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Developmental Dynamics : An Official... Jul 1995We describe a series of stages for development of the embryo of the zebrafish, Danio (Brachydanio) rerio. We define seven broad periods of embryogenesis--the zygote,...
We describe a series of stages for development of the embryo of the zebrafish, Danio (Brachydanio) rerio. We define seven broad periods of embryogenesis--the zygote, cleavage, blastula, gastrula, segmentation, pharyngula, and hatching periods. These divisions highlight the changing spectrum of major developmental processes that occur during the first 3 days after fertilization, and we review some of what is known about morphogenesis and other significant events that occur during each of the periods. Stages subdivide the periods. Stages are named, not numbered as in most other series, providing for flexibility and continued evolution of the staging series as we learn more about development in this species. The stages, and their names, are based on morphological features, generally readily identified by examination of the live embryo with the dissecting stereomicroscope. The descriptions also fully utilize the optical transparancy of the live embryo, which provides for visibility of even very deep structures when the embryo is examined with the compound microscope and Nomarski interference contrast illumination. Photomicrographs and composite camera lucida line drawings characterize the stages pictorially. Other figures chart the development of distinctive characters used as staging aid signposts.
Topics: Animals; Blastocyst; Cleavage Stage, Ovum; Embryo, Nonmammalian; Gastrula; Morphogenesis; Temperature; Time Factors; Zebrafish; Zygote
PubMed: 8589427
DOI: 10.1002/aja.1002030302 -
Cell Sep 2022In vitro cultured stem cells with distinct developmental capacities can contribute to embryonic or extraembryonic tissues after microinjection into pre-implantation...
In vitro cultured stem cells with distinct developmental capacities can contribute to embryonic or extraembryonic tissues after microinjection into pre-implantation mammalian embryos. However, whether cultured stem cells can independently give rise to entire gastrulating embryo-like structures with embryonic and extraembryonic compartments remains unknown. Here, we adapt a recently established platform for prolonged ex utero growth of natural embryos to generate mouse post-gastrulation synthetic whole embryo models (sEmbryos), with both embryonic and extraembryonic compartments, starting solely from naive ESCs. This was achieved by co-aggregating non-transduced ESCs, with naive ESCs transiently expressing Cdx2 or Gata4 to promote their priming toward trophectoderm and primitive endoderm lineages, respectively. sEmbryos adequately accomplish gastrulation, advance through key developmental milestones, and develop organ progenitors within complex extraembryonic compartments similar to E8.5 stage mouse embryos. Our findings highlight the plastic potential of naive pluripotent cells to self-organize and functionally reconstitute and model the entire mammalian embryo beyond gastrulation.
Topics: Animals; Cell Differentiation; Embryo, Mammalian; Embryonic Development; Embryonic Stem Cells; Endoderm; Gastrulation; Mammals; Mice
PubMed: 35988542
DOI: 10.1016/j.cell.2022.07.028 -
Nature Oct 2023The ability to study human post-implantation development remains limited owing to ethical and technical challenges associated with intrauterine development after...
The ability to study human post-implantation development remains limited owing to ethical and technical challenges associated with intrauterine development after implantation. Embryo-like models with spatially organized morphogenesis and structure of all defining embryonic and extra-embryonic tissues of the post-implantation human conceptus (that is, the embryonic disc, the bilaminar disc, the yolk sac, the chorionic sac and the surrounding trophoblast layer) remain lacking. Mouse naive embryonic stem cells have recently been shown to give rise to embryonic and extra-embryonic stem cells capable of self-assembling into post-gastrulation structured stem-cell-based embryo models with spatially organized morphogenesis (called SEMs). Here we extend those findings to humans using only genetically unmodified human naive embryonic stem cells (cultured in human enhanced naive stem cell medium conditions). Such human fully integrated and complete SEMs recapitulate the organization of nearly all known lineages and compartments of post-implantation human embryos, including the epiblast, the hypoblast, the extra-embryonic mesoderm and the trophoblast layer surrounding the latter compartments. These human complete SEMs demonstrated developmental growth dynamics that resemble key hallmarks of post-implantation stage embryogenesis up to 13-14 days after fertilization (Carnegie stage 6a). These include embryonic disc and bilaminar disc formation, epiblast lumenogenesis, polarized amniogenesis, anterior-posterior symmetry breaking, primordial germ-cell specification, polarized yolk sac with visceral and parietal endoderm formation, extra-embryonic mesoderm expansion that defines a chorionic cavity and a connecting stalk, and a trophoblast-surrounding compartment demonstrating syncytium and lacunae formation. This SEM platform will probably enable the experimental investigation of previously inaccessible windows of human early post implantation up to peri-gastrulation development.
