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MAPC culture condition supports the derivation of cells with nascent hypoblast features from bone marrow and blastocysts

ABSTRACT: Rat Extraembryonic Endoderm Precursor (XEN-P) cells resemble the nascent hypoblast of the blastocyst. Here we show that rat Multipotent Adult Progenitor Cell (rMAPC) lines derived from bone marrow (BM) exhibit salient nascent hypoblast characteristics. We could not identify cells with hypoblast features in rat BM, but they appeared after prolonged in vitro culture under rMAPC conditions. However, rMAPC culture conditions supported the very rapid isolation of new lines with nascent hypoblast features from rat blastocysts that were more homogenous than XEN-P cells isolated on feeders. The gene expression profile, growth factor requirement, Oct4 promoter methylation, and in vitro/vivo differentiation potential of cells isolated using rMAPC culture conditions from BM or blastocysts or XEN-P cells were highly similar. We conclude that the three cell populations have characteristics of hypoblast stem cells (HypoSC) and that rMAPC culture conditions support the isolation of XEN-P/HypoSC from the blastocyst and induction of HypoSC from BM. Comparison between bone marrow and blastocyst-derived Hypoblast Stem Cells (hypoSC) Three cell lines derived from bone marrow, three cell lines derived from blastocyst and one mesenchymal used, as control (for all of them 2 replicates)

ORGANISM(S): Rattus norvegicus

SUBMITTER: Joke Allemeersch

PROVIDER: E-GEOD-29114 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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MAPC culture condition supports the derivation of cells with nascent hypoblast features from bone marrow and blastocysts

Project description:Rat Extraembryonic Endoderm Precursor (XEN-P) cells resemble the nascent hypoblast of the blastocyst. Here we show that rat Multipotent Adult Progenitor Cell (rMAPC) lines derived from bone marrow (BM) exhibit salient nascent hypoblast characteristics. We could not identify cells with hypoblast features in rat BM, but they appeared after prolonged in vitro culture under rMAPC conditions. However, rMAPC culture conditions supported the very rapid isolation of new lines with nascent hypoblast features from rat blastocysts that were more homogenous than XEN-P cells isolated on feeders. The gene expression profile, growth factor requirement, Oct4 promoter methylation, and in vitro/vivo differentiation potential of cells isolated using rMAPC culture conditions from BM or blastocysts or XEN-P cells were highly similar. We conclude that the three cell populations have characteristics of hypoblast stem cells (HypoSC) and that rMAPC culture conditions support the isolation of XEN-P/HypoSC from the blastocyst and induction of HypoSC from BM.

2012-07-17 | GSE29114 | GEO

BMP and Activin treatment of mouse extraembryonic endoderm (XEN) cells

Project description:XEN cells are derived from the primitive endoderm of mouse blastocysts. In culture and in chimeras they exhibit properties of parietal endoderm. However, BMP signaling promotes XEN cells to form an epithelium and differentiate into visceral endoderm (VE). Of the several different subtypes of VE described, BMP induces a subtype that is most similar to the VE adjacent to the trophoblast-derived extraembryonic ectoderm. The experiment was performed to gain insight into genes regulated by BMP and activin in XEN cells, and also to more precisely define the VE subtypes formed in culture.

2011-11-10 | GSE32199 | GEO

BMP and Activin treatment of mouse extraembryonic endoderm (XEN) cells

2011-11-10 | E-GEOD-32199 | biostudies-arrayexpress

Primitive endoderm stem cell lines in mice

Project description:A blastocyst consists of three distinctive cell types: epiblast (EPI), trophoblast (TB), and primitive endoderm (PrE). Stem cell lines representing EPI and TB (embryonic stem (ES) cells and trophoblast stem (TS) cells) have been derived and they contribute to epiblast derivatives and trophoblast derivatives of stem cell-blastocyst chimeras, respectively. Although derived from PrE, extraembryonic endoderm (XEN) cells contribute to only a limited part of parietal endoderm (PE) but rarely to other PrE derivatives. Here we describe the establishment of primitive endoderm stem (PrES) cell lines in mice. PrES cells were derived and maintained in a serum-free media containing CHIR99021, FGF4, heparin, and PDGF-AA. RNA-seq analysis revealed that the transcriptome of PrES cells is globaly different from XEN cells: PrES cells express not only endoderm markers (Dab2, Gata4, Gata6, Sox17, etc.), but also pluripotent markers (Pou5f1, Cdh1, Nanog, Zfp42, etc.), and resembles in vivo founder PrE. PrES cells were rapidly and efficiently incorporated into PrE after blastocyst injection and efficiently contributed to all PrE derivatives, including PE, VE (visceral endoderm), and MZE (marginal zone endoderm) in chimeras. Importantly, PrES cells rescued embryonic lethality of PrE-depleted blastocysts by complementing all PrE derivatives. PrES cells thus not only contribute to understanding of the mechanisms of PrE specification but also provide a critical resource for artificial embryo reconstitution by stem cells alone.

2020-05-30 | GSE130957 | GEO

OSKM induce extraembryonic endoderm stem (iXEN) cells in parallel to iPS cells

Project description:While the reprogramming factors OCT4, SOX2, KLF4, and MYC (OSKM) can reactivate the pluripotency network in terminally differentiated cells, they also regulate expression of non-pluripotency genes in other contexts, such as the mouse primitive endoderm. The primitive endoderm is an extraembryonic lineage established alongside the pluripotent epiblast in the blastocyst, and is the progenitor pool for extraembryonic endoderm stem (XEN) cells. Several studies have shown that endodermal genes are upregulated in fibroblasts undergoing reprogramming, although whether endodermal genes promote or inhibit acquisition of pluripotency is unclear. We show that, in fibroblasts undergoing conventional reprogramming, OSKM-induced expression of endodermal genes leads to formation of induced XEN (iXEN) cells, which possess key properties of blastocyst-derived XEN cells, including morphology, transcription profile, self-renewal, and multipotency. Our data show that iXEN cells arise in parallel to iPS cells, indicating that OSKM are sufficient to drive cells to two distinct fates during reprogramming. Sequence-based mRNA transcriptional profiling of three different cell lines (MEF, XEN, iXEN) with multiple biological replicates, under two different growth medium conditions (ESC medium, XEN medium) for XEN and iXEN cells.

