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Miriam K Stock,
Linda Hammerich,
Nicole T do O, Marie-Luise Berres,
Muhammad Alsamman,
Daniel Heinrichs,
Andreas Nellen,
Christian Trautwein,
Frank Tacke,
Hermann E Wasmuth,
Hacer Sahin
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ABSTRACT: Fibrosis or scarring of the liver parenchyma is a mainstay of chronic liver diseases and is associated with increased morbidity and mortality. Since complete scarring of the liver develops over several decades, therapeutic intervention with the aim of ameliorating fibrosis is of great clinical interest. In a recent study, we could identify the chemokine receptor antagonist Met-CCL5 as a potential compound to inhibit fibrosis progression and accelerate its regression. In the current study we characterized immune changes during fibrosis regression associated with the treatment with the CCL5 (RANTES) chemokine receptor antagonist Met-CCL5 in an established mouse model of chronic liver damage. Met-CCL5 or PBS was given after fibrosis induction (8 weeks of CCl4) and mice were sacrificed three and seven days after peak fibrosis. Mouse livers were analyzed for immune cell infiltration and cytokine gene expression. The results show that overall monocyte recruitment was not affected by Met-CCL5, but there was a significant shift to a pro-inflammatory Gr1+ monocyte population in the livers of mice treated with Met-CCL5. These monocytes were mostly iNOS +, a phenomenon which was also evident when analyzing the overall gene expression profiles in the livers. Since a shift in monocyte subpopulations has recently been identified to contribute to fibrosis regression, our results help explaining the efficacy of CCL5 chemokine antagonism as a novel treatment option for fibrotic liver diseases.
International journal of clinical and experimental pathology 01/2013; 6(4):678-85. · 1.89 Impact Factor
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Hacer Sahin,
Erawan Borkham-Kamphorst,
Nicole T do O, Marie-Luise Berres,
Michaela Kaldenbach,
Petra Schmitz,
Ralf Weiskirchen,
Christian Liedtke,
Konrad L Streetz,
Kathrin Maedler,
Christian Trautwein,
Hermann E Wasmuth
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ABSTRACT: Aberrant expression of the chemokine CXCL10 has been linked to the severity of hepatitis C virus (HCV)-induced liver injury, but the underlying molecular mechanisms remain unclear. In this study, we describe a yet unknown pro-apoptotic effect of CXCL10 in hepatocytes, which is not mediated through its cognate chemokine receptor, but the LPS receptor TLR4. To this end, we investigated the link of CXCL10 expression with apoptosis in HCV-infected patients and in murine liver injury models. Mice were treated with CXCL10 or neutralizing antibody to systematically analyze effects on hepatocellular apoptosis in vivo. Direct pro-apoptotic functions of CXCL10 on different liver cell types were evaluated in detail in vitro. The results show that CXCL10 expression was positively correlated to liver cell apoptosis in humans and mice. Neutralization of CXCL10 ameliorated ConA-induced tissue injury in vivo which was strongly associated with reduced liver cell apoptosis. In vitro, CXCL10 mediated apoptosis of hepatocytes involving TLR4 but not CXCR3 signalling. Specifically, CXCL10 induced long-term Akt and JNK activation leading to hepatocyte apoptosis by Caspase-8, Caspase-3 and PAK-2 cleavage. Accordingly, systemic application of CXCL10 led to TLR4 induced liver cell apoptosis in vivo. In conclusion, the results identify CXCL10 and its non-cognate receptor TLR4 as a pro-apoptotic signalling cascade during liver injury. Antagonism of the CXCL10/TLR4 pathway might be a therapeutic option in liver diseases associated with increased apoptosis. (HEPATOLOGY 2012.).
