153 results on '"Zernicka-Goetz M"'
Search Results
2. Dynamic shapes of the zygote and two-cell mouse and human
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Graham, CF, Windsor, S, Ajduk, A, Trinh, T, Vincent, A, Jones, C, Coward, K, Kalsi, D, Zernicka-Goetz, M, Swann, K, Thomas, ALR, Graham, Chris F [0000-0002-6410-8171], Windsor, Shane [0000-0002-7597-4497], Ajduk, Anna [0000-0002-7262-1370], Trinh, Thanh [0000-0002-6973-9749], Kalsi, Dilraj [0000-0001-9300-1835], Zernicka-Goetz, Magdalena [0000-0002-7004-2471], Swann, Karl [0000-0002-4355-1449], Thomas, Adrian LR [0000-0003-0878-4113], and Apollo - University of Cambridge Repository more...
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Morphokinetics ,Cell Nucleus ,Cytoplasm ,QH301-705.5 ,Zygote ,Science ,Shape cycles ,Cytoplasm vortices ,General Biochemistry, Genetics and Molecular Biology ,Mice ,Pronuclear fading ,Animals ,Humans ,Biology (General) ,General Agricultural and Biological Sciences ,Mouse zygote ,Research Article ,Human - Abstract
Mouse zygote morphokinetics were measured during interphase, the mitotic period, cytokinesis, and two-cell stage. Sequences of rounder–distorted–rounder shapes were revealed, as were changing patterns of cross section area. A calcium chelator and an actin-disrupting agent inhibited the area changes that occurred between pronuclear envelope breakdown and cytokinesis. During cell division, two vortices developed in each nascent cell and they rotated in opposite directions at each end of the cell, a pattern that sometimes persisted for up to 10 h. Exchange with the environment may have been promoted by these shape and area cycles and persisting circulation in the cytoplasm may have a similar function between a cell's interior and periphery. Some of these movements were sporadically also seen in human zygotes with abnormal numbers of pronuclei and the two-cell stages that developed from these compromised human zygotes., Summary: Mouse zygote morphokinetics were measured during interphase, the mitotic period, cytokinesis, and two-cell stage. Sequences of rounder–distorted–rounder shapes were revealed, as were changing patterns of cross section area. Some of these movements were sporadically also seen in human zygotes with abnormal numbers of pronuclei and the two-cell stages that developed from these compromised human zygotes. more...
- Published
- 2021
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Catalog
3. O–257 Formation of distinct cell types in the mouse embryo
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Zernicka-Goetz, M.
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- 2011
4. RASSF1A uncouples Wnt from Hippo signalling and promotes YAP mediated differentiation via p73
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Papaspyropoulos, A, Bradley, L, Thapa, A, Leung, C, Toskas, K, Koennig, D, Pefani, D, Raso, C, Grou, C, Hamilton, G, Vlahov, N, Grawenda, A, Haider, S, Chauhan, J, Buti, L, Kanapin, A, Lu, X, Buffa, F, Dianov, G, von Kriegsheim, A, Matallanas, D, Samsonova, A, Zernicka-Goetz, M, and O'Neill, E more...
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Male ,endocrine system ,Science ,Cell Cycle Proteins ,Protein Serine-Threonine Kinases ,Article ,Basic Helix-Loop-Helix Transcription Factors ,Animals ,Humans ,Hippo Signaling Pathway ,lcsh:Science ,Embryonic Stem Cells ,beta Catenin ,Adaptor Proteins, Signal Transducing ,Tumor Suppressor Proteins ,Gene Expression Regulation, Developmental ,TEA Domain Transcription Factors ,Cell Differentiation ,Tumor Protein p73 ,YAP-Signaling Proteins ,Phosphoproteins ,DNA-Binding Proteins ,Mice, Inbred C57BL ,Wnt Proteins ,Mice, Inbred CBA ,lcsh:Q ,Female ,Octamer Transcription Factor-3 ,Signal Transduction ,Transcription Factors - Abstract
Transition from pluripotency to differentiation is a pivotal yet poorly understood developmental step. Here, we show that the tumour suppressor RASSF1A is a key player driving the early specification of cell fate. RASSF1A acts as a natural barrier to stem cell self-renewal and iPS cell generation, by switching YAP from an integral component in the β-catenin-TCF pluripotency network to a key factor that promotes differentiation. We demonstrate that epigenetic regulation of the Rassf1A promoter maintains stemness by allowing a quaternary association of YAP–TEAD and β-catenin–TCF3 complexes on the Oct4 distal enhancer. However, during differentiation, promoter demethylation allows GATA1-mediated RASSF1A expression which prevents YAP from contributing to the TEAD/β-catenin–TCF3 complex. Simultaneously, we find that RASSF1A promotes a YAP–p73 transcriptional programme that enables differentiation. Together, our findings demonstrate that RASSF1A mediates transcription factor selection of YAP in stem cells, thereby acting as a functional “switch” between pluripotency and initiation of differentiation., In development, the switch from pluripotency to differentiation is important but it is often unclear how it is regulated. Here, the authors show that the tumour suppressor RASSF1A mediates this switch by promoting YAP-p73 transcription, which in turn enables differentiation. more...
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- 2018
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5. Delivery of mtZFNs into Early Mouse Embryos
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McCann, B., Cox, A., Gammage, P., Stewart, J., Zernicka-Goetz, M., and Minczuk, M.
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Mitochondrial diseases often result from mutations in the mitochondrial genome (mtDNA). In most cases, mutant mtDNA coexists with wild-type mtDNA, resulting in heteroplasmy. One potential future approach to treat heteroplasmic mtDNA diseases is the specific elimination of pathogenic mtDNA mutations, lowering the level of mutant mtDNA below pathogenic thresholds. Mitochondrially targeted zinc-finger nucleases (mtZFNs) have been demonstrated to specifically target and introduce double-strand breaks in mutant mtDNA, facilitating substantial shifts in heteroplasmy. One application of mtZFN technology, in the context of heteroplasmic mtDNA disease, is delivery into the heteroplasmic oocyte or early embryo to eliminate mutant mtDNA, preventing transmission of mitochondrial diseases through the germline. Here we describe a protocol for efficient production of mtZFN mRNA in vitro, and delivery of these into 0.5 dpc mouse embryos to elicit shifts of mtDNA heteroplasmy. more...
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- 2018
6. Delayed APC/C activation extends the first mitosis of mouse embryos
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Ajduk, A, Strauss, B, Pines, J, Zernicka-Goetz, M, Zernicka-Goetz, Magdalena [0000-0002-7004-2471], and Apollo - University of Cambridge Repository
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Transcriptional Activation ,lcsh:R ,lcsh:Medicine ,Embryonic Development ,Gene Expression ,Gene Expression Regulation, Developmental ,Mitosis ,Article ,Anaphase-Promoting Complex-Cyclosome ,Mice ,Genes, Reporter ,Proteolysis ,Animals ,M Phase Cell Cycle Checkpoints ,lcsh:Q ,Cyclin B1 ,lcsh:Science ,Cyclin A2 ,Biomarkers - Abstract
The correct temporal regulation of mitosis underpins genomic stability because it ensures the alignment of chromosomes on the mitotic spindle that is required for their proper segregation to the two daughter cells. Crucially, sister chromatid separation must be delayed until all the chromosomes have attached to the spindle; this is achieved by the Spindle Assembly Checkpoint (SAC) that inhibits the Anaphase Promoting Complex/Cyclosome (APC/C) ubiquitin ligase. In many species the first embryonic M-phase is significantly prolonged compared to the subsequent divisions, but the reason behind this has remained unclear. Here, we show that the first M-phase in the mouse embryo is significantly extended due to a delay in APC/C activation. Unlike in somatic cells, where the APC/C first targets cyclin A2 for degradation at nuclear envelope breakdown (NEBD), we find that in zygotes cyclin A2 remains stable for a significant period of time after NEBD. Our findings that the SAC prevents cyclin A2 degradation, whereas over-expressed Plk1 stimulates it, support our conclusion that the delay in cyclin A2 degradation is caused by low APC/C activity. As a consequence of delayed APC/C activation cyclin B1 stability in the first mitosis is also prolonged, leading to the unusual length of the first M-phase. more...
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- 2017
7. Early Asymmetry and Development of Polarity in the Mouse Embryo
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Zernicka-Goetz, M., Wianny, F., Pedersen, R.A., and Weber, R.J.
