Gastruloid

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An example of a Gastruloid formed from Brachyury::GFP mouse ESCs, treated with a pulse of the Wnt/b-Catenin agonist CHIR99021 between 48 and 72h and imaged by wide-field fluorescence microscopy at 120h. Notice the polarised expression of Brachyury::GFP (Bra) at the elongating tip of the Gastruloid. Image from van den Brink et al. (2014), used with CC-BY licence. Example of a Gastruloid.png
An example of a Gastruloid formed from Brachyury::GFP mouse ESCs, treated with a pulse of the Wnt/β-Catenin agonist CHIR99021 between 48 and 72h and imaged by wide-field fluorescence microscopy at 120h. Notice the polarised expression of Brachyury::GFP (Bra) at the elongating tip of the Gastruloid. Image from van den Brink et al. (2014), used with CC-BY licence.

Gastruloids are three dimensional aggregates of embryonic stem cells (ESCs) that, when cultured in specific conditions, exhibit an organization resembling that of an embryo. They develop with three orthogonal axes and contain the primordial cells for various tissues derived from the three germ layers, without the presence of extraembryonic tissues. Notably, they do not possess forebrain, midbrain, and hindbrain structures. Gastruloids serve as a valuable model system for studying mammalian development, including human development, as well as diseases associated with it. They are a model system an embryonic organoid for the study of mammalian development (including humans) and disease. [1] [2] [3]

Background

The Gastruloid model system draws its origins from work by Marikawa et al.. [4] In that study, small numbers of mouse P19 embryonal carcinoma (EC) cells, were aggregated as embryoid bodies (EBs) and used to model and investigate the processes involved in anteroposterior polarity and the formation of a primitive streak region. [4] In this work, the EBs were able to organise themselves into structures with polarised gene expression, axial elongation/organisation and up-regulation of posterior mesodermal markers. This was in stark contrast to work using EBs from mouse ESCs, which had shown some polarisation of gene expression in a small number of cases but no further development of the multicellular system. [5] [6]

Following this study, the Martinez Arias laboratory in the Department of Genetics at the University of Cambridge demonstrated how aggregates of mouse embryonic stem cells (ESCs) were able to generate structures that exhibited collective behaviours with striking similarity to those during early development such as symmetry-breaking (in terms of gene expression), axial elongation and germ-layer specification. [1] [2] [3] To quote from the original paper: "Altogether, these observations further emphasize the similarity between the processes that we have uncovered here and the events in the embryo. The movements are related to those of cells in gastrulating embryos and for this reason we term these aggregates ‘gastruloids’". As noted by the authors of this protocol, a crucial difference between this culture method and previous work with mouse EBs was the use of small numbers of cells which may be important for generating the correct length scale for patterning, and the use of culture conditions derived from directed differentiation of ESCs in adherent culture [1] [7] [3] [2] [8]

Brachyury (T/Bra), a gene which marks the primitive streak and the site of gastrulation, is up-regulated in the Gastruloids following a pulse of the Wnt/β-Catenin agonist CHIR99021 [9] (Chi; other factors have also been tested [1] ) and becomes regionalised to the elongating tip of the Gastruloid. From or near the region expressing T/Bra, cells expressing the mesodermal marker tbx6 are extruded from the similar to cells in the gastrulating embryo; it is for this reason that these structures are called Gastruloids. [1]

Further studies revealed that the events that specify T/Bra expression in gastruloids mimic those in the embryo. [2] After seven days gastruloids exhibit an organization very similar to a midgestation embryo with spatially organized primordia for all mesodermal (axial, paraxial, intermediate, cardiac, cranial and hematopoietic) and endodermal derivatives as well as the spinal cord. [10] [11] [3] They also implement Hox gene expression with the spatiotemporal coordinates as the embryo. [3] Gastruloids lack brain as well as extraembryonic tissues but characterisation of the cellular complexity of gastruloids at the level of single cell and spatial transcriptomics, reveals that they contain representatives of the three germ layers including neural crest, Primordial Germ cells and placodal primordia. [12] [13]

