Induction of Rosette-to-Lumen stage embryoids using reprogramming paradigms in ESCs

Abstract

Blastocyst-derived stem cell lines were shown to self-organize into embryo-like structures in 3D cell culture environments. Here, we provide evidence that embryo-like structures can be generated solely based on transcription factor-mediated reprogramming of embryonic stem cells in a simple 3D co-culture system. Embryonic stem cells in these cultures self-organize into elongated, compartmentalized embryo-like structures reflecting aspects of the inner regions of the early post-implantation embryo. Single-cell RNA-sequencing reveals transcriptional profiles resembling epiblast, primitive-/visceral endoderm, and extraembryonic ectoderm of early murine embryos around E4.5–E5.5. In this stem cell-based embryo model, progression from rosette formation to lumenogenesis accompanied by progression from naïve- to primed pluripotency was observed within Epi-like cells. Additionally, lineage specification of primordial germ cells and distal/anterior visceral endoderm-like cells was observed in epiblast- or visceral endoderm-like compartments, respectively. The system presented in this study allows for fast and reproducible generation of embryo-like structures, providing an additional tool to study aspects of early embryogenesis.

Embryo model completes gastrulation to neurulation and organogenesis

“…Indeed, we and others reported a similar embryo model in which TS cells were exchanged with ES cells induced to express CDX2 57 , 58 . These studies add to a previous report on the generation of embryoids using ES cells to form the ExE lineage 9 . Embryoids in which the extraembryonic lineages are derived only from ES cells can also develop to neurulation stages.…”

Journey of the mouse primitive endoderm: from specification to maturation

“…Alpha-fetoprotein, a serum component, expressed throughout the VE starting at early post-implantation has been used as a reporter to track the fate of this lineage [39,40,165]. GATA6, which is expressed by VE cells [166,167], can induce VE-like cell clusters when expressed ectopically in mouse ES cells [168]. Mutants in Gata4, another VE marker [76,166,169], arrest at gastrulation and exhibit defects in yolk sac and cardiac formation in vivo [170,171] and VE differentiation in vitro [172].…”

“…GATA6, which is expressed by VE cells [166,167], can induce VE-like cell clusters when expressed ectopically in mouse ES cells [168]. Mutants in Gata4 , another VE marker [76,166,169], arrest at gastrulation and exhibit defects in yolk sac and cardiac formation in vivo [170,171] and VE differentiation in vitro [172].…”

Tissue-intrinsic Wnt signals antagonize Nodal-driven AVE differentiation

“…To investigate mechanisms of lineage crosstalk between the Epi and the VE, we used an experimental system where both lineages are established in reproducible proportions from a single starting population through cell-cell communication via FGF4. 9 This approach contrasts with previous studies, where bilayered aggregates have been formed by mixing ESCs with established XEN cell lines, 20 by mixing wildtype ESCs with GATA-inducible ESCs, 21 or by chemical conversion of ESCs towards the VE lineage. 22 Consistent with our results, these previous studies found that the Epi core induces an embryonic identity in the overlying VE.…”

“…22 Consistent with our results, these previous studies found that the Epi core induces an embryonic identity in the overlying VE. 22 They also reported expression of the AVE marker Lefty1 in the VE, 20,21 but whether these Lefty1expressing cells had acquired an AVE identity remained unclear. Our scRNAseq analysis demonstrates that a subset of VE cells in BELAs differentiate into AVE in the absence of an ExE.…”