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Polarity (Biology) -- Research ,Mice -- Research ,Embryology -- Research ,Biological sciences - Abstract
In most species the polarity of an embryo underlies the future body plan and is determined from that of the zygote. To determine whether asymmetry of the mouse egg is essential for polarity of the later embryo, we studied the development of eggs from which animal or vegetal pole cytoplasm was surgically removed. We extended these studies to determine whether axial determinants are localised at the 2-cell or 8-cell stage. Both series of experiments showed that polarity of the egg and early embryo are not critical for development, consistent with the view that the organisation of the early mouse embryo does not play a role in establishing polarity. Indeed, polarity is generally thought to develop significantly later, after implantation. It has not previously been possible, however, to relate the polarity of the preimplantation embryo to that of the later conceptus due to the lack of markers that endure long enough to follow lineages through implantation. To test whether early developmental events provide cues that predict the axes of the postimplantation embryo, we have injected mRNA encoding Mm GFP to trace the progeny of inner cell mass cells into the postimplantation visceral endoderm. This extraembryonic tissue plays a role in axis determination in adjacent embryonic tissue. We found that visceral endoderm cells that originated near the polar body (a marker of blastocyst symmetry) generally became distal as the egg cylinder formed, while those opposite the polar body tended to become proximal. It follows that, in normal development, bilateral symmetry of the blastocyst anticipates the polarity of the later conceptus. To test the consequences of eliminating the expression of candidate molecules that may have a role in establishing polarity as early as the blastocyst stage, we have developed a method for double stranded RNA interference (RNAi) in the mouse embryo. As a proof of principle, we have shown that RNAi can phenocopy the effects of disrupting genes expressed both zygotically and in the oocyte. We now apply this approach to study the importance of spatial patterns of gene expression in orchestrating early mouse development. more...
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- 2000
8. Polarity in Cell-Fate Acquisition in the Early Mouse Embryo
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DePamphilis, Melvin L., Leung, C. Y., Zhu, M., Zernicka-Goetz, M., DePamphilis, Melvin L., Leung, C. Y., Zhu, M., and Zernicka-Goetz, M.
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Establishing polarity is a fundamental part of embryogenesis and can be traced back to the earliest developmental stages. It can be achieved in one of two ways: through the preexisting polarization of germ cells before fertilization or via symmetry breaking after fertilization. In mammals, it seems to be the latter, and we will discuss the various cytological and molecular events that lead up to this event, its mechanisms and the consequences. In mammals, the first polarization event occurs in the preimplantation period, when the embryo is but a cluster of cells, free-floating in the oviduct. This provides a unique, autonomous system to study the de novo polarization that is essential to life. In this review, we will cover modern and past studies on the polarization of the early embryo, using the mouse as a model system, as well as hypothesizing the potential implications and functions of the biological events involved. more...
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- 2016
9. Pluripotency and differentiation in embryos and stem cells
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Ja, Adjaye, Ag, Byskov, Jb, Cibelli, De Maria R, Minger S, Maurilio Sampaolesi, Testa G, Verfaillie C, Zernicka-Goetz M, Schöler H, Boiani M, Crosetto N, and Ca, Redi
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DIFFERENTIATION ,Cancer ,Cell ,Differentiation ,Embryo ,Meeting report ,Pluripotency ,Stem ,Cell Differentiation ,Embryo, Mammalian ,Embryonic Stem Cells ,Forecasting ,Humans ,Pluripotent Stem Cells ,Stem Cells ,Embryology ,Developmental Biology ,Settore MED/04 - PATOLOGIA GENERALE ,EMBRYO ,CANCER ,PLURIPOTENCY ,STEM CELL ,MEETING REPORT - Published
- 2008
10. Novel gene expression patterns along the proximo-distal axis of the mouse embryo before gastrulation
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Frankenberg, S, Smith, L, Greenfield, A, Zernicka-Goetz, M, Frankenberg, S, Smith, L, Greenfield, A, and Zernicka-Goetz, M
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BACKGROUND: To date, the earliest stage at which the orientation of the anterior-posterior axis in the mouse embryo is distinguishable by asymmetric gene expression is shortly after E5.5. At E5.5, prospective anterior markers are expressed at the distal tip of the embryo, whereas prospective posterior markers are expressed more proximally, close to the boundary with the extraembryonic region. RESULTS: To contribute to elucidating the mechanisms underlying the events involved in early patterning of the mouse embryo, we have carried out a microarray screen to identify novel genes that are differentially expressed between the distal and proximal parts of the E5.5 embryo. Secondary screening of resulting candidates by in situ hybridisation at E5.5 and E6.5 revealed novel expression patterns for known and previously uncharacterised genes, including Peg10, Ctsz1, Cubilin, Jarid1b, Ndrg1, Sfmbt2, Gjb5, Talia and Plet1. The previously undescribed gene Talia and recently identified Plet1 are expressed specifically in the distal-most part of the extraembryonic ectoderm, adjacent to the epiblast, and are therefore potential candidates for regulating early patterning events. Talia and the previously described gene XE7 define a gene family highly conserved among metazoans and with a predicted protein structure suggestive of a post-transcriptional regulative function, whilst Plet1 appears to be mammal-specific and of unknown function. CONCLUSION: Our approach has allowed us to compare expression between dissected parts of the egg cylinder and has identified multiple genes with novel expression patterns at this developmental stage. These genes are potential candidates for regulating tissue interactions following implantation. more...
- Published
- 2007
11. INVITED SESSION, SESSION 64: EPIGENETICS AND EARLY EVENTS DURING MAMMALIAN DEVELOPMENT Wednesday 6 July 2011 12:00 - 13:00
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Zernicka-Goetz, M., primary, Patrat, C., additional, Okamoto, I., additional, Thepot, D., additional, Peynot, N., additional, Fauque, P., additional, Daniel, N., additional, Diabangouaya, P., additional, Renard, J., additional, Duranthon, V., additional, and Heard, E., additional more...
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- 2011
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12. RNA Interference by Production of Short Hairpin dsRNA in ES Cells, Their Differentiated Derivatives, and in Somatic Cell Lines
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Grabarek, J.B., primary, Wianny, F., additional, Plusa, B., additional, Zernicka-Goetz, M., additional, and Glover, D.M., additional
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- 2003
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13. Polarity of the mouse embryo is anticipated before implantation
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Weber, R. J., primary, Pedersen, R. A., additional, Wianny, F., additional, Evans, M. J., additional, and Zernicka-Goetz, M., additional
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- 1999
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14. Following cell fate in the living mouse embryo
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Zernicka-Goetz, M., primary, Pines, J., additional, Hunter, S. McLean, additional, Dixon, J. P. C., additional, Siemering, K. R., additional, Haseloff, J., additional, and Evans, M. J., additional
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- 1997
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15. An indelible lineage marker for Xenopus using a mutated green fluorescent protein
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Zernicka-Goetz, M., primary, Pines, J., additional, Ryan, K., additional, Siemering, K. R., additional, Haseloff, J., additional, Evans, M. J., additional, and Gurdon, J. B., additional
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- 1996
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16. Cytostatic factor inactivation is induced by a calcium-dependent mechanism present until the second cell cycle in fertilized but not in parthenogenetically activated mouse eggs
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Zernicka-Goetz, M., primary, Ciemerych, M.A., additional, Kubiak, J.Z., additional, Tarkowski, A.K., additional, and Maro, B., additional
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- 1995
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17. Blastomeres arising from the first cleavage division have distinguishable fates in normal mouse development.
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Piotrowska, K, Wianny, F, Pedersen, R A, and Zernicka-Goetz, M
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Two independent studies have recently suggested similar models in which the embryonic and abembryonic parts of the mouse blastocyst become separated already by the first cleavage division. However, no lineage tracing studies carried out so far on early embryos provide the support for such a hypothesis. Thus, to re-examine the fate of blastomeres of the two-cell mouse embryo, we have undertaken lineage tracing studies using a non-perturbing method. We show that two-cell stage blastomeres have a strong tendency to develop into cells that comprise either the embryonic or the abembryonic parts of the blastocyst. Moreover, the two-cell stage blastomere that is first to divide will preferentially contribute its progeny to the embryonic part. Nevertheless, we find that the blastocyst embryonic-abembryonic axis is not perfectly orthogonal to the first cleavage plane, but often shows some angular displacement from it. Consequently, there is a boundary zone adjacent to the interior margin of the blastocoel that is populated by cells derived from both earlier and later dividing blastomeres. The majority of cells that inhabit this boundary region are, however, derived from the later dividing two-cell stage blastomere that contributes predominantly to the abembryonic part of the blastocyst. Thus, at the two-cell stage it is already possible to predict which cell will contribute a greater proportion of its progeny to the abembryonic part of the blastocyst (region including the blastocyst cavity) and which to the embryonic part (region containing the inner cell mass) that will give rise to the embryo proper. more...
