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1.
Cells ; 10(2)2021 01 28.
Article in English | MEDLINE | ID: mdl-33525493

ABSTRACT

Macrophage migration inhibitory factor (MIF) is a pleiotropic inflammatory cytokine with anti-fibrotic properties in toxic liver injury models and anti-steatotic functions in non-alcoholic fatty liver disease (NAFLD) attributed to the CD74/AMPK signaling pathway. As NAFLD progression is associated with fibrosis, we studied MIF function during NAFLD-associated liver fibrogenesis in mice and men by molecular, histological and immunological methods in vitro and in vivo. After NASH diet feeding, hepatic Mif expression was strongly induced, an effect which was absent in Mif∆hep mice. In contrast to hepatotoxic fibrosis models, NASH diet-induced fibrogenesis was significantly abrogated in Mif-/- and Mif∆hep mice associated with a reduced accumulation of the pro-fibrotic type-I NKT cell subpopulation. In vitro, MIF skewed the differentiation of NKT cells towards the type-I subtype. In line with the murine results, expression of fibrosis markers strongly correlated with MIF, its receptors, and markers of NKT type-I cells in NASH patients. We conclude that MIF expression is induced during chronic metabolic injury in mice and men with hepatocytes representing the major source. In NAFLD progression, MIF contributes to liver fibrogenesis skewing NKT cell polarization toward a pro-fibrotic phenotype highlighting the complex, context-dependent role of MIF during chronic liver injury.


Subject(s)
Macrophage Migration-Inhibitory Factors/metabolism , Natural Killer T-Cells/immunology , Non-alcoholic Fatty Liver Disease/immunology , Non-alcoholic Fatty Liver Disease/pathology , Animals , Biomarkers/metabolism , Cell Polarity , Diet , Disease Progression , Fibrosis , Gene Expression Regulation , Hepatic Stellate Cells/metabolism , Hepatic Stellate Cells/pathology , Hepatocytes/metabolism , Hepatocytes/pathology , Humans , Liver/metabolism , Liver/pathology , Male , Mice, Inbred C57BL , Models, Biological , Non-alcoholic Fatty Liver Disease/genetics , Receptors, Immunologic/metabolism
2.
Front Immunol ; 11: 354, 2020.
Article in English | MEDLINE | ID: mdl-32218781

ABSTRACT

Background and Aims: Monocyte chemotactic protein-1 (MCP-1) is a potent chemoattractant for monocytes. It is involved in pathogenesis of several inflammatory diseases. Hepatic MCP-1 is a readout of macrophage activation. While inflammation is a major driver of liver disease progression, the origin and role of circulating MCP-1 as a biomarker remains unclear. Methods: Hepatic CC-chemokine ligand 2 (CCL2) expression and F4/80 staining for Kupffer cells were measured and correlated in a mouse model of chronic liver disease (inhalative CCl4 for 7 weeks). Next, hepatic RNA levels of CCL2 were measured in explanted livers of 39 patients after transplantation and correlated with severity of disease. Changes in MCP-1 were further evaluated in a rat model of experimental cirrhosis and acute-on-chronic liver failure (ACLF). Finally, we analyzed portal and hepatic vein levels of MCP-1 in patients receiving transjugular intrahepatic portosystemic shunt insertion for complications of portal hypertension. Results: In this mouse model of fibrotic hepatitis, hepatic expression of CCL2 (P = 0.009) and the amount of F4/80 positive cells in the liver (P < 0.001) significantly increased after induction of hepatitis by CCl4 compared to control animals. Moreover, strong correlation of hepatic CCL2 expression and F4/80 positive cells were seen (P = 0.023). Furthermore, in human liver explants, hepatic transcription levels of CCL2 correlated with the MELD score of the patients, and thus disease severity (P = 0.007). The experimental model of ACLF in rats revealed significantly higher levels of MCP-1 plasma (P = 0.028) and correlation of hepatic CCL2 expression (R = 0.69, P = 0.003). Particularly, plasma MCP-1 levels did not correlate with peripheral blood monocyte CCL2 expression. Finally, higher levels of MCP-1 were observed in the hepatic compared to the portal vein (P = 0.01) in patients receiving TIPS. Similarly, a positive correlation of MCP-1 with Child-Pugh score was observed (P = 0.018). Further, in the presence of ACLF, portal and hepatic vein levels of MCP-1 were significantly higher compared to patients without ACLF (both P = 0.039). Conclusion: Circulating levels of MCP-1 mainly derive from the injured liver and are associated with severity of liver disease. Therefore, liver macrophages contribute significantly to disease progression. Circulating MCP-1 may reflect the extent of hepatic macrophage activation.