Hepatology 09/2012; · 11.66 Impact Factor
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ABSTRACT: Although acute liver failure is a rare disease, its presence is associated with high morbidity and mortality in affected patients. While a contribution of the immune system to the outcome of toxic liver failure is anticipated, functionally relevant immune cell receptors for liver cell damage need to be better defined. We here investigate the relevance of the chemokine receptor CXCR3, which is important for hepatic immune cell infiltration, in a model of experimental acute liver failure. Liver injury was induced by a single intraperitoneal injection of carbon tetrachloride (CCl(4)) in CXCR3(-/-), CCR1(-/-), CCR5(-/-) and wild-type mice. In this model, CXCR3(-/-) mice displayed augmented liver damage compared with all other mouse strains as assessed by liver histology and serum transaminases 24 and 72 h after injury. Phenotypically, CXCR3(-/-) mice had significantly reduced intrahepatic NK and NKT cells after injury at all investigated time points (all P<0.05), but strongly elevated expression levels of IL1-β, TNF-α and IFN-γ. In line with a functional role of innate immune cells, wild-type mice depleted for NK cells with an anti-ASIALO GM1 antibody before liver injury also displayed increased liver injury after CCl(4) challenge. CXCR3(-/-) and NK cell-depleted mice show reduced apoptotic liver cells (TUNEL assay), but more necrotic hepatocytes. Functionally, the augmented liver cell necrosis in CXCR3(-/-) and NK cell-depleted mice was associated with increased expression of high mobility group 1 (HMGB1) protein and a consecutive enhanced infiltration of neutrophils into the liver. In conclusion, the results demonstrate a primarily unexpected beneficial role of CXCR3 in acute toxic liver injury. These findings should be taken into account when planning trials with CXCR3 antagonists.
Laboratory Investigation 03/2012; 92(5):724-34. · 3.64 Impact Factor
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Hacer Sahin,
Erawan Borkham-Kamphorst,
Christoph Kuppe,
Mirko Moreno Zaldivar,
Christoph Grouls,
Muhammad Al-samman,
Andreas Nellen,
Petra Schmitz,
Daniel Heinrichs, Marie-Luise Berres,
Dennis Doleschel,
David Scholten,
Ralf Weiskirchen,
Marcus J Moeller,
Fabian Kiessling,
Christian Trautwein,
Hermann E Wasmuth
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ABSTRACT: Recent data suggest that the chemokine receptor CXCR3 is functionally involved in fibroproliferative disorders, including liver fibrosis. Neoangiogenesis is an important pathophysiological feature of liver scarring, but a functional role of angiostatic CXCR3 chemokines in this process is unclear. We therefore investigated neoangiogenesis in carbon tetrachloride (CCl(4))-induced liver fibrosis in Cxcr3(-/-) and wildtype mice by histological, molecular, and functional imaging methods. Furthermore, we assessed the direct role of vascular endothelial growth factor (VEGF) overexpression on liver angiogenesis and the fibroproliferative response using a Tet-inducible bitransgenic mouse model. The feasibility of attenuation of angiogenesis and associated liver fibrosis by therapeutic treatment with the angiostatic chemokine Cxcl9 was systematically analyzed in vitro and in vivo. The results demonstrate that fibrosis progression in Cxcr3(-/-) mice was strongly linked to enhanced neoangiogenesis and VEGF/VEGFR2 expression compared with wildtype littermates. Systemic VEGF overexpression led to a fibrogenic response within the liver and was associated with a significantly increased Cxcl9 expression. In vitro, Cxcl9 displayed strong antiproliferative and antimigratory effects on VEGF-stimulated endothelial cells and stellate cells by way of reduced VEGFR2 (KDR), phospholipase Cγ (PLCγ), and extracellular signal-regulated kinase (ERK) phosphorylation, identifying this chemokine as a direct counter-regulatory molecule of VEGF signaling within the liver. Accordingly, systemic administration of Cxcl9 led to a strong attenuation of neoangiogenesis and experimental liver fibrosis in vivo. CONCLUSION: The results identify direct angiostatic and antifibrotic effects of the Cxcr3 ligand Cxcl9 in a model of experimental liver fibrosis. The amelioration of liver damage by systemic application of Cxcl9 might offer a novel therapeutic approach for chronic liver diseases associated with increased neoangiogenesis.
Hepatology 01/2012; 55(5):1610-9. · 11.66 Impact Factor
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ABSTRACT: The chemokine CCL5 is involved in the recruitment of immune cells and a subsequent activation of hepatic stellate cells (HSC) after liver injury. We here investigate whether inhibition of CCL5 oligomerization and glycosaminoglycan binding by a mutated CCL5 protein ((44)AANA(47)-CCL5) has the potential to ameliorate liver cell injury and fibrosis in vivo.
Liver injury was induced in C57BL/6 mice by intraperitoneal injection of carbon tetrachloride (CCl(4)) in an acute and a chronic liver injury model. Simultaneously, mice received either (44)AANA(47)-CCL5 or vehicle. Liver cell necrosis and fibrosis was analyzed by histology, and measurement of serum transaminases and hydroxyproline. Intrahepatic mRNA expression of fibrosis and inflammation related genes were determined by quantitative RT-PCR and infiltration of immune cells was assessed by FACS analysis and immunocytochemistry. In vitro, HSC were stimulated with conditioned media of T-cell enriched splenocytes.