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- 2001
18. Animal and vegetal poles of the mouse egg predict the polarity of the embryonic axis, yet are nonessential for development.
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Ciemerych, M A, Mesnard, D, and Zernicka-Goetz, M
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Recent studies suggest early (preimplantation) events might be important in the development of polarity in mammalian embryos. We report here lineage tracing experiments with green fluorescent protein showing that cells located either near to or opposite the polar body at the 8-cell stage of the mouse embryo retain their same relative positions in the blastocyst. Thus they come to lie on either end of an axis of symmetry of the blastocyst that has recently been shown to correlate with the anterior-posterior axis of the postimplantation embryo (see R. J. Weber, R. A. Pedersen, F. Wianny, M. J. Evans and M. Zernicka-Goetz (1999). Development 126, 5591-5598). The embryonic axes of the mouse can therefore be related to the position of the polar body at the 8-cell stage, and by implication, to the animal-vegetal axis of the zygote. However, we also show that chimeric embryos constructed from 2-cell stage blastomeres from which the animal or the vegetal poles have been removed can develop into normal blastocysts and become fertile adult mice. This is also true of chimeras composed of animal or vegetal pole cells derived through normal cleavage to the 8-cell stage. We discuss that although polarity of the postimplantation embryo can be traced back to the 8-cell stage and in turn to the organisation of the egg, it is not absolutely fixed by this time. more...
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- 2000
19. Fertile offspring derived from mammalian eggs lacking either animal or vegetal poles.
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Zernicka-Goetz, M
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In all animals so far tested, removing either pole of the undivided egg prevents normal development: embryos may arrest early, lack organs, or the adults may be sterile. These experiments have shown that spatial patterning of the egg is of utmost importance for subsequent development. However, the significance of spatial patterning in mammalian eggs is still controversial. To test the importance of egg polarity in the mouse a substantial amount of material either from the animal (polar body-associated) or the vegetal (opposite) pole of the fertilised egg was removed. One pole of the egg was cut away manually with a glass needle and the eggs were allowed to develop in vitro. Both kinds of surgical operation permit the development of blastocysts, which, after transfer to the uteri of pseudo-pregnant foster mothers, can produce viable offspring. Furthermore, these develop into fertile adult mice. I conclude that mouse eggs have no essential components that are localised uniquely to the animal or the vegetal pole and, therefore, do not rely for their axial development on maternal determinants that are so localised in the fertilised egg. Thus the mammalian egg appears to be very unusual in the animal kingdom in that it establishes the embryonic axes after the zygote has begun development. more...
- Published
- 1998
20. Following cell fate in the living mouse embryo
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Zernicka-Goetz, M., Pines, J., Hunter, S. M., Dixon, J. P. C., Siemering, K. R., Jim Haseloff, and Evans, M. J.
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It has been difficult to follow many of the dramatic changes in cell fate and cell migration during mouse development. This is because there has been no enduring marker that would allow cells to be recognised in the living embryo. We believe that we have overcome this problem by developing a novel form of green fluorescent protein, named MmGFP, that proves to be easily visible and non toxic to mouse cells and does not perturb embryogenesis. We show that synthetic mRNA encoding MmGFP can be injected into blastomeres to follow the fate of their progeny during preimplantation development. We have made a stable embryonic stem cell line that expresses MmGFP and introduced these fluorescent cells into mouse embryos. For the first time, we have been able to follow the fate of embryonic stem cells in living embryos and to observe directly the contribution of these cells to distinct lineages of the postimplantation embryo. This approach should lead to a more complete description of the dynamics of cell fate in the mouse. more...
21. Polarity of the mouse embryo is anticipated before implantation
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Weber, R. J., Pedersen, R. A., Florence WIANNY, Evans, M. J., and Zernicka-Goetz, M.
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embryonic structures - Abstract
In most species, the polarity of an embryo underlies the future body plan and is determined from that of the zygote. However, mammals are thought to be an exception to this; in the mouse, polarity is generally thought to develop significantly later, only after implantation. It has not been possible, however, to relate the polarity of the preimplantation mouse embryo to that of the later conceptus due to the lack of markers that endure long enough to follow lineages through implantation. To test whether early developmental events could provide cues that predict the axes of the postimplantation embryo, we have used the strategy of injecting mRNA encoding an enduring marker to trace the progeny of inner cell mass cells into the postimplantation visceral endoderm. This tissue, although it has an extraembryonic fate, plays a role in axis determination in adjacent embryonic tissue. We found that visceral endoderm cells that originated near the polar body (a marker of the blastocyst axis of symmetry) generally became distal as the egg cylinder formed, while those that originated opposite the polar body tended to become proximal. It follows that, in normal development, bilateral symmetry of the mouse blastocyst anticipates the polarity of the later conceptus. Moreover, our results show that transformation of the blastocyst axis of symmetry into the axes of the postimplantation conceptus involves asymmetric visceral endoderm cell movement. Therefore, even if the definitive axes of the mouse embryo become irreversibly established only after implantation, this polarity can be traced back to events before implantation. more...
22. An indelible lineage marker for Xenopus using a mutated green fluorescent protein
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Zernicka-Goetz, M., Jonathon Pines, Ryan, K., Siemering, K. R., Haseloff, J., Evans, M. J., and Gurdon, J. B.
- Abstract
We describe the use of a DNA construct (named GFP.RN3) encoding green fluorescent protein as a lineage marker for Xenopus embryos. This offers the following advantages over other lineage markers so far used in Xenopus. When injected as synthetic mRNA, its protein emits intense fluorescence in living embryos. It is non-toxic, and the fluorescence does not bleach when viewed under 480 nm light. It is surprisingly stable, being strongly visible up to the feeding tadpole stage (5 days), and in some tissues for several weeks after mRNA injection. We also describe a construct that encodes a blue fluorescent protein. We exemplify the use of this GFP.RN3 construct for marking the lineage of individual blastomeres at the 32- to 64-cell stage, and as a marker for single transplanted blastula cells. Both procedures have revealed that the descendants of one embryonic cell can contribute single muscle cells to nearly all segmental myotomes rather than predominantly to any one myotome. An independent aim of our work has been to follow the fate of cells in which an early regulatory gene has been temporarily overexpressed. For this purpose, we co-injected GFP.RN3 mRNA and mRNA for the early Xenopus gene Eomes, and found that a high concentration of Eomes results in ectopic muscle gene activation in only the injected cells. This marker may therefore be of general value in providing long term identification of those cells in which an early gene with ephemeral expression has been overexpressed. more...
23. Phospholipase C-ζ-induced Ca2+ oscillations cause coincident cytoplasmic movements in human oocytes that failed to fertilize after intracytoplasmic sperm injection.
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Swann K, Windsor S, Campbell K, Elgmati K, Nomikos M, Zernicka-Goetz M, Amso N, Lai FA, Thomas A, Graham C, Swann, Karl, Windsor, Shane, Campbell, Karen, Elgmati, Khalil, Nomikos, Michail, Zernicka-Goetz, Magdalena, Amso, Nazar, Lai, F Anthony, Thomas, Adrian, and Graham, Christopher more...
- Abstract
Objective: To evaluate the imaging of cytoplasmic movements in human oocytes as a potential method to monitor the pattern of Ca(2+) oscillations during activation.Design: Test of a laboratory technique.Setting: University medical school research laboratory.Patient(s): Donated unfertilized human oocytes from intracytoplasmic sperm injection (ICSI) cycles.Intervention(s): Microinjection of oocytes with phospholipase C (PLC) zeta (ζ) cRNA and a Ca(2+)-sensitive fluorescent dye.Main Outcome Measure(s): Simultaneous detection of oocyte cytoplasmic movements using particle image velocimetry (PIV) and of Ca(2+) oscillations using a Ca(2+)-sensitive fluorescent dye.Result(s): Microinjection of PLCζ cRNA into human oocytes that had failed to fertilize after ICSI resulted in the appearance of prolonged Ca(2+) oscillations. Each transient Ca(2+) concentration change was accompanied by a small coordinated movement of the cytoplasm that could be detected using PIV analysis.Conclusion(s): The occurrence and frequency of cytoplasmic Ca(2+) oscillations, a critical parameter in activating human zygotes, can be monitored by PIV analysis of cytoplasmic movements. This simple method provides a novel, noninvasive approach to determine in real time the occurrence and frequency of Ca(2+) oscillations in human zygotes. [ABSTRACT FROM AUTHOR] more...- Published
- 2012
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24. Primitive to visceral endoderm maturation is essential for mouse epiblast survival beyond implantation.