Subject(s)
Chemical and Drug Induced Liver Injury/immunology , Chemokine CCL2/blood , Liver Cirrhosis, Experimental/complications , Liver/immunology , Macrophage Activation , Acute-On-Chronic Liver Failure/complications , Acute-On-Chronic Liver Failure/immunology , Animals , Chemokine CCL2/analysis , Chemokine CCL2/genetics , Chemokine CCL2/physiology , Kupffer Cells/physiology , Liver Cirrhosis, Experimental/immunology , Male , Mice , Mice, Inbred C57BL
3.
Front Physiol ; 10: 326, 2019.
Article in English | MEDLINE | ID: mdl-30971954

ABSTRACT

Platelet factor 4 (PF4) is a pleiotropic inflammatory chemokine, which has been implicated in various inflammatory disorders including liver fibrosis. However, its role in acute liver diseases has not yet been elucidated. Here we describe an unexpected, anti-inflammatory role of PF4. Serum concentrations of PF4 were measured in patients and mice with acute liver diseases. Acute liver injury in mice was induced either by carbon tetrachloride or by D-galactosamine hydrochloride and lipopolysaccharide. Serum levels of PF4 were decreased in patients and mice with acute liver diseases. PF4-/- mice displayed increased liver damage in both models compared to control which was associated with increased apoptosis of hepatocytes and an enhanced pro-inflammatory response of liver macrophages. In this experimental setting, PF4-/- mice were unable to generate activated Protein C (APC), a protein with anti-inflammatory activities on monocytes/macrophages. In vitro, PF4 limited the activation of liver resident macrophages. Hence, the systemic application of PF4 led to a strong amelioration of experimental liver injury. Along with reduced liver injury, PF4 improved the severity of the pro-inflammatory response of liver macrophages and induced increased levels of APC. PF4 has a yet unidentified direct anti-inflammatory effect in two models of acute liver injury. Thus, attenuation of acute liver injury by systemic administration of PF4 might offer a novel therapeutic approach for acute liver diseases.

4.
Hepatol Commun ; 2(2): 152-154, 2018 02.
Article in English | MEDLINE | ID: mdl-29404523

ABSTRACT

Benign recurrent intrahepatic cholestasis (BRIC) is a peculiar familial disease caused by mutations of the genes encoding hepatocanalicular flippase for phosphatidylserine (ATP8B1; BRIC type 1) or the bile salt export pump (ABCB11; BRIC type 2). Here, we report on a patient with nasobiliary drainage-refractory BRIC type 2 who improved under plasma separation and anion absorption therapy. We also suggest that nasobiliary drainage might be an ineffective approach in carriers of severe loss-of-function mutations of the bile salt export pump ABCB11. (Hepatology Communications 2018;2:152-154).

5.
Liver Int ; 38(5): 875-884, 2018 05.
Article in English | MEDLINE | ID: mdl-29105936

ABSTRACT

BACKGROUND & AIMS: CXCR% ligands play an important role in hepatic injury, inflammation and fibrosis. While CXCL9 and CXCL11 are associated with survival in patients receiving transjugular intrahepatic portosystemic shunt (TIPS), the role of CXCL10 in severe portal hypertension remains unknown. METHODS: A total of 89 cirrhotic patients were analysed. CXCL10 protein levels were measured in portal and hepatic blood at TIPS insertion and 2 weeks later in 24 patients. CXCL10 and IL8 levels were assessed in portal, hepatic, cubital vein and right atrium blood in a further 25 patients at TIPS insertion. Furthermore, real-time PCR determined hepatic CXCL10-mRNA in 40 cirrhotic patients. RESULTS: Hepatic CXCL10 showed no association with decompensation. By contrast, circulating CXCL10-levels were higher in portal than in hepatic vein blood, suggesting an extrahepatic source of CXCL10 in cirrhosis. However, CXCL10 protein in blood samples from portal, hepatic, cubital veins and right atrium correlated excellently with each other and with IL-8 levels. Higher CXCL10 circulating levels were associated with presence of ascites and higher Child scores. Higher CXCL10 circulating protein levels were associated with acute decompensation, acute-on-chronic liver failure (ACLF) and independently with mortality. Moreover, a decrease in CXCL10 protein levels after TIPS insertion was associated with better survival in each cohort and analysed together. DISCUSSION: Circulating CXCL10 possibly reflects systemic inflammation and it is correlated with acute decompensation, ACLF and complications in patients with severe portal hypertension receiving TIPS. CXCL10 predicts survival in these patients and a decrease in CXCL10 after TIPS may be considered a good prognostic factor.


Subject(s)
Acute-On-Chronic Liver Failure/diagnosis , Chemokine CXCL10/blood , Hypertension, Portal/diagnosis , Liver Cirrhosis/surgery , Portasystemic Shunt, Transjugular Intrahepatic/adverse effects , Acute-On-Chronic Liver Failure/blood , Adult , Aged , Biomarkers/blood , Female , Humans , Hypertension, Portal/blood , Inflammation/metabolism , Liver Cirrhosis/complications , Male , Middle Aged , Portasystemic Shunt, Transjugular Intrahepatic/mortality , ROC Curve , Retrospective Studies , Survival Analysis
6.
Liver Int ; 36(3): 386-94, 2016 Mar.
Article in English | MEDLINE | ID: mdl-26212075

