title: CYLD deletion triggers nuclear factor-κB-signaling and increases cell death resistance in murine hepatocytes.
abstract: OBJECTIVE
  To analyze the role of CYLD for receptor-mediated cell death of murine hepatocytes in acute liver injury models.
  
  
  METHODS
  Hepatocyte cell death in CYLD knockout mice (CYLD(-/-) ) was analyzed by application of liver injury models for CD95- (Jo2) and tumor necrosis factor (TNF)-α- [D-GalN/lipopolysaccharide (LPS)] induced apoptosis. Liver injury was assessed by measurement of serum transaminases and histological analysis. Apoptosis induction was quantified by cleaved PARP staining and Western blotting of activated caspases. Nuclear factor (NF)-κB, ERK, Akt and jun amino-terminal kinases signaling were assessed. Primary Hepatocytes were isolated by two step-collagenase perfusion and treated with recombinant TNF-α and with the CD95-ligand Jo2. Cell viability was analyzed by MTT-assay.
  
  
  RESULTS
  Livers of CYLD(-/-) mice showed increased anti-apoptotic NF-κB signaling. In both applied liver injury models CYLD(-/-) mice showed a significantly reduced apoptosis sensitivity. After D-GalN/LPS treatment CYLD(-/-) mice exhibited significantly lower levels of alanine aminotransferase (ALT) (295 U/L vs 859 U/L, P < 0.05) and aspartate aminotransferase (AST) (560 U/L vs 1025 U/L, P < 0.01). After Jo injection CYLD(-/-) mice showed 2-fold lower ALT (50 U/L vs 110 U/L, P < 0.01) and lower AST (250 U/L vs 435 U/L, P < 0.01) serum-levels compared to WT mice. In addition, isolated CYLD(-/-) primary murine hepatocytes (PMH) were less sensitive towards death receptor-mediated apoptosis and showed increased levels of Bcl-2, XIAP, cIAP1/2, survivin and c-FLIP expression upon TNF- and CD95-receptor triggering, respectively. Inhibition of NF-κB activation by the inhibitor of NF-κB phosphorylation inhibitor BAY 11-7085 inhibited the expression of anti-apoptotic proteins and re-sensitized CYLD(-/-) PMH towards TNF- and CD95-receptor mediated cell death.
  
