration and can also be monitored employing trough levels requiring a lower dose to achieve the same drug exposition. These findings also suggest that parenteral TAC administration could be replaced by sl route. These findings could imply a significantly reduction on the cost of immunosuppression therapy.

 

A comparative analysis of the Spanish and Latin-American prospective drug-induced liver injury (DILI) networks.

Category: Hepatotoxicity
Descriptor: HO1. Pathogenesis and Mechanisms
F. Bessone, F.C. Tanno, Hospital Provincial del Centenario, Rosario, ARGENTINA; N. Hernández, A. Sanchez Ciceron, M. Di Pace, Hospital de Clínicas, Montevideo, URUGUAY; G. Gualano, Hospital Posadas , Buenos Aires, ARGENTINA; M. Arrese, A. Ruiz, Pontificia Universidad Católica de Chile, Santiago, CHILE; A. Loaeza del Castillo, Instituto Nacional de Ciencias Médicas y Nutrición “Salvador Zubirán”, Hospital General de México, Ciudad de Mexico , MEXICO; M.A. Girala, Hospital de Clínicas, Asunción, PARAGUAY; E. Carrera, Hospital de Especialidades Eugenio Espejo, Quito, ECUADOR; M. Lizarzábal, E. Mengual, Hospital Universitario de Maracaibo, Maracaibo, VENEZUELA, BOLIVARIAN REPUBLIC OF; J. Brahm, J.P. Arancibia, Hospital Clínico Universidad de Chile, Santiago, CHILE; M.B. Davalos-Moscol, Hospital Rebagliati, Facultad de Medicina, Universidad San Martin de Porres, Lima, PERU; R. Paraná, M.I. Schinoni, Hospital Universitario Prof. Edgard Santos, Universidad Federal da Bahía , Salvador de Bahía, BRAZIL; N. Méndez-Sanchéz, Fundación Clínica Médica Sur, Ciudad de Mexico, MEXICO; M.E. Garassini, Hospital Universitario de Caracas, Caracas, VENEZUELA, BOLIVARIAN REPUBLIC OF; R.L. Zapata, Universidad de Chile, Clínica Alemana de Santiago, Santiago, CHILE; I. Medina-Cáliz, A. González-Jiménez, M. Robles Diaz, C. Stephens, A. Ortega, J. Sanabria, M. García-Cortés , B. Garcia-Muñoz, M.I. Lucena, R.J. Andrade, Instituto de Investigación Biomédica de Málaga (IBIMA) , Hospital Universitario Virgen de la Victoria, Universidad de Málaga, CIBERehd , Málaga, SPAIN
Background: DILI characteristics concerning phenotype and involved drugs or other toxic compounds can vary between individuals and possibly between different geographic populations. We aimed to compare all DILI cases included in the ongoing Spanish and Latin-American DILI Network that share the same inclusion criteria and operational procedures. Material and methods: Demographics, clinical parameters and causative agents were compared between 200 Latin-American and 867 Spanish DILI cases.

Results: The mean age of DILI development differed between the two registries with 51 years in LatinAmerica and 54 years in Spain (p=0.02). Females predominated among the LatinAmerican cases (59%) compared to the Spanish cases (49%) (p=0.01). Duration of treatment and time to onset were higher in LatinAmerican cases (127 vs 88 days, p<0.001) and (116 vs 80 days, p=0.03), respectively. Jaundice was similar (67% and 68%) between registries. Although hepatocellular damage was the most frequent type of injury in both registries, the percentage of hepatocellular cases was significantly higher in the Spanish Registry (63% vs 54%, p=0.03) and the mean alkaline phosphatase value at onset was higher in the Latin American cases (2.5 vs 2.1, p<0.001). Severe cases (9% vs 8%) and fatal cases (liver-related death or liver transplantation) (4.6% vs 4%) did not differ. Antiinfectives ranked first in both registries, followed by nervous system and musculo-skeletal drugs in the Spanish network. Musculo-skeletal and sex hormones predominated in the Latin-American cohort. Amoxicillin-clavulanate, diclofenac, nimesulide, and nitrofurantoin were the most common causatives in Latin-America, while in Spain were amoxicillin-clavulanate, antituberculosis treatments, ibuprofen and atorvastatin. Herbal and dietary supplements for bodybuilding DILI were more represented in Latin-America (10% vs 6%, p=0.05).

 

Role of Fcgamma receptors in experimental Non-Alcoholic Steatohepatitis.

Category: Steatosis and Steatohepatitis
Descriptor: QO1. Steatohepatitis: Experimental
D. Cabrera, R. Cruz, P. Rojas de Santiago, M. Arrese, Departamento de Gastroenterología , Facultad de Medicina, Pontificia Universidad Católica de Chile , Santiago, CHILE; E. Jara, N. Muñoz-Durango, A. Kalergis, Departamento de Genética Molecular y Microbiología, Facultad de Ciencias Biológicas, Pontificia Universidad Católica de Chile, Santiago, CHILE; D. Cabrera, Departamento de Ciencias Químico-Biológicas, Universidad Bernardo O Higgins, Santiago, CHILE
Background: The role of innate immunity in Non-alcoholic steatohepatitis (NASH) development is emergent. Different populations of innate immune cells are present in the liver controlling tissue cytokine production through Fc gamma receptors (FcγRs), which are receptors for the Fc region of IgG antibodies. FcγRs cell-type specifically interact with various other receptors for selective amplification or inhibition of particular cytokines. Aim: To evaluate the role of the inhibitory (FcγRIIB) and activatory (FcγRIII) receptors in NASH pathogenesis.

Methods: Wild Type, FcγRIIb(-/-) and FcγRIII(-/-) mice were fed a methionine-choline deficient (MCD) diet for 5 weeks. Liver injury was assessed by measuring serum levels of alanine aminotransferase (ALT) and histologically. Hepatic triglyceride content (HTC) and hepatic mRNA levels of selected pro-inflammatory (TNF-α, IFN-γ, IL-1β, etc.), pro-fibrotic (TGF-β, CTGF, Collagen-1, etc.) and inflammasome (ASC, NLRP3, Caspase-1, etc.) genes were also assessed. Serum pro-inflammatory cytokine levels (TNF-α, IFN-γ, etc.) were determined by cromatographic bead assay (CBA). By flow cytometry was evaluated different populations of inflammatory cells infiltrated in the liver such as dendritic, neutrophils, macrophages and lymphocytes.

Results: FcγRIIb(-/-) MCD-fed mice developed a more robust hepatic inflammatory (decreased hepatic expression of TNFα and other cytokines) and fibrotic response (decreased hepatic expression of collagen I) in comparison with WT MCD-fed mice with no differences in HTC (Figure 1 A and B). No differences were found in liver cell populations of lymphoid and myeloid lineages. The main finding in FcγRIII(-/-) MCD-fed mice was a significant reduction in histological steatosis and HTC. (see figure 1C, below) likely related to reduced interleukin-10 production. Liver lymphoid and myeloid cell populations remained unchanged in these mice.

Conclusion: Our results suggest and important role of the FcγRs in NASH development. While the absence of FcγRIIb seems to promote NASH induction, the absence of FcγRIII strongly reduces liver steatosis. This is the first report that shows a direct role of FcγRs in the pathogenesis of NASH.(Grant support: FONDECYT 1150327 to M.A., PD3140396 to D.A.)

 

Multicentric Study of Liver Transplantation in patients with Hepatocellular Carcinoma in Brazil- Preliminary data of 913 patients.

Category: Hepatobiliary Neoplasia
Descriptor: GO3. Clinical: Hepatocellular Carcinoma and Cholangiocarcinoma
A. Chagas, L. Kikuchi, F.J. Carrilho, Division of Gastroenterology and Hepatology, Department of Gastroenterology, University of Sao Paulo School of Medicine, Sao Paulo, Sao Paulo, BRAZIL; A. Chagas, L. Kikuchi, L.C. D’Albuquerque, F.J. Carrilho, Sao Paulo Clinicas Liver Cancer Group, Sao Paulo, Sao Paulo, BRAZIL; G. Felga, M.D. Almeida, Liver Transplant, Hospital Israelita Albert Einstein, Sao Paulo, Sao Paulo, BRAZIL; A.A. Mattos, Department of Gastroenterology, Fundação Universidade Federal de Ciências da Saúde de Porto Alegre, Porto Alegre, BRAZIL; R.F. Silva, R.M. da Silva, Serviço de Gastroenterologia e Unidade de Transplante de Fígado, Hospital de Base – FUNFARME, Faculdade de Medicina de São José do Rio Preto, São José do Rio Preto, BRAZIL; F. Branco, Department of Gastroenterology, Santa Casa de Misericórdia de Porto Alegre, Porto Alegre, BRAZIL; I.F. Boin, Digestive Surgery Department, State University of Campinas, Campinas, BRAZIL; J.H. Garcia, Department of Surgery and Liver Transplantation, Federal University of Ceará, Ceará, BRAZIL; L.C. D’Albuquerque, Department of Gastroenterology, University of Sao Paulo School of Medicine, Sao Paulo, Sao Paulo, BRAZIL
Introduction: Liver transplantation (LT) is the treatment of choice for patients with cirrhosis and unresectable early hepatocellular carcinoma (HCC). The aim of this multicentric study was to analyze the demographic characteristics, clinical features and outcomes of patients submitted to liver transplantation with HCC in Brazil.

Methods: We conducted a national multicentric retrospective study to analyze the results of liver transplantation in patients with hepatocellular carcinoma (HCC). Medical records of 913 transplanted patients with hepatocellular carcinoma, between 2006 -2014, from 10 transplant centers in Brazil, were retrospectively analyzed. Patient and tumor characteristics, pathologic data and rate of tumor recurrence were collected.

Results: Of the 913 HCC transplanted patients, median age was 57 years and 81% were male. Etiology of cirrhosis was HCV in 60%. At diagnosis, most patients had uninodular HCC (57%) and median tumor burden was 30mm. During the waiting list period, HCC treatment was performed in 63%. Chemoembolization (TACE), ablation therapy (PEI/RF), surgery and combined therapy were used in 80%, 13%, 6% and 1% patients, respectively. In explant analysis, tumor was uninodular in 45% and moderately differentiated in the majority of cases (66%). Median HCC size was 28mm. Vascular invasion and satellite nodules were observed in 24.5% and 25% of patients, respectively. In 818 patients that survived beyond the immediate post-transplant period, mean follow-up was 27,7 months (±23,8), an overall survival was 70% in 5 years. Recurrence occurred in 8/818 (8%) cases, at a mean time of 15 months (1.5-76m). Sites of recurrence were 40% in liver, extrahepatic in 46% and both hepatic and extrahepatic in 14%. Vascular invasion and alpha-fetoprotein (AFP) level before liver transplantation were risk factors for tumor recurrence. The presence of HCC recurrence was directly related to poor survival. Female gender, need for re-transplantation, vascular invasion and explant outside the Milan criteria were also predictors of poor survival.

Conclusion: Liver transplantation for hepatocellular carcinoma in Brazil was associated with an overall survival of 70% in 5 years. HCC recurrence ocurred in 8% of patients. The presence of vascular invasion and AFP before liver transplantation were associated with increased risk of tumor recurrence. Female, vascular invasion, explant outside the Milan criteria and HCC recurrence were related to poor survival.

 

Baveno`s VI non-invasive approach for primary prophylaxis of variceal bleeding: can spleen stiffness help?