ABSTRACT

BACKGROUND & AIMS: Chemokines, such as CXCR3-ligands, have been identified to play an important role during hepatic injury, inflammation and fibrosis. While CXCL9 is associated with survival in patients receiving transjugular intrahepatic portosystemic shunt (TIPS), the role of CXCL11 in severe portal hypertension remains unknown. METHODS: CXCL11-levels were measured in 136 patients with liver diseases, and 63 healthy controls. In further 47 cirrhotic patients receiving TIPS, CXCL11 levels were measured in portal and hepatic veins at TIPS insertion by cytometric bead array. CXCL11-levels were measured in 23 patients in cubital vein and right atrium, whereas in 24 patients in portal and hepatic blood at an invasive reevaluation. RESULTS: CXCL11-levels were increased with the severity of liver fibrosis. CXCL11-levels from portal, hepatic and cubital veins and right atrium showed a highly significant correlation among each other in these patients. Furthermore, levels of CXCL11 from the right atrium were significantly higher than those from cubital vein. Interestingly, patients with alcoholic cirrhosis had significantly lower CXCL11-levels, than other aetiologies of cirrhosis. After TIPS, CXCL11 levels correlated with the degree of portal pressure and patients with higher CXCL11-levels in portal and hepatic veins showed higher mortality. Multivariate analysis revealed hepatic CXCL11-levels before TIPS, creatinine and age as independent predictors for survival in TIPS patients, whereas MELD score and low portal CXCL11-levels after TIPS predicted long-term survival. CONCLUSION: CXCL11 levels are mainly increased in patients with non-alcoholic cirrhosis and high portal pressure. Moreover, levels of CXCL11 might predict long-time survival of cirrhotic patients bearing TIPS.


Subject(s)
Chemokine CXCL11/blood , Hypertension, Portal/surgery , Liver Cirrhosis/complications , Portasystemic Shunt, Transjugular Intrahepatic , Adult , Aged , Biomarkers/blood , Case-Control Studies , Female , Humans , Hypertension, Portal/blood , Hypertension, Portal/etiology , Hypertension, Portal/mortality , Kaplan-Meier Estimate , Liver Cirrhosis/diagnosis , Liver Cirrhosis/mortality , Male , Middle Aged , Multivariate Analysis , Portasystemic Shunt, Transjugular Intrahepatic/adverse effects , Portasystemic Shunt, Transjugular Intrahepatic/mortality , Predictive Value of Tests , Proportional Hazards Models , Risk Factors , Severity of Illness Index , Time Factors , Treatment Outcome
7.
Medicine (Baltimore) ; 94(31): e1265, 2015 Aug.
Article in English | MEDLINE | ID: mdl-26252289

ABSTRACT

Chronic intestinal pseudoobstruction (CIP) can be a severe burden and even a life-threatening disorder. Typically, several years of uncertainty are passing before diagnosis. We are reporting the case of a young woman with a decade of severe, progressive gastrointestinal dysmotility. Unusually, she had also developed an autonomic neuropathy, and a stiff limb syndrome.In addition to achalasia and CIP the young woman also developed neuropathic symptoms: orthostatic intolerance, urinary retention, a Horner syndrome, and lower limb stiffness. Careful interdisciplinary diagnostics excluded underlying infectious, rheumatoid, metabolic or tumorous diseases.The detection of GAD (glutamic acid decarboxylase) antibodies, however, seemed to link CIP, autonomic neuropathy, and limb stiffness and pointed at an autoimmune origin of our patient's complaints. This was supported by the positive effects of intravenous immunoglobulin. In response to this therapy the body weight had stabilized, orthostatic tolerance had improved, and limb stiffness was reversed.The case suggested that GAD antibodies should be considered in CIP also in nondiabetic patients. This may support earlier diagnosis and immunotherapy.


Subject(s)
Antibodies/blood , Autonomic Nervous System Diseases/etiology , Glutamate Decarboxylase/immunology , Intestinal Pseudo-Obstruction/diagnosis , Intestinal Pseudo-Obstruction/etiology , Muscle Rigidity/etiology , Adult , Autonomic Nervous System Diseases/diagnosis , Autonomic Nervous System Diseases/therapy , Chronic Disease , Female , Humans , Intestinal Pseudo-Obstruction/therapy , Muscle Rigidity/diagnosis , Muscle Rigidity/therapy
8.
J Hepatol ; 62(2): 332-9, 2015 Feb.
Article in English | MEDLINE | ID: mdl-25457205

ABSTRACT

BACKGROUND & AIMS: Inflammation, collagen deposition and tissue remodelling are involved in the pathogenesis and complications of cirrhosis with portal hypertension. CXCL9 and other chemokines play an important role in these processes and have been associated with liver injury and complications of liver disease in humans. However, their predictive value in patients with cirrhosis and portal hypertension remains to be established. METHODS: 103 patients with liver cirrhosis who had received TIPS (transjugular intrahepatic portosystemic shunt) were included into this study. The TIPS indication was either refractory ascites or recurrent bleeding. Before and after the TIPS procedure portal and hepatic venous blood samples were obtained in 78 patients. In 25 patients blood samples were obtained from the portal vein, hepatic vein, right atrium and cubital vein at TIPS insertion. Serum levels of CXCL9 were measured by cytometric bead array and correlated with clinical parameters and overall outcome. RESULTS: Portal venous levels of CXCL9 decreased after TIPS. Child-Pugh score, refractory ascites, renal dysfunction and alcoholic aetiology of cirrhosis were associated with increased CXCL9 levels. Importantly, low levels of CXCL9 in portal and hepatic vein samples were prognostic factors for the survival of patients receiving TIPS during long-time follow-up. CONCLUSIONS: The CXCR3 ligand CXCL9 affects the liver and/or is released by the liver and thereby might contribute to hepatic and extrahepatic organ dysfunction. Elevated levels of CXCL9 are associated with shorter survival in cirrhotic patients with severe portal hypertension receiving TIPS. This chemokine should be further evaluated as a novel biomarker for the outcome in patients with cirrhosis and portal hypertension and its modulation as a new therapeutic strategy.