  
  CONCLUSIONS
  CYLD is a central regulator of apoptotic cell death in murine hepatocytes by controlling NF-κB dependent anti-apoptotic signaling.
authors.affiliation: Toni Urbanik, Bruno Christian Koehler, Laura Wolpert, Christin Elßner, Anna-Lena Scherr, Nicole Kautz, Stefan Welte, Dirk Jäger, Henning Schulze-Bergkamen, National Center for Tumor Diseases, Department of Medical Oncology, University Clinic of Heidelberg, 69120 Heidelberg, Baden-Wuerrtemberg, Germany.
authors.forename: Toni
authors.initials: T
authors.lastname: Urbanik
authors.affiliation: Toni Urbanik, Bruno Christian Koehler, Laura Wolpert, Christin Elßner, Anna-Lena Scherr, Nicole Kautz, Stefan Welte, Dirk Jäger, Henning Schulze-Bergkamen, National Center for Tumor Diseases, Department of Medical Oncology, University Clinic of Heidelberg, 69120 Heidelberg, Baden-Wuerrtemberg, Germany.
authors.forename: Bruno Christian
authors.initials: BC
authors.lastname: Koehler
authors.affiliation: Toni Urbanik, Bruno Christian Koehler, Laura Wolpert, Christin Elßner, Anna-Lena Scherr, Nicole Kautz, Stefan Welte, Dirk Jäger, Henning Schulze-Bergkamen, National Center for Tumor Diseases, Department of Medical Oncology, University Clinic of Heidelberg, 69120 Heidelberg, Baden-Wuerrtemberg, Germany.
authors.forename: Laura
authors.initials: L
authors.lastname: Wolpert
authors.affiliation: Toni Urbanik, Bruno Christian Koehler, Laura Wolpert, Christin Elßner, Anna-Lena Scherr, Nicole Kautz, Stefan Welte, Dirk Jäger, Henning Schulze-Bergkamen, National Center for Tumor Diseases, Department of Medical Oncology, University Clinic of Heidelberg, 69120 Heidelberg, Baden-Wuerrtemberg, Germany.
authors.forename: Christin
authors.initials: C
authors.lastname: Elßner
authors.affiliation: Toni Urbanik, Bruno Christian Koehler, Laura Wolpert, Christin Elßner, Anna-Lena Scherr, Nicole Kautz, Stefan Welte, Dirk Jäger, Henning Schulze-Bergkamen, National Center for Tumor Diseases, Department of Medical Oncology, University Clinic of Heidelberg, 69120 Heidelberg, Baden-Wuerrtemberg, Germany.
authors.forename: Anna-Lena
authors.initials: AL
authors.lastname: Scherr
authors.affiliation: Toni Urbanik, Bruno Christian Koehler, Laura Wolpert, Christin Elßner, Anna-Lena Scherr, Nicole Kautz, Stefan Welte, Dirk Jäger, Henning Schulze-Bergkamen, National Center for Tumor Diseases, Department of Medical Oncology, University Clinic of Heidelberg, 69120 Heidelberg, Baden-Wuerrtemberg, Germany.
authors.forename: Thomas
authors.initials: T
authors.lastname: Longerich
authors.affiliation: Toni Urbanik, Bruno Christian Koehler, Laura Wolpert, Christin Elßner, Anna-Lena Scherr, Nicole Kautz, Stefan Welte, Dirk Jäger, Henning Schulze-Bergkamen, National Center for Tumor Diseases, Department of Medical Oncology, University Clinic of Heidelberg, 69120 Heidelberg, Baden-Wuerrtemberg, Germany.
authors.forename: Nicole
authors.initials: N
authors.lastname: Kautz
authors.affiliation: Toni Urbanik, Bruno Christian Koehler, Laura Wolpert, Christin Elßner, Anna-Lena Scherr, Nicole Kautz, Stefan Welte, Dirk Jäger, Henning Schulze-Bergkamen, National Center for Tumor Diseases, Department of Medical Oncology, University Clinic of Heidelberg, 69120 Heidelberg, Baden-Wuerrtemberg, Germany.
authors.forename: Stefan
authors.initials: S
authors.lastname: Welte
authors.affiliation: Toni Urbanik, Bruno Christian Koehler, Laura Wolpert, Christin Elßner, Anna-Lena Scherr, Nicole Kautz, Stefan Welte, Dirk Jäger, Henning Schulze-Bergkamen, National Center for Tumor Diseases, Department of Medical Oncology, University Clinic of Heidelberg, 69120 Heidelberg, Baden-Wuerrtemberg, Germany.
authors.forename: Nadine
authors.initials: N
authors.lastname: Hövelmeyer
authors.affiliation: Toni Urbanik, Bruno Christian Koehler, Laura Wolpert, Christin Elßner, Anna-Lena Scherr, Nicole Kautz, Stefan Welte, Dirk Jäger, Henning Schulze-Bergkamen, National Center for Tumor Diseases, Department of Medical Oncology, University Clinic of Heidelberg, 69120 Heidelberg, Baden-Wuerrtemberg, Germany.
authors.forename: Dirk
authors.initials: D
authors.lastname: Jäger
authors.affiliation: Toni Urbanik, Bruno Christian Koehler, Laura Wolpert, Christin Elßner, Anna-Lena Scherr, Nicole Kautz, Stefan Welte, Dirk Jäger, Henning Schulze-Bergkamen, National Center for Tumor Diseases, Department of Medical Oncology, University Clinic of Heidelberg, 69120 Heidelberg, Baden-Wuerrtemberg, Germany.
authors.forename: Ari
authors.initials: A
authors.lastname: Waisman
authors.affiliation: Toni Urbanik, Bruno Christian Koehler, Laura Wolpert, Christin Elßner, Anna-Lena Scherr, Nicole Kautz, Stefan Welte, Dirk Jäger, Henning Schulze-Bergkamen, National Center for Tumor Diseases, Department of Medical Oncology, University Clinic of Heidelberg, 69120 Heidelberg, Baden-Wuerrtemberg, Germany.
authors.forename: Henning
authors.initials: H
authors.lastname: Schulze-Bergkamen
mesh_terms: D000818:Animals; D000906:Antibodies; D017209:Apoptosis / Q000187:drug effects; D015415:Biomarkers / Q000097:blood; D002470:Cell Survival; D002478:Cells, Cultured; D056486:Chemical and Drug Induced Liver Injury / Q000201:enzymology* / Q000209:etiology / Q000235:genetics / Q000276:immunology / Q000473:pathology / Q000517:prevention & control*; D003546:Cysteine Endopeptidases / Q000172:deficiency* / Q000235:genetics; D000074749:Deubiquitinating Enzyme CYLD; D004195:Disease Models, Animal; D005688:Galactosamine; D017353:Gene Deletion; D022781:Hepatocytes / Q000187:drug effects / Q000201:enzymology* / Q000276:immunology / Q000473:pathology; D008070:Lipopolysaccharides; D018345:Mice, Knockout; D016328:NF-kappa B / Q000037:antagonists & inhibitors / Q000378:metabolism*; D018124:Receptors, Tumor Necrosis Factor / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects; D013997:Time Factors; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism; D019014:fas Receptor / Q000276:immunology / Q000378:metabolism
chemical_list: D000906:Antibodies; D015415:Biomarkers; D008070:Lipopolysaccharides; D016328:NF-kappa B; D018124:Receptors, Tumor Necrosis Factor; D014409:Tumor Necrosis Factor-alpha; D019014:fas Receptor; D005688:Galactosamine; C511601:CYLD protein, mouse; D000074749:Deubiquitinating Enzyme CYLD; D003546:Cysteine Endopeptidases
keywords: Apoptosis; CD95; CYLD; Liver; Nuclear factor-κB; Tumor necrosis factor-α