(44)AANA(47)-CCL5 treated mice displayed a significantly reduced degree of acute liver injury (liver cell necrosis, transaminases) and fibrosis (Sirus red positive area and hydroxyproline content) compared to vehicle treated mice. Ameliorated fibrosis by (44)AANA(47)-CCL5 was associated with a decreased expression of fibrosis related genes, decreased α-smoth muscle antigen (αSMA) and a reduction of infiltrating immune cells. In the acute model, (44)AANA(47)-CCL5 treated mice displayed a reduced immune cell infiltration and mRNA levels of TNF, IL-1 and CCL3 compared to vehicle treated mice. In vitro, conditioned medium of T-cell enriched splenocytes of (44)AANA(47)-CCL5 treated mice inhibited the chemotaxis and proliferation of HSC.
The results provide evidence that inhibition of oligomerization and glycosaminoglycan binding of the chemokine CCL5 is a new therapeutic strategy for the treatment of acute and chronic liver injuries and represents an alternative to chemokine receptor antagonism.
PLoS ONE 01/2012; 7(5):e36614. · 4.09 Impact Factor
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Daniel Heinrichs,
Meike Knauel,
Christian Offermanns, Marie-Luise Berres,
Andreas Nellen,
Lin Leng,
Petra Schmitz,
Richard Bucala,
Christian Trautwein,
Christian Weber,
Jürgen Bernhagen,
Hermann E Wasmuth
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ABSTRACT: Macrophage migration inhibitory factor (MIF) is a pleiotropic inflammatory cytokine that has been implicated in various inflammatory diseases. Chronic inflammation is a mainstay of liver fibrosis, a leading cause of morbidity worldwide, but the role of MIF in liver scarring has not yet been elucidated. Here we have uncovered an unexpected antifibrotic role for MIF. Mice genetically deleted in Mif (Mif(-/-)) showed strongly increased fibrosis in two models of chronic liver injury. Pronounced liver fibrosis in Mif(-/-) mice was associated with alterations in fibrosis-relevant genes, but not by a changed intrahepatic immune cell infiltration. Next, a direct impact of MIF on hepatic stellate cells (HSC) was assessed in vitro. Although MIF alone had only marginal effects on HSCs, it markedly inhibited PDGF-induced migration and proliferation of these cells. The inhibitory effects of MIF were mediated by CD74, which we detected as the most abundant known MIF receptor on HSCs. MIF promoted the phosphorylation of AMP-activated protein kinase (AMPK) in a CD74-dependent manner and, in turn, inhibition of AMPK reversed the inhibition of PDGF-induced HSC activation by MIF. The pivotal role of CD74 in MIF-mediated antifibrotic properties was further supported by augmented liver scarring of Cd74(-/-) mice. Moreover, mice treated with recombinant MIF displayed a reduced fibrogenic response in vivo. In conclusion, we describe a previously unexplored antifibrotic function of MIF that is mediated by the CD74/AMPK signaling pathway in HSCs. The results imply MIF and CD74 as targets for treatment of liver diseases.
Proceedings of the National Academy of Sciences 10/2011; 108(42):17444-9. · 9.68 Impact Factor
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ABSTRACT: Progressive liver fibrosis is the main predictor of disease outcome in chronic hepatitis C viral (HCV) infection. Although the importance of the coagulation cascade has been suggested in liver fibrogenesis, the role of the fibrinolytic pathway is yet unclear.
We evaluated the association of serum levels of the fibrinolysis-associated soluble urokinase plasminogen activator receptor (suPAR) with the severity of liver fibrosis in HCV infection.
suPAR serum levels were assessed in 146 chronically HCV-infected patients of 2 independent cohorts (64 subjects in the screening cohort, 82 in the validation cohort) by enzyme-linked immunosorbent assay and correlated with biopsy-proven histologic stage of liver fibrosis and noninvasive liver fibrosis markers (aspartate transaminase to platelets ratio index score, transient elastography).
suPAR serum levels were strongly associated with the histologic stage of liver fibrosis in both cohorts (P<0.0001). Although mean suPAR levels in patients with F1 and F2 fibrosis were not different from healthy control subjects, they were significantly increased at higher stages of liver fibrosis (F3 and F4, P<0.0001). suPAR values had a high diagnostic specificity and sensitivity to differentiate mild/moderate fibrosis (F1/F2) from severe fibrosis (F3/F4) with an area under curve of 0.774 (P=0.0001) and for the differentiation of noncirrhosis from cirrhosis (F1/F2/F3 vs. F4, area under curve 0.791, P=0.0001). SuPAR serum levels were also strongly correlated to the noninvasive fibrosis markers aspartate transaminase to platelets ratio index score (r=0.52) and transient elastography (r=0.44, both P<0.0001).
