title: FGF2 and EGF Are Required for Self-Renewal and Organoid Formation of Canine Normal and Tumor Breast Stem Cells.
abstract: Recent studies suggest that human tumors are generated from cancer cells with stem cell (SC) properties. Spontaneously occurring cancers in dogs contain a diversity of cells that like for human tumors suggest that certain canine tumors are also generated from cancer stem cells (CSCs). CSCs, like normal SCs, have the capacity for self-renewal as mammospheres in suspension cultures. To understand how cells with SC properties contribute to canine mammary gland tumor development and progression, comparative analysis between normal SCs and CSCs, obtained from the same individual, is essential. We have utilized the property of sphere formation to develop culture conditions for propagating stem/progenitor cells from canine normal and tumor tissue. We show that cells from dissociated mammospheres retain sphere reformation capacity for several serial passages and have the capacity to generate organoid structures ex situ. Utilizing various culture conditions for passaging SCs and CSCs, fibroblast growth factor 2 (FGF2) and epidermal growth factor (EGF) were found to positively or negatively regulate mammosphere regeneration, organoid formation, and multi-lineage differentiation potential. The response of FGF2 and EGF on SCs and CSCs was different, with increased FGF2 and EGF self-renewal promoted in SCs and repressed in CSCs. Our protocol for propagating SCs from normal and tumor canine breast tissue will provide new opportunities in comparative mammary gland stem cell analysis between species and anticancer treatment and therapies for dogs. J. Cell. Biochem. 118: 570-584, 2017. © 2016 Wiley Periodicals, Inc.
authors.affiliation: Istituto di Tecnologie Biomediche, Consiglio Nazionale delle Ricerche, Via F.lli Cervi 93, 20090, Segrate, Milano, Italy.
authors.forename: Cinzia
authors.initials: C
authors.lastname: Cocola
authors.affiliation: Istituto di Tecnologie Biomediche, Consiglio Nazionale delle Ricerche, Via F.lli Cervi 93, 20090, Segrate, Milano, Italy.
authors.forename: Stefano
authors.initials: S
authors.lastname: Molgora
authors.affiliation: Istituto di Tecnologie Biomediche, Consiglio Nazionale delle Ricerche, Via F.lli Cervi 93, 20090, Segrate, Milano, Italy.
authors.forename: Eleonora
authors.initials: E
authors.lastname: Piscitelli
authors.affiliation: Department of Veterinary Medicine, Università degli Studi di Milano, Via Celoria 10, 20133, Milano, Italy.
authors.forename: Maria Cristina
authors.initials: MC
authors.lastname: Veronesi
authors.affiliation: Istituto di Tecnologie Biomediche, Consiglio Nazionale delle Ricerche, Via F.lli Cervi 93, 20090, Segrate, Milano, Italy.
authors.forename: Marianna
authors.initials: M
authors.lastname: Greco
authors.affiliation: Muscle Cell Biology Laboratory, Neuromuscular Diseases and Neuroimmunology Unit, Fondazione IRCCS Istituto Neurologico "C. Besta", Via Temolo 4, 20126, Milano, Italy.
authors.forename: Cinzia
authors.initials: C
authors.lastname: Bragato
authors.affiliation: Fondazione Istituto Nazionale di Genetica Molecolare 'Romeo ed Enrica Invernizzi', Via Francesco Sforza 35, 20122, Milan, Italy.
authors.forename: Monica
authors.initials: M
authors.lastname: Moro
authors.affiliation: Fondazione Istituto Nazionale di Genetica Molecolare 'Romeo ed Enrica Invernizzi', Via Francesco Sforza 35, 20122, Milan, Italy.
authors.forename: Mariacristina
authors.initials: M
authors.lastname: Crosti
authors.affiliation: Molecular Targeting Technologies, Inc., West Chester, Pennsylvania.
authors.forename: Brian
authors.initials: B
authors.lastname: Gray
authors.affiliation: Istituto di Tecnologie Biomediche, Consiglio Nazionale delle Ricerche, Via F.lli Cervi 93, 20090, Segrate, Milano, Italy.
authors.forename: Luciano
authors.initials: L
authors.lastname: Milanesi
authors.affiliation: Department of Veterinary Medicine, Università degli Studi di Milano, Via Celoria 10, 20133, Milano, Italy.
authors.forename: Valeria
authors.initials: V
authors.lastname: Grieco
authors.affiliation: Department of Health, Animal Science and Food Safety, Università degli Studi di Milano, 20133, Milano, Italy.
authors.forename: Gaia Cecilia
authors.initials: GC
authors.lastname: Luvoni
authors.affiliation: National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, Maryland, 20814.
authors.forename: James
authors.initials: J
authors.lastname: Kehler
authors.affiliation: Department of Biology, College of Science and Technology, Temple University, Philadelphia, Pennsylvania.|Sbarro Institute for Cancer Research and Molecular Medicine, College of Science and Technology, Temple University, Philadelphia, Pennsylvania.
authors.forename: Gianfranco
authors.initials: G
authors.lastname: Bellipanni
authors.affiliation: Istituto di Tecnologie Biomediche, Consiglio Nazionale delle Ricerche, Via F.lli Cervi 93, 20090, Segrate, Milano, Italy.
authors.forename: Rolland
authors.initials: R
authors.lastname: Reinbold
authors.affiliation: Istituto di Tecnologie Biomediche, Consiglio Nazionale delle Ricerche, Via F.lli Cervi 93, 20090, Segrate, Milano, Italy.
authors.forename: Ileana
authors.initials: I
authors.lastname: Zucchi
authors.affiliation: Department of Biology, College of Science and Technology, Temple University, Philadelphia, Pennsylvania.|Sbarro Institute for Cancer Research and Molecular Medicine, College of Science and Technology, Temple University, Philadelphia, Pennsylvania.
authors.forename: Antonio
authors.initials: A
authors.lastname: Giordano
mesh_terms: D000818:Animals; D049109:Cell Proliferation / Q000187:drug effects*; D004285:Dogs; D004815:Epidermal Growth Factor / Q000494:pharmacology*; D005260:Female; D016222:Fibroblast Growth Factor 2 / Q000494:pharmacology*; D015674:Mammary Neoplasms, Animal / Q000378:metabolism* / Q000473:pathology; D014411:Neoplastic Stem Cells / Q000378:metabolism* / Q000473:pathology; D009940:Organoids / Q000378:metabolism* / Q000473:pathology; D014407:Tumor Cells, Cultured
chemical_list: D016222:Fibroblast Growth Factor 2; D004815:Epidermal Growth Factor
keywords: ACINI; BREAST; CANCER; CANINES; FGF2; MAMMARY; MAMMOSPHERE; ORGANOIDS; STEM