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Weberling A, Siriwardena D, Penfold C, Christodoulou N, Boroviak TE, and Zernicka-Goetz M
- Abstract
The implantation of the mouse blastocyst initiates a complex sequence of tissue remodeling and cell differentiation events required for morphogenesis, during which the extraembryonic primitive endoderm transitions into the visceral endoderm. Through single-cell RNA sequencing of embryos at embryonic day 5.0, shortly after implantation, we reveal that this transition is driven by dynamic signaling activities, notably the upregulation of BMP signaling and a transient increase in Sox7 expression. Embryos deficient in Hepatocyte nuclear factor-1-beta (Hnf1b-/-), a gene critical for visceral endoderm differentiation, showed an interaction between visceral endoderm and epiblast, crucial for epiblast survival. Single-cell RNA profiling of Hnf1b-/- visceral endoderm shows developmental delays and severe dysregulation in several nutrient transport pathways. Impaired glucose uptake in Hnf1b-/- embryos suggests that the activation of nutrient transport mechanisms during the primitive-to-visceral endoderm transition may be vital for post-implantation epiblast development. These findings offer new insights into the molecular regulation of early mammalian development., Competing Interests: The Authors declare no competing interests., (© 2024 Published by Elsevier Inc.) more...
- Published
- 2024
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25. Temporal BMP4 effects on mouse embryonic and extraembryonic development.
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Hadas R, Rubinstein H, Mittnenzweig M, Mayshar Y, Ben-Yair R, Cheng S, Aguilera-Castrejon A, Reines N, Orenbuch AH, Lifshitz A, Chen DY, Elowitz MB, Zernicka-Goetz M, Hanna JH, Tanay A, and Stelzer Y
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- Animals, Female, Male, Mice, Pregnancy, Cell Differentiation, Cell Lineage, Chorion cytology, Chorion metabolism, Chorion embryology, Ectoderm cytology, Ectoderm metabolism, Ectoderm embryology, Gastrulation, Gene Expression Regulation, Developmental, Mesoderm cytology, Mesoderm embryology, Mesoderm metabolism, Placenta metabolism, Placenta cytology, Placenta embryology, Signal Transduction, Single-Cell Analysis, Time Factors, Trophoblasts cytology, Trophoblasts metabolism, Bone Morphogenetic Protein 4 metabolism, Embryo, Mammalian metabolism, Embryo, Mammalian cytology, Embryo, Mammalian embryology, Embryonic Development, Allantois cytology, Allantois embryology, Allantois metabolism
- Abstract
The developing placenta, which in mice originates through the extraembryonic ectoderm (ExE), is essential for mammalian embryonic development. Yet unbiased characterization of the differentiation dynamics of the ExE and its interactions with the embryo proper remains incomplete. Here we develop a temporal single-cell model of mouse gastrulation that maps continuous and parallel differentiation in embryonic and extraembryonic lineages. This is matched with a three-way perturbation approach to target signalling from the embryo proper, the ExE alone, or both. We show that ExE specification involves early spatial and transcriptional bifurcation of uncommitted ectoplacental cone cells and chorion progenitors. Early BMP4 signalling from chorion progenitors is required for proper differentiation of uncommitted ectoplacental cone cells and later for their specification towards trophoblast giant cells. We also find biphasic regulation by BMP4 in the embryo. The early ExE-originating BMP4 signal is necessary for proper mesoendoderm bifurcation and for allantois and primordial germ cell specification. However, commencing at embryonic day 7.5, embryo-derived BMP4 restricts the primordial germ cell pool size by favouring differentiation of their extraembryonic mesoderm precursors towards an allantois fate. ExE and embryonic tissues are therefore entangled in time, space and signalling axes, highlighting the importance of their integrated understanding and modelling in vivo and in vitro., (© 2024. The Author(s).) more...
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- 2024
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26. Proteome asymmetry in mouse and human embryos before fate specification.
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Iwamoto-Stohl LK, Petelski AA, Meglicki M, Fu A, Khan S, Specht H, Huffman G, Derks J, Jorgensen V, Weatherbee BAT, Weberling A, Gantner CW, Mandelbaum RS, Paulson RJ, Lam L, Ahmady A, Vasquez ES, Slavov N, and Zernicka-Goetz M more...
- Abstract
Pre-patterning of the embryo, driven by spatially localized factors, is a common feature across several non-mammalian species
1-4 . However, mammals display regulative development and thus it was thought that blastomeres of the embryo do not show such pre-patterning, contributing randomly to the three lineages of the blastocyst: the epiblast, primitive endoderm and trophectoderm that will generate the new organism, the yolk sac and placenta respectively4-6 . Unexpectedly, early blastomeres of mouse and human embryos have been reported to have distinct developmental fates, potential and heterogeneous abundance of certain transcripts7-12 . Nevertheless, the extent of the earliest intra-embryo differences remains unclear and controversial. Here, by utilizing multiplexed and label-free single-cell proteomics by mass-spectrometry13 , we show that 2-cell mouse and human embryos contain an alpha and a beta blastomere as defined by differential abundance of hundreds of proteins exhibiting strong functional enrichment for protein synthesis, transport, and degradation. Such asymmetrically distributed proteins include Gps1 and Nedd8, depletion or overexpression of which in one blastomere of the 2-cell embryo impacts lineage segregation. These protein asymmetries increase at 4-cell stage. Intriguingly, halved mouse zygotes display asymmetric protein abundance that resembles alpha and beta blastomeres, suggesting differential proteome localization already within zygotes. We find that beta blastomeres give rise to a blastocyst with a higher proportion of epiblast cells than alpha blastomeres and that vegetal blastomeres, which are known to have a reduced developmental potential, are more likely to be alpha. Human 2-cell blastomeres also partition into two clusters sharing strong concordance with clusters found in mouse, in terms of differentially abundant proteins and functional enrichment. To our knowledge, this is the first demonstration of intra-zygotic and inter-blastomere proteomic asymmetry in mammals that has a role in lineage segregation. more...- Published
- 2024
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27. HIF1A contributes to the survival of aneuploid and mosaic pre-implantation embryos.
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Sanchez-Vasquez E, Bronner ME, and Zernicka-Goetz M
- Abstract
Human fertility is suboptimal, partly due to error-prone divisions in early cleavage-stages that result in aneuploidy. Most human pre-implantation are mosaics of euploid and aneuploid cells, however, mosaic embryos with a low proportion of aneuploid cells have a similar likelihood of developing to term as fully euploid embryos. How embryos manage aneuploidy during development is poorly understood. This knowledge is crucial for improving fertility treatments and reducing developmental defects. To explore these mechanisms, we established a new mouse model of chromosome mosaicism to study the fate of aneuploid cells during pre-implantation development. We previously used the Mps1 inhibitor reversine to generate aneuploidy in embryos. Here, we found that treatment with the more specific Mps1 inhibitor AZ3146 induced chromosome segregation defects in pre-implantation embryos, similar to reversine. However, AZ3146-treated embryos showed a higher developmental potential than reversine-treated embryos. Unlike reversine-treated embryos, AZ3146-treated embryos exhibited transient upregulation of Hypoxia Inducible-Factor-1A (HIF1A) and lacked p53 upregulation. Pre-implantation embryos develop in a hypoxic environment in vivo , and hypoxia exposure in vitro reduced DNA damage in response to Mps1 inhibition and increased the proportion of euploid cells in the mosaic epiblast. Inhibiting HIF1A in mosaic embryos also decreased the proportion of aneuploid cells in mosaic embryos. Our work illuminates potential strategies to improve the developmental potential of mosaic embryos., Competing Interests: DECLARATION OF INTERESTS The authors declare that they have no conflicts of interest. more...
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- 2024
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28. Tead4 and Tfap2c generate bipotency and a bistable switch in totipotent embryos to promote robust lineage diversification.