Serum suPAR levels were robust markers of liver fibrosis in 2 cohorts with a comparable diagnostic accuracy for prediction of severe liver fibrosis as established noninvasive marker.
Journal of clinical gastroenterology 09/2011; 46(4):334-8. · 2.21 Impact Factor
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ABSTRACT: Chemokines are chemotactic mediators that are implicated in liver diseases. In viral hepatitis and primary biliary cirrhosis, a predominant chemokine receptor expressed in the liver is CXCR3, suggesting that its specific ligands are important in the progression of chronic liver diseases across different aetiologies.
We analysed the serum concentrations of the CXCR3 ligands, CXCL9 (monokine induced by interferon-γ), CXCL10 (interferon-γ-inducible protein 10) and CXCL11 (interferon-inducible T cell α chemo-attractant) in healthy controls (n=53), subjects with histologically determined liver fibrosis (n=109) and patients with different stages of cirrhosis (n=153) of various disease aetiologies. Chemokine concentrations were determined by cytometric bead assay or ELISA respectively. Serum concentrations of all three chemokines were significantly increased in patients with chronic liver diseases compared with healthy controls (P<0.001). In the biopsied fibrosis cohort, CXCL9 and CXCL10 were positively associated with the severity of liver fibrosis (histology and serum markers), while CXCL11 was not. In cirrhotic patients, CXCL9 was increased in early Child-Pugh stages, while CXCL11 was elevated only in Child B and C patients and CXCL10 across all stages. Notably, CXCR3 chemokines were also associated with the development of clinical complications of cirrhosis, especially portal hypertension. All chemokines significantly correlated with serum levels of the hepatoprotective cytokines interleukin (IL)-6 and IL-10, suggesting their involvement in a counter-regulatory response during the progression of liver disease, shedding new light on their involvement in the pathophysiology of chronic liver diseases.
CXCR3 chemokines are differentially expressed during chronic liver diseases across different disease stages and aetiologies. Their association with portal hypertension and hepatoprotective cytokines implies biological functions beyond immune cell recruitment, thereby provoking new diagnostic and therapeutic concepts.
Liver international: official journal of the International Association for the Study of the Liver 07/2011; 31(6):840-9. · 3.82 Impact Factor
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ABSTRACT: Chemokines are a family of small heparin-binding proteins that orchestrate the infiltration of immune cells into the liver, but also directly influence the biology of resident liver cells. A nonredundant role of different chemokines and their receptors has been identified within the last years in animal models of acute and chronic liver diseases. Chemokine receptors that have been shown to play an import pathophysiological role in the liver are CCR1, CCR2, CCR5, D6, CXCR2 and CXCR3. Interestingly, most of these receptors have already been targeted by specific antagonists in early human trials and a CCR5 antagonist is already licensed for use in HIV infection. Most of these trials have been performed in autoimmune and infectious human diseases, but no controlled clinical trials have yet been performed in patients with liver diseases. Nevertheless, in light of growing evidence that chemokines are important mediators of liver damage, these trials seem to be on the clinical horizon.
Expert Review of Clinical Pharmacology 07/2011; 4(4):503-13.
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ABSTRACT: Genetic host factors influence the progression of hepatitis C infection (HCV). Chemokines play important roles in HCV-induced liver fibrosis. Recently, a single nucleotide polymorphism in the Duffy antigen receptor for chemokines (DARC) was identified which strongly determines the serum concentrations of pivotal pro-fibrotic chemokines, including CCL2. We here tested the hypothesis that this genetic variant (rs12075 A/G) is a risk factor for liver fibrosis in HCV infection. Overall, 880 patients with HCV from three cohorts and 108 controls were genotyped for rs12075. Although serum CCL2 levels were associated with early liver fibrosis, rs12075 itself was not associated with HCV infection or the severity of liver disease in any of the cohorts. The lack of association was evident in qualitative and quantitative analysis despite sufficient statistical power. We conclude that gene variations that strongly determine serum concentrations of chemokines are not necessarily risk markers of the disease traits in which these molecules play pathophysiological roles.
