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In vitro generation of mouse polarized embryo-like structures from embryonic and trophoblast stem cells

Abstract

Mammalian embryogenesis requires the coordination of embryonic and extra-embryonic tissues to enable implantation into the uterus and post-implantation development to establish the body plan. Mouse embryonic stem cells (ESCs) are a useful tool for studying pluripotent embryonic tissue in vitro. However, they cannot undertake correct embryogenesis alone. Many attempts to model the early embryo in vitro involve the aggregation of ESCs into spheroids of variable size and cell number that undertake germ-layer specification but fail to recapitulate the characteristic architecture and arrangement of tissues of the early embryo. Here, we describe a protocol to generate the first embryo-like structures by directing the assembly of mouse ESCs and extra-embryonic trophoblast stem cells (TSCs) in a 3D extracellular matrix (ECM) into structures we call ‘polarized embryo-like structures’. By establishing the medium and culture conditions needed to support the growth of both stem cell types simultaneously, embryonic architecture is generated within 4 d of co-culture. This protocol can be performed by those proficient in standard ESC culture techniques and can be used in developmental studies to investigate the interactions between embryonic and extra-embryonic tissues during mammalian development.

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Fig. 1: Generation of polarized embryo-like system structures  from embryonic and extra-embryonic stem cells to form ETS embryos.
Fig. 2: Molecular markers indicating the lineage identity of the cell types in the embryonic and extra-embryonic compartments of polarized embryo-like structures (ETS embryos).

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Acknowledgements

We are grateful to our colleagues in the M.Z.-G. group and to M. Huch for advice. We are grateful to the Wellcome Trust for the Senior Research fellowship (grant no. 098287/Z/12/Z) and for a European Research Council grant (code: 669198) awarded to M.Z.-G. to fund this work. We are also grateful for the BBSRC DTP studentship that supports S.E.H. and to the Scientific and Technological Research Council of Turkey, which supports B.S.’

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S.E.H., B.S., and M.Z.-G. designed the protocol, carried out the work, analyzed the results, and prepared the manuscript.

Corresponding author

Correspondence to Magdalena Zernicka-Goetz.

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Competing interests

M.Z.-G. and S.E.H. declare that they are inventors on a patent application (1615343.9) submitted by Cell Guidance Systems (in which the University of Cambridge and the Wellcome Trust are beneficiaries) that covers the method and medium composition used to generate stem cell–derived embryos. B.S. declares no competing interests.

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Related links

1. Assembly of embryonic and extraembryonic stem cells to mimic embryogenesis in vitro: https://doi.org/10.1126/science.aal1810.

2. Self-organizing properties of mouse pluripotent cells initiate morphogenesis upon implantation: https://doi.org/10.1016/j.cell.2014.01.023.

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Harrison, S.E., Sozen, B. & Zernicka-Goetz, M. In vitro generation of mouse polarized embryo-like structures from embryonic and trophoblast stem cells. Nat Protoc 13, 1586–1602 (2018). https://doi.org/10.1038/s41596-018-0005-x

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