Janett Fischer

Genome-Wide Association Study Identifies Variants Associated With Autoimmune Hepatitis Type 1

BACKGROUND & AIMS Autoimmune hepatitis (AIH) is an uncommon autoimmune liver disease of unknown etiology. We used a genome-wide approach to identify genetic variants that predispose individuals to AIH. METHODS We performed a genome-wide association study of 649 adults in The Netherlands with AIH type 1 and 13,436 controls. Initial associations were further analyzed in an independent replication panel comprising 451 patients with AIH type 1 in Germany and 4103 controls. We also performed an association analysis in the discovery cohort using imputed genotypes of the major histocompatibility complex region. RESULTS We associated AIH with a variant in the major histocompatibility complex region at rs2187668 (P = 1.5 × 10(-78)). Analysis of this variant in the discovery cohort identified HLA-DRB1*0301 (P = 5.3 × 10(-49)) as a primary susceptibility genotype and HLA-DRB1*0401 (P = 2.8 × 10(-18)) as a secondary susceptibility genotype. We also associated AIH with variants of SH2B3 (rs3184504, 12q24; P = 7.7 × 10(-8)) and CARD10 (rs6000782, 22q13.1; P = 3.0 × 10(-6)). In addition, strong inflation of association signal was found with single-nucleotide polymorphisms associated with other immune-mediated diseases, including primary sclerosing cholangitis and primary biliary cirrhosis, but not with single-nucleotide polymorphisms associated with other genetic traits. CONCLUSIONS In a genome-wide association study, we associated AIH type 1 with variants in the major histocompatibility complex region, and identified variants of SH2B3and CARD10 as likely risk factors. These findings support a complex genetic basis for AIH pathogenesis and indicate that part of the genetic susceptibility overlaps with that for other immune-mediated liver diseases.

Combined effects of different interleukin‐28B gene variants on the outcome of dual combination therapy in chronic hepatitis C virus type 1 infection

In patients with chronic hepatitis C virus (HCV) infection, several variants of the interleukin‐28B (IL28B) gene have been shown to correlate significantly with a sustained virologic response (SVR). Recent evidence shows that determination of one single IL28B polymorphism, rs12979860, is sufficient for predicting treatment outcome. We examined whether the combined determination of the IL28B single‐nucleotide polymorphisms (SNPs), rs12979860, rs8099917, rs12980275, and rs8103142, might improve the prediction of SVR in patients with HCV. In the study cohort, 54% of 942 patients with chronic HCV type 1 infection had SVR. The IL28B SNPs, rs12979860CC and rs8099917TT, correlated significantly with SVR (68% and 62%). The SNPs, rs12980275 and rs8103142, were in strong linkage disequilibrium with rs12979860 and were not included in further analysis. In homozygous carriers of the rs12979860 responder allele C, additional genotyping of the rs8099917 SNP had no effect on response prediction, whereas in carriers of the rs12979860 nonresponder allele, the rs8099917 SNP improved the response prediction. In heterozygous carriers of the rs12979860 nonresponder T allele, SVR rates were 55% in the presence of the rs8099917TT genotype and 40% in patients carrying the rs8099917 TG or GG genotype. Analysis of an independent confirmation cohort of 377 HCV type 1–infected patients verified the significant difference in SVR rates between the combined genotypes, rs12979860CT/rs8099917TT and rs12979860CT/rs8099917TG (38% versus 21%; P = 0.018). Conclusion: Treatment outcome prediction could not be improved in homozygous carriers of the IL28B rs12979860 C responder allele by the additional determination of the rs8099917 SNP. There is evidence that a significant proportion of heterozygous carriers of the rs12979860 T nonresponder allele can profit with respect to SVR prediction by further determination of the rs8099917 SNP. (HEPATOLOGY 2012;55:1700–1710)

Evaluation of liver disease progression in the German hepatitis C virus (1b)‐contaminated anti‐D cohort at 35 years after infection

