title: Gene network activity in cultivated primary hepatocytes is highly similar to diseased mammalian liver tissue.
abstract: It is well known that isolation and cultivation of primary hepatocytes cause major gene expression alterations. In the present genome-wide, time-resolved study of cultivated human and mouse hepatocytes, we made the observation that expression changes in culture strongly resemble alterations in liver diseases. Hepatocytes of both species were cultivated in collagen sandwich and in monolayer conditions. Genome-wide data were also obtained from human NAFLD, cirrhosis, HCC and hepatitis B virus-infected tissue as well as mouse livers after partial hepatectomy, CCl4 intoxication, obesity, HCC and LPS. A strong similarity between cultivation and disease-induced expression alterations was observed. For example, expression changes in hepatocytes induced by 1-day cultivation and 1-day CCl4 exposure in vivo correlated with R = 0.615 (p < 0.001). Interspecies comparison identified predominantly similar responses in human and mouse hepatocytes but also a set of genes that responded differently. Unsupervised clustering of altered genes identified three main clusters: (1) downregulated genes corresponding to mature liver functions, (2) upregulation of an inflammation/RNA processing cluster and (3) upregulated migration/cell cycle-associated genes. Gene regulatory network analysis highlights overrepresented and deregulated HNF4 and CAR (Cluster 1), Krüppel-like factors MafF and ELK1 (Cluster 2) as well as ETF (Cluster 3) among the interspecies conserved key regulators of expression changes. Interventions ameliorating but not abrogating cultivation-induced responses include removal of non-parenchymal cells, generation of the hepatocytes' own matrix in spheroids, supplementation with bile salts and siRNA-mediated suppression of key transcription factors. In conclusion, this study shows that gene regulatory network alterations of cultivated hepatocytes resemble those of inflammatory liver diseases and should therefore be considered and exploited as disease models.
authors.affiliation: IfADo-Leibniz Research Centre for Working Environment and Human Factors, Technical University of Dortmund, Ardeystrasse 67, 44139, Dortmund, Germany. godoy@ifado.de.|Facultad de Ciencias Biológicas, Departamento de Fisiología, Universidad de Concepción, Concepción, Chile. godoy@ifado.de.
authors.forename: Patricio
authors.initials: P
authors.lastname: Godoy
authors.affiliation: IfADo-Leibniz Research Centre for Working Environment and Human Factors, Technical University of Dortmund, Ardeystrasse 67, 44139, Dortmund, Germany.
authors.forename: Agata
authors.initials: A
authors.lastname: Widera
authors.affiliation: Leibniz Institute for Natural Product Research and Infection Biology eV-Hans-Knöll Institute, Jena, Germany.
authors.forename: Wolfgang
authors.initials: W
authors.lastname: Schmidt-Heck
authors.affiliation: IfADo-Leibniz Research Centre for Working Environment and Human Factors, Technical University of Dortmund, Ardeystrasse 67, 44139, Dortmund, Germany.
authors.forename: Gisela
authors.initials: G
authors.lastname: Campos
authors.affiliation: Molecular Alcohol Research in Gastroenterology, Department of Medicine II, Faculty of Medicine at Mannheim, University of Heidelberg, Mannheim, Germany.
authors.forename: Christoph
authors.initials: C
authors.lastname: Meyer
authors.affiliation: IfADo-Leibniz Research Centre for Working Environment and Human Factors, Technical University of Dortmund, Ardeystrasse 67, 44139, Dortmund, Germany.
authors.forename: Cristina
authors.initials: C
authors.lastname: Cadenas
authors.affiliation: IfADo-Leibniz Research Centre for Working Environment and Human Factors, Technical University of Dortmund, Ardeystrasse 67, 44139, Dortmund, Germany.
authors.forename: Raymond
authors.initials: R
authors.lastname: Reif
authors.affiliation: IfADo-Leibniz Research Centre for Working Environment and Human Factors, Technical University of Dortmund, Ardeystrasse 67, 44139, Dortmund, Germany.
authors.forename: Regina
authors.initials: R
authors.lastname: Stöber
authors.affiliation: IfADo-Leibniz Research Centre for Working Environment and Human Factors, Technical University of Dortmund, Ardeystrasse 67, 44139, Dortmund, Germany.|Molecular Alcohol Research in Gastroenterology, Department of Medicine II, Faculty of Medicine at Mannheim, University of Heidelberg, Mannheim, Germany.|Department of Forensic and Veterinary Toxicology, Faculty of Veterinary Medicine, South Valley University, Qena, Egypt.
authors.forename: Seddik
authors.initials: S
authors.lastname: Hammad
authors.affiliation: IfADo-Leibniz Research Centre for Working Environment and Human Factors, Technical University of Dortmund, Ardeystrasse 67, 44139, Dortmund, Germany.
authors.forename: Larissa
authors.initials: L
authors.lastname: Pütter
authors.affiliation: IfADo-Leibniz Research Centre for Working Environment and Human Factors, Technical University of Dortmund, Ardeystrasse 67, 44139, Dortmund, Germany.
authors.forename: Kathrin
authors.initials: K
authors.lastname: Gianmoena
authors.affiliation: IfADo-Leibniz Research Centre for Working Environment and Human Factors, Technical University of Dortmund, Ardeystrasse 67, 44139, Dortmund, Germany.