Subject(s)
Chemokine CXCL9/blood , Hypertension, Portal/surgery , Liver Cirrhosis/blood , Portasystemic Shunt, Transjugular Intrahepatic/methods , Adult , Aged , Biomarkers/blood , Female , Follow-Up Studies , Humans , Hypertension, Portal/blood , Hypertension, Portal/etiology , Liver Cirrhosis/complications , Male , Middle Aged , Prognosis , Retrospective Studies , Time Factors , Treatment Outcome
9.
FASEB J ; 28(12): 5136-47, 2014 Dec.
Article in English | MEDLINE | ID: mdl-25122558

ABSTRACT

MIF is an inflammatory cytokine but is hepatoprotective in models of hepatotoxin-induced liver fibrosis. Hepatic fibrosis can also develop from metabolic liver disease, such as nonalcoholic fatty liver disease (NASH). We investigated the role of MIF in high-fat or methionine- and choline-deficient diet mouse models of NASH. Mif(-/-) mice showed elevated liver triglyceride levels (WT, 53±14 mg/g liver; Mif(-/-), 103±7 mg/g liver; P<0.05) and a 2-3-fold increased expression of lipogenic genes. Increased fatty degeneration in the livers of Mif(-/-) mice was associated with increased hepatic inflammatory cells (1.6-fold increase in F4/80(+) macrophages) and proinflammatory cytokines (e.g., 2.3-fold increase in Tnf-α and 2-fold increase in Il-6 expression). However, inflammatory cells and cytokines were decreased by 50-90% in white adipose tissue (WAT) of Mif(-/-) mice. Subset analysis showed that macrophage phenotypes in livers of Mif(-/-) mice were skewed toward M2 (e.g., 1.7-fold and 2.5-fold increase in Arg1 and Il-13, respectively, and 2.5-fold decrease in iNos), whereas macrophages were generally reduced in WAT of these mice (70% reduction in mRNA expression of F4/80(+) macrophages). The protective MIF effect was scrutinized in isolated hepatocytes. MIF reversed inflammation-induced triglyceride accumulation in Hepa1-6 cells and primary hepatocytes and also attenuated oleic acid-elicited triglyceride increase in 3T3-L1 adipocytes. Protection from fatty hepatocyte degeneration was paralleled by a 2- to 3-fold reduction by MIF of hepatocyte proinflammatory cytokine production. Blockade of MIF receptor cluster of differentiation 74 (CD74) but not of CXCR2 or CXCR4 fully reverted the protective effect of MIF, comparable to AMPK inhibition. In summary, we demonstrate that MIF mediates hepatoprotection through the CD74/AMPK pathway in hepatocytes in metabolic models of liver injury.


Subject(s)
Macrophage Migration-Inhibitory Factors/physiology , Non-alcoholic Fatty Liver Disease/prevention & control , Animals , Base Sequence , DNA Primers , Disease Models, Animal , Macrophage Migration-Inhibitory Factors/genetics , Mice , Mice, Knockout , Reverse Transcriptase Polymerase Chain Reaction
10.
PLoS One ; 8(6): e66106, 2013.
Article in English | MEDLINE | ID: mdl-23799074

ABSTRACT

Liver fibrosis is associated with infiltrating immune cells and activation of hepatic stellate cells. We here aimed to investigate the effects of the CC chemokine CCL3, also known as macrophage inflammatory protein-1α, in two different fibrosis models. To this end, we treated mice either with carbon tetrachloride or with a methionine- and choline-deficient diet to induce fibrosis in CCL3 deficient and wild-type mice. The results show that the protein expression of CCL3 is increased in wild-type mice after chronic liver injury. Deletion of CCL3 exhibited reduced liver fibrosis compared to their wild-type counterparts. We could validate these results by treating the two mouse groups with either carbon tetrachloride or by feeding a methionine- and choline-deficient diet. In these models, lack of CCL3 is functionally associated with reduced stellate cell activation and liver immune cell infiltration. In vitro, we show that CCL3 leads to increased proliferation and migration of hepatic stellate cells. In conclusion, our results define the chemokine CCL3 as a mediator of experimental liver fibrosis. Thus, therapeutic modulation of CCL3 might be a promising target for chronic liver diseases.


Subject(s)
Chemokine CCL3/physiology , Liver Cirrhosis/physiopathology , Animals , Carbon Tetrachloride/administration & dosage , Cell Proliferation , Enzyme-Linked Immunosorbent Assay , Lipogenesis , Liver Cirrhosis/pathology , Male , Mice , Mice, Inbred BALB C , Mice, Knockout , T-Lymphocytes/cytology
11.
Int J Clin Exp Pathol ; 6(4): 678-85, 2013.
Article in English | MEDLINE | ID: mdl-23573314

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 CCl(4)) 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.