A feature of gastruloids is a disconnect between the transcriptional programs and outlines and the morphogenesis. However, changes in the culture conditions can elicit morphogenesis, most significantly gastruloids have been shown to form somites [13] [12] and early cardiac structures. [14] In addition, interactions between gastruloids and extraembryonic tissues promote an anterior, brain-like polarised tissue. [15]

Gastruloids have recently been obtained from human ESCs, [16] which gives developmental biologists the ability to study early human development without needing human embryos. Importantly though, the human gastruloid model is not able to form a human embryo, meaning that is a non-intact, non-viable and non-equivalent to in vivo human embryos.

The term Gastruloid has been expanded to include self-organised human embryonic stem cell arrangements on patterned (micro patterns) that mimic early patterning events in development; [17] [18] these arrangements should be referred to as 2D gastruloids.

Related Research Articles

Developmental biology is the study of the process by which animals and plants grow and develop. Developmental biology also encompasses the biology of regeneration, asexual reproduction, metamorphosis, and the growth and differentiation of stem cells in the adult organism.

<span class="mw-page-title-main">Stem cell</span> Undifferentiated biological cells that can differentiate into specialized cells

In multicellular organisms, stem cells are undifferentiated or partially differentiated cells that can change into various types of cells and proliferate indefinitely to produce more of the same stem cell. They are the earliest type of cell in a cell lineage. They are found in both embryonic and adult organisms, but they have slightly different properties in each. They are usually distinguished from progenitor cells, which cannot divide indefinitely, and precursor or blast cells, which are usually committed to differentiating into one cell type.

<span class="mw-page-title-main">Mesoderm</span> Middle germ layer of embryonic development

The mesoderm is the middle layer of the three germ layers that develops during gastrulation in the very early development of the embryo of most animals. The outer layer is the ectoderm, and the inner layer is the endoderm.

<span class="mw-page-title-main">Gastrulation</span> Stage in embryonic development in which germ layers form

Gastrulation is the stage in the early embryonic development of most animals, during which the blastula, or in mammals the blastocyst, is reorganized into a two-layered or three-layered embryo known as the gastrula. Before gastrulation, the embryo is a continuous epithelial sheet of cells; by the end of gastrulation, the embryo has begun differentiation to establish distinct cell lineages, set up the basic axes of the body, and internalized one or more cell types including the prospective gut.

<span class="mw-page-title-main">Germ cell</span> Gamete-producing cell

A germ cell is any cell that gives rise to the gametes of an organism that reproduces sexually. In many animals, the germ cells originate in the primitive streak and migrate via the gut of an embryo to the developing gonads. There, they undergo meiosis, followed by cellular differentiation into mature gametes, either eggs or sperm. Unlike animals, plants do not have germ cells designated in early development. Instead, germ cells can arise from somatic cells in the adult, such as the floral meristem of flowering plants.

<span class="mw-page-title-main">Blastocyst</span> Structure formed around day 5 of mammalian embryonic development

The blastocyst is a structure formed in the early embryonic development of mammals. It possesses an inner cell mass (ICM) also known as the embryoblast which subsequently forms the embryo, and an outer layer of trophoblast cells called the trophectoderm. This layer surrounds the inner cell mass and a fluid-filled cavity or lumen known as the blastocoel. In the late blastocyst, the trophectoderm is known as the trophoblast. The trophoblast gives rise to the chorion and amnion, the two fetal membranes that surround the embryo. The placenta derives from the embryonic chorion and the underlying uterine tissue of the mother.

<span class="mw-page-title-main">Embryonic stem cell</span> Type of pluripotent blastocystic stem cell

Embryonic stem cells (ESCs) are pluripotent stem cells derived from the inner cell mass of a blastocyst, an early-stage pre-implantation embryo. Human embryos reach the blastocyst stage 4–5 days post fertilization, at which time they consist of 50–150 cells. Isolating the inner cell mass (embryoblast) using immunosurgery results in destruction of the blastocyst, a process which raises ethical issues, including whether or not embryos at the pre-implantation stage have the same moral considerations as embryos in the post-implantation stage of development.

