Abstract
Pluripotent stem cells within the inner cell mass and epiblast of mammalian embryos have the capacity to form all lineages in the adult organism, while multipotent trophoblast stem (TS) cells derived from the trophectoderm are capable of differentiating into fetal lineages of the placenta. While mouse embryonic stem (ES) cells and epiblast stem cells (EpiSCs) exhibit distinct expression patterns and utilize distinct external signaling pathways for self-renewal, because mouse EpiSCs resemble human ES cells they are a useful model to investigate mechanisms of human ES cell self-renewal and differentiation. Recent studies have shown that haploid embryos and ES cells can be generated from chemically-activated unfertilized mouse oocytes. However, it is unclear whether EpiSCs or TS cells can be derived from haploid embryos. Here, we describe the derivation of EpiSCs from haploid blastocyst-stage embryos using culture conditions that promote TS cell self-renewal. Maternal (parthenogenetic/gynogenetic) EpiSCs (maEpiSCs) functionally and morphologically resemble conventional EpiSCs. Established maEpiSCs and conventional EpiSCs are diploid and exhibit a normal number of chromosomes. Moreover, global expression analyses and epigenomic profiling revealed that maEpiSCs and conventional EpiSCs exhibit similarly primed transcriptional programs and epigenetic profiles, respectively. Altogether, our results describe a useful experimental model to generate EpiSCs from haploid embryos, provide insight into self-renewal mechanisms of EpiSCs, and suggest that FGF4 is not sufficient to derive TS cells from haploid blastocyst-stage embryos.
Highlights
Pluripotent stem cells originating from the inner cell mass (ICM) of pre-implantation blastocysts and epiblast stem cells (EpiSCs) derived from the epiblast of postimplantation embryos or pre-implantation embryos have the capacity to differentiate into cell types of the three embryonic germ layers
Because FGF4 has previously been shown to support the self-renewal of both mouse trophoblast stem (TS) cells and EpiSCs7,10, we investigated whether FGF4-culture conditions favors the generation of EpiSCs or TS cells from haploid mouse preimplantation-stage embryos
It is possible that the haploid genome state is unstable in lineage-committed cells, such as in TS cells, which are derived from the trophectoderm, which is the first lineage to emerge during development
Summary
Pluripotent stem cells originating from the inner cell mass (ICM) of pre-implantation blastocysts and epiblast stem cells (EpiSCs) derived from the epiblast of postimplantation embryos or pre-implantation embryos have the capacity to differentiate into cell types of the three embryonic germ layers. Mouse EpiSCs and human ES cells can both be maintained in culture indefinitely by activin/nodal and FGF signaling pathways[1,4,6]. Mouse trophoblast stem (TS) cells, which originate from preimplantation stage embryos, can be cultured indefinitely in the presence of activin/nodal and FGF signaling pathways[7]. The common signaling pathways that support self-renewal of pluripotent human ES cells, mouse EpiSCs and multipotent TS cells suggest that divergent fates of preimplantation-stage cells can be sustained in culture under similar conditions. We investigated whether culture of mouse haploid blastocyst-stage embryos generated from chemically-activated oocytes favors a pluripotent EpiSC or multipotent TS cell fate. Formation of TS cells from haploid blastocyst-stage embryos, we show that maternal (parthenogenetic/gynogenetic) EpiSCs (maEpiSCs) can be generated following culture of haploid mouse preimplantation-stage embryos in FGF4-culture conditions. Our results demonstrate the ability to generate maEpiSCs from haploid preimplantation-stage embryos
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