The natural course of HCV infection remains controversial. The German HCV (1b)‐contaminated anti‐D cohort provides an ideal population to investigate the natural course of HCV infection in a large, homogenous cohort of young women from the date of HCV inoculation. Our previous follow‐up studies at 20 and 25 years after infection suggested slow fibrosis progression rates in this unique cohort. The aim of our prospective, community‐based, multicenter study was to reevaluate the liver disease progression in 718 patients of the original anti‐D cohort at 35 years after infection. Patients with self‐limited HCV infection (n = 189) were compared to those who failed to eliminate the virus spontaneously (n = 529), comprising patients who were treatment naïve (n = 197) or achieved a sustained virological response (SVR; n = 149), respectively, failed to clear the virus (non‐SVR; n = 183) after antiviral therapy. In the overall cohort, 9.3% of patients showed clinical signs of liver cirrhosis at 35 years after infection. Liver disease progression largely depended on HCV infection status. The highest proportion of patients with clinical signs of end‐stage liver disease was observed in the non‐SVR group (15.3%), whereas decreased cirrhosis rates were detected in the SVR group (6%) and in patients with self‐limited HCV infection (1.1%; P = 6.2 × 10−6). Overall survival was significantly enhanced after SVR, compared to treatment‐naïve patients or non‐SVR (P = 0.027). Conclusion: The present study provides further evidence for a mild, but significant, disease progression at 35 years after infection in the German HCV (1b)‐contaminated anti‐D cohort. Patients with self‐limited HCV infection or SVR after antiviral treatment were protected from progressive liver disease and showed the best clinical long‐term outcome. (Hepatology 2014;58:49–57)

Transmembrane 6 superfamily member 2 gene E167K variant impacts on steatosis and liver damage in chronic hepatitis C patients

Steatosis and inherited host factors influence liver damage progression in chronic hepatitis C (CHC). The transmembrane 6 superfamily member 2 (TM6SF2) gene E167K variant increases liver fat and risk of progressive steatohepatitis by interfering with lipoprotein secretion. Our aim was to determine whether the E167K variant affects histological severity of steatosis, necroinflammation, and fibrosis in a cross‐sectional cohort of 815 Italian therapy‐naïve CHC patients. The association with clinically significant fibrosis was replicated in 645 Swiss/German patients. The TM6SF2 E167K variant was genotyped by TaqMan assays, steatosis graded according to the nonalcoholic fatty liver disease activity score, and necroinflammation and fibrosis graded and staged according to Ishak in Italian, and to Metavir in Swiss/German patients. The E167K variant was detected in 69 (9%) Italian patients and was associated with more severe steatosis, independently of confounders (P = 0.038). The association between E167K and steatosis severity was present in patients not infected by genotype 3 (G3) HCV (P = 0.031), but not in those infected by G3 HCV (P = 0.58). Furthermore, the E167K variant was associated with more severe necroinflammation (Ishak grade; adjusted P = 0.037) and nearly associated with more severe fibrosis (Ishak stage; adjusted P = 0.058). At multivariate logistic regression analysis, the E167K variant was independently associated with histologically probable or definite cirrhosis (Ishak stage S6; odds ratio [OR]: 2.19; 95% confidence interval [CI]: 1.18‐3.93; P = 0.010). After further conditioning for steatosis and necroinflammation, the E167K variant remained associated with cirrhosis (OR, 3.15; 95% CI: 1.60‐5.99; P < 0.001). In Swiss/German patients, the E167K variant was independently associated with clinically significant fibrosis Metavir stage F2‐F4 (OR, 1.81; 95% CI: 1.12‐3.02; P = 0.016). Conclusion: TM6SF2 E167K variant impacts on steatosis severity and is associated with liver damage and fibrosis in patients with CHC. (Hepatology 2015;62:111‐117)

Diverse impacts of the rs58542926 E167K variant in TM6SF2 on viral and metabolic liver disease phenotypes