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Zhu M, Meglicki M, Lamba A, Wang P, Royer C, Turner K, Jauhar MA, Jones C, Child T, Coward K, Na J, and Zernicka-Goetz M
- Subjects
- Animals, Mice, Humans, Signal Transduction, Cell Lineage, Gene Expression Regulation, Developmental, Muscle Proteins metabolism, Muscle Proteins genetics, Embryo, Mammalian metabolism, Embryo, Mammalian cytology, Hippo Signaling Pathway, Protein Serine-Threonine Kinases metabolism, Protein Serine-Threonine Kinases genetics, Embryonic Development genetics, Transcription Factor AP-2 metabolism, Transcription Factor AP-2 genetics, TEA Domain Transcription Factors, Transcription Factors metabolism, Transcription Factors genetics, DNA-Binding Proteins metabolism, DNA-Binding Proteins genetics
- Abstract
The mouse and human embryo gradually loses totipotency before diversifying into the inner cell mass (ICM, future organism) and trophectoderm (TE, future placenta). The transcription factors TFAP2C and TEAD4 with activated RHOA accelerate embryo polarization. Here we show that these factors also accelerate the loss of totipotency. TFAP2C and TEAD4 paradoxically promote and inhibit Hippo signaling before lineage diversification: they drive expression of multiple Hippo regulators while also promoting apical domain formation, which inactivates Hippo. Each factor activates TE specifiers in bipotent cells, while TFAP2C also activates specifiers of the ICM fate. Asymmetric segregation of the apical domain reconciles the opposing regulation of Hippo signaling into Hippo OFF and the TE fate, or Hippo ON and the ICM fate. We propose that the bistable switch established by TFAP2C and TEAD4 is exploited to trigger robust lineage diversification in the developing embryo., (© 2024. This is a U.S. Government work and not under copyright protection in the US; foreign copyright protection may apply.) more...
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- 2024
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29. Embryo models need consistent ethical oversight.
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Zernicka-Goetz M and Hyun I
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- Female, Humans, Pregnancy, Embryo, Mammalian cytology, Embryo, Mammalian embryology, Embryo Research ethics, Embryo Research legislation & jurisprudence, Models, Biological
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- 2024
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30. Basal delamination during mouse gastrulation primes pluripotent cells for differentiation.
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Sato N, Rosa VS, Makhlouf A, Kretzmer H, Sampath Kumar A, Grosswendt S, Mattei AL, Courbot O, Wolf S, Boulanger J, Langevin F, Wiacek M, Karpinski D, Elosegui-Artola A, Meissner A, Zernicka-Goetz M, and Shahbazi MN more...
- Subjects
- Animals, Mice, T-Box Domain Proteins metabolism, T-Box Domain Proteins genetics, Pluripotent Stem Cells cytology, Pluripotent Stem Cells metabolism, Primitive Streak cytology, Primitive Streak metabolism, Fetal Proteins metabolism, Fetal Proteins genetics, Wnt Signaling Pathway, Cell Proliferation, Gene Expression Regulation, Developmental, Embryo, Mammalian cytology, Embryo, Mammalian metabolism, Brachyury Protein, Gastrulation, Cell Differentiation, Germ Layers cytology, Germ Layers metabolism
- Abstract
The blueprint of the mammalian body plan is laid out during gastrulation, when a trilaminar embryo is formed. This process entails a burst of proliferation, the ingression of embryonic epiblast cells at the primitive streak, and their priming toward primitive streak fates. How these different events are coordinated remains unknown. Here, we developed and characterized a 3D culture of self-renewing mouse embryonic cells that captures the main transcriptional and architectural features of the early gastrulating mouse epiblast. Using this system in combination with microfabrication and in vivo experiments, we found that proliferation-induced crowding triggers delamination of cells that express high levels of the apical polarity protein aPKC. Upon delamination, cells become more sensitive to Wnt signaling and upregulate the expression of primitive streak markers such as Brachyury. This mechanistic coupling between ingression and differentiation ensures that the right cell types become specified at the right place during embryonic development., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2024 MRC Laboratory of Molecular Biology. Published by Elsevier Inc. All rights reserved.) more...
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- 2024
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31. Topical section: embryonic models (2023) for Current Opinion in Genetics & Development.
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Handford CE, Junyent S, Jorgensen V, and Zernicka-Goetz M
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- Animals, Female, Pregnancy, Organogenesis, Stem Cells, Mammals, Embryonic Development genetics, Embryo, Mammalian
- Abstract
Stem cell-based mammalian embryo models facilitate the discovery of developmental mechanisms because they are more amenable to genetic and epigenetic perturbations than natural embryos. Here, we highlight exciting recent advances that have yielded a plethora of models of embryonic development. Imperfections in these models highlight gaps in our current understanding and outline future research directions, ushering in an exciting new era for embryology., Competing Interests: Declaration of Competing Interest The authors are inventors on the following patents: 1. Patent applicant: Caltech. Inventors: Magdalena Zernicka-Goetz, Berna Sozen, and Victoria Jorgensen. Application number: 17/692,790. Specific aspect of the paper covered in patent application: Reconstructing human early embryogenesis in vitro with pluripotent stem cells. 2. Patent applicant: Caltech and Cambridge Enterprise Limited. Inventors: Magdalena Zernicka-Goetz, Gianluca Amadei, and Charlotte Handford. Application number: 63/397,630. Specific aspect of the paper covered in patent application: Synthetic embryos. 3. Patent Applicant: Caltech and Cambridge Enterprise Limited. Inventors: Magdalena Zernicka-Goetz, Bailey Weatherbee, and Carlos Gantner. Application number: 63/403,684. Specific aspect of the paper covered in patent application: Stem cell-derived model of the human embryo., (Copyright © 2023 Elsevier Ltd. All rights reserved.) more...
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- 2024
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32. Pluripotent stem cell-derived model of the post-implantation human embryo.
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Weatherbee BAT, Gantner CW, Iwamoto-Stohl LK, Daza RM, Hamazaki N, Shendure J, and Zernicka-Goetz M
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- Female, Humans, Pregnancy, Bone Morphogenetic Proteins, Cell Differentiation, Embryoid Bodies cytology, Germ Layers cytology, Germ Layers embryology, Human Embryonic Stem Cells cytology, Transcription Factors genetics, Transcription Factors metabolism, Embryo Implantation, Embryo, Mammalian cytology, Embryo, Mammalian embryology, Embryonic Development, Models, Biological, Pluripotent Stem Cells cytology
- Abstract
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
1 . 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., (© 2023. The Author(s).) more...- Published
- 2023
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33. Publisher Correction: Pluripotent stem cell-derived model of the post-implantation human embryo.
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Weatherbee BAT, Gantner CW, Iwamoto-Stohl LK, Daza RM, Hamazaki N, Shendure J, and Zernicka-Goetz M
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- 2023
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34. spinDrop: a droplet microfluidic platform to maximise single-cell sequencing information content.
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De Jonghe J, Kaminski TS, Morse DB, Tabaka M, Ellermann AL, Kohler TN, Amadei G, Handford CE, Findlay GM, Zernicka-Goetz M, Teichmann SA, and Hollfelder F
- Subjects
- Animals, Mice, RNA, Single-Cell Analysis methods, Microfluidics, Microfluidic Analytical Techniques methods
- Abstract
Droplet microfluidic methods have massively increased the throughput of single-cell sequencing campaigns. The benefit of scale-up is, however, accompanied by increased background noise when processing challenging samples and the overall RNA capture efficiency is lower. These drawbacks stem from the lack of strategies to enrich for high-quality material or specific cell types at the moment of cell encapsulation and the absence of implementable multi-step enzymatic processes that increase capture. Here we alleviate both bottlenecks using fluorescence-activated droplet sorting to enrich for droplets that contain single viable cells, intact nuclei, fixed cells or target cell types and use reagent addition to droplets by picoinjection to perform multi-step lysis and reverse transcription. Our methodology increases gene detection rates fivefold, while reducing background noise by up to half. We harness these properties to deliver a high-quality molecular atlas of mouse brain development, despite starting with highly damaged input material, and provide an atlas of nascent RNA transcription during mouse organogenesis. Our method is broadly applicable to other droplet-based workflows to deliver sensitive and accurate single-cell profiling at a reduced cost., (© 2023. Springer Nature Limited.) more...
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- 2023
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35. E-cadherin mediates apical membrane initiation site localisation during de novo polarisation of epithelial cavities.
