- Email: [email protected]
946 SHOTGUN PROTEOMICS IDENTIFIES SPECIFIC PROTEIN PROFILES IN APOPTOTIC BODIES FROM BILIARY EPITHELIAL CELLS
POSTERS that IFN-g orchestrates hepatocyte damage, whereas the correlation between Th17 immune responses and biochemical indices of cholestasis in AISC indicates that IL-17 is more likely to be involved in the bile duct damage characteristic of this condition. 946 SHOTGUN PROTEOMICS IDENTIFIES SPECIFIC PROTEIN PROFILES IN APOPTOTIC BODIES FROM BILIARY EPITHELIAL CELLS A. Lleo1 , W. Zhang2 , W.H. McDonald3 , D. Friedman3 , E.H. Seeley3 , P.S. Leung2 , P. Invernizzi1,2 , M.E. Gershwin2 . 1 Center for Autoimmune Liver Diseases, IRCCS Istituto Clinico Humanitas, Rozzano, Italy; 2 Division of Rheumatology, Allergy, and Clinical Immunology, University of California Davis, Davis, CA, 3 Mass Spectrometry Research Center and Department of Biochemistry, Vanderbilt University School of Medicine, Nashville, TN, USA E-mail: [email protected] Background and Aims: Primary biliary cirrhosis (PBC) is characterized by an autoimmune reaction against mitochondrial antigens and the destruction on small biliary epithelial cells (BEC). A major unanswered question regarding the pathogenesis of PBC is the specific targeting of the small biliary duct epithelial cell; even thou all nucleated cells have mitochondria, only small BECs are the targets of the autoimmune attack in PBC. We have reported that after apoptosis, human intrahepatic BECs translocate the E2 subunit of the pyruvate dehydrogenase complex (PDC-E2) immunologically intact into apoptotic bodies, forming an apotope, that leads to an innate immune activation. The aim of the study was to identify unique proteins that could induce the signature inflammatory cytokine response from PBC. Methods: We isolate apoptotic bodies from a primary culture of human intrahepatic biliary epithelial cells (HiBEC) (n = 4), and from two control primary epithelial cell types: human renal proximal tubular epithelial cells (n = 6) and human bronchial epithelial cells (n = 6). We then used shotgun proteomics to define the proteome of the apoptotic bodies. The data comparing the hepatic blebs to each of the control groups were individually plugged into a graphical interface software package that reads standard output from protein assemblies and analyzed with a aquasi-likelihood Generalized Linear Modeling. All reported proteins were identified with a minimum of 2 distinct peptides. The results were then sorted by a metric (quasi-P) that takes into account both the difference between the two groups and how consistent the values are within that group. Only p < 0.05 were considered significant. Results: Overall analysis identified a total of 40,843 distinct peptides and 6,160 protein groups; 13 proteins were identified to be specifically located within blebs from HiBEC. The pathway analysis of the identified proteins led in NF-kB activation pathway, ERK pathway, Notch signaling pathway, and IL8 and CXCR2-mediated signaling events. In conclusion, this study determined the proteome of apoptotic bodies of HiBEC and identified 13 specific proteins with a potential pathogenic role in PBC that suggest that they are more than an innocent victim in the pathogenesis of PBC. 947 POSSESSION OF HLA DR3 PREDICTS GOOD RESPONSE TO IMMUNOSUPPRESSIVE TREATMENT IN CHILDREN WITH AUTOIMMUNE LIVER DISEASE J.A. Underhill, S. Bansal, R.J. Thompson, D. Vergani, G. Mieli-Vergani, Y. Ma. Institute of Liver Studies, King’s College London School of Medicine at King’s College Hospital, London, UK E-mail: [email protected] Background: Childhood autoimmune liver disease (AILD) comprises autoimmune hepatitis type 1 (AIH-1), - type 2 (AIH-2) and autoimmune sclerosing cholangitis (ASC). It has been shown previously that HLA class II genes DRB1*03 (DR3), -*07 (DR7) and S390
