Christian Datz

Adenoma-linked barrier defects and microbial products drive IL-23/IL-17-mediated tumour growth.

Approximately 2% of colorectal cancer is linked to pre-existing inflammation known as colitis-associated cancer, but most develops in patients without underlying inflammatory bowel disease. Colorectal cancer often follows a genetic pathway whereby loss of the adenomatous polyposis coli (APC) tumour suppressor and activation of β-catenin are followed by mutations in K-Ras, PIK3CA and TP53, as the tumour emerges and progresses. Curiously, however, ‘inflammatory signature’ genes characteristic of colitis-associated cancer are also upregulated in colorectal cancer. Further, like most solid tumours, colorectal cancer exhibits immune/inflammatory infiltrates, referred to as ‘tumour-elicited inflammation’. Although infiltrating CD4+ TH1 cells and CD8+ cytotoxic T cells constitute a positive prognostic sign in colorectal cancer, myeloid cells and T-helper interleukin (IL)-17-producing (TH17) cells promote tumorigenesis, and a ‘TH17 expression signature’ in stage I/II colorectal cancer is associated with a drastic decrease in disease-free survival. Despite its pathogenic importance, the mechanisms responsible for the appearance of tumour-elicited inflammation are poorly understood. Many epithelial cancers develop proximally to microbial communities, which are physically separated from immune cells by an epithelial barrier. We investigated mechanisms responsible for tumour-elicited inflammation in a mouse model of colorectal tumorigenesis, which, like human colorectal cancer, exhibits upregulation of IL-23 and IL-17. Here we show that IL-23 signalling promotes tumour growth and progression, and development of a tumoural IL-17 response. IL-23 is mainly produced by tumour-associated myeloid cells that are likely to be activated by microbial products, which penetrate the tumours but not adjacent tissue. Both early and late colorectal neoplasms exhibit defective expression of several barrier proteins. We propose that barrier deterioration induced by colorectal-cancer-initiating genetic lesions results in adenoma invasion by microbial products that trigger tumour-elicited inflammation, which in turn drives tumour growth.

Regulation of iron homeostasis in anemia of chronic disease and iron deficiency anemia: diagnostic and therapeutic implications.

The anemia of chronic disease (ACD) is characterized by macrophage iron retention induced by cytokines and the master regulator hepcidin. Hepcidin controls cellular iron efflux on binding to the iron export protein ferroportin. Many patients, however, present with both ACD and iron deficiency anemia (ACD/IDA), the latter resulting from chronic blood loss. We used a rat model of ACD resulting from chronic arthritis and mimicked ACD/IDA by additional phlebotomy to define differing iron-regulatory pathways. Iron retention during inflammation occurs in macrophages and the spleen, but not in the liver. In rats and humans with ACD, serum hepcidin concentrations are elevated, which is paralleled by reduced duodenal and macrophage expression of ferroportin. Individuals with ACD/IDA have significantly lower hepcidin levels than ACD subjects, and ACD/IDA persons, in contrast to ACD subjects, were able to absorb dietary iron from the gut and to mobilize iron from macrophages. Circulating hepcidin levels affect iron traffic in ACD and ACD/IDA and are more responsive to the erythropoietic demands for iron than to inflammation. Hepcidin determination may aid to differentiate between ACD and ACD/IDA and in selecting appropriate therapy for these patients.

Peginterferon Alfa-2a and Ribavirin for 24 Weeks in Hepatitis C Type 1 and 4 Patients With Rapid Virological Response

