title: Activin/Nodal and FGF pathways cooperate to maintain pluripotency of human embryonic stem cells.
abstract: Maintenance of pluripotency is crucial to the mammalian embryo's ability to generate the extra-embryonic and embryonic tissues that are needed for intrauterine survival and foetal development. The recent establishment of embryonic stem cells from human blastocysts (hESCs) provides an opportunity to identify the factors supporting pluripotency at early stages of human development. Using this in vitro model, we have recently shown that Nodal can block neuronal differentiation, suggesting that TGFbeta family members are involved in cell fate decisions of hESCs, including preservation of their pluripotency. Here, we report that Activin/Nodal signalling through Smad2/3 activation is necessary to maintain the pluripotent status of hESCs. Inhibition of Activin/Nodal signalling by follistatin and by overexpression of Lefty or Cerberus-Short, or by the Activin receptor inhibitor SB431542, precipitates hESC differentiation. Nevertheless, neither Nodal nor Activin is sufficient to sustain long-term hESC growth in a chemically defined medium without serum. Recent studies have shown that FGF2 can also maintain long-term expression of pluripotency markers, and we find that inhibition of the FGF signalling pathway by the tyrosine kinase inhibitor SU5402 causes hESC differentiation. However, this effect of FGF on hESC pluripotency depends on Activin/Nodal signalling, because it is blocked by SB431542. Finally, long-term maintenance of in-vitro pluripotency can be achieved with a combination of Activin or Nodal plus FGF2 in the absence of feeder-cell layers, conditioned medium or Serum Replacer. These findings suggest that the Activin/Nodal pathway maintains pluripotency through mechanism(s) in which FGF acts as a competence factor and therefore provide further evidence of distinct mechanisms for preservation of pluripotency in mouse and human ESCs.
authors.affiliation: Department of Surgery and Cambridge Institute for Medical Research, Addenbrooke's Hospital, University of Cambridge, Hills Road, Cambridge CB2 2XY, UK. lv225@cam.ac.uk
authors.forename: Ludovic
authors.initials: L
authors.lastname: Vallier
authors.forename: Morgan
authors.initials: M
authors.lastname: Alexander
authors.forename: Roger A
authors.initials: RA
authors.lastname: Pedersen
mesh_terms: D030201:Activin Receptors, Type I / Q000037:antagonists & inhibitors / Q000378:metabolism; D028341:Activins / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D001549:Benzamides / Q000378:metabolism; D015415:Biomarkers / Q000378:metabolism; D002454:Cell Differentiation / Q000502:physiology; D002478:Cells, Cultured; D004149:Dioxoles / Q000378:metabolism; D004622:Embryo, Mammalian / Q000166:cytology* / Q000502:physiology; D004815:Epidermal Growth Factor / Q000235:genetics / Q000378:metabolism; D005346:Fibroblast Growth Factors / Q000378:metabolism*; D058851:GPI-Linked Proteins; D006801:Humans; D036341:Intercellular Signaling Peptides and Proteins; D008562:Membrane Glycoproteins / Q000235:genetics / Q000378:metabolism; D051379:Mice; D009363:Neoplasm Proteins / Q000235:genetics / Q000378:metabolism; D055457:Nodal Protein; D039904:Pluripotent Stem Cells / Q000166:cytology / Q000502:physiology*; D015398:Signal Transduction / Q000502:physiology*; D051785:Smad Proteins / Q000378:metabolism; D016212:Transforming Growth Factor beta / Q000235:genetics / Q000378:metabolism*; D053773:Transforming Growth Factor beta1
chemical_list: C459179:4-(5-benzo(1,3)dioxol-5-yl-4-pyridin-2-yl-1H-imidazol-2-yl)benzamide; D001549:Benzamides; D015415:Biomarkers; D004149:Dioxoles; D058851:GPI-Linked Proteins; D036341:Intercellular Signaling Peptides and Proteins; D008562:Membrane Glycoproteins; C526007:NODAL protein, human; D009363:Neoplasm Proteins; D055457:Nodal Protein; C526008:Nodal protein, mouse; D051785:Smad Proteins; C070636:TDGF1 protein, human; C508960:TGFB1 protein, human; C508961:Tgfb1 protein, mouse; D016212:Transforming Growth Factor beta; D053773:Transforming Growth Factor beta1; D028341:Activins; D005346:Fibroblast Growth Factors; D004815:Epidermal Growth Factor; D030201:Activin Receptors, Type I