-*13 (DR13) are the predisposition genes for AIH-1, AIH-2 and ASC respectively. Aim: To define whether in AILD there is an association between HLA predisposition genes and response to immunosuppressive treatment (steroids plus azathioprine). Patients and Methods: A total of 216 children with AILD were followed up for a median of 9.2 years (range 1 month to 35 years until August 2012). Patients were defined as: 1. responders (rapid and sustained remission); 2. non-responders (no remission leading to liver transplantation/death, or frequent relapsers after remission). HLA DRB antigens were defined by PCR/sequence specific primers (SSP) using kits obtained from Biotest (Dreieich, Germany). Results: Overall, 75/87 (86%) DR3 positive patients were responders, compared to 25/49 (51%, p < 0.0001) DR7 positive and 26/38 (68%, p < 0.05) DR13 positive patients. Amongst 91 patients with AIH-1, the proportion of responders (36/42, 86%) was higher within DR3 positive than DR7 positive patients (12/19, 63%, p = 0.046), but similar to that within DR13 positive patients (3/4, 75%, p = NS). Among 49 patients with AIH-2, the proportion of responders tended to be higher within DR3 positive (14/19, 74%) than DR7 positive patients (11/22, 50%, p = 0.12), but was similar to that within DR13 positive patients (3/4, 75%, p = NS). Among 76 ASC patients, the proportion of responders was higher (25/26, 96%) within DR3 positive than DR7 positive (4/10, 40%, p < 0.0002) and DR13 positive patients (11/19, 58%, p < 0.005). Conclusion: Though possession of at least one HLA DR3 allele is a risk factor for the development of autoimmune liver disease, it also predicts good response to immunosuppression in all three AILD subtypes. In contrast, possession of DR7 in all three subtypes and DR13 in ASC predicts a poor response to immunosuppressive treatment. 948 INCREASING INCIDENCE OF AUTOIMMUNE HEPATITIS M. Corouge, V. Mallet, A. Vallet-Pichard, J.-B. Trabut, S. Tripon, H. Fontaine, P. Sogni, S. Pol. Hepatology, Cochin Hospital, Paris, France E-mail: [email protected] Introduction: Autoimmune hepatitis is a rare disease, with an estimated annual incidence of nearly 1.9/100,000 inhabitants and a prevalence of 16.9/100,000. Over the past thirty years, an increase in the incidence of autoimmune diseases (Crohn’s disease, type 1 diabetes, multiple sclerosis) in industrialized countries has been reported. The aim of this study was to confirm our clinical feeling of a parallel increase in the incidence of autoimmune hepatitis. Patients and Methods: A 12 years (2001–2012)-retrospective study has been conducted in our Hepatology department: all patients diagnosed in the department with autoimmune hepatitis, according to the revised International Autoimmune Hepatitis Group, were included. Results: Between 2001 and 2012, 139 new cases of autoimmune hepatitis have been diagnosed, with a constant increase: 2001– 2002: 14, 2003–2004: 17, 2005–2006: 23, 2007–2008: 21, 2009– 2010: 33 et 2011-T3.2012: 31 (Figure 1). Over these 139 new cases, 101 (72.7%) were women (with a stable sex ratio over time), with a median age at diagnosis of 50 years (51 for women and 47.5 for men) and a trend of an older age at diagnosis, particularly during 2011–2012 with a median age at diagnosis of 59 years. The percentage of cirrhosis was 25% and the rate of overlap syndrome was 20.1%. Combined therapy with azathioprine and regressive dose of corticosteroids avoided liver transplantation and controlled hepatitis activity in 95.2% of patients. Most of cirrhosis had biopsy-proven cirrhosis reversal. Another autoimmune disease was observed in 39.8% patients. A comparison of this cohort with the 86 cases of autoimmune hepatitis which were diagnosed before 2001 will allow to determine whether or not clinical
Journal of Hepatology 2013 vol. 58 | S229–S407