BACKGROUND & AIMS This analysis reports the rate of sustained virological response (SVR) in patients infected with hepatitis C virus (HCV) genotype 1 or 4 who were assigned to 24 weeks of treatment with pegylated interferon (peginterferon) alfa-2a 180 mug/wk plus ribavirin 1000/1200 mg/day after achieving a rapid virological response (RVR; HCV RNA level <50 IU/mL) at week 4 in a prospective trial investigating response-guided therapy. METHODS Non-RVR patients with an early virological response were randomized to 48 or 72 weeks of therapy (this is a still-ongoing trial). RESULTS A total of 150 of 516 patients (29%) had an RVR, 143 of whom completed 24 weeks of treatment. Younger patients, leaner patients, and those with an HCV RNA level </=400,000 IU/mL and HCV genotype 4 infection were more likely to achieve an RVR; however, among patients with an RVR, no baseline factor predicted SVR. The SVR rate was 80.4% (115/143; 95% confidence interval [CI], 72.9-86.6) in patients who completed 24 weeks of treatment. The SVR rate was 86.7% (26/30; 95% CI, 69.3%-96.2%) in patients infected with genotype 4 and 78.8% in those infected with genotype 1 (89/113; 95% CI, 70.1%-85.9%; intent to treat: 89/120; 74.2%; 65.4-81.7%). Treatment was well tolerated. CONCLUSIONS This prospective study confirms that a 24-week regimen of peginterferon alfa-2a plus ribavirin 1000/1200 mg/day is appropriate in genotype 1 and 4 patients with a low baseline HCV RNA level who achieve an RVR by week 4 of therapy.

Genetic variation in the PNPLA3 gene is associated with alcoholic liver injury in caucasians.

A recent genome‐wide study revealed an association between variation in the PNPLA3 gene and liver fat content. In addition, the PNPLA3 single‐nucleotide polymorphism rs738409 (M148I) was reported to be associated with advanced alcoholic liver disease in alcohol‐dependent individuals of Mestizo descent. We therefore evaluated the impact of rs738409 on the manifestation of alcoholic liver disease in two independent German cohorts. Genotype and allele frequencies of rs738409 (M148I) were determined in 1,043 alcoholic patients with or without alcoholic liver injury and in 376 at‐risk drinkers from a population‐based cohort. Relative to alcoholic patients without liver damage (n = 439), rs738409 genotype GG was strongly overrepresented in patients with alcoholic liver cirrhosis (n = 210; OR 2.79; Pgenotype = 1.2 × 10−5; Pallelic = 1.6 × 10−6) and in alcoholic patients without cirrhosis but with elevated alanine aminotransferase levels (n = 219; OR 2.33; Pgenotype = 0.0085; Pallelic = 0.0042). The latter, biochemically defined association was confirmed in an independent population‐based cohort of at‐risk drinkers with a median alcohol intake of 300 g/week (OR 4.75; Pgenotype = 0.040; Pallelic = 0.022), and for aspartate aminotransferase (AST) levels. Frequencies of allele PNPLA3 rs738409(G) in individuals with steatosis and normal alanine aminotransferase (ALT) and AST levels were lower than in alcoholics without steatosis and normal ALT/AST (Pcombined = 0.03). The population attributable risk of cirrhosis in alcoholic carriers of allele PNPLA3 rs738409(G) was estimated at 26.6%. Conclusion: Genotype PNPLA3 rs738409(GG) is associated with alcoholic liver cirrhosis and elevated aminotransferase levels in alcoholic Caucasians. (HEPATOLOGY 2011)

Anti-tumor necrosis factor-alpha monoclonal antibody therapy in severe alcoholic hepatitis

BACKGROUND/AIMS Severe alcoholic hepatitis (AH) is associated with high mortality. Tumor necrosis factor-alpha (TNFalpha) has been demonstrated to play an important role in its pathophysiology. METHODS Twelve patients with biopsy-confirmed AH and a Maddrey discriminant factor >32 were treated with a single infusion of the anti-TNF monoclonal antibody Infliximab at a dose of 5mg/kg body weight. Serial measurements were made for various cytokines using specific enzyme-linked immunoassays (ELISA). In four patients, liver biopsy samples were available pretreatment and on day+28 of therapy. RESULTS Ten of the 12 patients are alive at a median of 15 (12-20) months. Two patients died within 30 days from septicemia. Serum bilirubin levels, Maddrey score, neutrophil count and C-reactive protein fell significantly within the first month. There was an early, though not significant, decrease in plasma levels of proinflammatory cytokines (interleukins (IL)-1beta, IL-6, IL-8, interferon-gamma), whereas plasma levels of TNFalpha remained near the sensitivity limit of the assay throughout the treatment course. While TNFalpha mRNA expression in the liver did not change, expression of IL-8, a cytokine regulated mainly by TNFalpha, was almost absent on day+28. CONCLUSIONS Our data suggest that randomized controlled trials of anti-TNF antibody in severe AH are warranted.