Category: Portal Hypertension and Other Complications of Cirrhosis
Descriptor: PO3. Varices and Bleeding
F.F. Fernandes, E.C. Castro Filho, R.A. Guimarães, L.M. Silva, C. Terra, F.F. Figueiredo, R.M. Perez, University of State of Rio de Janeiro, Rio de JAneiro, BRAZIL; R.M. Perez, Federal University of Rio de Janeiro, Rio de Janeiro, BRAZIL; F.F. Fernandes, G. Pereira, J. Pereira, Bonsucesso Federal Hospital, Rio de Janeiro, BRAZIL
Backgrounds: Gastroesophageal varices (GEV) are a turning point in natural course of cirrhosis as they represent significant portal hypertension and indicate primary prophylaxis for digestive hemorrhage. Recently, Baveno VI has proposed a combination of cut-off points of platelets (>150000) and liver stiffness (LS) (<20kPa) to obviate the use of endoscopy for screening GEV in cirrhotic patients. Spleen stiffness (SS) has already been proposed as an attractive complementary approach to LS in the assessment of GEV detection. Aims: to evaluate the performance of cut-off points of platelets and LS proposed by Baveno VI and the potential benefit of adding SS values to this non-invasive approach.

Methodology: Recently diagnosed cirrhotic patients were prospectively included and submitted to liver and spleen transient elastography (TE) (FibroScan® 502, Echosens, Paris, France) as well as endoscopy for GEV screening by experienced operators at the same day. Blood samples for platelets count and liver function evaluation were also obtained. TE results were expressed in kilopascals (kPa) and the median value of 10 acquisitions was considered for analysis. Only exams with a success rate > 60% and an IQR/Median < 30% were considered. SS was US guided.

Results: 99 patients were included: 63% women; age 59 (± 6.6) years; BMI 28 (± 4.8); MELD 9.7 (± 3.4); 80% Child-Pugh A. LS was invalid in 2 patients and SS in 19. Platelets was 136.000 (± 54.000), LS was 24 kPa (± 12) and SS was 53 kPa (± 18). Esophageal varices were detected in 54% of patients, 40% small and 14% large size. Gastric varices were present in 5% of patients, and GEV in 55%. Platelets (120.000 ± 45.000 vs 155.000 ± 57.000), LS (28.4 ± 13.1 vs 18.4 ± 7.6 kPa) and SS (59.5 ± 17.0 vs 43.3 ± 17.9 kPA) were significantly different among patients with or without GEV (p<0.05). In the AUROC analysis the better points for detection of GEV were: 136.000 platelets (AUROC 0.67, 95% CI: 0.57- 0.76), LS 18.6 kPa (AUROC 0.75, 95% CI: 0.65 to 0.83) and SS 55 kPa (AUROC 0.74, CI: 063 to 0.83). Accordingly to Baveno VI, among the 76 patients in whom endoscopy was indicated 48 (63%) really presented GEV and 21 did not. Additionaly, among the 21 patients without endoscopy indication, 15 (71%) actually did not have GEV and only 6 had GEV. All of this 6 patients, which had no prophylaxis evaluated, presented small varices. In two of then the SS was not feasible and in the other four the SS was 21.1, 26.3, 32.5 and 70.6 kPa.

Conclusion: Platelets and LS cut-offs proposed by Baveno VI performed well in evaluating the need for primary prophylaxis of GEV bleeding. SS was not helpful in obviating GEV screening endoscopy.

 

Heterogeneous IL28B/IFNL4 distribution and association of the C/TT/T haplotype with spontaneous clearance and less liver damage in Mexican patients with chronic hepatitis C virus infection.

Category: Hepatitis C
Descriptor: FO2. Diagnostics, Epidemiology, and Natural History
K. Gonzalez-Aldaco, S. Roman, N. Fierro, E. Martinez-Lopez, A. Panduro, University of Guadalajara, Jalisco, MEXICO; K. Gonzalez-Aldaco, S. Roman, N. Fierro, E. Martinez-Lopez, A. Panduro, Molecular Biology in Medicine, Civil Hospital of Guadalajara, Guadalajara, Jalisco, MEXICO; J.R. Pinho, K.G. Oliveira, L. Oyakawa, R.A. Santana, R. Sitnik, Hospital Israelita Albert Einstein, Sao Paulo, BRAZIL
Background & Aims: Recently, genetic polymorphisms of interleukin-28B (IL28B) (rs12979860 C allele and rs8099917 T allele) and interferon-lambda 4 (IFNL4) (ss469415590 TT allele) have been associated with spontaneous clearance (SC) of hepatitis C virus (HCV) infection among distinct populations worldwide. However, data regarding admixed and Amerindian populations from Latin America are lacking. This study aimed to analyze the genetic admixture of the West Mexico population based on the distribution of the IL28B (rs12979860C>T, rs8099917G>T) and IFNL4 (ss469415590∆G>TT) polymorphisms, and the association between the IL28B/IFNL4 haplotypes with SC and liver damage.

Methods: In a cross-sectional study, 711 unrelated individuals from West Mexico, including Amerindians (86 Nahuas/95 Huicholes) and Mestizo populations (32 from Villa Purificación (VP), 172 from Guadalajara, Jalisco and 326 from Tepic, Nayarit) were genotyped. In a case-control study, 234 treatment-naïve HCV-infected Mestizo patients (149 with chronic hepatitis C (CHC) and 85 with SC) were included for the association of haplotypes with SC and liver damage. A Real-Time PCR assay perfomed genotyping, and transitional elastography (FibroScan™) staged liver damage. Genetic relatedness was evaluated by pairwise comparisons (exact tests). Both, genetic distances (Fst) and haplotypes were inferred by using Arlequin software (version 3.1). Linkage disequilibrium (LD) was calculated by GDA software (version 1.0). All subjects signed an informed consent. The study protocol conformed to the Declaration of Helsinki and was approved by the Ethical Committee.

Results: Three different clusters (p<0.05) were found among Amerindians (Nahuas-Huicholes), Mestizos (from Guadalajara/Nayarit), and VP. The frequency of the respective beneficial (C, T, TT) alleles was higher in Mestizos than in Amerindians (p<0.05). LD for rs12979860 and ss469415590 polymorphisms was high (r2= 0.72-1) and moderate among ss469415590 and rs8099917 (r2= 0.45-0.92). A total of five haplotypes were identified in Mestizos, and two in Natives. The C/TT/T haplotype (rs12979860/ss469415590/rs8099917) was associated with SC (OR= 0.46, 95% IC 0.22-0.95, p=0.03) and with less liver damage (OR= 0.32, 95% IC 0.10-0.97, p=0.04) in Mestizos with CHC. The T/∆G/G haplotype predominated in Amerindians and secondly in Mestizos population.

Conclusion: The West Mexico population is genetically heterogenic based on the IL28B/IFNL4 polymorphisms. The potential T/∆G/G high-risk haplotype predominated in Amerindians and the alternative beneficial haplotype in Mestizos. The C/TT/T haplotype was associated with SC and less liver damage in CHC Mestizo patients.

 

Statins prevent liver tumor promoting effect of dioxin-like environmental toxins in different biological models.

Category: Hepatotoxicity
Descriptor: HO2. Drug Metabolism, Toxicity, and Therapeutics
A. Laura, Facultad de medicina UBA, Argentina, ARGENTINA
Hepatocellular carcinoma (HCC) is the third leading cause of cancer death worldwide. Hormonal imbalance plays a key role in the development of neoplasms. There is an open relationship between HCC and environmental toxins. Hexachlorobenzene (HCB) is an environmental pollutant, associated with a broad spectrum of harmful effects on human health as alterations in thyroid metabolism, neurotoxicity, developmental and carcinogenic effects in human and experimental animals. Statins reduce the incidence of various tumors. Their anti-tumor activity has been related to their pro-apoptotic and anti-angiogenic effect and the prevention of metastasis. However, the exact mechanism mediating the in vivo anti-tumor effect of statins has not been yet fully clarified. The objective of this study was to determine: key molecules involved in HCB promotion of liver preneoplastic foci in an initiation-promotion model in rats [diethylnitrosamine (DEN) (100 mg / kg bw) and HCB (100 mg / kg bw)]. 1- We evaluated in rat liver: a) Proliferating cell nuclear antigen levels (PCNA) in focal and non-focal areas, (Western blot); b) thyroid hormones (TH) concentration, c) deiodinase types I (DI) and III (DIII) mRNA levels; c) 3-Hydroxy-3-methylglutaryl-coenzyme A reductase (HMGCoAR) (RIA and RT-PCR), d) Thioredoxin 1 levels (TRX1) and d) serum cholesterol. 2-In Hep-G2 cells, we analyzed the ability of atorvastatin (AT) and simvastatin (SM) to reverse the effect of HCB on key molecules involved in the proliferative effect of HCB. Hep-G2 cells were treated with HCB (5 uM), in the presence and absence of AT (10, 20 and 30 µM) and SM (5, 10, 20 uM). We analyzed protein levels of: a) PCNA, b) pERK1/2, c) cyclin D1 (CD1); b) TRX1; d) TGF-β1 and HMGCoAR mRNA expression (RT-PCR).

Results: In vivo: HCB increased (60% p≤0,001) PCNA positive cells in focal areas (DEN + HCB) vs. DEN. HMGCoAR increased 31% (p≤0,01); T4 increased 38% (p≤0.01) and T3 decreased 37% (p≤0,01). DIII increased 30% (p≤0.01) and DI declined 41% (p≤0.01). TRX1 increased 29% (p≤0.01) in (DEN + HCB) (p≤0,01). Cholesterol increased 28% (p ≤ 0.05). In vitro, the proliferative effect of HCB decreased with AT or SM. HMGCoAR decreased 29% and 38% with AT (20 and 30 µM), and 20% and 31% with SM (10 to 20 µM) respectively. TRX 1 and TGF-β1 protein levels decreased with AT (30 µM) and SM (20 µM, 34%), p ≤ 0.05.

Conclusion: AT and SM reduced HCB-induced cell proliferation in Hep-G2 cells. HMGCoAR, TGF-β1, TRX1 and HT, may be potential target molecules for statins mechanism of action on HCC caused by environmental toxins.

 

A methyl donor, S-adenosylmethionine (SAM) simultaneously decreases HCV expression, enhances glutathione biosynthesis, modulates antioxidant enzyme systems and switches mat2/mat1 turnover in hepatoma cells expressing HCV proteins.

Category: Hepatitis C
Descriptor: FO1. Virology, Pathogenesis, and Immunology
S.A. Lozano-Sepulveda, E. Bautista-Osorio, P. Cordero, L.E. Muñoz, A.G. Rivas-Estilla, Bioquímica y Medicina Molecular, Universidad Autónoma de Nuevo León , Monterrey, Nuevo León, MEXICO
Purpose: Our aim was to elucidate the mechanism(s) by which SAM decreases HCV expression, using a hepatoma cell line expressing HCV non-structural proteins.

Methods: Huh7 HCV-replicon and parental cells were treated with 1mM SAM. Total glutathione level was evaluated at different time points by Ellman´s recycling method (0-24 h). ROS level was determined by dichlorofluorescein method (0-48 h), PDTC treatment was used as an antioxidant agent and hydrogen peroxide as a positive damage agent. Total RNA and protein were extracted (24-72h). cDNA was synthesized and real time-PCR was performed to quantify HCV-RNA, SOD1, SOD2, catalase, thioredoxin 1, MAT1 and MAT2 expression and RPS18 as endogenous gene. Cellular and viral protein expression was evaluated by western blot using antibodies vs. HCV-NS5A, SOD1, SOD2, catalase, thioredoxin-1, MAT1, MAT2, PKR, STAT1 and actin.

Results: SAM treatment decreased HCV-RNA levels 50-70% compared to untreated control (24-72h). Total glutathione levels increased in both cell lines about 50-60% compared to control without SAM (6h post-treatment in replicon cells and 2h post-treatment in parental cells). Transcriptional expression of SOD1, SOD2 and thioredoxin-1 was increased (0.5, 2.5 and 2 fold-times respectively compared to control) at different time points (24-72h). This effect was not observed for catalase. Interestingly, there was no significant change in ROS levels in both cell types upon SAM treatment at all times assessed, contrary to the observed with PDTC exposition where an average of 30% reduction was detected (0.5-24h). In other hand, MAT1 expression was increased (2.5 fold-times at 48h) and MAT2 was decreased upon SAM exposition (24h) compared with untreated cells at both transcriptional and translational level in both cell lines. SAM treatment does not modify STAT1 and PKR expression compared to untreated cells (24-72h), however both protein levels were increased upon IFN-a treatment used as a control.