-*13 (DR13) are the predisposition genes for AIH-1, AIH-2 and ASC respectively. Aim: To define whether in AILD there is an association between HLA predisposition genes and response to immunosuppressive treatment (steroids plus azathioprine). Patients and Methods: A total of 216 children with AILD were followed up for a median of 9.2 years (range 1 month to 35 years until August 2012). Patients were defined as: 1. responders (rapid and sustained remission); 2. non-responders (no remission leading to liver transplantation/death, or frequent relapsers after remission). HLA DRB antigens were defined by PCR/sequence specific primers (SSP) using kits obtained from Biotest (Dreieich, Germany). Results: Overall, 75/87 (86%) DR3 positive patients were responders, compared to 25/49 (51%, p < 0.0001) DR7 positive and 26/38 (68%, p < 0.05) DR13 positive patients. Amongst 91 patients with AIH-1, the proportion of responders (36/42, 86%) was higher within DR3 positive than DR7 positive patients (12/19, 63%, p = 0.046), but similar to that within DR13 positive patients (3/4, 75%, p = NS). Among 49 patients with AIH-2, the proportion of responders tended to be higher within DR3 positive (14/19, 74%) than DR7 positive patients (11/22, 50%, p = 0.12), but was similar to that within DR13 positive patients (3/4, 75%, p = NS). Among 76 ASC patients, the proportion of responders was higher (25/26, 96%) within DR3 positive than DR7 positive (4/10, 40%, p < 0.0002) and DR13 positive patients (11/19, 58%, p < 0.005). Conclusion: Though possession of at least one HLA DR3 allele is a risk factor for the development of autoimmune liver disease, it also predicts good response to immunosuppression in all three AILD subtypes. In contrast, possession of DR7 in all three subtypes and DR13 in ASC predicts a poor response to immunosuppressive treatment. 948 INCREASING INCIDENCE OF AUTOIMMUNE HEPATITIS M. Corouge, V. Mallet, A. Vallet-Pichard, J.-B. Trabut, S. Tripon, H. Fontaine, P. Sogni, S. Pol. Hepatology, Cochin Hospital, Paris, France E-mail: [email protected] Introduction: Autoimmune hepatitis is a rare disease, with an estimated annual incidence of nearly 1.9/100,000 inhabitants and a prevalence of 16.9/100,000. Over the past thirty years, an increase in the incidence of autoimmune diseases (Crohn’s disease, type 1 diabetes, multiple sclerosis) in industrialized countries has been reported. The aim of this study was to confirm our clinical feeling of a parallel increase in the incidence of autoimmune hepatitis. Patients and Methods: A 12 years (2001–2012)-retrospective study has been conducted in our Hepatology department: all patients diagnosed in the department with autoimmune hepatitis, according to the revised International Autoimmune Hepatitis Group, were included. Results: Between 2001 and 2012, 139 new cases of autoimmune hepatitis have been diagnosed, with a constant increase: 2001– 2002: 14, 2003–2004: 17, 2005–2006: 23, 2007–2008: 21, 2009– 2010: 33 et 2011-T3.2012: 31 (Figure 1). Over these 139 new cases, 101 (72.7%) were women (with a stable sex ratio over time), with a median age at diagnosis of 50 years (51 for women and 47.5 for men) and a trend of an older age at diagnosis, particularly during 2011–2012 with a median age at diagnosis of 59 years. The percentage of cirrhosis was 25% and the rate of overlap syndrome was 20.1%. Combined therapy with azathioprine and regressive dose of corticosteroids avoided liver transplantation and controlled hepatitis activity in 95.2% of patients. Most of cirrhosis had biopsy-proven cirrhosis reversal. Another autoimmune disease was observed in 39.8% patients. A comparison of this cohort with the 86 cases of autoimmune hepatitis which were diagnosed before 2001 will allow to determine whether or not clinical
Journal of Hepatology 2013 vol. 58 | S229–S407