Abstract:The early specification and rapid growth of extraembryonic membranes are distinctive hallmarks of primate embryogenesis. These complex tasks are resolved through an intricate combination of signals controlling the induction of extraembryonic lineages and, at the same time, safeguarding the pluripotent epiblast. Here, we delineate the signals orchestrating primate epiblast and amnion identity. We encapsulated marmoset pluripotent stem cells into agarose microgels and identified culture conditions for the develo… Show more
“…8b ). In line with this observation, a high BMP and low NODAL signalling environment has been implicated recently in amnion differentiation of marmoset ES cells 41 and in human ES cells during extraembryonic mesenchyme differentiation 34 , suggesting that similar dynamics may drive differentiation of these populations within inducible human embryo-like structures. Fig.…”
“…Indeed, immunofluorescence analysis demonstrated that the inner, SOX2-positive domain upregulated amnion markers, including CDX2 and ISL1 by day 6 post-aggregation. By day 8 post-aggregation, the inner domain expressed the mature amnion markers VTCN1 and HAND1 39 , 41 (Fig. 3d and Extended Data Fig.…”
Section: Differentiation Within Embryoidsmentioning
The human embryo undergoes morphogenetic transformations following implantation into the uterus, but our knowledge of this crucial stage is limited by the inability to observe the embryo in vivo. Models of the embryo derived from stem cells are important tools for interrogating developmental events and tissue–tissue crosstalk during these stages1. Here we establish a model of the human post-implantation embryo, a human embryoid, comprising embryonic and extraembryonic tissues. We combine two types of extraembryonic-like cell generated by overexpression of transcription factors with wild-type embryonic stem cells and promote their self-organization into structures that mimic several aspects of the post-implantation human embryo. These self-organized aggregates contain a pluripotent epiblast-like domain surrounded by extraembryonic-like tissues. Our functional studies demonstrate that the epiblast-like domain robustly differentiates into amnion, extraembryonic mesenchyme and primordial germ cell-like cells in response to bone morphogenetic protein cues. In addition, we identify an inhibitory role for SOX17 in the specification of anterior hypoblast-like cells2. Modulation of the subpopulations in the hypoblast-like compartment demonstrates that extraembryonic-like cells influence epiblast-like domain differentiation, highlighting functional tissue–tissue crosstalk. In conclusion, we present a modular, tractable, integrated3 model of the human embryo that will enable us to probe key questions of human post-implantation development, a critical window during which substantial numbers of pregnancies fail.
“…8b ). In line with this observation, a high BMP and low NODAL signalling environment has been implicated recently in amnion differentiation of marmoset ES cells 41 and in human ES cells during extraembryonic mesenchyme differentiation 34 , suggesting that similar dynamics may drive differentiation of these populations within inducible human embryo-like structures. Fig.…”
“…Indeed, immunofluorescence analysis demonstrated that the inner, SOX2-positive domain upregulated amnion markers, including CDX2 and ISL1 by day 6 post-aggregation. By day 8 post-aggregation, the inner domain expressed the mature amnion markers VTCN1 and HAND1 39 , 41 (Fig. 3d and Extended Data Fig.…”
Section: Differentiation Within Embryoidsmentioning
The human embryo undergoes morphogenetic transformations following implantation into the uterus, but our knowledge of this crucial stage is limited by the inability to observe the embryo in vivo. Models of the embryo derived from stem cells are important tools for interrogating developmental events and tissue–tissue crosstalk during these stages1. Here we establish a model of the human post-implantation embryo, a human embryoid, comprising embryonic and extraembryonic tissues. We combine two types of extraembryonic-like cell generated by overexpression of transcription factors with wild-type embryonic stem cells and promote their self-organization into structures that mimic several aspects of the post-implantation human embryo. These self-organized aggregates contain a pluripotent epiblast-like domain surrounded by extraembryonic-like tissues. Our functional studies demonstrate that the epiblast-like domain robustly differentiates into amnion, extraembryonic mesenchyme and primordial germ cell-like cells in response to bone morphogenetic protein cues. In addition, we identify an inhibitory role for SOX17 in the specification of anterior hypoblast-like cells2. Modulation of the subpopulations in the hypoblast-like compartment demonstrates that extraembryonic-like cells influence epiblast-like domain differentiation, highlighting functional tissue–tissue crosstalk. In conclusion, we present a modular, tractable, integrated3 model of the human embryo that will enable us to probe key questions of human post-implantation development, a critical window during which substantial numbers of pregnancies fail.
