Abstract
Mouse embryonic stem cells (ESCs) derive from the inner cell mass (ICM) of a blastocyst at E3.5 while mouse epiblast stem cells (EpiSCs) derive from the late epiblast of a post-implantation embryo at E5.5–E7.5. Both cells are able to differentiate into derivatives of the three germs layers but only ESCs are able to produce chimeras when they are introduced into a blastocyst. To support the naive state of pluripotency, ESC culture requires Leukemia inhibitory factor (Lif) and either serum or inhibitors of Erk and Gsk3 pathways (2i) while the primed pluripotency of EpiSCs is maintained using Activin A and Fibroblast Growth Factor 2 (FGF2). It is possible to obtain EpiSCs in vitro starting from ESCs but also to induce ESCs starting from EpiSCs even if this second process is very difficult and inefficient. In this protocol we describe how we perform the process of conversion from ESCs to EpiSCs.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Evans M, Kaufman MH (1981) Establishment in culture of pluripotential cells from mouse embryos. Nature 292:154–156
Brons IGM, Smithers LE, Trotter MWB et al (2007) Derivation of pluripotent epiblast stem cells from mammalian embryos. Nature 448(7150):191–195
Rossant J (2008) Stem cells and early lineage development. (Translated from eng). Cell 132(4):527–531
Niwa H, Burdon T, Chambers I et al (1998) Self-renewal of pluripotent embryonic stem cells is mediated via activation of STAT3. Genes Dev 12(13):2048–2060
Tesar PJ, Chenoweth JG, Brook FA et al (2007) New cell lines from mouse epiblast share defining features with human embryonic stem cells. Nature 448:196–199
Nichols J, Smith A (2012) Pluripotency in the embryo and in culture. (Translated from eng). Cold Spring Harb Perspect Biol 4(8):a008128
Veillard AC, Marks H, Bernardo AS et al (2014) Stable methylation at promoters distinguishes epiblast stem cells from embryonic stem cells and the in vivo epiblasts. (Translated from eng). Stem Cells Dev 23(17):2014–2029
Guo G, Yang J, Nichols J et al (2009) Klf4 reverts developmentally programmed restriction of ground state pluripotency. Development 136:1063–1069
Buecker C, Srinivasan R, Wu Z et al (2014) Reorganization of enhancer patterns in transition from naive to primed pluripotency. Cell Stem Cell 14(6):838–853
Bao S, Tang F, Li X et al (2009) Epigenetic reversion of post-implantation epiblast to pluripotent embryonic stem cells. Nature 461:1292–5
Factor Daniel C, Corradin O, Zentner Gabriel E et al (2014) Epigenomic comparison reveals activation of “seed” enhancers during transition from naive to primed pluripotency. Cell Stem Cell 14(6):854–863
Hayashi K, Ohta H, Kurimoto K et al (2011) Reconstitution of the mouse germ cell specification pathway in culture by pluripotent stem cells. (Translated from eng). Cell 146(4):519–532
Schulz EG, Meisig J, Nakamura T et al (2014) The two active X chromosomes in female ESCs block exit from the pluripotent state by modulating the ESC signaling network. (Translated from eng). Cell Stem Cell 14(2):203–216
Wray J, Kalkan T, Smith AG (2010) The ground state of pluripotency. (Translated from eng). Biochem Soc Trans 38(4):1027–1032
Marks H, Kalkan T, Menafra R et al (2012) The transcriptional and epigenomic foundations of ground state pluripotency. (Translated from eng). Cell 149(3):590–604
Habibi E, Brinkman AB, Arand J et al (2013) Whole-genome bisulfite sequencing of two distinct interconvertible DNA methylomes of mouse embryonic stem cells. (Translated from eng). Cell Stem Cell 13(3):360–369
Acknowledgments
We thank the ANR Programme Investissements d’Avenir REVIVE for their support. MT is a recipient of a PhD fellowship from DIM Biotherapies Ile-de-France.
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2015 Springer Science+Business Media New York
About this protocol
Cite this protocol
Tosolini, M., Jouneau, A. (2015). From Naive to Primed Pluripotency: In Vitro Conversion of Mouse Embryonic Stem Cells in Epiblast Stem Cells. In: Turksen, K. (eds) Embryonic Stem Cell Protocols. Methods in Molecular Biology, vol 1341. Humana Press, New York, NY. https://doi.org/10.1007/7651_2015_208
Download citation
DOI: https://doi.org/10.1007/7651_2015_208
Published:
Publisher Name: Humana Press, New York, NY
Print ISBN: 978-1-4939-2953-5
Online ISBN: 978-1-4939-2954-2
eBook Packages: Springer Protocols