Subject(s)
Chemokine CCL5/antagonists & inhibitors , Liver Cirrhosis/pathology , Liver Cirrhosis/prevention & control , Monocytes/drug effects , Monocytes/pathology , Receptors, CCR/antagonists & inhibitors , Animals , Carbon Tetrachloride/adverse effects , Cell Count , Chemokine CCL5/drug effects , Cytokines/metabolism , Disease Models, Animal , Liver/drug effects , Liver/metabolism , Liver/pathology , Liver Cirrhosis/chemically induced , Mice , Mice, Inbred C57BL , Nitric Oxide Synthase Type II/metabolism , Receptors, CCR/drug effects
12.
PLoS One ; 8(2): e56116, 2013.
Article in English | MEDLINE | ID: mdl-23437087

ABSTRACT

Hepatic stellate cells (HSCs) play a major role in the pathogenesis of liver fibrosis. Working on primary HSCs requires difficult isolation procedures; therefore we have generated and here characterize a mouse hepatic stellate cell line expressing GFP under control of the collagen 1(I) promoter/enhancer. These cells are responsive to pro-fibrogenic stimuIi, such as PDGF or TGF-ß1, and are able to activate intracellular signalling pathways including Smads and MAP kinases. Nevertheless, due to the basal level of activation, TGF-ß1 did not significantly induce GFP expression contrasting the TGF-ß1 regulated endogenous collagen I expression. We could demonstrate that the accessory TGF-ß-receptor endoglin, which is endogenously expressed at very low levels, has a differential effect on signalling of these cells when transiently overexpressed. In the presence of endoglin activation of Smad1/5/8 was drastically enhanced. Moreover, the phosphorylation of ERK1/2 was increased, and the expression of vimentin, α-smooth muscle actin and connective tissue growth factor was upregulated. Endoglin induced a slight increase in expression of the inhibitor of differentiation-2 while the amount of endogenous collagen type I was reduced. Therefore, this profibrogenic cell line with hepatic stellate cell origin is not only a promising novel experimental tool, which can be used in vivo for cell tracing experiments. Furthermore it allows investigating the impact of various regulatory proteins (e.g. endoglin) on profibrogenic signal transduction, differentiation and hepatic stellate cell biology.


Subject(s)
Hepatic Stellate Cells/metabolism , Intracellular Signaling Peptides and Proteins/metabolism , Signal Transduction , Transforming Growth Factor beta1/metabolism , Animals , Becaplermin , Biomarkers/metabolism , Cell Line, Transformed , Collagen/metabolism , Endoglin , Enzyme Activation/drug effects , Extracellular Signal-Regulated MAP Kinases/metabolism , Fluorescence , Green Fluorescent Proteins/metabolism , Hepatic Stellate Cells/drug effects , Hepatic Stellate Cells/enzymology , Hepatic Stellate Cells/pathology , Ligands , Liver Cirrhosis/enzymology , Liver Cirrhosis/pathology , Mice , Mice, Inbred C57BL , Proto-Oncogene Proteins c-sis/pharmacology , Rats , Rats, Sprague-Dawley , Signal Transduction/drug effects
13.
Hepatology ; 57(2): 797-805, 2013 Feb.
Article in English | MEDLINE | ID: mdl-22996399

ABSTRACT

UNLABELLED: Aberrant expression of the chemokine CXC chemokine ligand (CXCL)10 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 proapoptotic effect of CXCL10 in hepatocytes, which is not mediated through its cognate chemokine receptor, but the lipopolysaccharide receptor Toll-like receptor 4 (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 proapoptotic functions of CXCL10 on different liver cell types were evaluated in detail in vitro. The results showed that CXCL10 expression was positively correlated with liver cell apoptosis in humans and mice. Neutralization of CXCL10 ameliorated concanavalin A-induced tissue injury in vivo, which was strongly associated with reduced liver cell apoptosis. In vitro, CXCL10 mediated the apoptosis of hepatocytes involving TLR4, but not CXC chemokine receptor 3 signaling. Specifically, CXCL10 induced long-term protein kinase B and Jun N-terminal kinase activation, leading to hepatocyte apoptosis by caspase-8, caspase-3, and p21-activated kinase 2 cleavage. Accordingly, systemic application of CXCL10 led to TLR4-induced liver cell apoptosis in vivo. CONCLUSION: The results identify CXCL10 and its noncognate receptor, TLR4, as a proapoptotic signaling cascade during liver injury. Antagonism of the CXCL10/TLR4 pathway might be a therapeutic option in liver diseases associated with increased apoptosis.


Subject(s)
Apoptosis/drug effects , Chemokine CXCL10/pharmacology , Hepatocytes/pathology , Toll-Like Receptor 4/physiology , Animals , Carbon Tetrachloride Poisoning/pathology , Caspases/metabolism , Chemical and Drug Induced Liver Injury , Chemokine CXCL10/antagonists & inhibitors , Chemokine CXCL10/biosynthesis , Concanavalin A , Hepatitis C/pathology , Hepatocytes/drug effects , Hepatocytes/physiology , Humans , Liver/pathology , Mice , Receptors, CXCR3/physiology , Signal Transduction/physiology
14.
J Gastroenterol Hepatol ; 28(1): 153-60, 2013 Jan.
Article in English | MEDLINE | ID: mdl-22989351