<span class="mw-page-title-main">Embryoid body</span> Three-dimensional aggregate of pluripotent stem cells

Embryoid bodies (EBs) are three-dimensional aggregates formed by pluripotent stem cells. These include embryonic stem cells (ESC) and induced pluripotent stem cells (iPSC)

<span class="mw-page-title-main">Homeobox protein NANOG</span> Mammalian protein found in humans

Homeobox protein NANOG(hNanog) is a transcriptional factor that helps embryonic stem cells (ESCs) maintain pluripotency by suppressing cell determination factors. hNanog is encoded in humans by the NANOG gene. Several types of cancer are associated with NANOG.

<span class="mw-page-title-main">Stem-cell line</span> Culture of stem cells that can be propagated indefinitely

A stem cell line is a group of stem cells that is cultured in vitro and can be propagated indefinitely. Stem cell lines are derived from either animal or human tissues and come from one of three sources: embryonic stem cells, adult stem cells, or induced stem cells. They are commonly used in research and regenerative medicine.

The stem cell controversy concerns the ethics of research involving the development and use of human embryos. Most commonly, this controversy focuses on embryonic stem cells. Not all stem cell research involves human embryos. For example, adult stem cells, amniotic stem cells, and induced pluripotent stem cells do not involve creating, using, or destroying human embryos, and thus are minimally, if at all, controversial. Many less controversial sources of acquiring stem cells include using cells from the umbilical cord, breast milk, and bone marrow, which are not pluripotent.

<span class="mw-page-title-main">Organoid</span> Miniaturized and simplified version of an organ

An organoid is a miniaturised and simplified version of an organ produced in vitro in three dimensions that mimics the key functional, structural, and biological complexity of that organ. It is derived from one or a few cells from a tissue, embryonic stem cells, or induced pluripotent stem cells, which can self-organize in three-dimensional culture owing to their self-renewal and differentiation capacities. The technique for growing organoids has rapidly improved since the early 2010s, and The Scientist named it one of the biggest scientific advancements of 2013. Scientists and engineers use organoids to study development and disease in the laboratory, for drug discovery and development in industry, personalized diagnostics and medicine, gene and cell therapies, tissue engineering, and regenerative medicine.

Hemangioblasts are the multipotent precursor cells that can differentiate into both hematopoietic and endothelial cells. In the mouse embryo, the emergence of blood islands in the yolk sac at embryonic day 7 marks the onset of hematopoiesis. From these blood islands, the hematopoietic cells and vasculature are formed shortly after. Hemangioblasts are the progenitors that form the blood islands. To date, the hemangioblast has been identified in human, mouse and zebrafish embryos.

<span class="mw-page-title-main">Endothelial stem cell</span> Stem cell in bone marrow that gives rise to endothelial cells

Endothelial stem cells (ESCs) are one of three types of stem cells found in bone marrow. They are multipotent, which describes the ability to give rise to many cell types, whereas a pluripotent stem cell can give rise to all types. ESCs have the characteristic properties of a stem cell: self-renewal and differentiation. These parent stem cells, ESCs, give rise to progenitor cells, which are intermediate stem cells that lose potency. Progenitor stem cells are committed to differentiating along a particular cell developmental pathway. ESCs will eventually produce endothelial cells (ECs), which create the thin-walled endothelium that lines the inner surface of blood vessels and lymphatic vessels. The blood vessels include arteries and veins. Endothelial cells can be found throughout the whole vascular system and they also play a vital role in the movement of white blood cells

<span class="mw-page-title-main">Epiblast</span> Embryonic inner cell mass tissue that forms the embryo itself, through the three germ layers

In amniote embryonic development, the epiblast is one of two distinct cell layers arising from the inner cell mass in the mammalian blastocyst, or from the blastula in reptiles and birds, the other layer is the hypoblast. It drives the embryo proper through its differentiation into the three primary germ layers, ectoderm, mesoderm and endoderm, during gastrulation. The amniotic ectoderm and extraembryonic mesoderm also originate from the epiblast.