Topics: Humans; Embryo Implantation; Embryo, Mammalian; Embryonic Development; Fertilization; Gastrulation; Germ Layers; Human Embryonic Stem Cells; Trophoblasts; Yolk Sac; Giant Cells
PubMed: 37673118
DOI: 10.1038/s41586-023-06604-5 -
Nature Oct 2023The human embryo undergoes morphogenetic transformations following implantation into the uterus, but our knowledge of this crucial stage is limited by the inability to...
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 stages. 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 cells. 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, integrated 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.
Topics: Female; Humans; Pregnancy; Bone Morphogenetic Proteins; Cell Differentiation; Embryo Implantation; Embryo, Mammalian; Embryoid Bodies; Embryonic Development; Germ Layers; Human Embryonic Stem Cells; Models, Biological; Transcription Factors; Pluripotent Stem Cells
PubMed: 37369347
DOI: 10.1038/s41586-023-06368-y -
Cell Stem Cell Mar 2021Pluripotent cells emerge as a naive founder population in the blastocyst, acquire capacity for germline and soma formation, and then undergo lineage priming. Mouse...
Pluripotent cells emerge as a naive founder population in the blastocyst, acquire capacity for germline and soma formation, and then undergo lineage priming. Mouse embryonic stem cells (ESCs) and epiblast-derived stem cells (EpiSCs) represent the initial naive and final primed phases of pluripotency, respectively. Here, we investigate the intermediate formative stage. Using minimal exposure to specification cues, we derive stem cells from formative mouse epiblast. Unlike ESCs or EpiSCs, formative stem (FS) cells respond directly to germ cell induction. They colonize somatic tissues and germline in chimeras. Whole-transcriptome analyses show similarity to pre-gastrulation formative epiblast. Signal responsiveness and chromatin accessibility features reflect lineage capacitation. Furthermore, FS cells show distinct transcription factor dependencies, relying critically on Otx2. Finally, FS cell culture conditions applied to human naive cells or embryos support expansion of similar stem cells, consistent with a conserved staging post on the trajectory of mammalian pluripotency.
Topics: Animals; Blastocyst; Cell Differentiation; Embryonic Stem Cells; Germ Layers; Humans; Mice; Pluripotent Stem Cells
PubMed: 33271069
DOI: 10.1016/j.stem.2020.11.005 -
Cell Stem Cell Sep 2023Naive human pluripotent stem cells have the remarkable ability to self-organize into blastocyst-like structures ("blastoids") that model lineage segregation in the...
Naive human pluripotent stem cells have the remarkable ability to self-organize into blastocyst-like structures ("blastoids") that model lineage segregation in the pre-implantation embryo. However, the extent to which blastoids can recapitulate the defining features of human post-implantation development remains unexplored. Here, we report that blastoids cultured on thick three-dimensional (3D) extracellular matrices capture hallmarks of early post-implantation development, including epiblast lumenogenesis, rapid expansion and diversification of trophoblast lineages, and robust invasion of extravillous trophoblast cells by day 14. Extended blastoid culture results in the localized activation of primitive streak marker TBXT and the emergence of embryonic germ layers by day 21. We also show that the modulation of WNT signaling alters the balance between epiblast and trophoblast fates in post-implantation blastoids. This work demonstrates that 3D-cultured blastoids offer a continuous and integrated in vitro model system of human embryonic and extraembryonic development from pre-implantation to early gastrulation stages.
Topics: Humans; Gastrulation; Embryo Implantation; Embryo, Mammalian; Blastocyst; Epithelial Cells
PubMed: 37683602
DOI: 10.1016/j.stem.2023.08.005 -
Nature Oct 2023Investigating human development is a substantial scientific challenge due to the technical and ethical limitations of working with embryonic samples. In the face of...
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 vitro. 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 embryo 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 stage 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.
Topics: Female; Humans; Pregnancy; Cell Differentiation; Cell Lineage; Embryo Implantation; Embryonic Development; Germ Layers; Human Embryonic Stem Cells; Placenta; Pluripotent Stem Cells; Primitive Streak; Yolk Sac
PubMed: 37369348
DOI: 10.1038/s41586-023-06354-4 -
Cell Reports Jun 2022In human embryos, major zygotic genome activation (ZGA) initiates at the eight-cell (8C) stage. Abnormal ZGA leads to developmental defects and even contributes to the...
