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First published online March 7, 2005
doi: 10.1242/10.1242/dev.01715

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Imprinted X-inactivation in extra-embryonic endoderm cell lines from mouse blastocysts

Tilo Kunath^1,2,*, Danielle Arnaud³, Gary D. Uy⁴, Ikuhiro Okamoto⁵, Corinne Chureau³, Yojiro Yamanaka¹, Edith Heard⁵, Richard L. Gardner⁴, Philip Avner^3, and Janet Rossant^1,2,

¹ Samuel Lunenfeld Research Institute, Mount Sinai Hospital, 600 University Avenue, Toronto M5G 1X5, Canada
² Department of Molecular and Medical Genetics, University of Toronto, Toronto M5S 1A8, Canada
³ Unité de Génétique Moléculaire Murine, URA 2578 CNRS, Institut Pasteur, 25 rue du Docteur Roux, Paris 75015, France
⁴ Mammalian Development Laboratory, Department of Zoology, University of Oxford, South Parks Road, Oxford, OX1 3PS, UK
⁵ CNRS UMR218, Curie Institute, 26 rue d'Ulm, Paris 75005, France

Authors for correspondence (e-mail: rossant{at}mshri.on.ca and pavner{at}pasteur.fr)

Accepted 27 January 2005

The extra-embryonic endoderm lineage plays a major role in the nutritive support of the embryo and is required for several inductive events, such as anterior patterning and blood island formation. Blastocyst-derived embryonic stem (ES) and trophoblast stem (TS) cell lines provide good models with which to study the development of the epiblast and trophoblast lineages, respectively. We describe the derivation and characterization of cell lines that are representative of the third lineage of the blastocyst – extra-embryonic endoderm. Extra-embryonic endoderm (XEN) cell lines can be reproducibly derived from mouse blastocysts and passaged without any evidence of senescence. XEN cells express markers typical of extra-embryonic endoderm derivatives, but not those of the epiblast or trophoblast. Chimeras generated by injection of XEN cells into blastocysts showed exclusive contribution to extra-embryonic endoderm cell types. We used female XEN cells to investigate the mechanism of X chromosome inactivation in this lineage. We observed paternally imprinted X-inactivation, consistent with observations in vivo. Based on gene expression analysis, chimera studies and imprinted X-inactivation, XEN cell lines are representative of extra-embryonic endoderm and provide a new cell culture model of an early mammalian lineage.

Key words: Primitive endoderm, X-inactivation, Chimeras, Embryonic stem cells

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