Conclusions: A possible mechanism by which SAM decreases HCV expression could involve modulating antioxidant enzymes systems, biosynthesis of glutathione and switching MAT2/MAT1 turnover in Hepatitis C virus expressing cells. Proteins PKR and STAT1 were stimulated in the presence of IFN-a but not with SAM, suggesting that HCV down-regulation mediated by SAM is independent of IFN-a pathway. This may have clinical application in terms of understanding the pathophysiology of the disease. This work was supported by CONACYT-BASICA CB-2010-01155082.

Characteristics, indications and outcomes in cirrhotic patients with portal hypertension undergoing endoscopic retrograde cholangiography: a case-control study.
Category: Portal Hypertension and Other Complications of Cirrhosis
Descriptor: PO4. Encephalopathy and Other Complications
R. Macías-Rodríguez, J. Rodriguez-Garcia, A. Ruiz-Margáin, A. Torre, Gastroenterology, INCMNSZ, Mexico, DF, MEXICO
Background: Endoscopic retrograde cholangiography (ERC) is a useful tool for diagnostic and therapeutic biliary tract diseases. In patients with cirrhosis, portal hypertension leads to clinical and biochemical alterations which can exert higher risk for complications related to ERC. However, scarce data regarding ERC in cirrhosis are available in medical literature. Aim: to evaluate the indications, general characteristics and outcomes of ERC in patients with cirrhosis and portal hypertension.

Methods: This was a case-control study including 37 patients (71 procedures) with cirrhosis and portal hypertension (C) and 37 age and sex-matched controls without cirrhosis (NC), undergoing ERC and evaluated 30 days post-ERC. Characteristics related to ERC (cannulation, sphincterotomy, stenting and complication rate) and cirrhosis (Child-Pugh and MELD scores; presence of ascites, hepatic encephalopathy (HE) and esophageal varices) were recorded. Mann-Whitney´s U and X2 were used as appropriate.Logistic regression and ROC analysis with Youden index were used to analyze complications in cirrhotics.

Results: Main etiology of cirrhosis was hepatitis C infection (22%). Patients were classified as Child A/B/C (4/18/15), mean MELD was 17.8±6. Complications of cirrhosis were ascites (46%), esophageal varices (63%; large esophageal varices 43.7%) and HE (16%). Main differences in baseline characteristics between groups showed changes inherent to cirrhosis in biochemical parameters(higher bilirubin and lower PT, platelets, sodium and albumin). Main indication in C and NC group for ERC was choledocholithiasis in 46% and 48%, respectively p=0.816. Cannulation rate was the same in both groups, (97%, p=1.000). Biliary sphincterotomy was performed more frequently in NC patients compared to C (60 vs 35%, p=0.036); there was no difference in complications related to ERC between C and NC (10% vs 8%, p=0.677). Factors related to complications in cirrhosis were lower alkaline phosphatase levels(230± 71 vs 619± 550mg/dL) and sphincterotomy rate(71 vs 20% for complication and no complication subgroup, p=0.010). In the multivariable analysis only sphincterotomy was independently associated to the presence of complications (OR 9.8[1.7-56.3], p=0.011). ROC analysis yielded a MELD score>16 to best predict complications after ERC in cirrhosis (Se 71.4%, Sp 57.8%, AUC 0.616).

Conclusion: Outcomes after ERC in patients with cirrhosis are similar to non-cirrhotics despite of the alteration in coagulation parameters and the presence of disease-specific complications, however a more cautiously approach in patients with cirrhosis undergoing sphincterotomy and MELD>16 is needed.

 

Are the phenotypes of nonalcoholic liver disease in Latin american population determined by the expression of the polymorphism of PNPLA3?

Category: Steatosis and Steatohepatitis
Descriptor: QO2. Steatohepatitis: Clinical and Therapeutic
L.A. Martinez Rodriguez, A. Torre, D. Kershenobich, Gastrioenterology, Instituto nacional de ciencias medicas y nutricion salvador zubiran, Mexico city, MEXICO
Background: Non-alcoholic fatty liver disease (NAFLD) comprehends a wide range of conditions, encompassing from fatty liver or steatohepatitis with or without fibrosis, to cirrhosis and its complications. Patatin-like phospholipase domain-containing 3 (PNPLA3) displays anabolic and catabolic activities in lipid metabolism, and has been reported to be significantly associated with liver fat content. However, detailed demographic and ethnic characteristics of the I148M variant and its role in the wide range of conditions related to NAFLD have not been fully elucidated.

Objective: Investigate in a group of Mexican adult population the effects of PNPLA3 gene over to simple steatosis (SS) non-alcoholicsteatohepatitis (NASH) and fibrosis (FB) in NAFLD patients.

Methods: Peripheral blood DNA from 211 patients diagnosed with NAFLD ) was used to determine the PNPLA3 genotype by polymerase chain reaction (PCR) and direct sequencing. The relationship of SNPs and NAFLD-related markers of liver function, biopsy, imaging, anthropometric and clinical parameters were analyzed

Results: 189 (89.5%) patients were the G risk allele (CC: 23 (11%), GC: 73 (34.75%), GC: 115 (54.75%) GG), the calculated allelic frequency was 77% (allele variant frequencies: NASH: 71.87%; SS, 80.47% FB: 73.1). Analysis revealed that carriers of the GG genotype were at 3.81-fold (95% CI: 3.03 ~ 4.79) higher risk of have NASH and 2.32-fold (95% CI: 1.77 ~ 3.23) higher risk of have fibrosis. Compared with non-carriers (CC), PNPLA3 (GC, GG) was also found to be associated with: lower HDL colesterol levels (p 0.010), SS even being lean patient (p.007), insuline resistance determined by HOMA score (p 0.029), >0.675 NAFLD fibrosis score (p <0.001) and alterated serum levels of alanine aminotransferase (p <0.05).

Conclusion: Polymorphisms in the PNPLA3 gene in latin american is frequent and maybe a key in mediating the wide range of conditions related to NAFLD including the spectrum of SS, NASH and FB. PNPLA3 may play a determinant role in NAFLD pathogenesis.

 

The Antibody response to hepatitis B virus vaccination is not influenced by the hepatitis C virus viral load in patients with chronic hepatitis C

Category: Hepatitis C
Descriptor: FO1. Virology, Pathogenesis, and Immunology
R.P. Medeiros, M. Lopes, D.F. Mazo, C.P. Oliveira, P.M. Zitteli, J.R. Pinho, F.J. Carrilho, M.G. Pessoa, Gastroenterology, University of Sao Paulo School of Medicine, Sao Paulo, SP, BRAZIL
Background: Some immunogenicity studies of anti-HBV vaccine in patients with chronic HCV infection have demonstrated a diminished response ranging from 63.6% to 72.9% on seroconversion rate, compared to 90.9% to 93.9% in healthy controls. One possible explanation is the high HCV viral load in some patients. Aims: To evaluate the impact of HCV viral load on anti-HBV vaccination response in treatment naive chronic HCV patients without cirrhosis.

Methods: 110 chronic HCV adult patients without cirrhosis were randomized to receive anti-HBV vaccination regimen at standard 3 doses (0, 1 and 6 months) of 20ug, or higher dose of 40ug. Response to vaccination was measured by titers of anti-HBs 1 and 6 month after the last dose of anti-HBV vaccine. Healthy controls were negative to anti-HCV, anti-HBc, HBsAg and anti-HBs antibodies, and received standard 3 doses of 20ug at intervals of 0, 1 and 6 months, and were also evaluated for anti-HBs titers.

Results: Of the 110 HCV vaccinated patients, the seroconversion rate (anti-HBs ≥ 10IU/mL) was 74.5% (82/110). Out of the 45 healthy controls vaccinated with standard dose, we observed a seroconversion rate of 93.3% (42/45). Variables included in the logistic regression model were: Age, Gender, HCVRNA and Liver Fibrosis by Metavir (F0/F1 versus F2/F3). Age was the only variable that negatively influenced anti-HBs titers (p=0.003)

Conclusions: Our study had demonstrated that chronic HCV patients without cirrhosis presented impaired anti-HBV vaccine response compared to healthy controls, similar to data previously demonstrated in the literature. This impairment is apparently not influenced by HCV viral load.

 

Molecular Characterization of the Fecal Microbiome in Brazilian Obese NASH patients compared to lean healthy controls.

Category: Steatosis and Steatohepatitis
Descriptor: QO2. Steatohepatitis: Clinical and Therapeutic
C.P. Oliveira, J. Stefano, S.M. Duarte, L. Rodrigues, P.B. Campos, F.G. Costa, D.F. Mazo, F.J. Carrilho, Gastroenterology, University of São Paulo School of Medicine, São Paulo, São Paulo, BRAZIL; R.M. Ribeiro, E.C. Sabino, Institute of Tropical Medicine, University of São Paulo School of Medicine, São Paulo, BRAZIL
Background/Aim: The importance of the intestinal microbiota in the onset and clinical course of many diseases, including Nonalcoholic fatty liver disease (NAFLD) has been evaluated and it is not known if the pathophysiology and clinical-histological phenotype of the disease is similar in different parts of the world. The aim of the study was to compare the gut microbiome from Brazilian obese NASH patients versus Brazilian healthy controls.

Methods: We performed a cross sectional study comprising patients biopsied-proven NASH obese (n=6); and lean healthy controls (HC; n=6). Stool samples were prospectively collected, mixed with RNAlater and stored at -20°C. DNA was extracted from stool samples from 6 patients and 6 controls and PCR amplification was performed using primers for the V4 region of the 16S rRNA gene. The amplicons were sequenced using Ion PGM Torrent platform and data analyzed using QIIME software.

Results: The PCA using wheigted unifrac analysis showed a different distribution of the NASH cases versus control. The proportion of Firmicutes (p<0.01) and Tenericutes (p=0.02) were significantly higher among NASH cases, while Bacteriodetes (p=0.04) were higher among controls (Kruskal-Wallis test). At genus level, only Ruminococcus was significantly higher among NASH cases (p<0.01) (figure 1).

Conclusion: Our data suggest that Brazilian NASH patients have fecal dysbiosis compared with Brazilian lean healthy controls. Further studies are required to investigate the mechanism underlying the interaction between gut microbes and NASH in obese and lean subjects across the world.

 

Omega-3 fatty acids improve proteomic and lipidomic markers of endoplasmic reticulum (ER) stress and mitochondrial dysfunction in a randomized controlled trial in subjects with Nonalcoholic Steatohepatitis.

Category: Steatosis and Steatohepatitis
Descriptor: QO2. Steatohepatitis: Clinical and Therapeutic
L. Rodrigues, C.P. Oliveira, J. Stefano, M.A. Nogueira, F.J. Carrilho, D. Waitzberg, Gastroenterology, University of São Paulo School of Medicine, São Paulo, São Paulo, BRAZIL; I.D. Silva, Gynecology, Universidade Federal de São Paulo, São Paulo, BRAZIL; E.G. Lo Turco, Surgery/Urology, Universidade Federal de São Paulo, São Paulo, BRAZIL; V.F. Alves, Patology, University of São Paulo School of Medicine, São Paulo, BRAZIL; P. Puri, Gastroenterology, Hepatology and Nutrition, Virginia Commonwealth University-VCU, Richmond, Virginia, UNITED STATES
Background/Aims: There is no effective FDA approved treatment for nonalcoholic steatohepatitis (NASH). Increased n-6/n-3 polyunsaturated fatty acid (PUFA) ratio can induce endoplasmic reticulum (ER) stress and mitochondrial dysfunction that characterize NASH. We hypothesized that n-3 PUFA supplementation would improve these abnormalities. We aimed to define the hepatic proteomic and plasma lipidomic profiles following n-3 PUFA therapy in subjects with NASH and relate it to markers of ER stress and mitochondrial dysfunction.