2016-03-03 | E-GEOD-77550 | biostudies-arrayexpress

Regulation of extraembryonic endoderm stem cell differentiation by Nodal and EGF-CFC signaling

Project description:The signaling pathway for Nodal, a ligand of the transforming growth factor-beta (TGF-beta) superfamily, plays a central role in regulating the maintenance and/or differentiation of stem cell types that can be derived from the peri-implantation mouse embryo. Extraembryonic endoderm stem (XEN) cells are derived from the primitive endoderm of the blastocyst, which normally gives rise to the parietal and the visceral endoderm in vivo, but XEN cells do not contribute efficiently to the visceral endoderm in chimeric embryos. We have found that treatment of XEN cells with Nodal and/or Cripto, an EGF-CFC co-receptor for Nodal, results in up-regulation of markers for visceral endoderm as well as anterior visceral endoderm (AVE). Re-introduction of treated XEN cells into chimeric embryos by blastocyst injection or morula aggregation results in contribution to visceral endoderm and AVE. In culture, XEN cells do not express Cripto, but do express the related EGF-CFC co-receptor Cryptic and require Cryptic for Nodal signaling. Notably, the response to Nodal can be blocked by treatment with the ALK4/ALK5/ALK7 inhibitor SB431542, but Cripto treatment is unaffected, suggesting that its activity is independent of type I activin receptors. Gene set enrichment analysis of genome-wide expression signatures generated from XEN cells under these treatment conditions confirms the differing responses of Nodal- and Cripto-treated XEN cells to SB431542. Our findings define distinct pathways for Nodal and Cripto in the differentiation of visceral endoderm and AVE from XEN cells, and provide new insights into the specification of these cell types in vivo. Murine Xen-eyfg cell line treated with Alk4 inhibitor SB431542, Nodal and Cripto recombinant proteins.

2011-11-29 | E-GEOD-23675 | biostudies-arrayexpress

DNA methylation comparison of ntTS, TS and ES cells

Project description:Placental abnormalities occur frequently in cloned animals. To investigate DNA methylation profile of trophoblast cell lineage of somatic cell nuclear transferred (NT) embryos, we established TS cells from blastocysts produced by NT at the blastocyst stage. Three independent TS cell lines derived from NT embryos rom BDF1 background were used in this study. Two control lines, derived from native blastocysts of BDF1 background, were also used.

2010-05-16 | E-GEOD-17104 | biostudies-arrayexpress

Conversion of mES to cXEN cells

Project description:The inner cell mass of the mouse pre-implantation blastocyst is comprised of epiblast progenitor and primitive endoderm cells of which cognate embryonic (mESCs) or extra-embryonic (XEN) stem cell lines can be derived. Importantly, each stem cell type retains the defining properties and lineage restriction of their in vivo tissue of origin. Recently, we demonstrated that XEN-like cells arise within mESC cultures. This raises the possibility that mESCs can generate self-renewing XEN cells without the requirement for gene manipulation. We have developed a novel approach to convert mESCs to XEN cells (cXEN) using growth factors. We confirm that the down-regulation of the pluripotency transcription factor Nanog and the expression of primitive endoderm-associated genes Gata6, Gata4, Sox17 and Pdgfra are necessary for cXEN cell derivation. This approach highlights an important function for Fgf4 in cXEN cell derivation. Paracrine FGF-signalling compensates for the loss of endogenous Fgf4, which is necessary to exit mESC self-renewal, but not for XEN cell maintenance. Our cXEN protocol also reveals that distinct pluripotent stem cells respond uniquely to differentiation promoting signals. cXEN cells can be derived from mESCs cultured with Erk- and Gsk3-inhibitors (2i) and LIF, similarly to conventional mESCs. However, we find that epiblast stem cells (EpiSCs) derived from the post-implantation embryo are refractory to cXEN cell establishment, consistent with the hypothesis that EpiSCs represent a pluripotent state distinct from mESCs. In all, these findings suggest that the potential of mESCs includes the capacity to give rise to both extra-embryonic and embryonic lineages. A total of eight samples were analyzed. Three mouse embryonic stem cell (mESC) lines were used as a negative control, 2 embryo-derived extraembryonic endoderm (XEN) cell lines were used as a positive control, 3 converted XEN (cXEN) cells were used as the experiemental cell lines.

2012-06-04 | E-GEOD-38477 | biostudies-arrayexpress

Regulation of extraembryonic endoderm stem cell differentiation by Nodal and EGF-CFC signaling

2011-11-29 | GSE23675 | GEO

Transcription profiling of mouse trophoblast cell lineage of somatic cell nuclear transferred (NT) embryos

Project description:Placental abnormalities occur frequently in cloned animals. To investigate gene expression profile of trophoblast cell lineage of somatic cell nuclear transferred (NT) embryos, we established TS cells from blastocysts produced by NT at the blastocyst stage. Experiment Overall Design: Five independent TS cell lines derived from NT embryos were used in this study. Those included three lines from BDF1 background NT embryos and two lines from EGFP-Tg CD-1 background NT embryos. Two each of control lines, derived from native blastocysts of each background, were also used.

2009-08-07 | E-GEOD-17102 | biostudies-arrayexpress

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