ABSTRACT

BACKGROUND AND AIM: The development of end-stage graft disease is suspected to be partially determined by an individual genetic background. The aim of our study was to determine the prevalence of YKL-40-gene polymorphism in hepatitis C virus (HCV)-positive patients and its impact on the incidence of acute cellular rejection (ACR), graft fibrosis and antiviral treatment response. METHODS: A total of 149 patients, who underwent liver transplantation for HCV-induced liver disease, were genotyped for YKL-40 (rs4950928; G/C) by TaqMan Genotyping Assay. The results were correlated with 616 post-transplant graft biopsies regarding inflammation, fibrosis and evidence for ACR. RESULTS: No association of YKL-40-genotypes was observed regarding mean inflammation grade (P = 0.216) and antiviral treatment outcome (P = 0.733). However, the development of advanced fibrosis (F3-4) was significantly faster in patients with YKL-40-G-allele: t(CC) = 4.6 versus t(CG/GG) = 2.4 years; P = 0.006. Patients with lower fibrosis (F0-2) compared to advanced fibrosis (F3-4) received significantly more frequent dual immunosuppression (calcineurin inhibitors [CNIs]/mofetile mycophenolate [MMF] vs CNIs; P = 0.003). ACR-occurrence was associated with YKL-40-genotypes (ACR: CC = 60.4%, CG = 25.0% and GG = 14.6% vs non-ACR: CC = 74.2%, CG = 23.8% and GG = 2.0%; P = 0.009) and with gender compatibility between donor and recipient (P = 0.012). CONCLUSION: Fibrosis progression and ACR-incidence after transplantation for HCV-induced liver disease seem to be under genetic control. The negative impact of G-allele on post-transplant events observed in our study, deserves attention and should be verified in larger liver transplantation-cohorts.


Subject(s)
Adipokines/genetics , End Stage Liver Disease/virology , Graft Rejection/genetics , Graft Rejection/immunology , Hepatitis C/genetics , Lectins/genetics , Liver Cirrhosis/genetics , Adult , Antiviral Agents/therapeutic use , Chitinase-3-Like Protein 1 , Cyclosporine/therapeutic use , Disease Progression , Drug Therapy, Combination , End Stage Liver Disease/surgery , Female , Genotype , Graft Rejection/pathology , Hepacivirus , Hepatitis C/complications , Hepatitis C/pathology , Humans , Immunosuppressive Agents/therapeutic use , Interferon alpha-2 , Interferon-alpha/therapeutic use , Liver Cirrhosis/pathology , Liver Transplantation , Male , Middle Aged , Mycophenolic Acid/analogs & derivatives , Mycophenolic Acid/therapeutic use , Polyethylene Glycols/therapeutic use , Polymorphism, Single Nucleotide , Recombinant Proteins/therapeutic use , Ribavirin/therapeutic use , Sex Factors , Statistics, Nonparametric , Tacrolimus/therapeutic use , Time Factors , Young Adult
15.
Channels (Austin) ; 7(1): 38-42, 2013 Jan 01.
Article in English | MEDLINE | ID: mdl-23064163

ABSTRACT

Bile acid-sensitive ion channel (BASIC) is a member of the DEG/ENaC gene family of unknown function. Rat BASIC (rBASIC) is inactive at rest. We have recently shown that cholangiocytes, the epithelial cells lining the bile ducts, are the main site of BASIC expression in the liver and identified bile acids, in particular hyo- and chenodeoxycholic acid, as agonists of rBASIC. Moreover, it seems that extracellular divalent cations stabilize the resting state of rBASIC, because removal of extracellular divalent cations opens the channel. In this addendum, we demonstrate that removal of extracellular divalent cations potentiates the activation of rBASIC by bile acids, suggesting an allosteric mechanism. Furthermore, we show that rBASIC is strongly activated by the anticholestatic bile acid ursodeoxycholic acid (UDCA), suggesting that BASIC might mediate part of the therapeutic effects of UDCA.


Subject(s)
Ion Channels/metabolism , Rats/metabolism , Ursodeoxycholic Acid/metabolism , Animals , Calcium/metabolism , Ion Channels/genetics , Magnesium/metabolism , Rats/genetics
16.
J Hepatol ; 58(2): 335-41, 2013 Feb.
Article in English | MEDLINE | ID: mdl-23041308

ABSTRACT

BACKGROUND & AIMS: After myofibroblastic transdifferentiation, hepatic stellate cells (HSC), mainly involved in liver fibrosis by extracellular matrix production, exhibit an altered growth factor profile including increased expression of neuronal mediators. Here, we analyzed putative targets of neuronal microRNAs miR-9, miR-125b, and miR-128 by deep sequencing of the transcript population, interacting with the miRNA/Argonaute 2 (Ago2) complex in myofibroblastic HSC. METHODS: MicroRNA expression was quantified by real-time PCR in primary HSC, isolated from the rat or human liver. Myofibroblastic HSC were transfected either with mimics or inhibitors of miR-9, miR-125b, and miR-128. RNA from immunoprecipitated Ago2-miRNA/transcript complexes was purified and used for next generation sequencing. Additionally, gene expression was investigated in quiescent and activated primary HSC, treated with the miR-128 mimic or inhibitor, by microarray analysis. RESULTS: During myofibroblastic transdifferentiation of HSC, miR-9, miR-125b, and miR-128 expression was markedly increased. Transcriptome analysis of Ago2 bound mRNA by deep sequencing identified a broad spectrum of transcripts that interact with neuronal miRNAs in myofibroblastic HSC. In particular, in HSC overexpressing miR-128, many members of the chemokine family were bound to the Ago2 repression complex. Furthermore, a comprehensive profiling of gene expression demonstrates the high impact of neuronal miRNAs on the chemokine network. CONCLUSIONS: Ago2 immunoprecipitation followed by deep sequencing is a useful tool to identify novel miRNA targets. Upregulation of neuronal miR-9, miR-125b, and miR-128 during myofibroblastic transition and the identified interaction with a wide range of chemokines and chemokine receptors suggest a prominent role of neuronal miRNAs in the inflammatory response of HSC during fibrosis.


