Establishment of Trophoblast Stem Cells under Defined Culture Conditions in Mice

Ohinata, Yasuhide; Tsukiyama, Tomoyuki

doi:10.1371/journal.pone.0107308

Cited by 62 publications

(96 citation statements)

References 28 publications

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“…The rapid decrease in Eomes levels with 25 ng/ml FGF2 does not appear to be a result of differentiation, as its abundance was constant for the duration of the experiment and no morphological evidence of differentiation was observed after 6 D in culture, whereas differentiated cells were readily observed at ≤2.5 ng/ml FGF2. These data suggest that FGF2 can support TSC maintenance, which is consistent with a recent report by Ohinata and Tsukiyama () indicating that FGF2 can be used to derive TSCs from mouse blastocysts.…”

Section: Resultssupporting

confidence: 92%

“…These observations suggest that in addition to its role in Genome-wide analysis of ERF DNA-binding sites in embryo fibroblasts (Twigg et al, 2013) identified a strong interaction of ERF with Fgf2. FGF signaling plays a pivotal role in TSC differentiation: FGF4 expressed by the inner cell mass is required for trophectoderm maintenance (Arman, Haffner-Krausz, Chen, Heath, & Lonai, 1998), while maintenance of murine TSCs requires FGF4 (Tanaka et al, 1998) or FGF2 (Ohinata & Tsukiyama, 2014). Attenuation of FGF2 signaling is also required for the differentiation of TSCs in humans (Sudheer, Bhusan, Fauler, Lehrach, & Adjaye, 2012).…”

Section: Erf May Contribute To Syncytiotrophoblast Lineage Developmentmentioning

confidence: 99%

“…Extracellular signal‐regulated kinase (ERK) activation by FGF appears to be required for the emergence or survival of TSCs in humans and mice (Daoud et al, ; Yang et al, ). Recent studies indicate that FGF2 may have a role in placenta development, and can support the TSC phenotype and derivation more efficiently than FGF4 (Ohinata & Tsukiyama, ). Surprisingly, very little is currently known about the regulation of FGFs in placenta development.…”

Section: Introductionmentioning

confidence: 99%

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Suppression of Fgf2 by ETS2 repressor factor (ERF) is required for chorionic trophoblast differentiation

Vorgia

Ζαραγκούλιας

Peraki

et al. 2017

Molecular Reproduction Devel

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ETS2 repressor factor (ERF) is a ubiquitous transcriptional repressor regulated by Extracellular signal-regulated kinase 1/2 (ERK1/2) phosphorylation. Homozygous deletion of Erf in mice blocks chorionic trophoblast differentiation, resulting in the failure of chorioallantoic fusion and subsequent embryo death. Fibroblast growth factor (FGF) signaling is important for proper trophoblast stem cell (TSC) differentiation and development of the hemochorial placenta. Lack of Fgf2 promotes TSC differentiation, while FGF4 or FGF2 is required for murine TSC maintenance. Here, we show that low in vivo Fgf2 mRNA abundance occurs in patches of placental chorion cells and ex vivo in TSCs. This expression is repressed via direct interaction of ERF with the Fgf2 transcription unit is increased in the absence of ERF, and is decreased in the presence of an ERF mutant resistant to ERK phosphorylation. Thus, FGF2 inhibition by ERF appears to be necessary for proper chorionic TSC differentiation, and may account for the block of chorionic trophoblast differentiation in Erf-knockout animals. The differentiation of ERF-overexpressing TSC lines also suggests that ERF may have an FGF2-independent effect during the commitment towards syncytiotrophoblasts.

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Section: Resultssupporting

confidence: 92%

Section: Erf May Contribute To Syncytiotrophoblast Lineage Developmentmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Suppression of Fgf2 by ETS2 repressor factor (ERF) is required for chorionic trophoblast differentiation

Vorgia

Ζαραγκούλιας

Peraki

et al. 2017

Molecular Reproduction Devel

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“…However, hypomethylated patterns were unexpectedly found at the paternal allele in three of the four control TSC lines (TS-4, TS-5, and TS-6), ranging from 30.7% to 33.9%, although that of the SCNT-TSC lines remained hypermethylated (78.1% to 94.8%), probably in response to the SCNT procedure at this region (Table 1, Figure 4). We hypothesized that the reduction in DNA methylation at the paternal IG-DMR might have been caused by the use of serum-free conditioned medium (CDM) at their establishment [12], because it contains knockout serum supplement (KSR), known to induce hypomethylation of the IG-DMR in induced pluripotent stem cells (iPSCs) [13]. Thus, we next examined the IG-DMR of an additional series of TSCs (two IVF-TSC lines and three SCNT-TSC lines), which had been generated using the conventional TSC medium containing FGF4, heparin, and serum during the first 12-46 days of culture.…”

