Neural Induction Intermediates Exhibit Distinct Roles of Fgf Signaling  

Abstract

Formation of the neural plate is an intricate process in early mammalian embryonic development mediated by cells of the inner cell mass and involving a series of steps, including development of the epiblast. Here, we report on the creation of an embryonic stem (ES) cell-based system to isolate and identify neural induction intermediates with characteristics of epiblast cells and neural plate. We demonstrate that neural commitment requires prior differentiation of ES cells into epiblast cells that are indistinguishable from those derived from natural embryos. We also demonstrate that epiblast cells can be isolated and cultured as epiblast stem cell lines. Fgf signaling is shown to be required for the differentiation of ES cells into these epiblast cells. Fgf2, widely used for maintenance of both human ES cells and epiblast stem cells, inhibits formation of early neural cells by epiblast intermediates in a dose-dependent manner and is sufficient to promote transient self-renewal of epiblast stem cells. In contrast, Fgf8, the endogenous embryonic neural inducer, fails to promote epiblast self-renewal, but rather promotes more homogenous neural induction with transient self-renewal of early neural cells. Removal of Fgf signaling entirely from epiblast cells promotes rapid neural induction and subsequent neurogenesis. We conclude that Fgf signaling plays different roles during the differentiation of ES cells, with an initial requirement in epiblast formation and a subsequent role in self-renewal. Fgf2 and Fgf8 thus stimulate self-renewal in different cell types. STEM CELLS

Hes1 Desynchronizes Differentiation of Pluripotent Cells by Modulating STAT3 Activity

“…2C). This dip in Hes1 expression during differentiation corresponds to the time that cells are transiting to an epiblast-like state [43] and losing LIF responsiveness [17]. …”

Specific Glycosaminoglycans Modulate Neural Specification of Mouse Embryonic Stem Cells

“…This likely reflects the different regulatory factors involved in the formation of separate lineages. Sox1 expression initially requires upregulation of FGF-mediated signals [27,28,30,40]; whereas early hematopoiesis requires signaling via BMP4 and VEGF [41,42]. This demonstrates both how versatile oligosaccharides could be as culture additives, but also highlights the need for careful selection with a thorough knowledge of the specific pathways involved and the GAG structures needed to regulate them.…”

Distinct Developmental Ground States of Epiblast Stem Cell Lines Determine Different Pluripotency Features

“…While OG2 embryos derived at E5.5 still showed residual GFP expression, the GFP signal rapidly disappeared upon establishing the EpiSC lines (Supporting Information Fig. S3) [13,16]. Spontaneous GFP reactivation was not detected by routine monitoring using fluorescence-activated cell sorting (data not shown).…”

“…Some experiments were carried out in chemically defined N2B27 medium [11] in the presence of 10 ng/ml bFGF. For reversion experiments, EpiSCs were harvested using accutase (Invitrogen) and seeded as a single cell suspension at approximately 16 …”