Gut microbiome development along the colorectal adenoma–carcinoma sequence

Colorectal cancer, a commonly diagnosed cancer in the elderly, often develops slowly from benign polyps called adenoma. The gut microbiota is believed to be directly involved in colorectal carcinogenesis. The identity and functional capacity of the adenoma- or carcinoma-related gut microbe(s), however, have not been surveyed in a comprehensive manner. Here we perform a metagenome-wide association study (MGWAS) on stools from advanced adenoma and carcinoma patients and from healthy subjects, revealing microbial genes, strains and functions enriched in each group. An analysis of potential risk factors indicates that high intake of red meat relative to fruits and vegetables appears to associate with outgrowth of bacteria that might contribute to a more hostile gut environment. These findings suggest that faecal microbiome-based strategies may be useful for early diagnosis and treatment of colorectal adenoma or carcinoma.

Fibroblast-specific protein 1 identifies an inflammatory subpopulation of macrophages in the liver

Cirrhosis is the end result of chronic liver disease. Hepatic stellate cells (HSC) are believed to be the major source of collagen-producing myofibroblasts in cirrhotic livers. Portal fibroblasts, bone marrow-derived cells, and epithelial to mesenchymal transition (EMT) might also contribute to the myofibroblast population in damaged livers. Fibroblast-specific protein 1 (FSP1, also called S100A4) is considered a marker of fibroblasts in different organs undergoing tissue remodeling and is used to identify fibroblasts derived from EMT in several organs including the liver. The aim of this study was to characterize FSP1-positive cells in human and experimental liver disease. FSP1-positive cells were increased in human and mouse experimental liver injury including liver cancer. However, FSP1 was not expressed by HSC or type I collagen-producing fibroblasts. Likewise, FSP1-positive cells did not express classical myofibroblast markers, including αSMA and desmin, and were not myofibroblast precursors in injured livers as evaluated by genetic lineage tracing experiments. Surprisingly, FSP1-positive cells expressed F4/80 and other markers of the myeloid-monocytic lineage as evaluated by double immunofluorescence staining, cell fate tracking, flow cytometry, and transcriptional profiling. Similar results were obtained for bone marrow-derived and peritoneal macrophages. FSP1-positive cells were characterized by increased expression of COX2, osteopontin, inflammatory cytokines, and chemokines but reduced expression of MMP3 and TIMP3 compared with Kupffer cells/macrophages. These findings suggest that FSP1 is a marker of a specific subset of inflammatory macrophages in liver injury, fibrosis, and cancer.

Impact of IL28B Genotype on the Early and Sustained Virologic Response in Treatment-Naïve Patients With Chronic Hepatitis C