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Liang X, Weberling A, Hii CY, Zernicka-Goetz M, and Buckley CE
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- Animals, Mice, Cell Membrane metabolism, Cell Adhesion, Epithelial Cells metabolism, Cell Polarity, Cadherins genetics, Cadherins metabolism
- Abstract
Individual cells within de novo polarising tubes and cavities must integrate their forming apical domains into a centralised apical membrane initiation site (AMIS). This is necessary to enable organised lumen formation within multi-cellular tissue. Despite the well-documented importance of cell division in localising the AMIS, we have found a division-independent mechanism of AMIS localisation that relies instead on Cadherin-mediated cell-cell adhesion. Our study of de novo polarising mouse embryonic stem cells (mESCs) cultured in 3D suggests that cell-cell adhesion localises apical proteins such as PAR-6 to a centralised AMIS. Unexpectedly, we also found that mESC clusters lacking functional E-cadherin still formed a lumen-like cavity in the absence of AMIS localisation but did so at a later stage of development via a "closure" mechanism, instead of via hollowing. This work suggests that there are two, interrelated mechanisms of apical polarity localisation: cell adhesion and cell division. Alignment of these mechanisms in space allows for redundancy in the system and ensures the development of a coherent epithelial structure within a growing organ., (© 2022 The Authors. Published under the terms of the CC BY 4.0 license.) more...
- Published
- 2022
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36. Mouse embryo model derived exclusively from embryonic stem cells undergoes neurulation and heart development.
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Lau KYC, Rubinstein H, Gantner CW, Hadas R, Amadei G, Stelzer Y, and Zernicka-Goetz M
- Subjects
- Animals, Embryonic Development, Embryonic Stem Cells, Mice, Mouse Embryonic Stem Cells, Embryo, Mammalian, Neurulation
- Abstract
Several in vitro models have been developed to recapitulate mouse embryogenesis solely from embryonic stem cells (ESCs). Despite mimicking many aspects of early development, they fail to capture the interactions between embryonic and extraembryonic tissues. To overcome this difficulty, we have developed a mouse ESC-based in vitro model that reconstitutes the pluripotent ESC lineage and the two extraembryonic lineages of the post-implantation embryo by transcription-factor-mediated induction. This unified model recapitulates developmental events from embryonic day 5.5 to 8.5, including gastrulation; formation of the anterior-posterior axis, brain, and a beating heart structure; and the development of extraembryonic tissues, including yolk sac and chorion. Comparing single-cell RNA sequencing from individual structures with time-matched natural embryos identified remarkably similar transcriptional programs across lineages but also showed when and where the model diverges from the natural program. Our findings demonstrate an extraordinary plasticity of ESCs to self-organize and generate a whole-embryo-like structure., Competing Interests: Declaration of interests M.Z.-G. is an advisory board member of Cell Stem Cell. M.Z.-G. and G.A. wish to declare the filing of a patent on May 5(th), 2022 by Caltech and the University of Cambridge on the “Generation of synthetic embryos from multiple stem cell types.” The patent was filed under the following numbers: CIT File No.: CIT-8826-P and Serial Number: 63/344,251., (Copyright © 2022 The Author(s). Published by Elsevier Inc. All rights reserved.) more...
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- 2022
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37. Stem-cell-based human and mouse embryo models.
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Bao M, Cornwall-Scoones J, and Zernicka-Goetz M
- Subjects
- Animals, Embryonic Development genetics, Humans, Mammals, Mice, Morphogenesis, Embryo, Mammalian, Stem Cells
- Abstract
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., (Copyright © 2022. Published by Elsevier Ltd.) more...
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- 2022
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38. Stain-free detection of embryo polarization using deep learning.
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Shen C, Lamba A, Zhu M, Zhang R, Zernicka-Goetz M, and Yang C
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- Animals, Coloring Agents chemistry, Embryo, Mammalian chemistry, Embryo, Mammalian embryology, Embryonic Development, Fertilization in Vitro, Humans, Mice, Staining and Labeling, Cell Polarity, Deep Learning, Embryo, Mammalian cytology
- Abstract
Polarization of the mammalian embryo at the right developmental time is critical for its development to term and would be valuable in assessing the potential of human embryos. However, tracking polarization requires invasive fluorescence staining, impermissible in the in vitro fertilization clinic. Here, we report the use of artificial intelligence to detect polarization from unstained time-lapse movies of mouse embryos. We assembled a dataset of bright-field movie frames from 8-cell-stage embryos, side-by-side with corresponding images of fluorescent markers of cell polarization. We then used an ensemble learning model to detect whether any bright-field frame showed an embryo before or after onset of polarization. Our resulting model has an accuracy of 85% for detecting polarization, significantly outperforming human volunteers trained on the same data (61% accuracy). We discovered that our self-learning model focuses upon the angle between cells as one known cue for compaction, which precedes polarization, but it outperforms the use of this cue alone. By compressing three-dimensional time-lapsed image data into two-dimensions, we are able to reduce data to an easily manageable size for deep learning processing. In conclusion, we describe a method for detecting a key developmental feature of embryo development that avoids clinically impermissible fluorescence staining., (© 2022. The Author(s).) more...
- Published
- 2022
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39. Lima1 mediates the pluripotency control of membrane dynamics and cellular metabolism.
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Duethorn B, Groll F, Rieger B, Drexler HCA, Brinkmann H, Kremer L, Stehling M, Borowski MT, Mildner K, Zeuschner D, Zernicka-Goetz M, Stemmler MP, Busch KB, Vaquerizas JM, and Bedzhov I
- Subjects
- Animals, Blastocyst, Cell Proliferation, Embryonic Development physiology, Embryonic Stem Cells cytology, Female, Male, Mice, Pluripotent Stem Cells cytology, Cytoskeletal Proteins genetics, Cytoskeletal Proteins metabolism, Embryonic Stem Cells metabolism, Pluripotent Stem Cells metabolism
- Abstract
Lima1 is an extensively studied prognostic marker of malignancy and is also considered to be a tumour suppressor, but its role in a developmental context of non-transformed cells is poorly understood. Here, we characterise the expression pattern and examined the function of Lima1 in mouse embryos and pluripotent stem cell lines. We identify that Lima1 expression is controlled by the naïve pluripotency circuit and is required for the suppression of membrane blebbing, as well as for proper mitochondrial energetics in embryonic stem cells. Moreover, forcing Lima1 expression enables primed mouse and human pluripotent stem cells to be incorporated into murine pre-implantation embryos. Thus, Lima1 is a key effector molecule that mediates the pluripotency control of membrane dynamics and cellular metabolism., (© 2022. The Author(s).) more...
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- 2022
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40. Human embryo polarization requires PLC signaling to mediate trophectoderm specification.
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Zhu M, Shahbazi M, Martin A, Zhang C, Sozen B, Borsos M, Mandelbaum RS, Paulson RJ, Mole MA, Esbert M, Titus S, Scott RT, Campbell A, Fishel S, Gradinaru V, Zhao H, Wu K, Chen ZJ, Seli E, de Los Santos MJ, and Zernicka Goetz M more...
- Subjects
- Actins metabolism, Adult, Embryo Culture Techniques, Female, GATA3 Transcription Factor metabolism, Gene Expression Regulation, Developmental, Gene Expression Regulation, Enzymologic, Humans, Phosphoinositide Phospholipase C, Phospholipase C beta, Pregnancy, Signal Transduction, Time Factors, Young Adult, Body Patterning, Cell Differentiation, Cell Lineage, Cell Polarity, Embryo, Mammalian enzymology
- Abstract
Apico-basal polarization of cells within the embryo is critical for the segregation of distinct lineages during mammalian development. Polarized cells become the trophectoderm (TE), which forms the placenta, and apolar cells become the inner cell mass (ICM), the founding population of the fetus. The cellular and molecular mechanisms leading to polarization of the human embryo and its timing during embryogenesis have remained unknown. Here, we show that human embryo polarization occurs in two steps: it begins with the apical enrichment of F-actin and is followed by the apical accumulation of the PAR complex. This two-step polarization process leads to the formation of an apical domain at the 8-16 cell stage. Using RNA interference, we show that apical domain formation requires Phospholipase C (PLC) signaling, specifically the enzymes PLCB1 and PLCE1, from the eight-cell stage onwards. Finally, we show that although expression of the critical TE differentiation marker GATA3 can be initiated independently of embryo polarization, downregulation of PLCB1 and PLCE1 decreases GATA3 expression through a reduction in the number of polarized cells. Therefore, apical domain formation reinforces a TE fate. The results we present here demonstrate how polarization is triggered to regulate the first lineage segregation in human embryos., Competing Interests: MZ, MS, AM, CZ, BS, MB, RM, RP, MM, ME, ST, RS, AC, SF, VG, HZ, KW, ZC, ES, Md, MZ No competing interests declared, (© 2021, Zhu et al.) more...