Subject(s)
Argonaute Proteins/genetics , Argonaute Proteins/metabolism , Chemokines/metabolism , Hepatic Stellate Cells/metabolism , MicroRNAs/metabolism , Neurons/metabolism , Transcriptome/genetics , Animals , Cell Differentiation , Cells, Cultured , Disease Models, Animal , Hepatic Stellate Cells/pathology , Humans , Immunoprecipitation , Liver Cirrhosis/metabolism , Liver Cirrhosis/pathology , Myofibroblasts/metabolism , Myofibroblasts/pathology , Rats , Up-Regulation
17.
Am J Pathol ; 182(1): 107-17, 2013 Jan.
Article in English | MEDLINE | ID: mdl-23141925

ABSTRACT

Platelet-derived growth factors (PDGF) are key mediators of organ fibrosis. We investigated whether PDGF-C(-/-) mice or mice treated with neutralizing PDGF-C antibodies are protected from bile duct ligation-induced liver fibrosis, and we compared the effects with those of PDGF-C deficiency or neutralization on kidney fibrosis induced by unilateral ureteral obstruction. Unexpectedly, and in contrast to kidney fibrosis, PDGF-C deficiency or antagonism did not protect from liver fibrosis or functional liver impairment. Furthermore, the hepatic infiltration of monocytes/macrophages/dendritic cells and chemokine mRNA expression (CC chemokine ligand [CCL]5, CCL2, and CC chemokine receptor 2 [CCR2]) remained unchanged. Transcript expression of PDGF ligands increased in both liver and kidney fibrosis and was not affected by neutralization of PDGF-C. In kidney fibrosis, PDGF-C deficiency or antagonism led to reduced expression and signaling of PDGF-receptor (R)-α- and PDGFR-ß-chains. In contrast, in liver fibrosis there was either no difference (PDGF-C(-/-) mice) or even an upregulation of PDGFR-ß and signaling (anti-PDGF-C group). Finally, in vitro studies in portal myofibroblasts pointed to a predominant role of PDGF-B and PDGF-D signaling in liver fibrosis. In conclusion, our study revealed significant differences between kidney and liver fibrosis in that PDGF-C mediates kidney fibrosis, whereas antagonism of PDGF-C in liver fibrosis appears to be counteracted by significant upregulation and increased PDGFR-ß signaling. PDGF-C antagonism, therefore, may not be effective to treat liver fibrosis.


Subject(s)
Kidney/pathology , Liver Cirrhosis/metabolism , Lymphokines/physiology , Platelet-Derived Growth Factor/physiology , Receptors, Platelet-Derived Growth Factor/physiology , Animals , Cells, Cultured , Disease Models, Animal , Fibrosis , Liver Cirrhosis/etiology , Liver Cirrhosis/prevention & control , Lymphokines/antagonists & inhibitors , Lymphokines/deficiency , Male , Mice , Mice, Inbred C57BL , Mice, Knockout , Myofibroblasts/metabolism , Platelet-Derived Growth Factor/antagonists & inhibitors , Platelet-Derived Growth Factor/deficiency , Rats , Rats, Sprague-Dawley , Receptors, Platelet-Derived Growth Factor/metabolism , Signal Transduction/physiology , Up-Regulation/physiology , Ureteral Obstruction/complications
18.
Biochim Biophys Acta ; 1832(7): 1041-8, 2013 Jul.
Article in English | MEDLINE | ID: mdl-23159607

ABSTRACT

Fibrosis or scarring of diverse organs and tissues is considered as a pathologic consequence of a chronically altered wound healing response which is tightly linked to inflammation and angiogenesis. The recruitment of immune cells, local proliferation of fibroblasts and the consecutive accumulation of extracellular matrix proteins are common pathophysiological hallmarks of tissue fibrosis, irrespective of the organ involved. Chemokines, a family of chemotactic cytokines, appear to be central mediators of the initiation as well as progression of these biological processes. Traditionally chemokines have only been considered to play a critical role in orchestrating the influx of immune cells to sites of tissue injury. However, within the last years, further aspects of chemokine biology including fibroblast activation and angiogenesis have been deciphered in tissue fibrosis of many different organs. Interestingly, certain chemokines appear to mediate common effects in liver, kidney, lung, and skin of various animal models, while others mediate tissue specific effects. These aspects have to be kept in mind when extrapolating data of animal studies to early human trials. Nevertheless, the further understanding of chemokine effects in tissue fibrosis might be an attractive approach for identifying novel therapeutic targets in chronic organ damage associated with high morbidity and mortality. This article is part of a Special Issue entitled: Fibrosis: Translation of basic research to human disease.