authors.forename: Rosemarie
authors.initials: R
authors.lastname: Marchan
authors.affiliation: IfADo-Leibniz Research Centre for Working Environment and Human Factors, Technical University of Dortmund, Ardeystrasse 67, 44139, Dortmund, Germany.|Department of Forensic and Veterinary Toxicology, Faculty of Veterinary Medicine, South Valley University, Qena, Egypt.
authors.forename: Ahmed
authors.initials: A
authors.lastname: Ghallab
authors.affiliation: IfADo-Leibniz Research Centre for Working Environment and Human Factors, Technical University of Dortmund, Ardeystrasse 67, 44139, Dortmund, Germany.
authors.forename: Karolina
authors.initials: K
authors.lastname: Edlund
authors.affiliation: BG Trauma Center, Siegfrid Weller Insitut, Eberhard Karls University Tübingen, Tübingen, Germany.
authors.forename: Andreas
authors.initials: A
authors.lastname: Nüssler
authors.affiliation: Center for Liver Cell Research, Department of General, Visceral, Transplantation, Vascular and Thorax Surgery, Grosshadern Hospital, Ludwig Maximilians University, Munich, Germany.
authors.forename: Wolfgang E
authors.initials: WE
authors.lastname: Thasler
authors.affiliation: Department of General, Visceral and Transplantation Surgery, Charité University Medicine Berlin, Berlin, Germany.
authors.forename: Georg
authors.initials: G
authors.lastname: Damm
authors.affiliation: Department of General, Visceral and Transplantation Surgery, Charité University Medicine Berlin, Berlin, Germany.
authors.forename: Daniel
authors.initials: D
authors.lastname: Seehofer
authors.affiliation: Center for Liver Cell Research, University Children Hospital (KUNO), Regensburg University Hospital, Regensburg, Germany.
authors.forename: Thomas S
authors.initials: TS
authors.lastname: Weiss
authors.affiliation: Institute of Pathology, Friedrich-Schiller-University of Jena, Jena, Germany.
authors.forename: Olaf
authors.initials: O
authors.lastname: Dirsch
authors.affiliation: Experimental Transplantation Surgery, Department of General, Visceral and Vascular Surgery, University Hospital Jena, Friedrich-Schiller-University of Jena, Jena, Germany.
authors.forename: Uta
authors.initials: U
authors.lastname: Dahmen
authors.affiliation: Institute of Biochemistry, Faculty of Medicine, University of Leipzig, Leipzig, Germany.
authors.forename: Rolf
authors.initials: R
authors.lastname: Gebhardt
authors.affiliation: Institute of Neurophysiology, Medical Faculty, University of Cologne, Cologne, Germany.
authors.forename: Umesh
authors.initials: U
authors.lastname: Chaudhari
authors.affiliation: Institute of Neurophysiology, Medical Faculty, University of Cologne, Cologne, Germany.|Department of Developmental Biology, Washington University School of Medicine, St. Louis, MO, USA.
authors.forename: Kesavan
authors.initials: K
authors.lastname: Meganathan
authors.affiliation: Institute of Neurophysiology, Medical Faculty, University of Cologne, Cologne, Germany.
authors.forename: Agapios
authors.initials: A
authors.lastname: Sachinidis
authors.affiliation: InSphero AG, Wagistrasse 27, 8952, Schlieren, Switzerland.
authors.forename: Jens
authors.initials: J
authors.lastname: Kelm
authors.affiliation: Dr. Margarete Fischer-Bosch Institute of Clinical Pharmacology, University of Tuebingen, Auerbachstrasse 112, 70376, Stuttgart, Germany.
authors.forename: Ute
authors.initials: U
authors.lastname: Hofmann
authors.affiliation: Leibniz-Institut für Analytische Wissenschaften - ISAS - e.V., Dortmund, Germany.
authors.forename: René P
authors.initials: RP
authors.lastname: Zahedi
authors.affiliation: Leibniz Institute for Natural Product Research and Infection Biology eV-Hans-Knöll Institute, Jena, Germany.
authors.forename: Reinhard
authors.initials: R
authors.lastname: Guthke
authors.affiliation: Institute of Pathology, Charité-Universitätsmedizin Berlin, Berlin, Germany.
authors.forename: Nils
authors.initials: N
authors.lastname: Blüthgen
authors.affiliation: Molecular Alcohol Research in Gastroenterology, Department of Medicine II, Faculty of Medicine at Mannheim, University of Heidelberg, Mannheim, Germany.
authors.forename: Steven
authors.initials: S
authors.lastname: Dooley
authors.affiliation: IfADo-Leibniz Research Centre for Working Environment and Human Factors, Technical University of Dortmund, Ardeystrasse 67, 44139, Dortmund, Germany. hengstler@ifado.de.
authors.forename: Jan G
authors.initials: JG
authors.lastname: Hengstler
mesh_terms: D000818:Animals; D002478:Cells, Cultured; D053263:Gene Regulatory Networks; D055106:Genome-Wide Association Study; D022781:Hepatocytes / Q000276:immunology / Q000378:metabolism*; D006801:Humans; D008107:Liver Diseases / Q000209:etiology / Q000235:genetics* / Q000276:immunology / Q000473:pathology; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D061251:Primary Cell Culture; D013045:Species Specificity; D059467:Transcriptome
keywords: Bioinformatics; Differentiation; Gene arrays; Inflammation; Metabolism