<span class="mw-page-title-main">Induced pluripotent stem cell</span> Pluripotent stem cell generated directly from a somatic cell

Induced pluripotent stem cells are a type of pluripotent stem cell that can be generated directly from a somatic cell. The iPSC technology was pioneered by Shinya Yamanaka and Kazutoshi Takahashi in Kyoto, Japan, who together showed in 2006 that the introduction of four specific genes, collectively known as Yamanaka factors, encoding transcription factors could convert somatic cells into pluripotent stem cells. Shinya Yamanaka was awarded the 2012 Nobel Prize along with Sir John Gurdon "for the discovery that mature cells can be reprogrammed to become pluripotent."

<span class="mw-page-title-main">Cell potency</span> Ability of a cell to differentiate into other cell types

Cell potency is a cell's ability to differentiate into other cell types. The more cell types a cell can differentiate into, the greater its potency. Potency is also described as the gene activation potential within a cell, which like a continuum, begins with totipotency to designate a cell with the most differentiation potential, pluripotency, multipotency, oligopotency, and finally unipotency.

Directed differentiation is a bioengineering methodology at the interface of stem cell biology, developmental biology and tissue engineering. It is essentially harnessing the potential of stem cells by constraining their differentiation in vitro toward a specific cell type or tissue of interest. Stem cells are by definition pluripotent, able to differentiate into several cell types such as neurons, cardiomyocytes, hepatocytes, etc. Efficient directed differentiation requires a detailed understanding of the lineage and cell fate decision, often provided by developmental biology.

A blastoid is an embryoid, a stem cell-based embryo model which, morphologically and transcriptionally resembles the early, pre-implantation, mammalian conceptus, called the blastocyst. The first blastoids were created by the Nicolas Rivron laboratory by combining mouse embryonic stem cells and mouse trophoblast stem cells. Upon in vitro development, blastoids generate analogs of the primitive endoderm cells, thus comprising analogs of the three founding cell types of the conceptus, and recapitulate aspects of implantation on being introduced into the uterus of a compatible female. Mouse blastoids have not shown the capacity to support the development of a foetus and are thus generally not considered as an embryo but rather as a model. As compared to other stem cell-based embryo models, blastoids model the preimplantation stage and the integrated development of the conceptus including the embryo proper and the two extraembryonic tissues. The blastoid is a model system for the study of mammalian development and disease. It might be useful for the identification of therapeutic targets and preclinical modelling.

<span class="mw-page-title-main">Jacob Hanna</span> Israeli Arab researcher in molecular biochemistry

JacobH. Hanna is a Palestinian Arab-Israeli biologist who is working as a professor in the Department of Molecular Genetics at the Weizmann Institute of Science in Rehovot, Israel. An expert in embryonic stem cell research, he is most recognized for developing the first bona fide synthetic embryo models from stem cells in the petri dish in mice and humans.