A genome‐wide exome association study has identified the transmembrane 6 superfamily member 2 (TM6SF2) rs58542926 variant encoding an E167K substitution as a genetic determinant of hepatic steatosis in nonalcoholic fatty liver disease (NAFLD). The roles of this variant across a spectrum of liver diseases and pathologies and on serum lipids comparing viral hepatitis to NAFLD and viral load in chronic viral hepatitis, as well as its intrahepatic molecular signature, have not been well characterized. We undertook detailed analyses in 3260 subjects with viral and nonviral liver diseases and in healthy controls. Serum inflammatory markers and hepatic expression of TM6SF2 and genes regulating lipid metabolism were assessed in a subset with chronic hepatitis C (CHC). The rs58542926 T allele was more prevalent in 502 NAFLD patients than controls (P = 0.02) but not different in cohorts with CHC (n = 2023) and chronic hepatitis B (n = 507). The T allele was associated with alterations in serum lipids and hepatic steatosis in all diseases and with reduced hepatic TM6SF2 and microsomal triglyceride transfer protein expression. Interestingly, the substitution was associated with reduced CHC viral load but increased hepatitis B virus DNA. The rs58542926 T allele had no effect on inflammation, impacted ≥F2 fibrosis in CHC and NAFLD assessed cross‐sectionally (odds ratio = 1.39, 95% confidence interval 1.04‐1.87, and odds ratio = 1.62, 95% confidence interval 1.03‐2.52, respectively; P < 0.03 for both), but had no effect on fibrosis progression in 1174 patients with CHC and a known duration of infection. Conclusion: The TM6SF2 E167K substitution promotes steatosis and lipid abnormalities in part by altering TM6SF2 and microsomal triglyceride transfer protein expression and differentially impacts CHC and chronic hepatitis B viral load, while effects on fibrosis are marginal. (Hepatology 2016;64:34–46)

MBOAT7 rs641738 increases risk of liver inflammation and transition to fibrosis in chronic hepatitis C

Cirrhosis likely shares common pathophysiological pathways despite arising from a variety of liver diseases. A recent GWAS identified rs641738, a polymorphism in the MBOAT7 locus, as being associated with the development of alcoholic cirrhosis. Here we explore the role of this variant on liver inflammation and fibrosis in two cohorts of patients with chronic hepatitis C. In 2,051 patients, rs641738 associated with severe hepatic inflammation and increased risk of fibrosis, as well as fast fibrosis progression. At functional level, rs641738 associated with MBOAT7 transcript and protein levels in liver and blood, and with serum inflammatory, oxidative stress and macrophage activation markers. MBOAT7 was expressed in immune cell subsets, implying a role in hepatic inflammation. We conclude that the MBOAT7 rs641738 polymorphism is a novel risk variant for liver inflammation in hepatitis C, and thereby for liver fibrosis.

IFN-λ3, not IFN-λ4, likely mediates IFNL3–IFNL4 haplotype–dependent hepatic inflammation and fibrosis

Genetic variation in the IFNL3–IFNL4 (interferon-λ3–interferon-λ4) region is associated with hepatic inflammation and fibrosis. Whether IFN-λ3 or IFN-λ4 protein drives this association is not known. We demonstrate that hepatic inflammation, fibrosis stage, fibrosis progression rate, hepatic infiltration of immune cells, IFN-λ3 expression, and serum sCD163 levels (a marker of activated macrophages) are greater in individuals with the IFNL3–IFNL4 risk haplotype that does not produce IFN-λ4, but produces IFN-λ3. No difference in these features was observed according to genotype at rs117648444, which encodes a substitution at position 70 of the IFN-λ4 protein and reduces IFN-λ4 activity, or between patients encoding functionally defective IFN-λ4 (IFN-λ4–Ser70) and those encoding fully active IFN-λ4–Pro70. The two proposed functional variants (rs368234815 and rs4803217) were not superior to the discovery SNP rs12979860 with respect to liver inflammation or fibrosis phenotype. IFN-λ3 rather than IFN-λ4 likely mediates IFNL3–IFNL4 haplotype–dependent hepatic inflammation and fibrosis.

Influence of the CXCL1 rs4074 A Allele on Alcohol Induced Cirrhosis and HCC in Patients of European Descent