BACKGROUND & AIMS Single nucleotide polymorphisms (SNPs) in the gene that encodes interleukin (IL)-28B predict response of patients with chronic hepatitis C to antiviral therapy. We investigated the roles of polymorphisms rs12979860 and rs8099917 on the early virologic response of treatment-naïve patients. METHODS SNPs were identified by real-time polymerase chain reaction analysis of samples from 682 patients (genotype [GT]1=372, GT2/3=208, GT4=102) who were treated with 180 μg pegylated interferon-α2a and 400 or 800 mg (GT2/3, depending on the protocol) or 1000-1200 mg (GT1/4) ribavirin/day. The duration of treatment was 24 (GT2/3) or 24-72 weeks (GT1/4). RESULTS Patients with C/C also had higher rates of rapid virologic response (RVR) (GT1, 38.3% vs 11.6%; GT4, 76.5% vs 23.5%; both P<.001) and sustained virologic responses (SVRs) (GT1, 79.1% vs 43.2%; GT4, 85.3% vs 44.1%; both P<.001). In patients with GT2/3, the RVR was more frequent in carriers of C/C (75.3% vs 52.6%, P<.01), but SVR rates were similar between those with C/C and T (80.5% vs 74.4%, P=.31). Results for rs8099917 were comparable. The positive predictive value of rs12979860 C/C for SVR was higher than that of rs8099917 T/T (80.5% vs 71.6%). Overall, RVR was the best predictor of SVR. In patients who did not have GT1, IL28B polymorphisms did not affect the SVR if RVR data were included in the multivariate analysis. CONCLUSIONS An early virologic response to pegylated interferon and ribavirin is more likely among carriers of rs12979860 C/C and rs8099917 T/T, which might underlie their high rates of SVR. Determination of the IL28B genotype and whether patients have an RVR might be used in future studies of patients with hepatitis C virus genotype 1 or 4.

The relation of iron status and hemochromatosis gene mutations in patients with chronic hepatitis C

BACKGROUND & AIMS Elevated hepatic iron concentration may affect the response to antiviral therapy in chronic hepatitis C. This study explored the contribution of genetic hemochromatosis to iron accumulation in chronic hepatitis C. METHODS HFE mutations (C282Y and H63D) were assessed in 184 patients with chronic hepatitis C virus and 487 controls. Liver biopsy specimens were available in 149 patients. Hepatic iron content was measured in 114 patients by atom-absorption spectrophotometry. RESULTS The C282Y and H63D allele frequencies were 7.06 and 11.6 in patients and 4.83 and 11.09 in controls, respectively. Eight patients were homozygotes (5 C282Y [2.7%] and 3 H63D [1.6%]), 2 compound heterozygotes (1%), and 49 heterozygotes (14 C282Y [7.6%] and 35 H63D [19%]). Biochemical evidence of iron overload was more common in patients with HFE mutations (28 of 47) than in those without (34 of 102; P = 0.0045). Histological iron grading and hepatic iron content overlapped among patients with or without mutations. A hepatic iron index of >1.9 was observed only in 1 of the 4 C282Y homozygotes and 1 of the 3 H63D homozygotes. CONCLUSIONS HFE mutations contribute to but do not fully explain hepatic iron accumulation in chronic hepatitis C. Furthermore, C282Y or H63D homozygosity in chronic hepatitis C is not necessarily associated with a high hepatic iron content.

Diagnostic Value of Quantitative Hepatic Copper Determination in Patients With Wilson’s Disease

BACKGROUND & AIMS A 5-fold increase of hepatic copper concentration is considered as the best available test for diagnosis of hepatic Wilsons disease (WD). However, the sensitivity and specificity of this test have never been fully investigated. METHODS Copper content was measured by flame atomic absorption spectroscopy in 114 liver biopsies obtained at diagnosis of WD, in 219 patients with noncholestatic liver diseases (including 144 with chronic hepatitis C and 44 with nonalcoholic fatty liver disease), and in 26 without evidence of liver disease. RESULTS Liver copper content was >250 microg/g in 95 WD patients (83.3%), between 50 and 250 microg/g in 15, and below 50 microg/g in 4. It did not correlate with age (r(2) = .003), the grade of fibrosis, or the presence of stainable copper. Liver copper content was >250 or between 50 and 250 microg/g in 3 (1.4%) and 20 (9.1%) of 219 patients with noncholestatic liver diseases, respectively. By lowering the cutoff from 250 to 75 microg/g, the sensitivity of liver copper content to diagnose WD increased from 83.3% (95% confidence interval, 75.2%-89.6%) to 96.5% (91.3%-99.1%), but the specificity decreased from 98.6% (96.0%-99.7%) to 95.4% (91.8%-97.8%). CONCLUSIONS There is no gold standard for the diagnosis of WD. Liver copper content is a useful parameter, but a value below 250 microg/g does not exclude WD. Diagnosis requires the combination of a variety of clinical and biochemical tests.