References

  1. 1 2 3 4 5 Brink, Susanne C. van den; Baillie-Johnson, Peter; Balayo, Tina; Hadjantonakis, Anna-Katerina; Nowotschin, Sonja; Turner, David A.; Arias, Alfonso Martinez (2014-11-15). "Symmetry breaking, germ layer specification and axial organisation in aggregates of mouse embryonic stem cells". Development. 141 (22): 4231–4242. doi:10.1242/dev.113001. ISSN   0950-1991. PMC   4302915 . PMID   25371360.
  2. 1 2 3 4 Turner, David A.; Girgin, Mehmet; Alonso-Crisostomo, Luz; Trivedi, Vikas; Baillie-Johnson, Peter; Glodowski, Cherise R.; Hayward, Penelope C.; Collignon, Jérôme; Gustavsen, Carsten (2017-11-01). "Anteroposterior polarity and elongation in the absence of extra-embryonic tissues and of spatially localised signalling in gastruloids: mammalian embryonic organoids". Development. 144 (21): 3894–3906. doi:10.1242/dev.150391. ISSN   0950-1991. PMC   5702072 . PMID   28951435.
  3. 1 2 3 4 5 Beccari, Leonardo; Moris, Naomi; Girgin, Mehmet; Turner, David A.; Baillie-Johnson, Peter; Cossy, Anne-Catherine; Lutolf, Matthias P.; Duboule, Denis; Arias, Alfonso Martinez (October 2018). "Multi-axial self-organization properties of mouse embryonic stem cells into gastruloids". Nature. 562 (7726): 272–276. Bibcode:2018Natur.562..272B. doi:10.1038/s41586-018-0578-0. ISSN   0028-0836. PMID   30283134. S2CID   52915553.
  4. 1 2 Marikawa, Yusuke; Tamashiro, Dana Ann A.; Fujita, Toko C.; Alarcón, Vernadeth B. (2009-02-01). "Aggregated P19 mouse embryonal carcinoma cells as a simple in vitro model to study the molecular regulations of mesoderm formation and axial elongation morphogenesis". Genesis. 47 (2): 93–106. doi:10.1002/dvg.20473. ISSN   1526-968X. PMC   3419260 . PMID   19115346.
  5. ↑ Leahy, Amy; Xiong, Jing-Wei; Kuhnert, Frank; Stuhlmann, Heidi (1999). "Use of developmental marker genes to define temporal and spatial patterns of differentiation during embryoid body formation". Journal of Experimental Zoology. 284 (1): 67–81. Bibcode:1999JEZ...284...67L. doi:10.1002/(SICI)1097-010X(19990615)284:1<67::AID-JEZ10>3.0.CO;2-O. ISSN   1097-010X. PMID   10368935.
  6. ↑ ten Berge, Derk; Koole, Wouter; Fuerer, Christophe; Fish, Matt; Eroglu, Elif; Nusse, Roel (November 2008). "Wnt Signaling Mediates Self-Organization and Axis Formation in Embryoid Bodies". Cell Stem Cell. 3 (5): 508–518. doi:10.1016/j.stem.2008.09.013. PMC   2683270 . PMID   18983966.
  7. ↑ Baillie-Johnson, Peter; Brink, Susanne Carina van den; Balayo, Tina; Turner, David Andrew; Arias, Alfonso Martinez (2015). "Generation of Aggregates of Mouse Embryonic Stem Cells that Show Symmetry Breaking, Polarization and Emergent Collective Behaviour In Vitro". Journal of Visualized Experiments (105): e53252. doi:10.3791/53252. PMC   4692741 . PMID   26650833.
  8. ↑ Girgin, Mehmet; Turner, David Andrew; Baillie-Johnson, Peter; Cossy, Anne-Catherine; Beccari, Leonardo; Moris, Naomi; Lutolf, Matthias; Duboule, Denis; Martinez Arias, Alfonso (2018-10-12). "Generating Gastruloids from Mouse Embryonic Stem Cells". Protocol Exchange. doi: 10.1038/protex.2018.094 . ISSN   2043-0116.
  9. ↑ Ring, David B.; Johnson, Kirk W.; Henriksen, Erik J.; Nuss, John M.; Goff, Dane; Kinnick, Tyson R.; Ma, Sylvia T.; Reeder, John W.; Samuels, Isa (2003-03-01). "Selective glycogen synthase kinase 3 inhibitors potentiate insulin activation of glucose transport and utilization in vitro and in vivo". Diabetes. 52 (3): 588–595. doi: 10.2337/diabetes.52.3.588 . ISSN   0012-1797. PMID   12606497.