- Published
- 2021
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41. Reconstructing aspects of human embryogenesis with pluripotent stem cells.
- Author
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Sozen B, Jorgensen V, Weatherbee BAT, Chen S, Zhu M, and Zernicka-Goetz M
- Subjects
- Adaptor Proteins, Signal Transducing genetics, Adaptor Proteins, Signal Transducing metabolism, Biomarkers metabolism, Blastocyst metabolism, Cell Lineage genetics, Embryo, Mammalian anatomy & histology, Embryo, Mammalian metabolism, GATA3 Transcription Factor genetics, GATA3 Transcription Factor metabolism, Gene Expression, Humans, Phospholipase C beta genetics, Phospholipase C beta metabolism, Pluripotent Stem Cells metabolism, SOXB1 Transcription Factors genetics, SOXB1 Transcription Factors metabolism, SOXF Transcription Factors genetics, SOXF Transcription Factors metabolism, Sequence Analysis, RNA, Single-Cell Analysis, Blastocyst cytology, Cell Culture Techniques, Embryo, Mammalian cytology, Embryonic Development genetics, Models, Biological, Pluripotent Stem Cells cytology
- Abstract
Understanding human development is of fundamental biological and clinical importance. Despite its significance, mechanisms behind human embryogenesis remain largely unknown. Here, we attempt to model human early embryo development with expanded pluripotent stem cells (EPSCs) in 3-dimensions. We define a protocol that allows us to generate self-organizing cystic structures from human EPSCs that display some hallmarks of human early embryogenesis. These structures mimic polarization and cavitation characteristic of pre-implantation development leading to blastocyst morphology formation and the transition to post-implantation-like organization upon extended culture. Single-cell RNA sequencing of these structures reveals subsets of cells bearing some resemblance to epiblast, hypoblast and trophectoderm lineages. Nevertheless, significant divergences from natural blastocysts persist in some key markers, and signalling pathways point towards ways in which morphology and transcriptional-level cell identities may diverge in stem cell models of the embryo. Thus, this stem cell platform provides insights into the design of stem cell models of embryogenesis., (© 2021. The Author(s).) more...
- Published
- 2021
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42. Modelling the impact of decidual senescence on embryo implantation in human endometrial assembloids.
- Author
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Rawlings TM, Makwana K, Taylor DM, Molè MA, Fishwick KJ, Tryfonos M, Odendaal J, Hawkes A, Zernicka-Goetz M, Hartshorne GM, Brosens JJ, and Lucas ES
- Subjects
- Coculture Techniques, Decidua physiology, Female, Humans, Organoids, Pregnancy, Cellular Senescence, Embryo Implantation physiology, Endometrium cytology, Stromal Cells cytology
- Abstract
Decidual remodelling of midluteal endometrium leads to a short implantation window after which the uterine mucosa either breaks down or is transformed into a robust matrix that accommodates the placenta throughout pregnancy. To gain insights into the underlying mechanisms, we established and characterized endometrial assembloids, consisting of gland-like organoids and primary stromal cells. Single-cell transcriptomics revealed that decidualized assembloids closely resemble midluteal endometrium, harbouring differentiated and senescent subpopulations in both glands and stroma. We show that acute senescence in glandular epithelium drives secretion of multiple canonical implantation factors, whereas in the stroma it calibrates the emergence of anti-inflammatory decidual cells and pro-inflammatory senescent decidual cells. Pharmacological inhibition of stress responses in pre-decidual cells accelerated decidualization by eliminating the emergence of senescent decidual cells. In co-culture experiments, accelerated decidualization resulted in entrapment of collapsed human blastocysts in a robust, static decidual matrix. By contrast, the presence of senescent decidual cells created a dynamic implantation environment, enabling embryo expansion and attachment, although their persistence led to gradual disintegration of assembloids. Our findings suggest that decidual senescence controls endometrial fate decisions at implantation and highlight how endometrial assembloids may accelerate the discovery of new treatments to prevent reproductive failure., Competing Interests: TR, KM, DT, MM, KF, MT, JO, AH, MZ, GH, JB, EL No competing interests declared, (© 2021, Rawlings et al.) more...
- Published
- 2021
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43. An in vitro stem cell model of human epiblast and yolk sac interaction.
- Author
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Mackinlay KM, Weatherbee BA, Souza Rosa V, Handford CE, Hudson G, Coorens T, Pereira LV, Behjati S, Vallier L, Shahbazi MN, and Zernicka-Goetz M
- Subjects
- Animals, Cell Line, Ectoderm growth & development, Embryonic Development, Humans, Mice, Germ Layers growth & development, Pluripotent Stem Cells metabolism, Yolk Sac growth & development
- Abstract
Human embryogenesis entails complex signalling interactions between embryonic and extra-embryonic cells. However, how extra-embryonic cells direct morphogenesis within the human embryo remains largely unknown due to a lack of relevant stem cell models. Here, we have established conditions to differentiate human pluripotent stem cells (hPSCs) into yolk sac-like cells (YSLCs) that resemble the post-implantation human hypoblast molecularly and functionally. YSLCs induce the expression of pluripotency and anterior ectoderm markers in human embryonic stem cells (hESCs) at the expense of mesoderm and endoderm markers. This activity is mediated by the release of BMP and WNT signalling pathway inhibitors, and, therefore, resembles the functioning of the anterior visceral endoderm signalling centre of the mouse embryo, which establishes the anterior-posterior axis. Our results implicate the yolk sac in epiblast cell fate specification in the human embryo and propose YSLCs as a tool for studying post-implantation human embryo development in vitro., Competing Interests: KM, BW, VS, CH, GH, TC, LP, SB, LV, MS, MZ No competing interests declared, (© 2021, Mackinlay et al.) more...
- Published
- 2021
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44. A single cell characterisation of human embryogenesis identifies pluripotency transitions and putative anterior hypoblast centre.
- Author
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Molè MA, Coorens THH, Shahbazi MN, Weberling A, Weatherbee BAT, Gantner CW, Sancho-Serra C, Richardson L, Drinkwater A, Syed N, Engley S, Snell P, Christie L, Elder K, Campbell A, Fishel S, Behjati S, Vento-Tormo R, and Zernicka-Goetz M more...
- Subjects
- Bone Morphogenetic Protein 1 antagonists & inhibitors, Cell Lineage, Cells, Cultured, Embryo Implantation physiology, Embryo, Mammalian, Fibroblast Growth Factors metabolism, Gastrulation physiology, Germ Layers cytology, Humans, Image Processing, Computer-Assisted, Multigene Family, Nodal Protein antagonists & inhibitors, RNA-Seq, Spatio-Temporal Analysis, Embryo Implantation genetics, Embryonic Development, Gastrulation genetics, Gene Expression Regulation, Developmental genetics, Germ Layers metabolism, Single-Cell Analysis methods, Wnt Signaling Pathway
- Abstract
Following implantation, the human embryo undergoes major morphogenetic transformations that establish the future body plan. While the molecular events underpinning this process are established in mice, they remain unknown in humans. Here we characterise key events of human embryo morphogenesis, in the period between implantation and gastrulation, using single-cell analyses and functional studies. First, the embryonic epiblast cells transition through different pluripotent states and act as a source of FGF signals that ensure proliferation of both embryonic and extra-embryonic tissues. In a subset of embryos, we identify a group of asymmetrically positioned extra-embryonic hypoblast cells expressing inhibitors of BMP, NODAL and WNT signalling pathways. We suggest that this group of cells can act as the anterior singalling centre to pattern the epiblast. These results provide insights into pluripotency state transitions, the role of FGF signalling and the specification of anterior-posterior axis during human embryo development. more...
- Published
- 2021
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45. Unifying synthetic embryology.
- Author
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Cornwall-Scoones J and Zernicka-Goetz M
- Subjects
- Animals, Embryo, Mammalian cytology, Embryo, Mammalian physiology, Humans, Stem Cells cytology, Stem Cells physiology, Embryology, Synthetic Biology
- Abstract
Competing Interests: Declaration of Competing interests The authors declare no competing interests.
- Published
- 2021
- Full Text
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46. Modeling human embryo development with embryonic and extra-embryonic stem cells.