Methods: A 6-month double blind randomized controlled trial in subjects with biopsy proven NASH was conducted with n-3 PUFA (945 mg [α linolenic acid/ 64%, eicosapentaenoic acid (EPA)/16% and docosahexaenoic acid (DHA)/21%]), 3 capsules/day and matched placebo. A 6-month follow up liver biopsy was performed per IRB protocol. Hepatic proteomics from formalin-fixed paraffin embedded liver tissue and plasma lipidomics were performed at baseline and 6-months in the n-3 PUFA group using mass spectrometry. The proteins and lipids were matched to UniProt and LIPID MAPS database respectively. Cytoscape software was used to analyze functional pathways.

Results: Age and gender matched 60 NASH subjects (n=32, n-3 PUFA treatment group; n=28, placebo group) completed the study. A 6-month n-3 PUFA therapy significantly (all p<0.05) and favorably modulated 3 different hepatic pathways: (i) it inhibited PGRMC2 and induced FABPL which regulate lipid metabolism by lowering cellular lipotoxicity potential, (ii) reduced ERE proteasome degradation of proteins (HSPD1, EEF1A2, STUB1, EEF2) and induced chaperones (RS27A, RL40, UBB), (iii) cellular energy homeostasis by inhibiting glycolysis and gluconeogenesis (PPIA, TPI1, ALDOB, GAPDH, PGM1, ENO3, KPYR, PCKGM, LDHC, HNRPU , C1TC, ATP1A4, ATP1A1) and activation of mitochondrial beta-oxidation (CATA, ATPA, CLUD1, CLUD2, FASTKD, CP2A6). Plasma lipidomics analysis showed an increase in n-3 PUFA and reduced arachidonic acid (n-6 PUFA) that would favorably impact n6/n3-PUFA ratio. Also, n-3PUFA supplementation increased Glycerophospholipids subclasses (LPG, GLE) indicating phospholipids membrane homeostasis and regulation of ER.

Conclusion: 1) The Omega-3 PUFA treatment induced proteins associated with the activation of mitochondrial biogenesis and function, and regulation of the endoplasmic reticulum suggesting favorable regulation of cellular energy homeostasis, 2) Increased Glycerophospholipids indicate modulation of lipids biosynthesis by the ER towards reversal of the lipidomic phenotype of NASH.

 

Relevance of the Oxidative Stress in Obese Patients with Nonalcoholic Fatty Liver Disease and Obstructive Sleep Apnea.

Category: Steatosis and Steatohepatitis
Descriptor: QO2. Steatohepatitis: Clinical and Therapeutic
Y.P. Oliveira, H.P. Cotrim, C. Daltro, Programa de Pós Graduação em Medicina e Saúde Medicina e Saúde, Salvador, Bahia, BRAZIL; E. Alves, R.K. Barros, Nucleo de Tratamento e Cirurgia da Obesidade, Salvador, BRAZIL; L.R. Freitas, A.A. Noronha-Dutra, Centro de Pesquisas Gonçalo Moniz, Salvador, BRAZIL; S.R. Figueiredo, A. Vasconcelos, M. Vieira, D. Velame, NASH study group, Salvador, BRAZIL
Background/Aim: The relationship between obstructive sleep apnea (OSA) and nonalcoholic fatty liver disease (NAFLD) severity has been discussed. Some studies have suggested that fatty acids increased inside the mitochondria can lead to generation of Reactive Oxygen Species (ROS) and consequent (oxidative stress) OXS. Nocturnal hypoxemic episodes during OSA have been implicated in the pathogenesis and progression of NAFLD and it may be influenced by insulin resistance (IR) and OXS.The study aimed to evaluate the relevance of the ROS and OXS on NAFLD severity in severe obese patients with OSA.

Methodology: Obese patients (BMI ≥35Kg/m2) who underwent bariatric surgery (BAS) and liver biopsy from September/2013 to April/2014were included.NAFLD/NASH diagnosis was done according NASH Clinical Research Network (Kleiner/2011).The study participants were selected according histological diagnosis in 2 groups: G1- patients with steatosis; G2- patients with NASH. The ROS production in fresh whole blood of the patients and was measured by chemiluminescence in real time using a sensitive photon counterand L-012 (Wako Pure Chemical Industries Osaka, Japan) used as secondary emitter light. The nature of ROS was characterized using specific inhibitors: hydralazine and desferroxamine as inhibitors of peroxynitrite; superoxide dismutase and sodium aside as inhibitors of myeloperoxidase.The OSA diagnosis was done by polysomnography parameters. Patients with an apnea-hypopnea index (AIH) ≥5 events/hour were diagnosis with OSA. For sample analysis, the patients were included in 2 groups according with index (AIH)>15 e ≤15.

Results: The sample included 37severe obese patients. The mean age was 38.1(SD=9.8) and 51.4% were women. Steatosis was observed in 43.2% (16) of them and NASH with and without fibrosis in 57% (21). Comparing the 2 groups, patients with NASH presented higher levels of ROS [Md1: 11621 (IIQ: 954-30033) versus Md2: 2485 (IIQ: 1206-5564) photons, p<0,001], they had higher BMI [Md1: 42.4 (IIQ: 34.0-58.5) versus Md2: 41.0 (IIQ: 37.6-46.8) kg/m2, p< 0,001] and they also had higher apnea index (76.2% versus 18.8%, p< 0,001).

Conclusions: In obese patients the severity of nonalcoholic fatty liver disease was positively associated with reactive oxygen species, obstructive sleep apnea and BMI index.

 

Prevalence of non-alcoholic fatty liver disease (NAFLD) and its subtypes obtained by liver biopsy in patients with type 2 diabetes not selected by ultrasound and/or aminotransferase levels.

Category: Steatosis and Steatohepatitis
Descriptor: QO2. Steatohepatitis: Clinical and Therapeutic
H. Perazzo, National Institute of Infectious Diseases Evandro Chagas, Rio de Janeiro, BRAZIL; L.M. Silva, F.F. Figueiredo, M.Q. Miguez, C. Terra, R.M. Perez, Liver Unit, University of the State of Rio de Janeiro, Rio de Janeiro, BRAZIL; F.F. Figueiredo, Internal Medicine, Federal University of Rio de Janeiro, Rio de Janeiro, BRAZIL; F.F. Campos, Pathology, University of the State of Rio de Janeiro, Rio de Janeiro, BRAZIL; V.P. Lanzoni, Pathology, Federal University of São Paulo, São Paulo, BRAZIL; M.B. Gomes, Diabetology, University of the State of Rio de Janeiro, Rio deJaneiro, BRAZIL; R.M. Perez, D’Or Institute for Research and Education , Rio de Janeiro, BRAZIL
Background: Non-alcoholic fatty liver disease (NAFLD) and its severe form, steatohepatitis (NASH), have been described as highly prevalent in patients with metabolic diseases. The aims were: (i) to estimate the prevalence of NAFLD and its spectrum by liver biopsy in type 2 diabetes patients; (ii) to quantify steatosis and fibrosis; (iii) to identify variables predictive of NASH and fibrosis. Patients and methods: Patients with type 2 diabetes were submitted to percutaneous liver biopsy regardless of alanine aminotransferase (ALT) levels or abdominal ultrasound results. The exclusion criteria were presence of other chronic liver disease or serious systemic diseases; coagulation disturbances; morbid obesity; alcohol abuse; use of hepatotoxic drugs; pregnancy; or refusal to participate. Slides of liver biopsy were evaluated by two independent pathologists using the NASH Activity Score. Factors associated with NASH and significant fibrosis (stage F≥2) were defined by logistic regression analysis.

Results: 85 patients [82% female, mean age=54 yrs, mean BMI=31 Kg/m², 61% with normal ALT, 53% treated by insulin] were included. Median (range) liver specimen length was 30 (25-75) mm. Prevalence of NAFLD in type 2 diabetes was 92% [n=78/85], 54% (n=42) of whom with simple steatosis and 46% (n=36) with NASH. Severe steatosis (>33% of hepatocytes) was more prevalent in patients with NASH compared to those with simple steatosis (65% vs 7%; p<0.001). Patients with simple steatosis had no fibrosis (stage F=0) and significant fibrosis was present in patients 44% (n=16) of patients with NASH (Figure 1). In a multivariate model, BMI [OR=1.16 (95%IC 1.03-1.30)] and ALT [1.09 (1.05-1.13)] were independently associated with NASH and ALT [1.08 (1.05-1.13)] was independently associated with significant fibrosis.

Conclusion: Liver biopsy proven NAFLD and NASH were highly prevalent in patients with type 2 diabetes. Severity of steatosis was associated with NASH. BMI was independently associated with NASH and ALT level was independently associated with NASH and significant fibrosis.

 

Sequential assessment of Acute-on-Chronic Liver Failure and Hyponatremia in cirrhotic patients: correlation of reversibility and evolution with short term survival.

Category: Portal Hypertension and Other Complications of Cirrhosis
Descriptor: PO5. Infections and Acute on Chronic Liver Failure
L.B. Victor, C.B. Souza, C.M. Alcântara, T. Valdeolivas, V.L. Zenatti, Z.D. Veiga, D.M. Mariz, E. Ahmed, F.F. Fernandes, G. Pereira, Hospital Geral de Bonsucesso, Rio de Janeiro, BRAZIL
Acute-on-Chronic Liver Failure (ACLF) is a syndrome characterized by organ failure and high mortality. Hyponatremia conveys poor prognosis in cirrhosis and is not part of CLIF C-OF, a score used for diagnosis of ACLF. Few studies have evaluated the prognostic capability of sequential assessment of ACLF and Hyponatremia.

Objective: evaluate changes in ACLF and hyponatremia and their interaction in predicting survival.

Methods: prospective study with consecutive hospitalized cirrhotic patients. Presence and grading of ACLF and Hyponatremia were evaluated at day 1 (D1) and 7 (D7). Both were classified as persistent, transient, de novo or absent.

Results: 272 patients (56±14 years, 56% male) were included. Ascites and hepatic encephalopathy were present at admission in 68% and 34% of patients. Child and MELD were 9±2 and 17±7. At inclusion, ACLF was diagnosed in 64 patients, and was most commonly classified as grade I (86%). Mean CLIF C-OF was 7.6±1.3 (vs. 5.5±0.8 for patients without ACLF, P<0.001). Throughout the first week, ACLF was reversed in 28 and persisted in 36 patients and de novo ACLF was diagnosed in 9 patients. ACLF at day 7 was more severe, as evidenced by higher prevalence of grade II/III ACLF (37% vs. 14% at D1) and high CLIF C-OF (mean values for persistent, de novo, transient and no ACLF 10±3, 11±3, 7±1 and 6±1 respectively, p<0.001). Hyponatremia was present at inclusion in 42 patients and was classified as persistent in 20 and transient in 22. Serum sodium values for these groups were 123±6 and 126±3. In addition, 23 patients developed de novo hyponatremia. Mean serum sodium changes from D1 to D7 for persistent, transient, de novo, and no hyponatremia was 2±6, 9±6, -7±4 and 0.6±5 mEq/L (p<0.001). Severe hyponatremia (Na+ <125mEq/L) was less frequent at D7 (21% vs. 38%. at D1, p<0.01). 3-month survival was 69%. On multivariate analysis, ACLF at D7 and Hyponatremia at D1 as well as ascites and leucocyte count, were independent predictors of survival. Survival for patients with D7 ACLF was 23% (versus 59% for transient and 82% for no ACLF, p<0.001). Patients with persistent and transient hyponatremia had similar survival (27 and 36% respectively), which was significantly lower than for de novo or no hyponatremia (74 and 76%, p<0.001).

Conclusion: ACLF and Hyponatremia are frequent and associated with poor survival in cirrhosis. ACLF at D7 is strongly correlated with mortality, independently of being persistent or having developed during hospitalization. Hyponatremia retains most of this prognostic capability at hospital admission. Combination of hyponatremia at D1 and ACLF at D7 identify patients at higher risk of short-term mortality.

