title: Small molecule and RNAi induced phenotype transition of expanded and primary colonic epithelial cells.
abstract: Recent progress in mammalian intestinal epithelial cell culture led to novel concepts of tissue modeling. Especially the development of phenotypically stable cell lines from individual animals enables an investigation of distinct intestinal loci and disease states. We here report primary and prolonged culture of normal porcine epithelial cells from colon for cell line development. In addition, a novel primary three-dimensional intestinal culture system is presented, which generated organoids composed of a highly polarized epithelial layer lining a core of subepithelial tissue. Cellular characterization of monolayer cell lines revealed epithelial identity and pointed to a proliferative crypt cell phenotype. We evaluated both RNAi and chemical approaches to induce epithelial differentiation in generated cell lines by targeting promoters of epithelial to mesenchymal transition (EMT). By in silico prediction and ectopic expression, miR-147b was proven to be a potent trigger of intestinal epithelial cell differentiation. Our results outline an approach to generate phenotypically stable cell lines expanded from primary colonic epithelial cultures and demonstrate the relevance of miR-147b and chemical inhibitors for promoting epithelial differentiation features.
authors.affiliation: Institute of Veterinary-Biochemistry, Freie Universitaet Berlin, Oertzenweg 19 b, 14163 Berlin, Germany.
authors.forename: Jutta
authors.initials: J
authors.lastname: Sharbati
authors.affiliation: Institute of Veterinary-Biochemistry, Freie Universitaet Berlin, Oertzenweg 19 b, 14163 Berlin, Germany.
authors.forename: Carlos
authors.initials: C
authors.lastname: Hanisch
authors.affiliation: Institute of Animal Nutrition, Freie Universitaet Berlin, Koenigin-Luise-Str. 49, 14195 Berlin, Germany.
authors.forename: Robert
authors.initials: R
authors.lastname: Pieper
authors.affiliation: Institute of Veterinary-Biochemistry, Freie Universitaet Berlin, Oertzenweg 19 b, 14163 Berlin, Germany.
authors.forename: Ralf
authors.initials: R
authors.lastname: Einspanier
authors.affiliation: Institute of Veterinary-Biochemistry, Freie Universitaet Berlin, Oertzenweg 19 b, 14163 Berlin, Germany.
authors.forename: Soroush
authors.initials: S
authors.lastname: Sharbati
mesh_terms: D000818:Animals; D002454:Cell Differentiation / Q000187:drug effects*; D003106:Colon / Q000166:cytology / Q000378:metabolism*; D004847:Epithelial Cells / Q000166:cytology / Q000378:metabolism*; D058750:Epithelial-Mesenchymal Transition / Q000187:drug effects*; D007413:Intestinal Mucosa / Q000166:cytology / Q000378:metabolism*; D035683:MicroRNAs / Q000378:metabolism; D034741:RNA, Small Interfering / Q000494:pharmacology*; D013552:Swine
chemical_list: D035683:MicroRNAs; D034741:RNA, Small Interfering