Reduction of N-glycolylneuraminic acid in human induced pluripotent stem cells generated or cultured under feeder- and serum-free defined conditions.

<h4>Background</h4>The successful establishment of human induced pluripotent stem cells (hiPSCs) has increased the possible applications of stem cell research in biology and medicine. In particular, hiPSCs are a promising source of cells for regenerative medicine and pharmacology. Howeve...

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Autores principales: Yohei Hayashi, Techuan Chan, Masaki Warashina, Masakazu Fukuda, Takashi Ariizumi, Koji Okabayashi, Naoya Takayama, Makoto Otsu, Koji Eto, Miho Kusuda Furue, Tatsuo Michiue, Kiyoshi Ohnuma, Hiromitsu Nakauchi, Makoto Asashima
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Publicado: Public Library of Science (PLoS) 2010
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spelling oai:doaj.org-article:c18b4106c0c149c0bbff7552180f87012021-11-18T07:36:36ZReduction of N-glycolylneuraminic acid in human induced pluripotent stem cells generated or cultured under feeder- and serum-free defined conditions.1932-620310.1371/journal.pone.0014099https://doaj.org/article/c18b4106c0c149c0bbff7552180f87012010-11-01T00:00:00Zhttps://www.ncbi.nlm.nih.gov/pmc/articles/pmid/21124894/?tool=EBIhttps://doaj.org/toc/1932-6203<h4>Background</h4>The successful establishment of human induced pluripotent stem cells (hiPSCs) has increased the possible applications of stem cell research in biology and medicine. In particular, hiPSCs are a promising source of cells for regenerative medicine and pharmacology. However, one of the major obstacles to such uses for hiPSCs is the risk of contamination from undefined pathogens in conventional culture conditions that use serum replacement and mouse embryonic fibroblasts as feeder cells.<h4>Methodology/principal findings</h4>Here we report a simple method for generating or culturing hiPSCs under feeder- and serum-free defined culture conditions that we developed previously for human embryonic stem cells. The defined culture condition comprises a basal medium with a minimal number of defined components including five highly purified proteins and fibronectin as a substrate. First, hiPSCs, which were generated using Yamanaka's four factors and conventional undefined culture conditions, adapted to the defined culture conditions. These adapted cells retained the property of self renewal as evaluated morphologically, the expression of self-renewal marker proteins, standard growth rates, and pluripotency as evaluated by differentiation into derivatives of all three primary germ layers in vitro and in vivo (teratoma formation in immunodeficient mice). Moreover, levels of nonhuman N-glycolylneuraminic acid (Neu5Gc), which is a xenoantigenic indicator of pathogen contamination in human iPS cell cultures, were markedly decreased in hiPSCs cultured under the defined conditions. Second, we successfully generated hiPSCs using adult dermal fibroblast under the defined culture conditions from the reprogramming step. For a long therm culture, the generated cells also had the property of self renewal and pluripotency, they carried a normal karyotype, and they were Neu5Gc negative.<h4>Conclusion/significance</h4>This study suggested that generation or adaption culturing under defined culture conditions can eliminate the risk posed by undefined pathogens. This success in generating hiPSCs using adult fibroblast would be beneficial for clinical application.Yohei HayashiTechuan ChanMasaki WarashinaMasakazu FukudaTakashi AriizumiKoji OkabayashiNaoya TakayamaMakoto OtsuKoji EtoMiho Kusuda FurueTatsuo MichiueKiyoshi OhnumaHiromitsu NakauchiMakoto AsashimaPublic Library of Science (PLoS)articleMedicineRScienceQENPLoS ONE, Vol 5, Iss 11, p e14099 (2010)
institution DOAJ
collection DOAJ
language EN
topic Medicine
R
Science
Q
spellingShingle Medicine
R
Science
Q
Yohei Hayashi
Techuan Chan
Masaki Warashina
Masakazu Fukuda
Takashi Ariizumi
Koji Okabayashi
Naoya Takayama
Makoto Otsu
Koji Eto
Miho Kusuda Furue
Tatsuo Michiue
Kiyoshi Ohnuma
Hiromitsu Nakauchi
Makoto Asashima
Reduction of N-glycolylneuraminic acid in human induced pluripotent stem cells generated or cultured under feeder- and serum-free defined conditions.
