Tsukiyama Tomoyuki, Ohinata Yasuhide
PRESTO, Japan Science and Technology Agency (JST), Kawaguchi, Saitama, Japan; Laboratory for Pluripotent Stem Cell Studies, RIKEN Center for Developmental Biology (CDB), Kobe, Hyogo, Japan.
PRESTO, Japan Science and Technology Agency (JST), Kawaguchi, Saitama, Japan; Laboratory for Pluripotent Stem Cell Studies, RIKEN Center for Developmental Biology (CDB), Kobe, Hyogo, Japan; Life Science Experimental Facility, Department of Biotechnology, Faculty of Life and Environmental Science, University of Yamanashi, Kofu, Yamanashi, Japan.
PLoS One. 2014 Apr 15;9(4):e95329. doi: 10.1371/journal.pone.0095329. eCollection 2014.
Embryonic stem cells (ESCs) can contribute to the tissues of chimeric animals, including the germline. By contrast, epiblast stem cells (EpiSCs) barely contribute to chimeras. These two types of cells are established and maintained under different culture conditions. Here, we show that a modified EpiSC culture condition containing the GSK3 inhibitor CHIR99021 can support a germline-competent pluripotent state that is intermediate between ESCs and EpiSCs. When ESCs were cultured under a modified condition containing bFGF, Activin A, and CHIR99021, they converted to intermediate pluripotent stem cells (INTPSCs). These INTPSCs were able to form teratomas in vivo and contribute to chimeras by blastocyst injection. We also induced formation of INTPSCs (iINTPSCs) from mouse embryonic fibroblasts by exogenous expression of four reprogramming factors, Oct3/4, Sox2, Klf4, and c-Myc, under the INTPSC culture condition. These iINTPSCs contributed efficiently to chimeras, including the germline, by blastocyst injection. The INTPSCs exhibited several characteristic properties of both ESCs and EpiSCs. Our results suggest that the modified EpiSC culture condition can support growth of cells that meet the most stringent criteria for pluripotency, and that germline-competent pluripotency may depend on the activation state of Wnt signaling.
胚胎干细胞（ESCs）可分化为嵌合体动物的组织，包括生殖系。相比之下，上胚层干细胞（EpiSCs）几乎不参与形成嵌合体。这两种细胞是在不同的培养条件下建立和维持的。在此，我们表明，含有GSK3抑制剂CHIR99021的改良EpiSC培养条件可支持一种介于ESCs和EpiSCs之间的具有生殖系能力的多能状态。当ESCs在含有碱性成纤维细胞生长因子（bFGF）、激活素A和CHIR99021的改良条件下培养时，它们会转化为中间多能干细胞（INTPSCs）。这些INTPSCs能够在体内形成畸胎瘤，并通过囊胚注射参与形成嵌合体。我们还在INTPSC培养条件下，通过外源表达四种重编程因子Oct3/4、Sox2、Klf4和c-Myc，从小鼠胚胎成纤维细胞诱导形成了INTPSCs（iINTPSCs）。通过囊胚注射，这些iINTPSCs有效地参与形成嵌合体，包括生殖系。INTPSCs表现出ESCs和EpiSCs的几种特征特性。我们的结果表明，改良的EpiSC培养条件可支持符合最严格多能性标准的细胞生长，并且具有生殖系能力的多能性可能取决于Wnt信号的激活状态。