In human embryos, major zygotic genome activation (ZGA) initiates at the eight-cell (8C) stage. Abnormal ZGA leads to developmental defects and even contributes to the failure of human blastocyst formation or implantation. An in vitro cell model mimicking human 8C blastomeres would be invaluable to understanding the mechanisms regulating key biological events during early human development. Using the non-canonical promoter of LEUTX that putatively regulates human ZGA, we developed an 8C::mCherry reporter, which specifically marks the 8C state, to isolate rare 8C-like cells (8CLCs) from human preimplantation epiblast-like stem cells. The 8CLCs express a panel of human ZGA genes and have a unique transcriptome resembling that of the human 8C embryo. Using the 8C::mCherry reporter, we further optimize the chemical-based culture condition to increase and maintain the 8CLC population. Functionally, 8CLCs can self-organize to form blastocyst-like structures. The discovery and maintenance of 8CLCs provide an opportunity to recapitulate early human development.
Topics: Blastocyst; Embryonic Development; Gene Expression Regulation, Developmental; Genome; Humans; Zygote
PubMed: 35732112
DOI: 10.1016/j.celrep.2022.110994 -
Epigenetics & Chromatin Dec 2021Dynamic changes of histone posttranslational modifications are important contexts of epigenetic reprograming after fertilization in pre-implantation embryos. Recently,...
BACKGROUND
Dynamic changes of histone posttranslational modifications are important contexts of epigenetic reprograming after fertilization in pre-implantation embryos. Recently, lactylation has been reported as a novel epigenetic modification that regulates various cellular processes, but its role during early embryogenesis has not been elucidated.
RESULTS
We examined nuclear accumulation of H3K23la, H3K18la and pan histone lactylation in mouse oocytes and pre-implantation embryos by immunofluorescence with specific antibodies. All of the three modifications were abundant in GV stage oocytes, and both H3K23la and pan histone lactylation could be detected on the condensed chromosomes of the MII oocytes, while H3K18la were not detected. After fertilization, the nuclear staining of H3K23la, H3K18la and pan histone lactylation was faint in zygotes but homogeneously stained both of the parental pronuclei. The signal remained weak in the early cleavage stage embryos and increased remarkably in the blastocyst stage embryos. Comparison of the embryos cultured in four different conditions with varying concentrations of oxygen found that H3K23la, H3K18la and pan histone lactylation showed similar and comparable staining pattern in embryos cultured in atmospheric oxygen concentration (20% O), gradient oxygen concentration (5% O to 2% O) and embryos obtained from in vivo, but the modifications were greatly reduced in embryos cultured in hypoxic condition (2% O). In contrast, nuclear accumulation of H3K18ac or H3K23ac was not significantly affected under hypoxic condition. Moreover, the developmental rate of in vitro cultured embryo was significantly reduced by low oxygen concentration and small molecule inhibition of LDHA activity led to decreased lactate production, as well as reduced histone lactylation and compromised developmental rate.
CONCLUSIONS
We provided for the first time the dynamic landscape of H3K23la, H3K18la and pan histone lactylation in oocytes and pre-implantation embryos in mice. Our data suggested that histone lactylation is subjected to oxygen concentration in the culture environment and hypoxic in vitro culture reduces histone lactylation, which in turn compromises developmental potential of pre-implantation embryos in mice.
Topics: Animals; Blastocyst; Embryo, Mammalian; Embryonic Development; Female; Histones; Mice; Oocytes; Pregnancy; Zygote
PubMed: 34930415
DOI: 10.1186/s13072-021-00431-6 -
Current Opinion in Genetics &... Oct 2022Synthetic embryology aims to develop embryo-like structures from stem cells to provide new insight into early stages of mammalian development. Recent advances in... (Review)
Review
Synthetic embryology aims to develop embryo-like structures from stem cells to provide new insight into early stages of mammalian development. Recent advances in synthetic embryology have highlighted the remarkable capacity of stem cells to self-organize under certain biochemical or biophysical stimulations, generating structures that recapitulate the fate and form of early mouse/human embryos, in which symmetry breaking, pattern formation, or proper morphogenesis can be observed spontaneously. Here we review recent progress on the design principles for different types of embryoids and discuss the impact of different biochemical and biophysical factors on the process of stem-cell self-organization. We also offer our thoughts about the principal future challenges.
Topics: Animals; Embryo, Mammalian; Embryonic Development; Humans; Mammals; Mice; Morphogenesis; Stem Cells
PubMed: 35988317
DOI: 10.1016/j.gde.2022.101970