 

Systemic inflammatory response syndrome (SIRS) in cirrhotic patients with and without bacterial infections: clinical characteristics and correlation with liver and kidney function and prognosis.

Category: Portal Hypertension and Other Complications of Cirrhosis
Descriptor: PO5. Infections and Acute on Chronic Liver Failure
C.B. Souza, L.B. Victor, C.M. Alcântara, T. Valdeolivas, V.L. Zenatti, D.M. Mariz, Z.D. Veiga, E. Ahmed, F.F. Fernandes, G. Pereira, Hospital Geral de Bonsucesso, Rio de Janeiro, BRAZIL
SIRS has been associated with morbidity and mortality in cirrhosis. Although commonly described in infections, it´s also observed in patients without it. Differences in clinical data, incidence of complications and survival between these two groups have not been studied so far.

Objective: Analyze differences in clinical characteristics, diagnostic criteria, liver and kidney function, complications and survival in cirrhotics with SIRS associated or not with infections.

Methods: 256 patients were evaluated. Diagnosis of SIRS was established within 48 hours of admission. Patients were classified as having sepsis (SIRS associated with infections) or SIRS without infection. Additionally we compared these groups with patients with bacterial infections without SIRS and a control group, composed of cirrhotic patients without neither SIRS nor infections.

Results: Prevalence of SIRS was 35%. SIRS was diagnosed at admission in 51 and developed within 48h in 39 patients. The majority of patients fulfilled 2 diagnostic criteria (83%) of which leucocytes and temperature were the most common (69 and 62%). Sepsis was diagnosed in 55 patients and SIRS without infections in the remaining 35. Patients with sepsis more frequently had 3 or more diagnostic criteria for SIRS (22 vs. 9%, p=0.1) and higher leucocyte count (9.6 ±8.1 vs 6.2±4.6, p=0.006). Patients with sepsis had worst liver and kidney function, as evidenced by lower albumin and higher INR and creatinine values. Complications of cirrhosis were equally distributed in both groups, except for higher frequency of gastrointestinal bleeding in SIRS group (29 vs 6%, p=0.003). Acute-on-Chronic Liver Failure (ACLF) was more frequent in sepsis group (46 vs 13%, p=0.001). The probability of survival at 28 and 90 days was 79 and 68%. Patients with Sepsis had lower survival than patients with SIRS both at 28 days (62 vs. 86%) and 90 days (53% vs. 74%), p<0.05. On multivariate analysis, presence of ACLF and ascites were independent predictors of survival. Patients with infections without SIRS had liver and kidney function, frequency of complications, ACLF and mortality comparable to sepsis group. Conversely, control group had the survival probabilities similar to SIRS without infection.

Conclusions: SIRS is frequent and develops in absence of bacterial infections in almost 40% of patients. Bacterial infections discriminates two groups respect to clinical characteristics and prognosis. Sepsis is related to more severe systemic inflammation, worst liver and kidney function and higher frequency of ACLF and mortality. Survival in patients with SIRS is mainly dependent on presence of bacterial infection and development of ACLF.

 

Six minute walking test performance predicts cardiac dysfunction and mortality in liver cirrhosis.

Category: Portal Hypertension and Other Complications of Cirrhosis
Descriptor: PO4. Encephalopathy and Other Complications
C.F. Pimentel, A.A. Feldner, M. Ferraz, M. Kondo, Gastroenterology, Federal University of Sao Paulo, Sao Paulo, BRAZIL; C.F. Pimentel, Liver Transplant, Euryclides de J Zerbini Transplant Hospital, Sao Paulo, BRAZIL; M. Lai, Gastroenterology and Hepatology, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, Massachusetts, UNITED STATES; W. Mathias, Heart Institute , University of Sao Paulo, Sao Paulo, BRAZIL; M.D. Aguiar, G.D. Branco, M. Superbia, Echocardiology, Sao Luiz Hospital, Sao Paulo, BRAZIL
Background: Patients with cirrhosis are at risk of cirrhotic cardiomyopathy, with resulting cardiac dysfunction and exercise limitations. Six minute walking test (6MWT) is a simple exercise test, which assesses functional status and predicts morbidity and mortality in pulmonary and cardiac diseases. We analyzed 6MWT performance in patients with liver cirrhosis and its relationship with cardiac dysfunctions and mortality. Method: A cohort of 103 cirrhotic outpatients with portal hypertension was evaluated with echocardiogram and 6MWT. Cardiac impairment was defined by diastolic dysfunction represented as E/e’>8 and E/A<0.8 or systolic dysfunction as myocardial strain < -18%. The 6MWT was performed indoors, along a flat, straight corridor with distance in meters recorded at the end of 6 minutes (6MWD). Pearson’s correlation, t-tests, ANOVA, Kaplan-Meier curve and logistics regression were performed to evaluate the relationship between abnormal walk distance and cardiac dysfunction and mortality.

Results: This cohort had a mean age of 52 years and 55% male; patients were staged as Child A in 66%, B 21.1% and C 12.3%. Diastolic dysfunction features were demonstrated in 24% of patients and systolic dysfunction in 5%. Moderate or severe intrapulmonary shunting were present in 26.2% of patients. Walk distance inversely correlated with Child scores (p=0.01, r=-0.3), and was significantly reduced with worse Child stages (A 570m, B 504m and C 471m). Distances were statistically shorter in patients with evidence of cardiac dysfunction or liver decompensation (ascites or encephalopathy) (see Table). Both Child scores and 6MWD independently predicted mortality in logistics regression with bootstrap analyses (p=0.001, p=0.01, respectively).There were no differences between 6MWD and intrapulmonary shunts in this study. Survival after 8 month was 100% for patients who achieved predicted 6MWD and 88.7% for those who did not. Among 6 deaths during this period, none of them achieved the predicted 6MWD.

Conclusion: The 6MWT is a very simple, inexpensive, noninvasive test whose result is related to Child scores and cardiac dysfunction and predicts mortality independent of the Child Class. It is a promising prognostic marker in patients with liver cirrhosis.

 

Myocardial strain as a noninvasive measure of circulatory dysfunction and systolic function in liver cirrhosis.

Category: Portal Hypertension and Other Complications of Cirrhosis
Descriptor: PO4. Encephalopathy and Other Complications
C.F. Pimentel, M. Ferraz, A.A. Feldner, M. Kondo, Gastroenterology, Federal University of Sao Paulo, Sao Paulo, BRAZIL; M.D. Aguiar, G.D. Branco, M. Superbia, Echocardiology, Sao Luiz Hospital, Sao Paulo, BRAZIL; W. Mathias, Heart Institute, University of Sao Paulo, Sao Paulo, BRAZIL; C.F. Pimentel, Liver Transplant, Euryclides de J Zerbini Transplant Hospital , Sao Paulo, BRAZIL; M. Lai, Gastroenterology and Hepatology, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, Massachusetts, UNITED STATES
Cirrhosis is related to hyperdynamic circulation with insufficient organ perfusion, activation of RAAS and compensatory increase in cardiac output. Left ventricular longitudinal myocardial strain is a non-invasive echocardiographic marker of myocardial contractility, able to identify subclinical systolic dysfunction (SD). We evaluated cirrhotic patients and measured strain rate, parameters for cardiac dysfunction and serum renin levels and analyzed them by Child Classes.

Method: A cohort of 103 cirrhotic outpatients was studied. SD was defined as strain>-18% or ejection fraction (EF)< 55%. and diastolic dysfunction (DD) findings as E/A<0.8 or E/e’>8.

Results: Non-alcoholic liver cirrhosis was the most common etiology (70%). Child stages were A (66%), B (21%) and C(13%). Alcohol consumption was unrelated to any parameters. SD was more frequently diagnosed by strain rate than EF (5% vs. 0.9%). All patients had normal ventricular measurements and 24% evidence of DD. These patients had strain rates consistent with decreased myocardial contractility, even without anatomical abnormalities. Both strain rate and serum renin levels were significantly correlated with Child scores (p=0.002, r=-0.3; p=0.03, r=0.3, respectively), showing increased myocardial contractility and progressive RAAS activation with increasing Child scores. Mean strain rates and mean serum renin levels were statistically different among Child classes (see figure).

Conclusions: Myocardial strain rate and serum renin level correlated with Child stages consistent with a progressively more hyperdynamic circulatory state with worsening liver decompensation. Strain rate was more sensitive than ejection fraction in diagnosing SD and is a promising tool for early diagnosis of subclinical systolic dysfunction and circulatory dysfunction in cirrhosis.

 

Autonomic cardiac dysfunctions are common in cirrhotic patients and may be associated with an increased mortality.

Category: Portal Hypertension and Other Complications of Cirrhosis
Descriptor: PO4. Encephalopathy and Other Complications
C.F. Pimentel, A.A. Feldner, M. Ferraz, M. Kondo, Gastroenterology, Federal University of Sao Paulo, Sao Paulo, BRAZIL; R. Salvadori, Arrhythmology, Sao Luiz Hospital, Sao Paulo, BRAZIL; M.D. Aguiar, G.D. Branco, M. Superbia, Echocardiology, Sao Luiz Hospital, Sao Paulo, BRAZIL; W. Mathias, Heart Institute, University of Sao Paulo, Sao Paulo, BRAZIL; C.F. Pimentel, Liver Transplant, Euryclides de J Zerbini Transplant Hospital, Sao Paulo, BRAZIL; M. Lai, Gastroenterology and Hepatology, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, Massachusetts, UNITED STATES
Background: Although autonomic cardiac dysfunction has been shown to be associated with liver cirrhosis, the prevalence and prognostic implications are unclear. Whereas prolonged QT interval is a well-known complication of cirrhosis, abnormal heart rate variability (HRV) has not been investigated in cirrhosis. Certain high-risk abnormal HRV parameters, measured by 24-hours Holter monitoring, have been found to predict increased mortality and malignant arrhythmias after acute myocardial infarction (MI) such as: Standard deviation of all normal to normal R-R (NN) intervals (SDNN) <100ms, Standard deviation of 5-minute average NN intervals (SDANN)<55ms and Square root of the mean of the squares of successive NN interval differences (rMSSD)<15. We evaluated the prevalence of these high risk HRV parameters in a cohort of cirrhotic patients.

Method: A cohort of 103 cirrhotic outpatients with portal hypertention was evaluated and followed for 8 months. All patients had laboratory, electrocardiogram and 24-hours Holter monitoring within 2 months of enrollment. In addition, corrected QT interval (QTc) was computed, and considered prolonged if >440ms. T-tests, one-way ANOVA, Pearson’s correlation tests and Kaplan-Meier analysis were performed.

Results: Non-alcoholic liver cirrhosis was the most common etiology (69.8%); Child stages were A (66.0%), B(21.1%) and C(12.3%). Alcohol consumption was unrelated to any of HRV parameters. High risk HRV parameters were prevalent in this cohort, with 64% of the cohort having at least one high-risk parameter. Table 1 shows prevalence of high risk HRV parameters. SDNN and SDANN correlated with Child scores (p<0.0001, r=0.406 and 0.407, respectively), also, means were statistically different between Child stages (p=0.002 for both), tending to decrease as Child increases (110, 93 and 73ms for SDNN; and 97, 81 and 63ms for SDANN). During 8 months follow-up, there were 6 deaths, all in patients with at least one high-risk HRV parameter. This study was not powered to detect differences in mortality.

Conclusion: High-risk abnormal HRV parameters are associated with increased overall mortality in patients post-MI and are prevalent among cirrhotic patients. There is a need for more careful cardiac evaluation of cirrhotic patients and further studies to determine whether abnormal HRV is associated with increased overall mortality in cirrhotic patients.

 

Combination of serum HA, CK18 and TIMP-1 predicts advanced fibrosis in nonalcoholic fatty liver disease.

