title: Feeder-free generation and long-term culture of human induced pluripotent stem cells using pericellular matrix of decidua derived mesenchymal cells.
abstract: Human ES cells (hESCs) and human induced pluripotent stem cells (hiPSCs) are usually generated and maintained on living feeder cells like mouse embryonic fibroblasts or on a cell-free substrate like Matrigel. For clinical applications, a quality-controlled, xenobiotic-free culture system is required to minimize risks from contaminating animal-derived pathogens and immunogens. We previously reported that the pericellular matrix of decidua-derived mesenchymal cells (PCM-DM) is an ideal human-derived substrate on which to maintain hiPSCs/hESCs. In this study, we examined whether PCM-DM could be used for the generation and long-term stable maintenance of hiPSCs. Decidua-derived mesenchymal cells (DMCs) were reprogrammed by the retroviral transduction of four factors (OCT4, SOX2, KLF4, c-MYC) and cultured on PCM-DM. The established hiPSC clones expressed alkaline phosphatase, hESC-specific genes and cell-surface markers, and differentiated into three germ layers in vitro and in vivo. At over 20 passages, the hiPSCs cultured on PCM-DM held the same cellular properties with genome integrity as those at early passages. Global gene expression analysis showed that the GDF3, FGF4, UTF1, and XIST expression levels varied during culture, and GATA6 was highly expressed under our culture conditions; however, these gene expressions did not affect the cells' pluripotency. PCM-DM can be conveniently prepared from DMCs, which have a high proliferative potential. Our findings indicate that PCM-DM is a versatile and practical human-derived substrate that can be used for the feeder-cell-free generation and long-term stable maintenance of hiPSCs.
authors: Division of Regenerative Medicine, Institute for Clinical Research, Osaka National Hospital, National Hospital Organization, Osaka, Japan.
authors: Hayato
authors: H
authors: Fukusumi
authors: Tomoko
authors: T
authors: Shofuda
authors: Daisuke
authors: D
authors: Kanematsu
authors: Atsuyo
authors: A
authors: Yamamoto
authors: Hiroshi
authors: H
authors: Suemizu
authors: Masato
authors: M
authors: Nakamura
authors: Mami
authors: M
authors: Yamasaki
authors: Masatoshi
authors: M
authors: Ohgushi
authors: Yoshiki
authors: Y
authors: Sasai
authors: Yonehiro
authors: Y
authors: Kanemura
mesh_terms: D000704:Analysis of Variance; D018929:Cell Culture Techniques / Q000379:methods*; D002454:Cell Differentiation / Q000502:physiology; D003656:Decidua / Q000166:cytology*; D053595:Embryonic Stem Cells / Q000166:cytology*; D005109:Extracellular Matrix / Q000378:metabolism*; D005260:Female; D005434:Flow Cytometry; D020869:Gene Expression Profiling; D006801:Humans; D007150:Immunohistochemistry; D007621:Karyotyping; D000090062:Kruppel-Like Factor 4; D051741:Kruppel-Like Transcription Factors / Q000235:genetics; D059630:Mesenchymal Stem Cells / Q000378:metabolism*; D046228:Microarray Analysis; D050814:Octamer Transcription Factor-3 / Q000235:genetics; D039904:Pluripotent Stem Cells / Q000166:cytology*; D016271:Proto-Oncogene Proteins c-myc / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D055748:SOXB1 Transcription Factors / Q000235:genetics; D017422:Sequence Analysis, DNA; D018709:Statistics, Nonparametric; D014161:Transduction, Genetic
chemical_list: C000715467:KLF4 protein, human; C000715468:Klf4 protein, mouse; D000090062:Kruppel-Like Factor 4; D051741:Kruppel-Like Transcription Factors; C489427:MYC protein, human; D050814:Octamer Transcription Factor-3; C494173:POU5F1 protein, human; D016271:Proto-Oncogene Proteins c-myc; C527782:SOX2 protein, human; D055748:SOXB1 Transcription Factors
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