Human immunology 03/2011; 72(3):273-7. · 2.55 Impact Factor
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ABSTRACT: The recurrence of liver fibrosis after liver transplantation (LT) for hepatitis C virus (HCV) infection is responsible for graft loss and patient mortality. Although the contribution of the immune system to fibrosis recurrence is anticipated, systematic studies evaluating immune parameters as predictive markers of allograft fibrosis are lacking. The infiltration of immune cells into the graft is governed by chemokines. Here we assessed the predictive value of serum levels of chemokines [chemokine (C-X-C motif) ligand 9 (CXCL9), CXCL10, CXCL11, and chemokine (C-C motif) ligand 2 (CCL2)] with respect to fibrosis recurrence after LT in 90 HCV-infected organ recipients. Chemokines were determined within the first and third years after LT and were correlated with histological fibrosis progression in protocol biopsy samples at 1, 3, 5, and 7 years (median follow-up = 3 years). The association of chemokines with fibrosis progression was assessed by univariate and multivariate analyses and by Cox regression analysis. The results for the analyzed chemokines showed that CXCL10 levels in the first year after LT were strongly associated with early fibrosis recurrence (P = 0.005) independently of risk confounders (including the donor age, HCV viral load, HCV genotype, acute rejection, and inflammatory activity). As assessed by Cox regression analysis, a CXCL10 serum level ≤ 140 pg/mL was significantly predictive of the absence of F2 fibrosis (P = 0.001), whereas a level ≤ 220 pg/mL early after LT predicted the absence of F3 fibrosis during follow-up (P = 0.035). CONCLUSION: CXCL10 is an independent biomarker of the recurrence of significant fibrosis after LT for HCV infection. These results might guide patients' care after transplantation and help us to select optimal candidates for antiviral therapy post-LT.
Hepatology 02/2011; 53(2):596-603. · 11.66 Impact Factor
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Marie-Luise Berres,
Rory R Koenen,
Anna Rueland,
Mirko Moreno Zaldivar,
Daniel Heinrichs,
Hacer Sahin,
Petra Schmitz,
Konrad L Streetz,
Thomas Berg,
Nikolaus Gassler,
Ralf Weiskirchen,
Amanda Proudfoot,
Christian Weber,
Christian Trautwein,
Hermann E Wasmuth
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ABSTRACT: Activation of hepatic stellate cells in response to chronic inflammation represents a crucial step in the development of liver fibrosis. However, the molecules involved in the interaction between immune cells and stellate cells remain obscure. Herein, we identify the chemokine CCL5 (also known as RANTES), which is induced in murine and human liver after injury, as a central mediator of this interaction. First, we showed in patients with liver fibrosis that CCL5 haplotypes and intrahepatic CCL5 mRNA expression were associated with severe liver fibrosis. Consistent with this, we detected Ccl5 mRNA and CCL5 protein in 2 mouse models of liver fibrosis, induced by either injection of carbon tetrachloride (CCl4) or feeding on a methionine and choline-deficient (MCD) diet. In these models, Ccl5-/- mice exhibited decreased hepatic fibrosis, with reduced stellate cell activation and immune cell infiltration. Transplantation of Ccl5-deficient bone marrow into WT recipients attenuated liver fibrosis, identifying infiltrating hematopoietic cells as the main source of Ccl5. We then showed that treatment with the CCL5 receptor antagonist Met-CCL5 inhibited cultured stellate cell migration, proliferation, and chemokine and collagen secretion. Importantly, in vivo administration of Met-CCL5 greatly ameliorated liver fibrosis in mice and was able to accelerate fibrosis regression. Our results define a successful therapeutic approach to reduce experimental liver fibrosis by antagonizing Ccl5 receptors.
The Journal of clinical investigation 10/2010; 120(11):4129-40. · 15.39 Impact Factor
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ABSTRACT: Chemokines are small chemotactic molecules which regulate the infiltration of immune cells to sites of inflammatory injury. In recent years their contribution to the initiation and perpetuation of liver injury has been better defined. However, the role of chemokines in liver diseases related to the metabolic syndrome still needs to be elucidated in detail.
Chemokines were mostly detected at the mRNA level in the liver and as proteins in the serum of patients with non-alcoholic steatohepatitis (NASH) or fatty liver. Animals with targeted deletion of chemokines have recently been subjected to NASH models to functionally dissect the role of chemokines in fatty liver diseases.
In human liver with features of NASH, different CC and CXC chemokines have been detected at elevated mRNA levels in comparison to healthy subjects. Some of these chemokines have also been associated with NASH by demonstrating higher serum levels in affected patients. Until now, only a few animal models have been analyzed with respect to the functional role of these molecules. However, data from CCL2 and CXCR3 knockout mice suggest that these pathways are important in liver injury. CCL2 seems to influence the infiltration of macrophages to adipose tissue and thereby modulate insulin resistance.