Category: Steatosis and Steatohepatitis
Descriptor: QO2. Steatohepatitis: Clinical and Therapeutic
C.F. Pimentel, T. Otsubo, A. Francescucci, M. Lai, Gastroenterology and Hepatology, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, Massachusetts, UNITED STATES; C.F. Pimentel, Gastroenterology, Federal University of Sao Paulo, Sao Paulo, BRAZIL; T.L. Challies, I. Nasser, Pathology, Beth Israel Deaconess Medical Center, Boston, Massachusetts, UNITED STATES
Background: Non-alcoholic fatty liver disease (NAFLD) is a prevalent disease (15-40% of U.S. population), with a spectrum ranging from simple steatosis to nonalcoholic steatohepatitis (NASH) with steatosis, inflammation, hepatocyte injury, leading to fibrosis. Patients with NASH can progress to cirrhosis, liver cancer and liver failure. Identifying those with advanced fibrosis at high risk for liver cancer and liver failure is important for clinical management. The current gold standard for identifying these patients is a liver biopsy, which is invasive with known sampling variability. Many noninvasive serum markers of fibrosis in NAFLD have been explored as an alternative to the liver biopsy. While some of these individual serum markers show promise, they do not perform well enough on their own to replace the liver biopsy. We believe that a combination of serum biomarkers would perform better to identify advanced fibrosis in NAFLD patients. Method: We evaluated a combination of serum fibrosis markers in a cohort of 180 NAFLD patients from a single-center NAFLD patient registry, which excluded those with other chronic liver diseases. Clinical, laboratory and serum were collected within 3 months of a percutaneous liver biopsy. Advanced fibrosis was defined as fibrosis stages 3-4, NASH as NAS score>5. Hyaluronic acid (HA), Tissue inhibitor of metalloproteinase 1 (TIMP-1), Cytokeratin-18 (CK18) and YKL40 were evaluated. These biomarkers were combined in a logistics regression model to predict advanced fibrosis; influence of variables was assessed with backward stepwise Wald method. The best fit model was chosen according to statistical significance, odds ratio, overall accuracy and area under ROC curve.

Results: This cohort had a mean age of 50 years (SD 12.6), was 60% male and 86.7% non-Hispanic. 48.9% had NASH, 18.9% advanced fibrosis. Logistic regression analysis of biomarkers indicated that HA, CK18 and TIMP1 were independent predictor of advanced fibrosis (odds ratio [OR] HA=1.057, CK18=1.002, TIMP1=0.99, 95% CI, p<0.001) and a mathematical model was constructed including all three (R2= 0.56, overall accuracy of 88.4%), giving 88.2% sensitivity and 84.1% specificity, and negative and positive predictive values of 96.8% and 60%, respectively. Clinical variables, such as age and gender, did not significant interfere in the model. The Area under ROC curve (AUROC) built with the model predicted values was 0.90, compared to 0.40, 0.73 and 0.82 for TIMP-1, CK18 and HA alone, respectively.

Conclusion: The combination of HA, CK18 and TIMP-1 performs well as a noninvasive biomarker combination to predict advanced fibrosis in NAFLD.

 

Mitochondrial Mutator-Phenotype May Be Related to Pathogenesis of Nonalcoholic Fatty Liver Disease: Insights from Deep Sequencing of Liver Mitochondrial Genomes.

Category: Steatosis and Steatohepatitis
Descriptor: QO2. Steatohepatitis: Clinical and Therapeutic
C.J. Pirola, R. Scian, Molecular Genetics and Biology of Complex Diseases , IDIM-CONICET, Buenos Aires, ARGENTINA; R. Scian, S. Sookoian, Clinical and Molecular Hepatology, IDIM-CONICET, Buenos Aires, ARGENTINA; C.O. Rohr, H. Dopazo, Biomedical Genomics and Evolution Laboratory, CONICET, Buenos Aires, ARGENTINA; G.O. Castaño, Medicine and Surgery Department, Liver Unit, Hospital Abel Zubizarreta, Buenos Aires, ARGENTINA
Background: Mitochondrial (mt) dysfunction is involved in the development of NAFLD; normal activity of mitochondria critically determines fatty acid beta-oxidation, OXPHOS and insulin signaling. In addition, liver mitochondrial biogenesis is reduced in NAFLD. The mt-genome is highly polymorphic and variants in mtDNA affect mt function. A small proportion of mtDNA belongs to the control region involved in mtDNA duplication (D-loop). To understand the clinical implications of mtDNA-variation in the pathogenesis of NAFLD, we sequenced whole liver mtDNA-genomes of 28 individuals, including patients with NAFLD (n = 20) and age and sex-matched controls (n = 8). In addition, we sequenced the entire nuclear POLG and POLG2 genes, which are involved in mtDNA-replication.

Methods: Liver mtDNA was first amplified by long-range PCR; deep next generation sequencing was further performed.

Results: We achieved an average read depth >800 per individual; mtDNA sequencing revealed 689 variants, 525 (76%) of them were observed in NAFLD showing an enrichment of 1.28-fold mutation fraction compared to controls (p = 0.0056). Ten of 16 base positions containing heteroplasmic variants were highly polymorphic (>0.1) and were observed in NAFLD. The mutation fraction of liver mtDNA-genomes in controls compared with that in patients with simple steatosis (NAFL) shows no significant differences. Remarkably, patients with NASH compared with controls harbored a significantly higher number of mutations in mitochondrially encoded ATP synthase 6 (p = 0.033), cytochrome b (MT-CYB) (p = 0.011), and members of the NADH-dehydrogenase complex (p = 4.5 E-5). The comparison of liver mtDNA-diversity among patients with NAFL and NASH showed that the disease severity was associated with increased number of variants in the NADH-complex (p = 7.4 E-3). For variants predicted to be deleterious, an allelic association analysis was further conducted; we found a nonsynonymous variant in MT-CYB (m.15326) and two mutations in D-loop (m.146 and m.16298) that were significantly associated with the disease severity. Overall, mutations located in the NADH-complex were significantly associated with liver-related phenotypes and arterial hypertension. The missense p.Gln1236His variant in POLG was associated with liver mtDNA-copy number (p = 0.01) and the POLG2- rs7223078 was associated with the degree of histological steatosis (p = 0.036).

Conclusion: The burden of mutations in liver mt-genomes may contribute to the pathogenesis of NAFLD and metabolic syndrome-associated comorbidities explaining part of the “missing heritability”. NASH development may be associated with an OXPHOS-negative phenotype.

 

The dual and opposite role of the TM6SF2-rs58542926 Variant in Protecting against Cardiovascular Disease and Conferring Risk for Non-alcoholic fatty liver: A meta-analysis.

Category: Steatosis and Steatohepatitis
Descriptor: QO2. Steatohepatitis: Clinical and Therapeutic
S. Sookoian, Clinical and Molecular Hepatology, IDIM-CONICET, Ciudad Autonoma de Buenos Aires, Buenos Aires, ARGENTINA; C.J. Pirola, Molecular Genetics and Biology of Complex Diseases, IDIM CONICET, Buenos Aires, ARGENTINA
Background: The nonsynonymous p.Glu167Lys (rs58542926 C/T, E167K) variant, located in TM6SF2 (transmembrane 6 superfamily member 2) gene, was associated not only with blood lipid levels, including serum total cholesterol (TC), low-density lipoprotein cholesterol (LDL-C) and triglycerides (TG), but also myocardial infarction risk and NAFLD susceptibility. This variant, which has a relatively low frequency not only in Caucasian population (0.09) but globally (0.07), presents a clinical paradox, as the C (Glu167) allele was shown to be associated with increased CVD risk, while the T allele (Lys167) was associated with NAFLD.

Objective: to examine the evidence provided in the available literature to estimate the strength of the effect of rs58542926 on both circulating lipid traits and NAFLD across different populations. In addition, we assessed whether associations are consistent across studies in magnitude and direction for all explored traits. Methods/design: We performed a systematic review by a meta-analysis; literature searches identified eleven studies.

Results: The rs58542926 exerts a significant role in modulating lipid traits, including TC, LDL-C, TG, and NAFLD. However, this influence on lipids and NAFLD is opposite between genotypes in the dominant model of inheritance. Pooled estimates of random effects in 193,931 individuals showed that, compared with carriers of the minor K allele (EK+KK individuals), subjects homozygous for the ancestral C allele (EE genotype) are at risk of having higher levels of TC, LDL-C and TG; the differences in mean±SE (mg/dL) are 7.4±2.3, 3.7±0.9 and 9.4±2.1, respectively. The rs58542926 was not associated with HDL-C in a large sample (n =91,937). Of note, the EE genotype is associated with a relatively large effect on blood levels of TC (about 2-3% increase on an upper limit of 200 mg/dL), LDL-C (about 1-2% increase on an upper limit of 150 mg/dL), and TG (about 6.4% increase on an upper limit of 150 mg/dL) for a single variant influencing a polygenic trait. In contrast, homozygous EE subjects appear to be protected against NAFLD (OR 0.469, 95% CI 0.300-0.734, p = 0.0009, n = 3273), while carriers of the K allele show about~ 2.2% higher lipid fat content when compared with homozygous EE (n = 3,413), and have 2.13-fold higher risk of developing NAFLD.

Conclusion: The rs58542926 appears to be an important modifier of blood lipid traits in different populations. As a challenge for the personalized medicine, the “major” C-allele, which has a frequency as high as 93%, is associated with CVD risk, while the “minor”-low frequency T allele confers risk for NAFLD; in turn, CVD and NAFLD are strongly related outcomes.

 

Bioelectrical impedance vector analysis (BIVA) is an objective method for nutritional assessment in patients with cirrhosis.

Category: Portal Hypertension and Other Complications of Cirrhosis
Descriptor: PO4. Encephalopathy and Other Complications
A. Ruiz-Margáin, R. Macías-Rodríguez, L.A. Chi-Cervera, S.L. Ríos-Torres, P.A. Rodríguez-Córdova, G. Osalde-Solís, A. Torre, Gastroenterology, Instituto Nacional de Ciencias Médicas y Nutrición “Salvador Zubiran”, Mexico City, MEXICO; A. Espinosa-Cuevas, Nephrology and Mineral Metabolism, Instituto Nacional de Ciencias Médicas y Nutrición “Salvador Zubirán”, Mexico City, MEXICO; A. Duarte-Rojo, Division of Gastroenterology and Hepatology, University of Arkansas for Medical Sciences , Little Rock, Arkansas, UNITED STATES
Background: Malnutrition in cirrhosis is related to poor prognosis. There are few reliable methods for nutritional assessment since most of them are biased by fluid retention and synthetic liver dysfunction, and useful tools such as CT-scan are not easy accessible. Bioelectrical impedance vector analysis (BIVA) is a bedside non-expensive method to evaluate body composition and hydration status, however, data on its usefulness in cirrhosis are limited. The aim of this study was to evaluate the efficacy of BIVA for nutritional assessment in patients with cirrhosis.

Methods: Prospective cohort study. BIVA, mid-arm muscle circumference (MAMC), triceps skinfold thickness (TSF), body mass index with ascites cutoffs (BMIa), and bioelectrical impedance (BIA) were used for nutritional assessment. Malnutrition with BIVA was defined as a vector outside the 75th percentile in the 4th quadrant of the RXc graph. Clinical (ascites and edema) and biochemical evaluations were performed at baseline. T test, Mann Whitney’s , and Xi2 were used as appropriate. Hotelling´s T test was used to compare Rxc graph groups. Kaplan-Meier curves and Cox regression models were used for survival.

