title: Establishment and characterisation of a novel bovine SV40 large T-antigen-transduced foetal hepatocyte-derived cell line.
abstract: Due to lack of in vitro models for bovine hepatocytes apart from primary cells, there is demand for a bovine hepatocyte-derived cell line. Transduction of bovine foetal hepatocytes with SV40 large T-antigen was performed using the vector pRetro-E2 SV40. Phase contrast microscopy was carried out to evaluate morphology. Immunofluorescence staining was conducted to study expression of keratins, tight junction proteins zona occludens-1 and claudin-1, glucose transporter-2 and P-glycoprotein as well as phosphoenolpyruvate carboxykinase. Urea and triglyceride production was quantified photometrically. Histochemical staining of glycogen by Periodic acid-Schiff stain and of lipids with Oil red O was performed after 24 h incubation with 20 mM glucose and 85 μM palmitic acid, respectively. Gene expression analysis of hepatocyte-typical genes was conducted by reverse transcription PCR. We obtained a SV40LTAg-transduced extended passage cell line, referred to as BFH12. Polygonal growth, keratins, tight junction proteins zona occludens-1 and claudin-1 and glucose transporter-2 as well as P-glycoprotein and phosphoenolpyruvate carboxykinase were attested positively. Urea production calculated as cell-specific rate was 14.2 ± 2.0 fmol/h (early passage) and 17.6 ± 3.7 fmol/h (late passage). Cell-specific triglyceride production was 1.6 ± 0.5 fmol/h (early passage) and 2.1 ± 0.3 fmol/h (late passage). Additionally, cells were positive for glycogen and lipid storage and showed a gene expression pattern resembling foetal hepatocytes. With the properties described here, the novel cell line BFH12 is a hepatocyte-derived cell line which can be used as an in vitro whole cell model.
authors.affiliation: Institute of Veterinary Physiological Chemistry, Faculty of Veterinary Medicine, University of Leipzig, Leipzig, Germany.
authors.forename: Alexander
authors.identifier: http://orcid.org/0000-0002-8584-9155
authors.initials: A
authors.lastname: Gleich
authors.affiliation: Institute of Veterinary Physiology, Faculty of Veterinary Medicine, University of Leipzig, Leipzig, Germany.
authors.forename: Bastian
authors.initials: B
authors.lastname: Kaiser
authors.affiliation: Clinic for Anaesthesiology and Surgical Intensive Care, University Hospital Halle (Saale), Halle (Saale), Germany.
authors.forename: Julia
authors.initials: J
authors.lastname: Schumann
authors.affiliation: Institute of Veterinary Physiological Chemistry, Faculty of Veterinary Medicine, University of Leipzig, Leipzig, Germany. fuhrmann@vmf.uni-leipzig.de.
authors.forename: Herbert
authors.initials: H
authors.lastname: Fuhrmann
mesh_terms: D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000378:metabolism; D000818:Animals; D000952:Antigens, Polyomavirus Transforming / Q000378:metabolism*; D002417:Cattle; D002460:Cell Line; D062445:Claudin-1 / Q000378:metabolism; D033042:Fetal Research; D005333:Fetus / Q000166:cytology*; D005455:Fluorescent Antibody Technique / Q000662:veterinary; D020869:Gene Expression Profiling; D051273:Glucose Transporter Type 2 / Q000378:metabolism; D022781:Hepatocytes / Q000166:cytology* / Q000378:metabolism; D007633:Keratins / Q000378:metabolism; D008858:Microscopy, Phase-Contrast; D010730:Phosphoenolpyruvate Carboxylase / Q000378:metabolism; D016133:Polymerase Chain Reaction; D014161:Transduction, Genetic; D014280:Triglycerides / Q000378:metabolism; D014508:Urea / Q000378:metabolism; D062826:Zonula Occludens-1 Protein / Q000378:metabolism
chemical_list: D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D000952:Antigens, Polyomavirus Transforming; D062445:Claudin-1; D051273:Glucose Transporter Type 2; D014280:Triglycerides; D062826:Zonula Occludens-1 Protein; D007633:Keratins; D014508:Urea; D010730:Phosphoenolpyruvate Carboxylase
keywords: Bovine; Cattle; Cell line; Hepatocytes; SV40 large T-antigen