Subject(s)
Chemokines , Fibrosis , Animals , Chemokines/metabolism , Fibroblasts/metabolism , Humans , Liver Cirrhosis/metabolism
19.
Lab Invest ; 92(12): 1726-37, 2012 Dec.
Article in English | MEDLINE | ID: mdl-22986785

ABSTRACT

The prognosis of liver failure is often determined by infectious and cholestatic complications. As HGF/c-Met and interleukin (IL)-6/gp130 control hepatic cytoprotective pathways, we here investigated their cooperative role during the onset of cholestatic liver injury. Conditional hepatocyte-specific ((Δhepa)) c-Met, gp130 and c-Met/gp130 knockout mice (Cre-loxP system) were subjected to bile duct ligation (BDL) and lipopolysaccharide (LPS) stimulation. gp130(Δhepa) and c-Met/gp130(Δhepa) mice displayed increased lethality associated with severe bacteraemia early after BDL, whereas c-Met(Δhepa) and wild-type mice showed normal survival. Analysis of the innate immune response and the regulation of hepatic antibacterial pathways showed that the LPS-triggered hepatocellular response via the Toll-like receptor-4 pathway was regulated differentially by HGF/c-Met and IL-6/gp130. Activation of p38MAPK, c-Jun N-terminal kinase and signalling transducer and activator of transcription-3 was impaired in gp130(Δ) and c-Met(Δhepa) livers. In addition, the acute-phase response (APR) was reduced in c-Met(Δhepa) livers, whereas gp130(Δhepa) displayed a completely abolished APR. In contrast, TNF-α-dependent NF-κB activation was enhanced in gp130(Δhepa) and c-Met(Δhepa) mice and it was associated with a higher rate of apoptosis and inflammation. Moreover, expression of the neutrophil produced and secreted cathelin-related antimicrobial peptide and of genes related to the inflammasome complex correlated with the strength of the bacterial infection and with TNF-α expression. In conclusion, Gp130 and c-Met are involved in the hepatic antibacterial and innate immune response, control the APR and thus prevent sepsis and liver injury during cholestatic conditions.


Subject(s)
Bacteremia/metabolism , Bile Ducts/metabolism , Bile Ducts/surgery , Cytokine Receptor gp130/deficiency , Liver/metabolism , Proto-Oncogene Proteins c-met/deficiency , Acute-Phase Reaction/metabolism , Animals , Antimicrobial Cationic Peptides , Apoptosis/physiology , Bacteremia/microbiology , Bacterial Load , Bile Ducts/microbiology , Cathelicidins/genetics , Cathelicidins/metabolism , Cell Proliferation , Cholestasis/metabolism , Cytokine Receptor gp130/genetics , Cytokine Receptor gp130/metabolism , Immunity, Innate/physiology , Kaplan-Meier Estimate , Ki-67 Antigen/genetics , Ki-67 Antigen/metabolism , Ligation , Lipopolysaccharides/pharmacology , Liver/injuries , Liver/microbiology , Liver/pathology , Mice , Mice, Knockout , Myeloid Differentiation Factor 88/genetics , Myeloid Differentiation Factor 88/metabolism , Proto-Oncogene Proteins c-met/genetics , Proto-Oncogene Proteins c-met/metabolism , Toll-Like Receptor 4/genetics , Toll-Like Receptor 4/metabolism
20.
Antivir Ther ; 17(6): 1049-58, 2012.
Article in English | MEDLINE | ID: mdl-22892524

ABSTRACT

BACKGROUND: Tenofovir disoproxil fumarate (TDF), an acyclic nucleotide analogue was shown to be effective in many HBV-infected patients with resistance to adefovir dipivoxil (ADV). This observation is intriguing because in vitro studies show that HBV mutations selected by ADV confer cross-resistance to TDF. To assess the clinical relevance of this cross-resistance, we studied the evolution of HBV polymerase gene variants in patients with genotypic resistance against ADV (rtN236T and/or rtA181V/T) during TDF treatment. METHODS: In 10 HBV-monoinfected patients (9 male, mean age 47 ±11 [range 27-67] years, 6 hepatitis B e antigen-positive) with virological breakthrough during ADV treatment associated with the mutations rtN236T and/or rtA181T/V, HBV polymerase gene variants were studied during up to 24 months of consecutive monotherapy with TDF by population sequencing, line probe assay and clonal analysis. RESULTS: In all patients, switching to TDF resulted in a continuous reduction of HBV DNA from a median of 7.6 (4.6-9.4) log(10) copies/ml to 3.3 (2-5) log(10) copies/ml, remaining in 7 patients >400 copies/ml at 12 months. ADV-resistance mutations remained detectable throughout the whole observation period in most patients. Apart from an M204Q mutation in one sample, no new HBV polymerase gene mutations were found. In two patients with low level viraemia after 72 weeks of TDF, adding lamivudine led to a complete response within a few weeks. CONCLUSIONS: ADV-resistant HBV variants may further become selected during TDF treatment, however they cause only a mild decrease in TDF susceptibility.


Subject(s)
Adenine/analogs & derivatives , Antiviral Agents/pharmacology , Evolution, Molecular , Gene Products, pol/genetics , Hepatitis B/drug therapy , Organophosphonates/pharmacology , Adenine/pharmacology , Adult , Aged , Cloning, Molecular , DNA, Viral/blood , Drug Resistance, Viral , Drug Substitution , Escherichia coli/genetics , Escherichia coli/metabolism , Female , Follow-Up Studies , Gene Products, pol/metabolism , Hepatitis B/metabolism , Hepatitis B/virology , Hepatitis B e Antigens/blood , Hepatitis B virus/drug effects , Hepatitis B virus/genetics , Hepatitis B virus/pathogenicity , Humans , Lamivudine/pharmacology , Male , Middle Aged , Mutation , Tenofovir , Viremia/drug therapy , Viremia/metabolism , Viremia/virology
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