Results: 308 patients were followed for 26.6 ± 9.8 months. Child-Pugh stage was A in 36%, B in 46%, and C in 18%, and MELD score was 11.5 ± 4.1. Nutritional parameters showed a MAMC of 23.5 ± 3.9, TSF of 18.8 ± 8.4, and of BMI 27.12 ± 5.24. The frequency of malnutrition was 14.7% for BIA, 25% for BIVA, 32% for MAMC, 13% for TSF, and 14 for % BMIa, In the Rxc graph only patients with clinical ascites displayed fluid retention by BIVA (55.1% vs 44.9% without ascites, p=0.005). Also, higher stages of Child-Pugh were significantly associated with a higher rate of malnutrition and fluid retention (data not shown). Kaplan-Meier curves disclosed a higher mortality in the malnourished group identified by BIVA (39.7% vs. 21.4% in non malnourished, p=<0.004), but malnutrition defined by BIA, MAMC, TSF, or BMIa was not associated to mortality. In univariate regression only BIVA, Child-Pugh and MELD score were associated to mortality. In multivariate analysis malnutrition by BIVA remained associated to mortality when controlled for severity scales (HR=1.68, 1.02-2.76).

Conclusions: BIVA is an objective method for nutritional assessment that can identify both malnutrition and fluid overload in patients with cirrhosis. Notably, only malnutrition defined by BIVA was associated with mortality in this cohort of cirrhotics, and it remained a significant prognostic factor when adjusted by liver disease severity scores. Both the rate of malnutrition and fluid overload increased in the severest stages of cirrhosis.

 

Renal Function Improvement in Early Conversion to Sirolimus with Similar Patient Survival and Metabolic Side Effects of Continued CNI Immunosuppression.

Category: Liver Transplantation and Liver Surgery
Descriptor: LO3. Immunosuppression, Outcomes, and Complications
A. Silvera, P. Cordero, L.E. Muñoz, Liver Unit, Universidad Autonoma de Nuevo Leon, Mexico, Nuevo Leon, MEXICO; C. Sanchez, Department of Nephrology, Universidad Autonoma de Nuevo Leon, Monterrey, MEXICO; M. Escobedo, H.A. Zapata, E. Perez-Rodriguez, A. Guevara, M.A. Hernandez-Guedea, Transplantation Service, Universidad Autonoma de Nuevo Leon, Monterrey, MEXICO; G. Alarcon-Galvan, Department of Pathology, Universidad Autonoma de Nuevo Leon, Monterrey, MEXICO
Aim: The aim of this study was to evaluate the efficacy and safety of sirolimus (SRL) conversion compared with continued calcineurin inhibitor (CNI)-based immunosuppression in patients with orthotopic liver transplant (OLT) on a long-term follow-up. PATIENTS AND

Methods: In this retrospective study 64 patients were analyzed: CNI group (n=31), 6 patients on cyclosporine A (CsA), and 21 patients on tacrolimus (TAC); CNI/SRL group (n=33), initially on CsA or TAC converted to SRL early (≤12 months) or late (>12 months) after OLT.

Results: Patients had a mean follow-up of: CNI 58 ± 48 months and CNI/SRL 68 ± 40 months. CNI/SRL had lower glomerular filtration rate (GFR) by MDRD-6 (109.09 ± 52.64 vs 76.27 ± 52.96 mL/min, p=0.02) at baseline. GFR was significantly higher in CNI at six months (p=0.02) and 12 months (p=0.009). Long-term follow-up showed no significant difference in GFR between groups (p=0.44). Patients converted to SRL due to renal dysfunction (n=27) were subanalyzed according to time to conversion. Patients converted to SRL ≤12 months after OLT had a significant recovery of renal function at last follow-up (38.43 ± 15.08 mL/min vs. 63.12 ± 22.50 mL/min, p=0.05), compared to patients converted at a later time (45.91 ± 14.74 mL/min vs. 49.14 ± 26.49 mL/min, p=0.09). CNI/SRL had higher CHOL, TG and LDL-C at 6 months, however at 12 months only TG remained elevated (p<0.005). Lipid profile was not significantly different on a long-term follow-up, with adequate control of dyslipidemia in both groups. Blood glucose was higher in CNI (p<0.05) but there was no difference in new onset type 2 diabetes mellitus between groups at the end of follow-up. Biopsy proven acute rejection was seen in 2 (6%) patients in the CNI/SRL group and 5 (16%) in the CNI group. A total of 8 (24%) patients died in CNI/SRL group. Main causes of death included hepatocellular carcinoma recurrence (n=3), infections (n=3), cardiovascular events (n=1) and CKD secondary to diabetic nephropathy and recurrence of cirrhosis due to NASH (n=1). In CNI group, 9 (29%) patients died. Causes of death were recurrence of biliary cirrhosis (n=1), secondary sclerosing cholangitis (n=1), CKD secondary to diabetic nephropathy (n=2), chronic graft rejection (n=2) and infections (n=3). Immunosuppression reduction was achieved in 9 (27%) patients receiving SRL without evidence of rejection.

Conclusion: Early conversion to SRL preserves renal function and avoids irreversible kidney damage, with no difference in graft rejection patient survival, and metabolic side effects compared with continued CNIs.

 

Intrahepatic Cholestasis of Pregnancy (ICP) in U.S. Latinas and Chileans: Ancestry Analysis, Admixture Mapping, and Biochemical Features.

Category: Human Cholestatic and Autoimmune Liver Diseases
Descriptor: IO1. PBC/PSC and Other Cholestatic Disease
L. Bull, S. Shah, L. Temple, J. Melgar, M. Geiser, U. Sanford, Medicine, San Francisco General Hospital, UCSF, San Francisco, California, UNITED STATES; L. Bull, D. Hu, S. Shah, L. Temple, S. Huntsman, J. Melgar, M. Geiser, U. Sanford, E. Ziv, Institute for Human Genetics, UCSF, San Francisco, California, UNITED STATES; D. Hu, S. Huntsman, E. Ziv, Medicine, UCSF, San Francisco, California, UNITED STATES; K. Silva, J.P. Kusanovic, Center for Research and Innovation in Maternal-Fetal Medicine, Hospital Dr. Sótero del Río, Puente Alto, CHILE; K. Silva, J. Ortiz, J.P. Kusanovic, Obstetrics and Gynecology, Hospital Dr. Sótero del Río, Puente Alto, CHILE; J. Ortiz, Obstetrics and Gynecology, Clínica Santa María, Santiago, CHILE; R. Lee, Obstetrics and Gynecology, Los Angeles County + University of Southern California Medical Center, Los Angeles, California, UNITED STATES; J. Vargas, Obstetrics, Gynecology and Reproductive Sciences, UCSF, San Francisco, California, UNITED STATES; J. Vargas, Radiology, UCSF, San Francisco, California, UNITED STATES
Purpose: In the Americas, women with Indigenous American ancestry are at increased risk of intrahepatic cholestasis of pregnancy (ICP). We hypothesized that ancestry-related genetic factors contribute to this increased risk, and that the statistically powerful genetic admixture mapping approach could allow mapping of an ICP susceptibility locus. We considered that diagnosis, features, and management of ICP might differ between U.S. and Chilean clinical sites.

Methods: We collected patient data and performed biochemical assays and SNP genotyping on samples from U.S. Latinas and Chilean women, with and without ICP. The study sample included 198 women with ICP (90 from the U.S. and 108 from Chile) and 174 pregnant control women (69 from the U.S. and 105 from Chile). We compared overall genetic ancestry between cases and controls, and performed genome-wide admixture mapping, taking country of enrollment into account, to screen for ICP susceptibility loci. We characterized features of ICP at the U.S. and Chilean clinical sites.

Results: Cases had a greater proportion of Indigenous American ancestry than did controls (p=0.034); when U.S. and Chilean study samples were analyzed separately, this difference was apparent in the U.S., but not the Chilean, study sample, potentially due to differences in population history. Admixture mapping allowed identification of one locus for which Native American ancestry was associated with increased risk of ICP at a genome-wide level of significance (P = 3.1 x 10-5, Pcorrected = 0.035). This locus, on chromosome 2, has an odds ratio of 4.48 (95% CI: 2.21-9.06) for 2 versus zero Indigenous American chromosomes; the 10 Mb 95% confidence interval does not contain any previously identified hereditary ‘cholestasis genes.’ ICP appears biochemically more severe in the U.S., than Chilean, study sample. Differences in management include 1) ursodeoxycholate was prescribed to a greater proportion of U.S. than Chilean patients (48% vs 1%); 2) most U.S. cases were delivered by 37 weeks gestation (or at time of diagnosis if later), while induction of labor is not usually performed until 40 weeks at the Chilean site; and 3) cesarean birth is more common in Chilean cases than controls, while rates are similar between U.S. cases and controls.

Conclusions: Our results indicate that genetic factors contribute to risk of developing ICP in the Americas, and support the utility of clinical and genetic studies of ethnically mixed populations for increasing our understanding of ICP. Greater characterization of diagnosis, management and outcomes of ICP in different countries may improve our understanding of ICP, and its potential subtypes.

 

Improvement in Hepatic Steatosis in Nonalcoholic Steatohepatitis in Response to Synbiotic Supplementation.

Category: Steatosis and Steatohepatitis
Descriptor: QO2. Steatohepatitis: Clinical and Therapeutic
S.M. Ferolla, C.A. Couto, G.N. Armiliato, L.C. Silva, E.C. Lima, Q.C. Lisboa, F.S. Martins, M.A. Ferrari, A.S. Cunha, C.A. Pereira, T.C. Ferrari, Federal University of Minas Gerais, Belo Horizonte, Minas Gerais, BRAZIL
Background: Nonalcoholic fatty liver disease (NAFLD) is one of the most prevalent chronic liver diseases in the world, which can progress to a severe form of nonalcoholic steatohepatitis (NASH), cirrhosis and even hepatocellular carcinoma. Oral supplementation with a synbiotic has been proposed as an effective treatment of NAFLD as these agents influence the gut-liver axis improving inflammatory parameters.

Objective: We aim to evaluate the effects of supplementation with a synbiotic on the grade of hepatic steatosis, intestinal bacterial overgrowth, and serum levels of lipopolysaccharide (LPS) and adiponectin in NASH patients. Design: In a randomized, controlled clinical trial, 50 patients with biopsy-proven NASH were supplemented twice daily for three months with a synbiotic or not receive any supplementation. Both groups were advised to follow an energy-balanced diet and physical activity recommendations. The anthropometric and metabolic parameters, grade of steatosis (evaluated by magnetic resonance [MR] techniques), glucose hydrogen breath test, and levels of LPS and adiponectin were evaluated before and after the treatment.

Results: At the end of the study, the following significant differences were observed in the synbiotic group when compared to the control individuals: decrease in the hepatic proton density fat fraction measured by MR (-3.4%; p=0.027); loss of body weight (-1.3kg; 1.5%; p=0.06); reduction in BMI (-0.4kg/m2; 1.2%; p=0.005) and reduction in the waist circumference measurement (-1.9cm; 1.8%;p=0.001). Food consumption (total daily calorie intake) was not different between groups.

Conclusions: Synbiotic supplementation in addition to nutritional counseling seems superior to nutritional counseling alone for the treatment of NAFLD, as the intervention group presented attenuation of steatosis, and reduction of body weight, BMI and waist circumference. Whether these effects will be sustained with longer treatment durations remains to be determined.

 

Circulating interleukin 6, 10 and 17 as prognostic markers in patients with liver cirrhosis.

Category: Portal Hypertension and Other Complications of Cirrhosis
Descriptor: PO5. Infections and Acute on Chronic Liver Failure
J. Fischer, T.E. Silva, P.E. Soares e Silva, B.S. Colombo, E.B. Dantas-Correa, J.L. Narciso-Schiavon, L.L. Schiavon, Internal Medicine, Division of Gastroenterology, Federal University of Santa Catarina, Florianópolis, Santa Catarina, BRAZIL; L.M. Wildner, M. Bazzo, T.S. Frode, S. Vigil de Melo, J.S. Rosa, Department of Clinical Analysis, Federal University of Santa Catarina, Florianopolis, BRAZIL
Introduction: Activation of inflammatory system is present from the early stages of cirrhosis and it is associated with elevated levels of cytokines. However, data about the prognostic significance of circulating cytokines in liver cirrhosis is still lacking. We sought to investigate the prognostic significance of IL-6, IL-10 and IL-17 in patients with stable cirrhosis and in subjects admitted for acute decompensation (AD) of cirrhosis.

