title: Bovine mammary epithelial cells retain stem-like phenotype in long-term cultures.
abstract: The detection and characterization of bovine mammary stem cells may give a better understanding of the cyclic characteristic of mammary gland development. In turn, this could potentially offer techniques to manipulate lactation yield and for regenerative medicine. We previously demonstrated that adult stem cells reside in the bovine mammary gland and possess an intrinsic regenerative potential. In vitro maintenance and expansion of this primitive population is a challenging task that could make easier the study of adult mammary stem cells. The aim of this study is to investigate this possibility. Different subpopulations of mammary epithelial cells emerge when they are cultured in two defined culture conditions. Specific cell differentiation markers as cytokeratin 18 (CK18) and cytokeratin 14 (CK14) were expressed with significant differences according to culture conditions. Vimentin, a well-known fibroblast marker was observed to increase significantly (P < 0.5) only after day 20. In both conditions, after prolonged culture (25 days) a subset of cells still retained regenerative capabilities. These cells were able to form organized pseudo-alveoli when transplanted in immunodeficient mice as shown by the expression of cytokeratin 14 (CK14), cytokeratin 18 (CK18), p63 (a mammary basal cell layer marker) and Epithelial Cell Adhesion Molecule (EpCAM). We also were able to observe the presence of milk proteins signal in these regenerated structures, which is a specific marker of functional mammary alveoli. Progenitor content was also analyzed in vitro through Colony-Forming Cell (CFC) assays with no substantial differences among culture conditions and time points. These results demonstrate that long-term culture of a multipotent cell subpopulation with intrinsic regenerative potential is possible.
authors.forename: Diego
authors.initials: D
authors.lastname: Cravero
authors.affiliation: Department of Veterinary Science, University of Turin, Via Leonardo da Vinci 44, 10095 Grugliasco, TO, Italy.
authors.forename: Cravero
authors.initials: C
authors.lastname: Diego
authors.forename: Eugenio
authors.initials: E
authors.lastname: Martignani
authors.forename: Martignani
authors.initials: M
authors.lastname: Eugenio
authors.forename: Silvia
authors.initials: S
authors.lastname: Miretti
authors.forename: Miretti
authors.initials: M
authors.lastname: Silvia
authors.forename: Elisabetta
authors.initials: E
authors.lastname: Macchi
authors.forename: Macchi
authors.initials: M
authors.lastname: Elisabetta
authors.forename: Paolo
authors.initials: P
authors.lastname: Accornero
authors.forename: Accornero
authors.initials: A
authors.lastname: Paolo
authors.forename: Mario
authors.initials: M
authors.lastname: Baratta
authors.forename: Baratta
authors.initials: B
authors.lastname: Mario
mesh_terms: D000822:Animal Husbandry / Q000379:methods; D000818:Animals; D002417:Cattle; D002454:Cell Differentiation; D002478:Cells, Cultured; D004847:Epithelial Cells / Q000166:cytology* / Q000378:metabolism; D005260:Female; D053547:Keratin-14 / Q000378:metabolism; D053538:Keratin-18 / Q000378:metabolism; D007774:Lactation / Q000378:metabolism; D008321:Mammary Glands, Animal / Q000166:cytology* / Q000378:metabolism; D051379:Mice; D016688:Mice, Inbred NOD; D016513:Mice, SCID; D039902:Multipotent Stem Cells / Q000166:cytology* / Q000378:metabolism; D010641:Phenotype; D014183:Transplantation, Heterologous / Q000662:veterinary
chemical_list: D053547:Keratin-14; D053538:Keratin-18
keywords: Bovine; Cell differentiation; Mammary gland; Stem cell