TY - JOUR
T1 - Modeling early stages of endoderm development in epiblast stem cell aggregates with supply of extracellular matrices
AU - Inamori, Sachiko
AU - Fujii, Mai
AU - Satake, Sayaka
AU - Iida, Hideaki
AU - Teramoto, Machiko
AU - Sumi, Tomoyuki
AU - Meno, Chikara
AU - Ishii, Yasuo
AU - Kondoh, Hisato
N1 - Funding Information:
We thank Hiroshi Sasaki for granting the use of the knockin mice. This study was supported by Grants‐in‐Aid for Scientific Research, JP19K16184 to MT and JP17H03680 to HK from MEXT Japan, Private University Research Branding Project of MEXT, and Kyoto Sangyo University Research Fund for Institute for Protein Dynamics. Foxa2‐Egfp
Publisher Copyright:
© 2020 The Authors. Development, Growth & Differentiation published by John Wiley & Sons Australia, Ltd on behalf of Japanese Society of Developmental Biologists
PY - 2020/5/1
Y1 - 2020/5/1
N2 - Endoderm precursors expressing FoxA2 and Sox17 develop from the epiblast through the gastrulation process. In this study, we developed an experimental system to model the endoderm-generating gastrulation process using epiblast stem cells (EpiSCs). To this end, we established an EpiSC line i22, in which enhanced green fluorescent protein is coexpressed with Foxa2. Culturing i22 EpiSCs as aggregates for a few days was sufficient to initiate Foxa2 expression, and further culturing of the aggregates in Matrigel promoted the sequential activation of transcription factor genes involved in endoderm precursor development, e.g., Eomes, Gsc, and Sox17. In aggregation culture of i22 cells for 3 days, all cells expressed POU5F1, SOX2, and E-cadherin, a signature of the epiblast, whereas expression of GATA4 and SOX17 was also activated moderately in dispersed cells, suggesting priming of these cells to endodermal development. Embedding the aggregates in Matrigel for further 3 days elicited migration of the cells into the lumen of laminin-rich matrices covering the aggregates, in which FOXA2 and SOX17 were expressed at a high level with the concomitant loss of E-cadherin, indicating the migratory phase of endodermal precursors. Prolonged culturing of the aggregates generated three segregating cell populations found in post-gastrulation stage embryos: (1) definitive endoderm co-expressing high SOX17, GATA4, and E-cadherin, (2) mesodermal cells expressing a low level of GATA4 and lacking E-cadherin, and (3) primed epiblast cells expressing POU5F1, SOX2 without E-cadherin. Thus, aggregation of EpiSCs followed by embedding of aggregates in the laminin-rich matrix models the gastrulation-dependent endoderm precursor development.
AB - Endoderm precursors expressing FoxA2 and Sox17 develop from the epiblast through the gastrulation process. In this study, we developed an experimental system to model the endoderm-generating gastrulation process using epiblast stem cells (EpiSCs). To this end, we established an EpiSC line i22, in which enhanced green fluorescent protein is coexpressed with Foxa2. Culturing i22 EpiSCs as aggregates for a few days was sufficient to initiate Foxa2 expression, and further culturing of the aggregates in Matrigel promoted the sequential activation of transcription factor genes involved in endoderm precursor development, e.g., Eomes, Gsc, and Sox17. In aggregation culture of i22 cells for 3 days, all cells expressed POU5F1, SOX2, and E-cadherin, a signature of the epiblast, whereas expression of GATA4 and SOX17 was also activated moderately in dispersed cells, suggesting priming of these cells to endodermal development. Embedding the aggregates in Matrigel for further 3 days elicited migration of the cells into the lumen of laminin-rich matrices covering the aggregates, in which FOXA2 and SOX17 were expressed at a high level with the concomitant loss of E-cadherin, indicating the migratory phase of endodermal precursors. Prolonged culturing of the aggregates generated three segregating cell populations found in post-gastrulation stage embryos: (1) definitive endoderm co-expressing high SOX17, GATA4, and E-cadherin, (2) mesodermal cells expressing a low level of GATA4 and lacking E-cadherin, and (3) primed epiblast cells expressing POU5F1, SOX2 without E-cadherin. Thus, aggregation of EpiSCs followed by embedding of aggregates in the laminin-rich matrix models the gastrulation-dependent endoderm precursor development.
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U2 - 10.1111/dgd.12663
DO - 10.1111/dgd.12663
M3 - Article
C2 - 32277710
AN - SCOPUS:85084378346
VL - 62
SP - 243
EP - 259
JO - Development Growth and Differentiation
JF - Development Growth and Differentiation
SN - 0012-1592
IS - 4
ER -