title: Establishment and Characterization of an SV40 Large T Antigen-Transduced Porcine Colonic Epithelial Cell Line.
abstract: Continuous cell lines have become indispensable tools that have enabled investigations into cellular mechanisms by increasing experimental reproducibility and sample availability, and decreasing the use of experimental animals. To facilitate studies of epithelial barrier function of the porcine colon, we aimed to establish an epithelial cell line with an extended replicative capacity. Cells were isolated from the proximal colon of a 3-week-old piglet and transduced using a recombinant retroviral vector construct containing the simian virus 40 large T antigen (SV40 TAg). We established a clonal epithelial cell line, referred to as PoCo83-3, that stably expressed the SV40 TAg, verified at mRNA and protein levels. PoCo83-3 showed epithelial cell-specific features, such as cobblestone-like morphology, dome structure formation, the presence of apical microvilli, and the expression of keratin 18, E-cadherin and the tight junction-associated proteins zonula occludens-1, occludin, and claudin-1. To validate PoCo83-3 as an in vitro model in epithelial barrier research, proinflammatory cytokine-inducible alterations in barrier integrity were demonstrated by incubating the cells with TNF-α and IFN-γ for 48 h. These cytokine treatments promoted a decreased transepithelial electrical resistance. In summary, PoCo83-3 exhibited an extended life span and a differentiated phenotype while maintaining epithelial characteristics. Based on these results, we present this cell line as a valuable in vitro model for investigations of epithelial barrier function in the porcine colon.
authors.affiliation: Institute of Veterinary Physiology, Faculty of Veterinary Medicine, Leipzig University, Leipzig, Germany.
authors.forename: Bastian
authors.initials: B
authors.lastname: Kaiser
authors.forename: Martina
authors.initials: M
authors.lastname: Böttner
authors.forename: Thilo
authors.initials: T
authors.lastname: Wedel
authors.forename: Ronald M
authors.initials: RM
authors.lastname: Brunner
authors.forename: Tom
authors.initials: T
authors.lastname: Goldammer
authors.forename: Szilvia
authors.initials: S
authors.lastname: Lesko
authors.forename: Gotthold
authors.initials: G
authors.lastname: Gäbel
authors.forename: Alexander
authors.initials: A
authors.lastname: Gleich
authors.forename: Helga
authors.initials: H
authors.lastname: Pfannkuche
mesh_terms: D000818:Animals; D000957:Antigens, Viral, Tumor / Q000235:genetics*; D018929:Cell Culture Techniques / Q000379:methods*; D002460:Cell Line; D002469:Cell Separation / Q000379:methods; D002470:Cell Survival; D002478:Cells, Cultured; D003106:Colon / Q000166:cytology* / Q000378:metabolism; D015925:Cryopreservation / Q000379:methods; D004847:Epithelial Cells / Q000166:cytology* / Q000378:metabolism; D005822:Genetic Vectors / Q000235:genetics; D059785:Karyotype; D008297:Male; D013552:Swine; D014161:Transduction, Genetic
chemical_list: D000957:Antigens, Viral, Tumor
keywords: Cell immortalization; Cell line; Epithelial barrier; Pig; Porcine colon epithelium; Retroviral transduction; Swine; Transepithelial electrical resistance