“…In PSC populations, PAC1 formed a self-interacting source of FGF2 , but far fewer FGF2 -centric interactions were observed in 4i, with CAC2/3 instead appearing to act as a source for FGF1 . Because of differences in the relative proportions of individual clusters in the PSC/4i/PreME samples, this suggests a general decrease in FGF2/3/4 -related interactions in PreME and 4i populations versus PSC, an observation that correlates with previous studies in humans ( Penfold et al, 2018 Preprint ) and rabbits ( Kobayashi et al, 2021 ) suggesting an antagonistic for FGF in PGC competence, with additional roles in PSC proliferation ( Kobayashi et al, 2021 ) and ablation of amnion formation ( Munger et al, 2022 ).…”
Human germline–soma segregation occurs during weeks 2–3 in gastrulating embryos. Although direct studies are hindered, here, we investigate the dynamics of human primordial germ cell (PGCs) specification using in vitro models with temporally resolved single-cell transcriptomics and in-depth characterisation using in vivo datasets from human and nonhuman primates, including a 3D marmoset reference atlas. We elucidate the molecular signature for the transient gain of competence for germ cell fate during peri-implantation epiblast development. Furthermore, we show that both the PGCs and amnion arise from transcriptionally similar TFAP2A-positive progenitors at the posterior end of the embryo. Notably, genetic loss of function experiments shows that TFAP2A is crucial for initiating the PGC fate without detectably affecting the amnion and is subsequently replaced by TFAP2C as an essential component of the genetic network for PGC fate. Accordingly, amniotic cells continue to emerge from the progenitors in the posterior epiblast, but importantly, this is also a source of nascent PGCs.
“…We next investigated the specification of epiblast-derived cell populations, specifically focusing on amnion patterning, which forms before gastrulation in primates. Previous studies of non-human primates and human stem cells suggest that amnion induction is dependent on BMP 30 – 33 , but many functional and molecular features of this lineage remain unclear. Morphologically, hEEs frequently contained ISL1 + cells (a marker of primate amnion 34 ) by D4 to D6, with efficiency ranging from 46.79% to 70.86% depending on the genetic background (Fig.…”
Section: Emergence Of Late Amnion-like Cellsmentioning
Investigating human development is a substantial scientific challenge due to the technical and ethical limitations of working with embryonic samples. In the face of these difficulties, stem cells have provided an alternative to experimentally model inaccessible stages of human development in vitro1–13. Here we show that human pluripotent stem cells can be triggered to self-organize into three-dimensional structures that recapitulate some key spatiotemporal events of early human post-implantation embryonic development. Our system reproducibly captures spontaneous differentiation and co-development of embryonic epiblast-like and extra-embryonic hypoblast-like lineages, establishes key signalling hubs with secreted modulators and undergoes symmetry breaking-like events. Single-cell transcriptomics confirms differentiation into diverse cell states of the perigastrulating human embryo14,15 without establishing placental cell types, including signatures of post-implantation epiblast, amniotic ectoderm, primitive streak, mesoderm, early extra-embryonic endoderm, as well as initial yolk sac induction. Collectively, our system captures key features of human embryonic development spanning from Carnegie stage16 4–7, offering a reproducible, tractable and scalable experimental platform to understand the basic cellular and molecular mechanisms that underlie human development, including new opportunities to dissect congenital pathologies with high throughput.
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