- Author
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Weatherbee BAT, Cui T, and Zernicka-Goetz M
- Subjects
- Animals, Embryo, Mammalian cytology, Embryoid Bodies cytology, Embryoid Bodies metabolism, Embryonic Stem Cells cytology, Germ Layers cytology, Humans, Stem Cells cytology, Trophoblasts cytology, Embryo, Mammalian metabolism, Embryonic Development, Embryonic Stem Cells metabolism, Stem Cells metabolism
- Abstract
Early human post-implantation development involves extensive growth combined with a series of complex morphogenetic events. The lack of precise spatial and temporal control over these processes leads to pregnancy loss. Given the ethical and technical limitations in studying the natural human embryo, alternative approaches are needed to investigate mechanisms underlying this critical stage of human development. Here, we present an overview of the different stem cells and stem cell-derived models which serve as useful, albeit imperfect, tools in understanding human embryogenesis. Current models include stem cells that represent each of the three earliest lineages: human embryonic stem cells corresponding to the epiblast, hypoblast-like stem cells and trophoblast stem cells. We also review the use of human embryonic stem cells to model complex aspects of epiblast morphogenesis and differentiation. Additionally, we propose that the combination of both embryonic and extra-embryonic stem cells to form three-dimensional embryo models will provide valuable insights into cell-cell chemical and mechanical interactions that are essential for natural embryogenesis., (Copyright © 2020 Elsevier Inc. All rights reserved.) more...
- Published
- 2021
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47. The dynamics of morphogenesis in stem cell-based embryology: Novel insights for symmetry breaking.
- Author
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Sozen B, Cornwall-Scoones J, and Zernicka-Goetz M
- Subjects
- Animals, Body Patterning, Embryo, Mammalian cytology, Embryo, Mammalian metabolism, Humans, Mice, Signal Transduction, Stem Cells cytology, Embryonic Development, Morphogenesis
- Abstract
Breaking embryonic symmetry is an essential prerequisite to shape the initially symmetric embryo into a highly organized body plan that serves as the blueprint of the adult organism. This critical process is driven by morphogen signaling gradients that instruct anteroposterior axis specification. Despite its fundamental importance, what triggers symmetry breaking and how the signaling gradients are established in time and space in the mammalian embryo remain largely unknown. Stem cell-based in vitro models of embryogenesis offer an unprecedented opportunity to quantitatively dissect the multiple physical and molecular processes that shape the mammalian embryo. Here we review biochemical mechanisms governing early mammalian patterning in vivo and highlight recent advances to recreate this in vitro using stem cells. We discuss how the novel insights from these model systems extend previously proposed concepts to illuminate the extent to which embryonic cells have the intrinsic capability to generate specific, reproducible patterns during embryogenesis., (Published by Elsevier Inc.) more...
- Published
- 2021
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- View/download PDF
48. Machine learning-assisted high-content analysis of pluripotent stem cell-derived embryos in vitro.
- Author
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Guo J, Wang P, Sozen B, Qiu H, Zhu Y, Zhang X, Ming J, Zernicka-Goetz M, and Na J
- Subjects
- Animals, Bone Morphogenetic Protein 4 metabolism, Cell Polarity drug effects, Cytokines metabolism, Ectoderm cytology, Embryo Implantation drug effects, Endoderm cytology, Gene Expression Regulation, Developmental drug effects, Induced Pluripotent Stem Cells drug effects, Induced Pluripotent Stem Cells metabolism, Mice, Morphogenesis drug effects, Morphogenesis genetics, Small Molecule Libraries pharmacology, Transcriptome genetics, Trophoblasts cytology, Trophoblasts drug effects, Trophoblasts metabolism, Embryo, Mammalian cytology, Induced Pluripotent Stem Cells cytology, Machine Learning
- Abstract
Stem cell-based embryo models by cultured pluripotent and extra-embryonic lineage stem cells are novel platforms to model early postimplantation development. We showed that induced pluripotent stem cells (iPSCs) could form ITS (iPSCs and trophectoderm stem cells) and ITX (iPSCs, trophectoderm stem cells, and XEN cells) embryos, resembling the early gastrula embryo developed in vivo. To facilitate the efficient and unbiased analysis of the stem cell-based embryo model, we set up a machine learning workflow to extract multi-dimensional features and perform quantification of ITS embryos using 3D images collected from a high-content screening system. We found that different PSC lines differ in their ability to form embryo-like structures. Through high-content screening of small molecules and cytokines, we identified that BMP4 best promoted the morphogenesis of the ITS embryo. Our study established an innovative strategy to analyze stem cell-based embryo models and uncovered new roles of BMP4 in stem cell-based embryo models., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.) more...
- Published
- 2021
- Full Text
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49. Embryo size regulates the timing and mechanism of pluripotent tissue morphogenesis.
- Author
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Orietti LC, Rosa VS, Antonica F, Kyprianou C, Mansfield W, Marques-Souza H, Shahbazi MN, and Zernicka-Goetz M
- Subjects
- Amnion embryology, Animals, Apoptosis, Cell Aggregation, Embryo, Mammalian cytology, Embryonic Stem Cells cytology, Embryonic Stem Cells metabolism, Female, Mice, Inbred C57BL, Mice, Inbred CBA, Organ Size, Time Factors, Mice, Embryo, Mammalian anatomy & histology, Morphogenesis
- Abstract
Mammalian embryogenesis is a paradigm of regulative development as mouse embryos show plasticity in the regulation of cell fate, cell number, and tissue morphogenesis. However, the mechanisms behind embryo plasticity remain largely unknown. Here, we determine how mouse embryos respond to an increase in cell numbers to regulate the timing and mechanism of embryonic morphogenesis, leading to the formation of the pro-amniotic cavity. Using embryos and embryonic stem cell aggregates of different size, we show that while pro-amniotic cavity formation in normal-sized embryos is achieved through basement membrane-induced polarization and exocytosis, cavity formation of increased-size embryos is delayed and achieved through apoptosis of cells that lack contact with the basement membrane. Importantly, blocking apoptosis, both genetically and pharmacologically, alters pro-amniotic cavity formation but does not affect size regulation in enlarged embryos. We conclude that the regulation of embryonic size and morphogenesis, albeit concomitant, have distinct molecular underpinnings., (Copyright © 2020 The Authors. Published by Elsevier Inc. All rights reserved.) more...
- Published
- 2021
- Full Text
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50. Integrin β1 coordinates survival and morphogenesis of the embryonic lineage upon implantation and pluripotency transition.
- Author
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Molè MA, Weberling A, Fässler R, Campbell A, Fishel S, and Zernicka-Goetz M
- Subjects
- Actomyosin metabolism, Amides pharmacology, Animals, Embryo Implantation, Embryo, Mammalian cytology, Embryo, Mammalian embryology, Embryo, Mammalian metabolism, Embryonic Development, Female, Fibroblast Growth Factor 2 pharmacology, Germ Layers growth & development, Glycogen Synthase Kinase 3 antagonists & inhibitors, Glycogen Synthase Kinase 3 metabolism, Humans, Insulin-Like Growth Factor I pharmacology, Integrin beta1 genetics, Mice, Mice, Inbred C57BL, Mice, Inbred CBA, Mouse Embryonic Stem Cells cytology, Mouse Embryonic Stem Cells metabolism, Pyridines pharmacology, Signal Transduction, rho-Associated Kinases antagonists & inhibitors, rho-Associated Kinases metabolism, Integrin beta1 metabolism, Morphogenesis drug effects
- Abstract
At implantation, the embryo establishes contacts with the maternal endometrium. This stage is associated with a high incidence of preclinical pregnancy losses. While the maternal factors underlying uterine receptivity have been investigated, the signals required by the embryo for successful peri-implantation development remain elusive. To explore these, we studied integrin β1 signaling, as embryos deficient for this receptor degenerate at implantation. We demonstrate that the coordinated action of pro-survival signals and localized actomyosin suppression via integrin β1 permits the development of the embryo beyond implantation. Failure of either process leads to developmental arrest and apoptosis. Pharmacological stimulation through fibroblast growth factor 2 (FGF2) and insulin-like growth factor 1 (IGF1), coupled with ROCK-mediated actomyosin inhibition, rescues the deficiency of integrin β1, promoting progression to post-implantation stages. Mutual exclusion between integrin β1 and actomyosin seems to be conserved in the human embryo, suggesting the possibility that these mechanisms could also underlie the transition of the human epiblast from pre- to post-implantation., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.) more...
- Published
- 2021
- Full Text
- View/download PDF
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