Background and Aims CXCL1 (CXC chemokine-ligand-1) is a ligand for CXC chemokine receptor 2 expressed on hepatic stellate cells (HSC). Thus, CXCL1 might contribute to HSC activation and fibrogenesis. In the present study, we investigated the influence of the CXCL1 rs4074 polymorphism on the occurrence of alcohol induced liver cirrhosis and hepatocellular carcinoma (HCC). Methods The study involved 458 patients with alcoholic cirrhosis (170 with HCC), 115 alcoholics without liver disease and 342 healthy controls. All subjects were genotyped for the CXCL1 rs4074 polymorphism and CXCL1 serum levels of 132 patients were measured. In vitro CXCL1 secretion in TLR-transfected cell lines were studied by ELISA. Results Distribution of the CXCL1 genotypes (GG/GA/AA) was 159/219/80 in patients with alcoholic cirrhosis, 52/44/19 in alcoholic controls and 158/140/44 in healthy controls. Patients with alcohol-induced cirrhosis were significantly more often carriers of the CXCL1 rs4074 A allele (65.3%) than alcoholics without liver disease (54.8%, OR=1.55; 95%CI=1.025-2.350; p=0.04) and healthy controls (53.8%, OR=1.62; 95%CI=1.212-2.151; p=0.001). Accordingly, the frequency of the CXCL1 rs4074 A allele was significantly higher in the cirrhotic patients than in the subjects without cirrhosis (41.4% vs. 33.9%, OR=1.38, 95% CI:1.14–1.66, p=0.001). Furthermore cirrhotic carriers of the CXCL1 rs4074 A allele had significantly higher CXCL1 serum levels than carriers of the GG genotype. In contrast to sera from healthy controls, sera from patients with alcoholic cirrhosis induced CXCL1 secretion in TLR2- (p=0.016) and TLR4- (p=0.008) transfected HEK293 cells. This finding indicates that sera from patients with alcoholic cirrhosis contain soluble ligands that can induce CXCL1 production via stimulation of TLRs. Conclusion The enhanced CXCL1 serum levels in carriers of the rs4074 A allele together with their increased frequency in patients with alcohol induced cirrhosis suggest the CXCL1 rs4074 A allele as a genetic risk factor for alcoholic cirrhosis.

The membrane-bound O-acyltransferase domain-containing 7 variant rs641738 increases inflammation and fibrosis in chronic hepatitis B

Chronic hepatitis B (CHB) is characterized by hepatic inflammation that promotes progression to cirrhosis and predisposes to the development of hepatocellular carcinoma (HCC). Subtle interindividual genetic variation as well as viral and environmental factors interact to determine disease progression between individuals. Recently, the rs641738 membrane‐bound O‐acyltransferase domain‐containing 7 (MBOAT7) polymorphism was demonstrated to influence histological liver damage in alcoholic liver disease, nonalcoholic fatty liver disease, and hepatitis C, but no data are available for CHB. We evaluated rs641738 influence on disease severity in a cohort of 1,101 patients with CHB. Forty‐two patients underwent gene expression analysis to assess the functional consequences of rs641738 on hepatic MBOAT7 expression. The minor allele (T) of rs641738 was associated with greater inflammation (odds ratio [OR], 1.45; 95% confidence interval [CI], 1.06‐1.95; P = 0.001) and fibrosis (OR = 1.31; 95% CI, 1.19‐1.92; P = 0.01). Risk allele frequency in whites (0.43) was greater than in Chinese (0.24), translating to a larger size effect in the former. The rs641738 (T) allele was associated with lower hepatic MBOAT7 expression (P = 0.008), and the latter was associated with serum liver enzymes and inflammation. Neither patatin‐like phospholipase domain‐containing protein 3 rs738409 nor transmembrane 6 superfamily member 2 rs58542926 polymorphisms influenced disease severity. Conclusion: In patients with CHB, MBOAT7 rs641738 influences hepatic inflammation and fibrosis stage. (Hepatology 2017;65:1840‐1850).

Further evidence for the relevance of TLR2 gene variants in spontaneous bacterial peritonitis

Editor: ad with interest the recent study on the association n Toll-like receptor (TLR) 2 gene polymorphisms and neous bacterial peritonitis (SBP) by Hans Dieter Nischalke lleagues published in the Journal of Hepatology [1]. As a ane-bound pattern recognition receptor that is expressed riety of immune cells, TLR2 can either act as a homodimer heterodimers with TLR1 or TLR6 to recognize an ed spectrum of microbial lipopeptides from Gram-positive am-negative bacteria [2]. Nischalke et al. reported a higher SBP in patients with increasing numbers of tandem GT in intron 2 of the TLR2 gene, which correlated with the ygous TLR2 promoter variant 16934A>T (rs4696480). y, they also observed a trend toward increased risk of presence of the TLR2 Arg753Gln variant (rs5743708) that t reach statistical significance (p = 0.089). Despite its low frequency in Caucasian subjects (3%) [1], the TLR2 Gln polymorphism has recently been found to be involved riety of infectious diseases, including endocarditis [3], uriact infections and recurrent bacterial infections in children reexisting infections in critically ill patients admitted to ensive care unit [6], and infection recurrence in patients ram-positive bacterial infections after liver transplantation