Methods: This prospective study included two cohorts: (1) stable cirrhosis attended in the Outpatient Clinic (n = 118), and (2) subjects hospitalized for AD (n = 130). Thirty healthy subjects served as control group. The acute-on-chronic liver failure (ACLF) criteria were applied according to the EASL-CLIF Consortium.

Results: IL-6 and IL-10 levels were higher in both groups of patients with cirrhosis as compared to control group and also in patients with AD in relation to stable cirrhosis (P < 0.05). In stable cirrhosis, during a median follow-up of 17 months, an event (hospitalization, death or liver transplantation) occurred in 26 patients and was associated with higher IL-6 (3.56 pg/mL vs. 2.13 pg/mL, P = 0.013) and IL-10 (0.54 pg/mL vs. 0.22 pg/mL, P = 0.021), but not IL-17 levels. In the hospitalized cohort, 39 patients died after 90 days of follow-up. Logistic regression analysis showed that death in AD cohort was independently associated with ascites (OR 6.286, 95% CI 1.826 – 21.635; P = 0.004), MELD (OR 1.300, 95% CI 1.175 – 1.439; P < 0.001) and IL-6 (OR 1.002, 95% CI 1.000 – 1.004, P = 0.029). The AUROC of IL-6 to predict 90-day mortality was 0.779 ± 0.046 and the Kaplan–Meier survival probability was 90.0% for IL-6 < 21 pg/mL and 46.7% for IL-6 ≥ 21 pg/mL (P < 0.001). Cytokine levels were evaluated for the prediction of bacterial infection. Regression analysis showed that bacterial infection diagnosed during the first 48 hours after admission was associated with IL-6, CRP and ascites. IL-6 exhibited higher AUROC than CRP for predicting bacterial infection (0.831 ± 0.043 vs. 0.763 ± 0.048, respectively). Higher IL-6 levels were observed in ACLF patients even in the absence of bacterial infection whereas IL-10 was higher only in subjects with infection-related ACLF. Cytokines levels were reassessed at the third day of hospitalization in 74 subjects. No differences between admission and third-day levels were noted for IL-6. Lower IL-10 levels were observed at third day regardless of the presence of ACLF or death during follow-up. However, IL-17 levels dropped significantly only in those who died during follow-up.

Conclusion: Circulating IL-6, IL-10 and IL-17 are of prognostic value in patients with cirrhosis.

 

NFkB inhibition by Andrographolide ameliorates inflammation and fibrogenesis through inflammasome substrate depletion in experimental Non-Alcoholic Steatohepatitis (NASH).

Category: Steatosis and Steatohepatitis
Descriptor: QO1. Steatohepatitis: Experimental
D. Cabrera, N. Solis, M. Pizarro, P. Rojas de Santiago, M. Arrese, Departamento de Gastroenterología, Facultad de Medicina Pontificia Universidad Católica de Chile , Santiago, CHILE; D. Cabrera, Departamento de Ciencias Químico-Biológicas, Universidad Bernardo O Higgins, Santiago, CHILE; A. Wree, D. Povero, A.E. Feldstein, Department of Pediatrics, University of California San Diego, San Diego, California, UNITED STATES; C. Cabello-Verrugio, Departamento de Ciencias Biológicas, Universidad Andres Bello, Santiago, CHILE; H. Moshage, Department of Gastroenterology and Hepatology, University of Groningen, Santiago, CHILE; E. Brandan, Departamenti de Biologia Celular y Molecular, Pontificia Universidad Católica de Chile, Santiago, CHILE; J. Torres, Departamento de Anatomia Patologica, Pontificia Universidad Católica de Chile, Santiago, CHILE
Background: Options to treat NASH are limited. Andrographolide (AP), the bioactive component of Andrographis paniculata, has potent anti-inflammatory activity related to NFκB inhibition. NFkB is a crucial factor in Interleukin-1β (IL-1β) expression, the main substrate for the inflammasome. Aim: To evaluate the effects of AP in experimental NASH and its influence in inflammasome activity.

Methods: C57bl6 mice were fed a choline-deficient-amino-acid–defined (CDAA) diet (22 weeks) with or without AP (1 mg/Kg, 3 times/week, intraperitoneally). Serum levels of alanine aminotransferase (ALT) and hepatic steatosis, inflammation, and fibrosis were assessed. Hepatic triglyceride content (HTC) and hepatic mRNA levels of selected pro-inflammatory and pro-fibrotic genes as well as those related to inflammasome were also measured. Direct AP effect on IL-1β expression and inflammasome activity was evaluated in HepG2 cells loaded with a mixture of FFAs during 24h.

Results: AP decreased HTC and attenuated hepatic inflammation and fibrosis in CDAA-fed mice reducing serum ALT activity and hepatic collagen deposition. AP treatment was associated with a strong reduction in hepatic macrophage infiltration and of hepatic mRNA levels of both pro-inflammatory and pro-fibrotic genes. In addition, mice treated with AP showed reduced expression of inflammasome genes (-60% reduction in inflammasome adaptor ASC hepatic mRNA levels, and -40% reduction in NLRP3 hepatic mRNA levels). Finally, AP inhibited IL-1β expression induced by LPS through NFκB inhibition in fat laden HepG2 cells (figure).

Conclusion: AP administration prevented liver damage in NASH. Inflammasome inactivation by NFκB-dependent mechanism involving IL-1β expression inhibition is likely involved in the therapeutic effects of AP. (Fondecyt 1150327 to M.A. and FONDECYT PD3140396 to D.C.)

 

Hepatitis E infection in immunocompromised persons in Argentina.

Category: Liver Transplantation and Liver Surgery
Descriptor: LO4. Viral Hepatitis
J. Debes, University of MInnesota, Minneapolis, Minnesota, UNITED STATES; M. Martinez Wassaf , Área de Virología y Biología Molecular, LACE Laboratorios, Cordoba, ARGENTINA; M. Pisano, M. Lotto, V. Re, Instituto de Virología “Dr.J.M.Vanella”, Facultad de Ciencias Médicas, Universidad Nacional de Córdoba, Cordoba, ARGENTINA; L. Marianelli, Hospital Rawson, Cordoba, ARGENTINA; D. Balderramo, H. Coseano, Hospital Privado, Cordoba, ARGENTINA
Background: Hepatitis E virus (HEV) is a single-stranded RNA virus that can cause hepatitis in an epidemic fashion. In immunocompetent individuals, infection with HEV usually leads to silent seroconversion or to acute self-limited disease. Several reports suggest an increased rate of seroprevalence of HEV in immunosuppressed individuals, particularly those undergoing solid-organ transplantation. In this setting, HEV can develop into a chronic infection. The data is less clear in patients infected with human immunodeficiency virus (HIV). Moreover, information about prevalence of HEV in immunocopromised subjects outside of Europe or North America is scarce. In this study we addressed the seroprevalence of HEV in immunocompromised subjects in Argentina and associated risk factors.

Methods: We performed third generation enzyme immunoassay for determination of IgG specific antibodies against HEV in 95 subjects infected with HIV, 81 subjects on hemodialysis (HD) and 58 solid-organ transplant recipients. On those samples that were positive for HEV IgG, further assessment of HEV IgM levels. HEV PCR was performed in all samples. Subjects on HD and transplant recipients were evaluated regarding social habits and potential risk factors. Results were compared to 433 HIV-negative, immunocompetent controls from our center.

Results: In our entire HIV-positive group we found 8 of 95 samples to be positive for HEV IgG (8.8%), compared to 19 of 433 samples (4.4%) in the control group. Interestingly, in a subgroup of patients (N27) with low CD4 counts 18% were positive for HEV IgG (p=0.03, compared to controls). This group had a much lower CD4 count when compared to the general HIV cohort (median CD4 count of 43/mm3 vs 543/mm3 respectively). Eight out of 81 subjects (9.8%) on HD and 5 of 58 (8.6%) of transplant recipients were positive for HEV IgG. Interestingly, half of HEV seropositive patients in the HD group had positive IgM for HEV. There was no association between HEV serostatus and consumption of pork, fish, potable water or history of blood transfusion. Only 1 sample showed a positive PCR for HEV, within the HIV group. This patient had recent history of vomiting and diarrhea and likely experienced acute HEV infection.

Conclusions: Our study found an increased seroprevalence of HEV IgG in subjects infected with HIV, on HD and solid-organ transplant recipients in Argentina. However, the only significant difference compared to controls was on HIV-infected patients with low CD4 counts.

 

Serum Lipidomic Profiling for Screening Potential Biomarkers of Liver Cirrhosis among Patients with Chronic Hepatitis C.

Category: Liver Fibrogenesis and Non-Parenchymal Cell Biology
Descriptor: KO2. Imaging and Noninvasive Markers of Liver Disease
A. Passos-Castilho, C. Granato, Division of Infectious Diseases, Federal University of Sao Paulo, Sao Paulo, BRAZIL; M. Ferraz, Department of Gastroenterology, Federal University of Sao Paulo, Sao Paulo, BRAZIL; V.M. Carvalho, K.H. Cardozo, C. Granato, Fleury SA Group, Sao Paulo, BRAZIL; L. Kikuchi, A. Chagas, J.R. Pinho, M.S. Gomes-Gouvêa, F. Malta, F.J. Carrilho, Department of Gastroenterology, University of Sao Paulo School of Medicine, Sao Paulo, BRAZIL
Background: Liver cirrhosis (LC) is the 14th most common cause of death in adults worldwide, resulting in 1.03 million deaths per year. Chronic hepatitis C (CHC) is one of the main causes of LC. Non-invasive markers of fibrosis have been increasingly studied as they may represent a more informative, safe and accessible diagnostic and/or screening tool for at-risk patients. Aim: To explore the serum lipidome profiles of LC to identify potential biomarkers.

Methods: A total of 182 subjects were enrolled from 2011 to 2014. An ultraperformance liquid chromatography–mass spectrometry (UPLC-MS) lipidomic method was used to characterize serum profiles from LC (n=59), CHC (n=65) and healthy subjects (n=58). Patients were diagnosed by clinical, laboratory and imaging evidence of LC while healthy controls had normal liver function and no infectious diseases. Reversed-phased analysis was performed on a Waters ACQUITY IClass UPLC system coupled to a Waters Synapt-MS hidrid quadrupole-time of flight mass spectrometer operating in the positive ion electrospray mode with a mass scan range 200–1200 m/z for data acquisition in continuous mode. All data were processed with Progenesis software (Waters). The resulting multivariate data set was analyzed with MetaboAnalyst (TMIC) using supervised partial least-squares discriminate analysis (PLS-DA). Potential biomarkers were selected according to the variable importance in the projection (VIP) and statistical significance was evaluated using Mann-Whitney test. The receiver operating characteristic (ROC) curve was performed to assess the accuracy of selected biomarkers in LC diagnosis. To identify the potential biomarkers, the HMDB and Lipid Maps databases were queried with the exact mass.

Results: The UPLC–MS-based serum lipidomic profile detected 51 of 59 LC cases, performing with a diagnostic accuracy of 87%. The 3 potential biomarkers differentiated LC from CHC individually with 76 to 88% sensitivity and 72 to 74% specificity. They all presented with fold change higher than 2 and p < 0.0001 in univariate analyses. The combination of the 3 potential biomarkers resulted in an area under the curve of 0.95, 86% sensitivity and 88% specificity. The model was validated with 1000 permutation tests (p < 0.001) to prevent overfitting. 3-Hydroxy-cis-5-tetradecenocylcarnitine, a hydroxy fatty acid involved in many metabolic and proinflammatory pathways, exhibited a significant increase in LC and a decrease in CHC and was proposed as an important indicator of LC.

Conclusions: UPLC-MS lipid profiling proved to be an efficient and convenient tool for diagnosis and screening of LC in an at-risk population.