title: Peculiarity of Porcine Amniotic Membrane and Its Derived Cells: A Contribution to the Study of Cell Therapy from a Large Animal Model.
abstract: The aim of this work was to provide, for the first time, a protocol for isolation and characterization of stem cells from porcine amniotic membrane in view of their potential uses in regenerative medicine. From three samples of allanto-amnion recovered at delivery, the amniotic membrane was stripped from overlying allantois and digested with trypsin and collagenase to isolate epithelial (amniotic epithelial cells [AECs]) and mesenchymal cells, respectively. Proliferation, differentiation, and characterization studies by molecular biology and flow cytometry were performed. Histological examination revealed very few mesenchymal cells in the stromal layer, and a cellular yield of AECs of 10 × 10(6)/gram of digested tissue was achieved. AECs readily attached to plastic culture dishes displaying typical cuboidal morphology and, although their proliferative capacity decreased to the fifth passage, AECs showed a mean doubling time of 24.77 ± 6 h and a mean frequency of one fibroblast colony-forming unit (CFU-F) for every 116.75 plated cells. AECs expressed mesenchymal stem cell (MSC) mRNA markers (CD29, CD166, CD90, CD73, CD117) and pluripotent markers (Nanog and Oct 4), whereas they were negative for CD34 and MHCII. Mesodermic, ectodermic, and endodermic differentiation was confirmed by staining and expression of specific markers. We conclude that porcine amniotic membrane can provide an attractive source of stem cells that may be a useful tool for biomedical research.
authors.affiliation: 1 Large Animal Hospital, Reproduction Unit, Università degli Studi di Milano , Lodi, Italy .
authors.forename: Anna
authors.initials: A
authors.lastname: Lange-Consiglio
authors.affiliation: 2 Department of Life and Environmental Sciences, Università Politecnica delle Marche , Ancona, Italy .
authors.forename: Bruna
authors.initials: B
authors.lastname: Corradetti
authors.affiliation: 1 Large Animal Hospital, Reproduction Unit, Università degli Studi di Milano , Lodi, Italy .
authors.forename: Sabrina
authors.initials: S
authors.lastname: Bertani
authors.affiliation: 2 Department of Life and Environmental Sciences, Università Politecnica delle Marche , Ancona, Italy .
authors.forename: Valentina
authors.initials: V
authors.lastname: Notarstefano
authors.affiliation: 1 Large Animal Hospital, Reproduction Unit, Università degli Studi di Milano , Lodi, Italy .
authors.forename: Claudia
authors.initials: C
authors.lastname: Perrini
authors.affiliation: 2 Department of Life and Environmental Sciences, Università Politecnica delle Marche , Ancona, Italy .
authors.forename: Maria Giovanna
authors.initials: MG
authors.lastname: Marini
authors.affiliation: 3 Department of Health, Animal Science and Food Safety, Università degli Studi di Milano , Milan, Italy .
authors.forename: Silvana
authors.initials: S
authors.lastname: Arrighi
authors.affiliation: 3 Department of Health, Animal Science and Food Safety, Università degli Studi di Milano , Milan, Italy .
authors.forename: Giampaolo
authors.initials: G
authors.lastname: Bosi
authors.affiliation: 4 Large Animal Hospital, Clinic for Ruminants and Pigs, Università degli Studi di Milano , Lodi, Italy .
authors.forename: Angelo
authors.initials: A
authors.lastname: Belloli
authors.affiliation: 4 Large Animal Hospital, Clinic for Ruminants and Pigs, Università degli Studi di Milano , Lodi, Italy .
authors.forename: Davide
authors.initials: D
authors.lastname: Pravettoni
authors.affiliation: 4 Large Animal Hospital, Clinic for Ruminants and Pigs, Università degli Studi di Milano , Lodi, Italy .
authors.forename: Valentina
authors.initials: V
authors.lastname: Locatelli
authors.affiliation: 1 Large Animal Hospital, Reproduction Unit, Università degli Studi di Milano , Lodi, Italy .|3 Department of Health, Animal Science and Food Safety, Università degli Studi di Milano , Milan, Italy .
authors.forename: Fausto
authors.initials: F
authors.lastname: Cremonesi
authors.affiliation: 2 Department of Life and Environmental Sciences, Università Politecnica delle Marche , Ancona, Italy .
authors.forename: Davide
authors.initials: D
authors.lastname: Bizzaro
mesh_terms: D000650:Amnion / Q000166:cytology / Q000473:pathology*; D000818:Animals; D018929:Cell Culture Techniques; D002454:Cell Differentiation; D049109:Cell Proliferation; D064987:Cell- and Tissue-Based Therapy / Q000379:methods*; D004847:Epithelial Cells / Q000166:cytology; D005434:Flow Cytometry; D020869:Gene Expression Profiling; D059630:Mesenchymal Stem Cells / Q000166:cytology; D023421:Models, Animal; D050814:Octamer Transcription Factor-3 / Q000378:metabolism; D009841:Oligonucleotides / Q000235:genetics; D012333:RNA, Messenger / Q000378:metabolism; D060888:Real-Time Polymerase Chain Reaction; D013234:Stem Cells / Q000166:cytology*; D013552:Swine
chemical_list: D050814:Octamer Transcription Factor-3; D009841:Oligonucleotides; D012333:RNA, Messenger