The further elucidation of the pathophysiology of NASH will lead to new therapeutic options to halt or reverse progressive liver disease. In this respect, chemokines are attractive target molecules, as they influence immunologic and metabolic pathways and the first oral chemokine receptor antagonists have already been licensed for humans.
Digestive Diseases 01/2010; 28(1):192-6. · 2.37 Impact Factor
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Mirko Moreno Zaldivar,
Katrin Pauels,
Philipp von Hundelshausen, Marie-Luise Berres,
Petra Schmitz,
Jörg Bornemann,
M Anna Kowalska,
Nikolaus Gassler,
Konrad L Streetz,
Ralf Weiskirchen,
Christian Trautwein,
Christian Weber,
Hermann E Wasmuth
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ABSTRACT: Liver fibrosis is a major cause of morbidity and mortality worldwide. Platelets are involved in liver damage, but the underlying molecular mechanisms remain elusive. Here, we investigate the platelet-derived chemokine (C-X-C motif) ligand 4 (CXCL4) as a molecular mediator of fibrotic liver damage. Serum concentrations and intrahepatic messenger RNA of CXCL4 were measured in patients with chronic liver diseases and mice after toxic liver injury. Platelet aggregation in early fibrosis was determined by electron microscopy in patients and by immunohistochemistry in mice. Cxcl4(-/-) and wild-type mice were subjected to two models of chronic liver injury (CCl(4) and thioacetamide). The fibrotic phenotype was analyzed by histological, biochemical, and molecular analyses. Intrahepatic infiltration of immune cells was investigated by fluorescence-activated cell sorting, and stellate cells were stimulated with recombinant Cxcl4 in vitro. The results showed that patients with advanced hepatitis C virus-induced fibrosis or nonalcoholic steatohepatitis had increased serum levels and intrahepatic CXCL4 messenger RNA concentrations. Platelets were found directly adjacent to collagen fibrils. The CCl(4) and thioacetamide treatment led to an increase of hepatic Cxcl4 levels, platelet activation, and aggregation in early fibrosis in mice. Accordingly, genetic deletion of Cxcl4 in mice significantly reduced histological and biochemical liver damage in vivo, which was accompanied by changes in the expression of fibrosis-related genes (Timp-1 [tissue inhibitor of matrix metalloproteinase 1], Mmp9 [matrix metalloproteinase 9], Tgf-beta [transforming growth factor beta], IL10 [interleukin 10]). Functionally, Cxcl4(-/-) mice showed a strongly decreased infiltration of neutrophils (Ly6G) and CD8(+) T cells into the liver. In vitro, recombinant murine Cxcl4 stimulated the proliferation, chemotaxis, and chemokine expression of hepatic stellate cells. Conclusion: The results underscore an important role of platelets in chronic liver damage and imply a new target for antifibrotic therapies.
Hepatology 11/2009; 51(4):1345-53. · 11.66 Impact Factor
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ABSTRACT: The chemokine decoy receptor D6 is a promiscuous chemokine receptor lacking classical signaling functions. It negatively regulates inflammation by targeting CC chemokines to cellular internalization and degradation. Here we analyze the function of D6 in acute CCl(4)-induced liver damage in constitutive D6(-/-) and wild-type mice. The degree of liver injury was assessed by liver histology, serum transaminases, IL-6, and TNFalpha mRNA expression. Protein levels of D6 ligands (CCL2, CCL3, CCL5) and the non-D6-ligand CXCL9 within the livers were determined by ELISAs. The intrahepatic infiltration of immune cells was characterized by FACS. Genetic deletion of D6 led to prolonged liver damage after acute CCl(4) administration. The augmented liver damage in D6(-/-) mice was associated with increased protein levels of intrahepatic inflammatory chemokines CCL2, CCL3, and CCL5 after 48 h, whereas CXCL9 was not different between knockout and wild-type mice. Functionally, increased intra-hepatic CC chemokine concentrations led to increased infiltration of CD45(+) leukocytes, which were mainly identified as T and NK cells. In conclusion, the chemokine scavenger receptor D6 has a non-redundant role in acute toxic liver injury in vivo. These results support the importance of post-translational chemokine regulation and describe a new mechanism of immune modulation within the liver.