description <h4>Background</h4>The successful establishment of human induced pluripotent stem cells (hiPSCs) has increased the possible applications of stem cell research in biology and medicine. In particular, hiPSCs are a promising source of cells for regenerative medicine and pharmacology. However, one of the major obstacles to such uses for hiPSCs is the risk of contamination from undefined pathogens in conventional culture conditions that use serum replacement and mouse embryonic fibroblasts as feeder cells.<h4>Methodology/principal findings</h4>Here we report a simple method for generating or culturing hiPSCs under feeder- and serum-free defined culture conditions that we developed previously for human embryonic stem cells. The defined culture condition comprises a basal medium with a minimal number of defined components including five highly purified proteins and fibronectin as a substrate. First, hiPSCs, which were generated using Yamanaka's four factors and conventional undefined culture conditions, adapted to the defined culture conditions. These adapted cells retained the property of self renewal as evaluated morphologically, the expression of self-renewal marker proteins, standard growth rates, and pluripotency as evaluated by differentiation into derivatives of all three primary germ layers in vitro and in vivo (teratoma formation in immunodeficient mice). Moreover, levels of nonhuman N-glycolylneuraminic acid (Neu5Gc), which is a xenoantigenic indicator of pathogen contamination in human iPS cell cultures, were markedly decreased in hiPSCs cultured under the defined conditions. Second, we successfully generated hiPSCs using adult dermal fibroblast under the defined culture conditions from the reprogramming step. For a long therm culture, the generated cells also had the property of self renewal and pluripotency, they carried a normal karyotype, and they were Neu5Gc negative.<h4>Conclusion/significance</h4>This study suggested that generation or adaption culturing under defined culture conditions can eliminate the risk posed by undefined pathogens. This success in generating hiPSCs using adult fibroblast would be beneficial for clinical application.
format article
author Yohei Hayashi
Techuan Chan
Masaki Warashina
Masakazu Fukuda
Takashi Ariizumi
Koji Okabayashi
Naoya Takayama
Makoto Otsu
Koji Eto
Miho Kusuda Furue
Tatsuo Michiue
Kiyoshi Ohnuma
Hiromitsu Nakauchi
Makoto Asashima
author_facet Yohei Hayashi
Techuan Chan
Masaki Warashina
Masakazu Fukuda
Takashi Ariizumi
Koji Okabayashi
Naoya Takayama
Makoto Otsu
Koji Eto
Miho Kusuda Furue
Tatsuo Michiue
Kiyoshi Ohnuma
Hiromitsu Nakauchi
Makoto Asashima
author_sort Yohei Hayashi
title Reduction of N-glycolylneuraminic acid in human induced pluripotent stem cells generated or cultured under feeder- and serum-free defined conditions.
title_short Reduction of N-glycolylneuraminic acid in human induced pluripotent stem cells generated or cultured under feeder- and serum-free defined conditions.
title_full Reduction of N-glycolylneuraminic acid in human induced pluripotent stem cells generated or cultured under feeder- and serum-free defined conditions.
title_fullStr Reduction of N-glycolylneuraminic acid in human induced pluripotent stem cells generated or cultured under feeder- and serum-free defined conditions.
title_full_unstemmed Reduction of N-glycolylneuraminic acid in human induced pluripotent stem cells generated or cultured under feeder- and serum-free defined conditions.
title_sort reduction of n-glycolylneuraminic acid in human induced pluripotent stem cells generated or cultured under feeder- and serum-free defined conditions.
publisher Public Library of Science (PLoS)
publishDate 2010
url https://doaj.org/article/c18b4106c0c149c0bbff7552180f8701
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