  10. ↑ Hashmi, Ali; Tlili, Sham; Perrin, Pierre; Martinez-Arias, Alfonso; Lenne, Pierre-François (2020-05-24). "Cell-state transitions and collective cell movement generate an endoderm-like region in gastruloids". doi: 10.1101/2020.05.21.105551 . hdl: 10230/53528 . S2CID   218891815.{{cite journal}}: Cite journal requires |journal= (help)
  11. ↑ Vianello, Stefano; Lutolf, Matthias P. (2020-06-09). "In vitro endoderm emergence and self-organisation in the absence of extraembryonic tissues and embryonic architecture". doi:10.1101/2020.06.07.138883. S2CID   219604191.{{cite journal}}: Cite journal requires |journal= (help)
  12. 1 2 Veenvliet, Jesse V; Bolondi, Adriano; Kretzmer, Helene; Haut, Leah; Scholze-Wittler, Manuela; Schifferl, Dennis; Koch, Frederic; Pustet, Milena; Heimann, Simon; Buschow, Rene; Wittler, Lars (2020-03-04). "Mouse embryonic stem cells self-organize into trunk-like structures with neural tube and somites". doi:10.1101/2020.03.04.974949. hdl: 21.11116/0000-0007-AB69-3 .{{cite journal}}: Cite journal requires |journal= (help)
  13. 1 2 van den Brink, Susanne C.; Alemany, Anna; van Batenburg, Vincent; Moris, Naomi; Blotenburg, Marloes; Vivié, Judith; Baillie-Johnson, Peter; Nichols, Jennifer; Sonnen, Katharina F.; Martinez Arias, Alfonso; van Oudenaarden, Alexander (June 2020). "Single-cell and spatial transcriptomics reveal somitogenesis in gastruloids". Nature. 582 (7812): 405–409. Bibcode:2020Natur.582..405V. doi:10.1038/s41586-020-2024-3. hdl: 10230/56609 . ISSN   1476-4687. PMID   32076263. S2CID   211194842.
  14. ↑ Rossi, Giuliana; Boni, Andrea; Guiet, Romain; Girgin, Mehmet; Kelly, Robert G.; Lutolf, Matthias P. (2019-10-14). "Embryonic organoids recapitulate early heart organogenesis": 802181. doi: 10.1101/802181 . S2CID   208588024.{{cite journal}}: Cite journal requires |journal= (help)
  15. ↑ Bérenger-Currias, Noémie M. L. P.; Mircea, Maria; Adegeest, Esmée; van den Berg, Patrick R.; Feliksik, Marleen; Hochane, Mazène; Idema, Timon; Tans, Sander J.; Semrau, Stefan (2020-02-14). "Early neurulation recapitulated in assemblies of embryonic and extraembryonic cells". doi:10.1101/2020.02.13.947655. S2CID   238251910.{{cite journal}}: Cite journal requires |journal= (help)
  16. ↑ Moris, Naomi; Anlas, Kerim; van den Brink, Susanne C.; Alemany, Anna; Schröder, Julia; Ghimire, Sabitri; Balayo, Tina; van Oudenaarden, Alexander; Martinez Arias, Alfonso (June 2020). "An in vitro model of early anteroposterior organization during human development". Nature. 582 (7812): 410–415. Bibcode:2020Natur.582..410M. doi:10.1038/s41586-020-2383-9. ISSN   0028-0836. PMID   32528178. S2CID   219567725.
  17. ↑ Etoc, Fred; Metzger, Jakob; Ruzo, Albert; Kirst, Christoph; Yoney, Anna; Ozair, M. Zeeshan; Brivanlou, Ali H.; Siggia, Eric D. (2016). "A Balance between Secreted Inhibitors and Edge Sensing Controls Gastruloid Self-Organization". Developmental Cell. 39 (3): 302–315. doi:10.1016/j.devcel.2016.09.016. PMC   5113147 . PMID   27746044.
  18. ↑ Warmflash, Aryeh; Sorre, Benoit; Etoc, Fred; Siggia, Eric D; Brivanlou, Ali H (2014). "A method to recapitulate early embryonic spatial patterning in human embryonic stem cells". Nature Methods. 11 (8): 847–854. doi:10.1038/nmeth.3016. PMC   4341966 . PMID   24973948.
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