Biological Chemistry 08/2009; 390(10):1039-45. · 2.96 Impact Factor
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Hermann E Wasmuth,
Frank Lammert,
Mirko Moreno Zaldivar,
Ralf Weiskirchen,
Claus Hellerbrand,
David Scholten, Marie-Luise Berres,
Henning Zimmermann,
Konrad L Streetz,
Frank Tacke,
Sonja Hillebrandt,
Petra Schmitz,
Hildegard Keppeler,
Thomas Berg,
Edgar Dahl,
Nikolaus Gassler,
Scott L Friedman,
Christian Trautwein
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ABSTRACT: Fibrosis is the hallmark of chronic liver diseases, yet many aspects of its mechanism remain to be defined. Chemokines are ubiquitous chemotactic molecules that mediate many acute and chronic inflammatory conditions, and CXC chemokine genes colocalize with a locus previously shown to include fibrogenic genes. We investigated the roles of the chemokine CXCL9 and its receptor CXCR3 in liver fibrosis.
The effects of CXCL variants on fibrogenesis were analyzed using samples from patients with hepatitis C virus infection and by induction of fibrosis in CXCR3(-/-) and wild-type mice. In mice, intrahepatic immune cell subsets were investigated and interferon gamma messenger RNA levels were measured at baseline and after injury. Human serum CXCL9 levels were measured and correlated with CXCL9 variant and fibrosis severity. The effects of stimulation with CXCL9 were investigated on human hepatic stellate cells (LX-2).
Specific CXCL9 variants were associated with liver fibrosis in mice and humans; CXCL9 serum concentrations correlated with genotypes and levels of fibrosis in patients. In contrast to other chemokines, CXCL9 exerted antifibrotic effects in vitro, suppressing collagen production in LX-2 cells. CXCR3(-/-) mice had increased liver fibrosis; progression was associated with decreased numbers of intrahepatic interferon gamma-positive T cells and reduced interferon gamma messenger RNA, indicating that CXCL9-CXCR3 regulates Th1-associated immune pathways.
This is the first description of a chemokine-based antifibrotic pathway in the liver; antifibrotic therapies might be developed to modulate CXC chemokine levels.
Gastroenterology 03/2009; 137(1):309-19, 319.e1-3. · 11.68 Impact Factor
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Marie-Luise Berres,
Sven Papen,
Katrin Pauels,
Petra Schmitz,
Mirko Moreno Zaldivar,
Claus Hellerbrand,
Tobias Mueller,
Thomas Berg,
Ralf Weiskirchen,
Christian Trautwein,
Hermann E Wasmuth
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ABSTRACT: YKL-40 is a chitinase-like protein involved in matrix remodelling and a non-invasive fibrosis marker. We assessed whether a functional promoter polymorphism in CHI3L1, encoding YKL-40, is associated with HCV-induced liver fibrosis and influences YKL-40 serum concentrations.
The CHI3L1 -131G-->C promoter polymorphism was genotyped in two cohorts of HCV infected patients (n=440) by 5'-endonuclease assays. Histological fibrosis scores and YKL-40 serum levels (ELISA) were associated with CHI3L1 -131G-->C by quantitative and qualitative genetic analyses and corrected by multivariate analysis.
CHI3L1 -131G-->C genotype was strongly associated with the stage of liver fibrosis in the screening (n=265, P=0.001) and validation cohort (n=175, P=0.009). Homozygous carriers of the G allele were protected from severe fibrosis (F3/F4). This association was confirmed after correction for age and gender. Functionally, the G allele was associated with reduced serum levels of YKL-40 in HCV infected patients (P=0.002).
The CHI3L1 promoter polymorphism -131G-->C determines YKL-40 serum levels and is associated with the severity of HCV-induced liver fibrosis. These results suggest a functional role of YKL-40 in liver fibrogenesis and should be taken into account when using YKL-40 as a non-invasive fibrosis marker.
Journal of Hepatology 12/2008; 50(2):370-6. · 9.26 Impact Factor
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ABSTRACT: Critical illness in cirrhotic patients is associated with a poor prognosis and increased susceptibility to infections. Monocyte HLA-DR expression is decreased in cirrhotic patients, but its prognostic value has not been investigated prospectively.
Thirty-eight critically ill patients with decompensated liver cirrhosis were included in this prospective study. On admission to the intensive care unit (ICU), inflammatory parameters (C-reactive protein, procalcitonin and lipopolysaccharide-binding protein), interleukin (IL)-10, interferon (IFN)-gamma serum levels, tumour necrosis factor (TNF)-alpha ex vivo stimulation (whole blood assay) and HLA-DR expression on monocytes (FACS analysis) were determined. Immune parameters were furthermore measured every third day until discharge from the ICU or death of the patients.
