title: Derivation of naive human embryonic stem cells.
abstract: The naïve pluripotent state has been shown in mice to lead to broad and more robust developmental potential relative to primed mouse epiblast cells. The human naïve ES cell state has eluded derivation without the use of transgenes, and forced expression of OCT4, KLF4, and KLF2 allows maintenance of human cells in a naïve state [Hanna J, et al. (2010) Proc Natl Acad Sci USA 107(20):9222-9227]. We describe two routes to generate nontransgenic naïve human ES cells (hESCs). The first is by reverse toggling of preexisting primed hESC lines by preculture in the histone deacetylase inhibitors butyrate and suberoylanilide hydroxamic acid, followed by culture in MEK/ERK and GSK3 inhibitors (2i) with FGF2. The second route is by direct derivation from a human embryo in 2i with FGF2. We show that human naïve cells meet mouse criteria for the naïve state by growth characteristics, antibody labeling profile, gene expression, X-inactivation profile, mitochondrial morphology, microRNA profile and development in the context of teratomas. hESCs can exist in a naïve state without the need for transgenes. Direct derivation is an elusive, but attainable, process, leading to cells at the earliest stage of in vitro pluripotency described for humans. Reverse toggling of primed cells to naïve is efficient and reproducible.
authors.affiliation: Institute for Stem Cell and Regenerative Medicine, Departments of Comparative Medicine, Biology, and Biochemistry, and Divisions of Hematology, Metabolism, Endocrinology and Nutrition, and Medical Genetics, Department of Medicine, University of Washington, Seattle, WA 98195.
authors.forename: Carol B
authors.initials: CB
authors.lastname: Ware
authors.forename: Angelique M
authors.initials: AM
authors.lastname: Nelson
authors.forename: Brigham
authors.initials: B
authors.lastname: Mecham
authors.forename: Jennifer
authors.initials: J
authors.lastname: Hesson
authors.forename: Wenyu
authors.initials: W
authors.lastname: Zhou
authors.forename: Erica C
authors.initials: EC
authors.lastname: Jonlin
authors.forename: Antonio J
authors.initials: AJ
authors.lastname: Jimenez-Caliani
authors.forename: Xinxian
authors.initials: X
authors.lastname: Deng
authors.forename: Christopher
authors.initials: C
authors.lastname: Cavanaugh
authors.forename: Savannah
authors.initials: S
authors.lastname: Cook
authors.forename: Paul J
authors.initials: PJ
authors.lastname: Tesar
authors.forename: Jeffrey
authors.initials: J
authors.lastname: Okada
authors.forename: Lilyana
authors.initials: L
authors.lastname: Margaretha
authors.forename: Henrik
authors.initials: H
authors.lastname: Sperber
authors.forename: Michael
authors.initials: M
authors.lastname: Choi
authors.forename: C Anthony
authors.initials: CA
authors.lastname: Blau
authors.forename: Piper M
authors.initials: PM
authors.lastname: Treuting
authors.forename: R David
authors.initials: RD
authors.lastname: Hawkins
authors.forename: Vincenzo
authors.initials: V
authors.lastname: Cirulli
authors.forename: Hannele
authors.initials: H
authors.lastname: Ruohola-Baker
mesh_terms: D000818:Animals; D019070:Cell Lineage; D002478:Cells, Cultured; D053595:Embryonic Stem Cells / Q000166:cytology* / Q000378:metabolism; D020869:Gene Expression Profiling; D038362:Glycogen Synthase Kinase 3 / Q000037:antagonists & inhibitors; D056572:Histone Deacetylase Inhibitors / Q000494:pharmacology; D006801:Humans; D000090062:Kruppel-Like Factor 4; D051379:Mice; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D019076:Transgenes; D049951:X Chromosome Inactivation
chemical_list: D056572:Histone Deacetylase Inhibitors; C000715467:KLF4 protein, human; C000715468:Klf4 protein, mouse; D000090062:Kruppel-Like Factor 4; D047428:Protein Kinase Inhibitors; D038362:Glycogen Synthase Kinase 3