Section: Resultsmentioning

confidence: 99%

Aberrant imprinting in mouse trophoblast stem cells established from somatic cell nuclear transfer-derived embryos

et al. 2018

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Although phenotypic abnormalities frequently appear in the placenta following somatic cell nuclear transfer (SCNT), mouse trophoblast stem cells (TSCs) established from SCNT embryos reportedly show no distinct abnormalities compared with those derived from normal fertilization. In this study, we reexamined SCNT-TSCs to identify their imprinting statuses. Placenta-specific maternally imprinted genes (Gab1, Slc38a4, and Sfmbt2) consistently showed biallelic expression in SCNT-TSCs, suggesting their loss of imprinting (LOI). The LOI of Gab1 was associated with decreased DNA methylation, and that of Sfmbt2 was associated with decreased DNA methylation and histone H3K27 trimethylation. The maternal allele of the intergenic differentially methylated region (IG-DMR) was aberrantly hypermethylated following SCNT, even though this region was prone to demethylation in TSCs when established in a serum-free chemically defined medium. These findings indicate that the development of cloned embryos is associated with imprinting abnormalities specifically in the trophoblast lineage from its initial stage, which may affect subsequent placental development.

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“…Thus, a portion of the population of trophoblast cell lines maintains AP activity; however, the significance of this cell population is obscure in the trophoblastic cell lineage. Moreover, RT-PCR showed that these cells express trophoblast-specific genes, such as ELF5 , CDX2 , and SOX2 , which were also expressed in mouse TSCs [42, 43]. In addition, biTBCs express IFN-τ , an antiluteolytic factor responsible for preventing the regression of the maternal corpus luteum and supporting the sustainability of pregnancy for cattle uteri [7, 44].…”

Section: Discussionmentioning

confidence: 99%

Derivation of Induced Trophoblast Cell Lines in Cattle by Doxycycline-Inducible piggyBac Vectors

et al. 2016

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Trophectoderm lineage specification is one of the earliest differentiation events in mammalian development. The trophoblast lineage, which is derived from the trophectoderm, mediates implantation and placental formation. However, the processes involved in trophoblastic differentiation and placental formation in cattle remain unclear due to interspecies differences when compared with other model systems and the small repertoire of available trophoblast cell lines. Here, we describe the generation of trophoblast cell lines (biTBCs) from bovine amnion-derived cells (bADCs) using an induced pluripotent stem cell technique. bADCs were introduced with piggyBac vectors containing doxycycline (Dox)-inducible transcription factors (Oct3⁄4(POU5F1), Sox2, Klf4, and c-Myc). Colonies that appeared showed a flattened epithelial-like morphology similar to cobblestones, had a more definite cell boundary between cells, and frequently formed balloon-like spheroids similar to trophoblastic vesicles (TVs). biTBCs were propagated for over 60 passages and expressed trophoblast-related (CDX2, ELF5, ERRβ, and IFN-τ) and pluripotency-related genes (endogenous OCT3/4, SOX2, KLF4, and c-MYC). Furthermore, when biTBCs were induced to differentiate by removing Dox from culture, they formed binucleate cells and began to express pregnancy-related genes (PL, PRP1, and PAG1). This is the first report demonstrating that the induction of pluripotency in bovine amniotic cells allows the generation of trophoblastic cell lines that possess trophoblast stem cell-like characteristics and have the potential to differentiate into the extra-embryonic cell lineage. These cell lines can be a new cell source as a model for studying trophoblast cell lineages and implantation processes in cattle.

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Establishment of Trophoblast Stem Cells under Defined Culture Conditions in Mice

Cited by 62 publications

References 28 publications

Suppression of Fgf2 by ETS2 repressor factor (ERF) is required for chorionic trophoblast differentiation

Suppression of Fgf2 by ETS2 repressor factor (ERF) is required for chorionic trophoblast differentiation

Aberrant imprinting in mouse trophoblast stem cells established from somatic cell nuclear transfer-derived embryos

Derivation of Induced Trophoblast Cell Lines in Cattle by Doxycycline-Inducible piggyBac Vectors

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