Intensive care unit mortality of the cirrhotic patients was 34.2%. During admission, TNF ex vivo, IFN-gamma and HLA-DR expression were lower in non-survivors (all P<0.05), while IL-10 levels were increased in non-survivors compared with survivors (P=0.001). However, individual values clearly overlapped between groups. Prospective analysis revealed that monocyte HLA-DR expression remained stable or increased in survivors, but decreased in non-survivors (P=0.002). A decrease in HLA-DR expression between admission and day 3 was strongly associated with decreased IFN-gamma levels and increased ICU mortality (hazard ratio 3.36, P=0.008), mostly owing to late sepsis. This association was independent of the sequential organ failure assessment and model for end-stage liver disease score.
Here we establish the relative HLA-DR expression (admission/day 3) as a prognostic marker for ICU mortality in critically ill cirrhotic patients. These results may guide the evaluation of immune-modulating therapies in these patients.
Liver international: official journal of the International Association for the Study of the Liver 10/2008; 29(4):536-43. · 3.82 Impact Factor
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ABSTRACT: Chronic hepatitis C (HCV) represents one of the most common chronic infections worldwide and is a major indication for liver transplantation. Liver inflammation is the main predictor of advanced fibrosis in HCV. Inflammatory cells are recruited to the liver by chemokines. Recently, a novel class of chemokine receptors has been characterized that lack signaling functions and are termed scavenger receptors. We determine here whether genetic variations of the scavenger receptor D6 contribute to the grade of liver inflammation in HCV. Four haplotype tagging single nucleotide polymorphisms (SNPs) were identified from HapMap that cover the genetic information of D6 (CCBP2). Among these SNPs, rs4683336 was associated with liver inflammation in qualitative (p = 0.003) and quantitative (p = 0.0086) genotype analysis. This association was confirmed in an independent cohort of HCV-infected patients (p = 0.006 for qualitative and p = 0.0046 for quantitative analysis, respectively). Furthermore, the haplotype that is tagged by marker rs4683336 was significantly correlated with liver inflammation when compared with the most common D6 haplotype (p = 0.014). The importance of genetic variations in D6 was supported through the demonstration of an association of D6 mRNA expression with histologic inflammation in liver biopsies and a considerable range of D6 mRNA expression in isolated human hepatocytes. In conclusion, we demonstrate that variations in a chemokine scavenging receptor are significantly correlated with clinical inflammatory phenotypes such as HCV infection.
Human Immunology 10/2008; 69(12):861-6. · 2.84 Impact Factor
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Hermann E Wasmuth,
Mirko Moreno Zaldivar, Marie-Luise Berres,
Alexa Werth,
David Scholten,
Sonja Hillebrandt,
Frank Tacke,
Petra Schmitz,
Edgar Dahl,
Tonio Wiederholt,
Claus Hellerbrand,
Thomas Berg,
Ralf Weiskirchen,
Christian Trautwein,
Frank Lammert
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ABSTRACT: The chemokine receptor CX3CR1 and its specific ligand fractalkine (CX3CL1) are known to modulate inflammatory and fibroproliferative diseases. Here we investigate the role of CX3CR1/fractalkine in HCV-induced liver fibrosis.
A genotype analysis of CX3CR1 variants was performed in 211 HCV-infected patients. Hepatic expression of CX3CR1 was studied in HCV-infected livers and isolated liver cell populations by RT-PCR and immunohistochemistry. The effects of fractalkine on mRNA expression of profibrogenic genes were determined in isolated hepatic stellate cells (HSC) and CX3CR1 genotypes were related to intrahepatic TIMP-1 mRNA levels.
The intrahepatic mRNA expression of CX3CR1 correlates with the stage of HCV-induced liver fibrosis (P=0.03). The CX3CR1 coding variant V249I is associated with advanced liver fibrosis, independent of the T280M variant (P=0.009). CX3CR1 is present on primary HSC and fractalkine leads to a suppression of tissue inhibitor of metalloproteinase (TIMP)-1 mRNA in HSC (P=0.03). Furthermore, CX3CR1 genotypes are associated with TIMP-1 mRNA expression in HCV-infected liver (P=0.03).
The results identify the fractalkine receptor CX3CR1 as susceptibility a gene for hepatic fibrosis in HCV infection. The modulation of TIMP-1 expression by fractalkine and CX3CR1 genotypes provides functional support for the observed genotype-phenotype association.
Journal of Hepatology 03/2008; 48(2):208-15. · 9.26 Impact Factor
