title: Partial maintenance and long-term expansion of murine skin epithelial stem cells by Wnt-3a in vitro.
abstract: CD49f(+)CD34(+) cells, a skin epithelial stem cell (EpSC)-rich population, were prepared from adult mouse skin and cultured in the presence of Wnt-3a without feeder cells. CD34 expression was retained in about 10% of the cells, which had proliferated about 1,000-fold by day 10, although completely lost by day 14. CD49f(+)CD34(+) cells sorted on day 10 retained canonical Wnt-responsiveness, proliferated markedly in the presence of Wnt-3a, maintained undifferentiated epithelial cell marker expression, and promoted hair follicle development in vivo. Those were subjected to a second 10-day culture with Wnt-3a and sorted, and then the same procedures were repeated a total of 15 times. CD49f(+)CD34(+) cells obtained from each of those cultures retained the same EpSC characteristics as the original cells. CD34(+) and CD34(-) cells were found to produce Wnt-3a and Wnt/β-catenin inhibitors, respectively. CD34(+) cells resided as small cellular clusters surrounded by a large amount of CD34(-) cells. Furthermore, we found that exogenous Wnt-3a delayed the conversion of CD34(+) cells to CD34(-) cells and induced CD34(-) cells to suppress the production of Wnt/β-catenin inhibitors, likely leading to generation of a microenvironment favorable for maintaining EpSCs. Our results suggest the possibility of partial long-term maintenance of EpSCs in vitro by Wnt-3a.
authors.affiliation: Department of Pathogen, Infection and Immunity, Nara Medical University, Kashihara, Japan; Program in Tissue Engineering, Department of Parasitology, Nara Medical University, Kashihara, Japan. Electronic address: oujix@naramed-u.ac.jp.
authors.forename: Yukiteru
authors.initials: Y
authors.lastname: Ouji
authors.affiliation: Program in Tissue Engineering, Department of Parasitology, Nara Medical University, Kashihara, Japan.
authors.forename: Shigeaki
authors.initials: S
authors.lastname: Ishizaka
authors.affiliation: Department of Pathogen, Infection and Immunity, Nara Medical University, Kashihara, Japan.
authors.forename: Fukumi
authors.initials: F
authors.lastname: Nakamura-Uchiyama
authors.affiliation: DNA-chip Development Center for Infectious Diseases, Research Institute for Microbial Diseases, Osaka University, Suita, Osaka, Japan.
authors.forename: Daisuke
authors.initials: D
authors.lastname: Okuzaki
authors.affiliation: Department of Pathogen, Infection and Immunity, Nara Medical University, Kashihara, Japan; Program in Tissue Engineering, Department of Parasitology, Nara Medical University, Kashihara, Japan. Electronic address: myoshika@naramed-u.ac.jp.
authors.forename: Masahide
authors.initials: M
authors.lastname: Yoshikawa
mesh_terms: D000818:Animals; D018952:Antigens, CD34 / Q000378:metabolism; D002454:Cell Differentiation; D049109:Cell Proliferation; D004847:Epithelial Cells / Q000166:cytology*; D005260:Female; D018859:Hair Follicle / Q000378:metabolism; D039503:Integrin alpha6 / Q000378:metabolism; D051379:Mice; D008807:Mice, Inbred BALB C; D008809:Mice, Inbred C3H; D008819:Mice, Nude; D020411:Oligonucleotide Array Sequence Analysis; D011994:Recombinant Proteins / Q000494:pharmacology; D015398:Signal Transduction; D013234:Stem Cells / Q000166:cytology*; D060509:Wnt3A Protein / Q000494:pharmacology*; D051176:beta Catenin / Q000378:metabolism
chemical_list: D018952:Antigens, CD34; D039503:Integrin alpha6; D011994:Recombinant Proteins; D060509:Wnt3A Protein; C555511:Wnt3a protein, mouse; D051176:beta Catenin