Bulk cell density and Wnt/TGFbeta signalling regulate mesendodermal patterning of human pluripotent stem cells

In vitro differentiation of human pluripotent stem cells (hPSCs) recapitulates early aspects of human embryogenesis, but the underlying processes are poorly understood and controlled. Here we show that modulating the bulk cell density (BCD: cell number per culture volume) deterministically alters an...

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Main Authors:	Kempf, Henning, Olmer, Ruth, Haase, Alexandra, Franke, Annika, Bolesani, Emiliano, Schwanke, Kristin, Robles-Diaz, Diana, Coffee, Michelle, Göhring, Gudrun, Dräger, Gerald, Pötz, Oliver, Joos, Thomas, Martinez-Hackert, Erik, Haverich, Axel, Buettner, Falk F. R., Martin, Ulrich, Zweigerdt, Robert
Format:	Online
Language:	English
Published:	Nature Publishing Group 2016
Online Access:	https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5155150/

id	pubmed-5155150
recordtype	oai_dc
spelling	pubmed-51551502016-12-21 Bulk cell density and Wnt/TGFbeta signalling regulate mesendodermal patterning of human pluripotent stem cells Kempf, Henning Olmer, Ruth Haase, Alexandra Franke, Annika Bolesani, Emiliano Schwanke, Kristin Robles-Diaz, Diana Coffee, Michelle Göhring, Gudrun Dräger, Gerald Pötz, Oliver Joos, Thomas Martinez-Hackert, Erik Haverich, Axel Buettner, Falk F. R. Martin, Ulrich Zweigerdt, Robert Article In vitro differentiation of human pluripotent stem cells (hPSCs) recapitulates early aspects of human embryogenesis, but the underlying processes are poorly understood and controlled. Here we show that modulating the bulk cell density (BCD: cell number per culture volume) deterministically alters anteroposterior patterning of primitive streak (PS)-like priming. The BCD in conjunction with the chemical WNT pathway activator CHIR99021 results in distinct paracrine microenvironments codifying hPSCs towards definitive endoderm, precardiac or presomitic mesoderm within the first 24 h of differentiation, respectively. Global gene expression and secretome analysis reveals that TGFß superfamily members, antagonist of Nodal signalling LEFTY1 and CER1, are paracrine determinants restricting PS progression. These data result in a tangible model disclosing how hPSC-released factors deflect CHIR99021-induced lineage commitment over time. By demonstrating a decisive, functional role of the BCD, we show its utility as a method to control lineage-specific differentiation. Furthermore, these findings have profound consequences for inter-experimental comparability, reproducibility, bioprocess optimization and scale-up. Nature Publishing Group 2016-12-09 /pmc/articles/PMC5155150/ /pubmed/27934856 http://dx.doi.org/10.1038/ncomms13602 Text en Copyright © 2016, The Author(s) http://creativecommons.org/licenses/by/4.0/ This work is licensed under a Creative Commons Attribution 4.0 International License. The images or other third party material in this article are included in the article's Creative Commons license, unless indicated otherwise in the credit line; if the material is not included under the Creative Commons license, users will need to obtain permission from the license holder to reproduce the material. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/
repository_type	Open Access Journal
institution_category	Foreign Institution
institution	US National Center for Biotechnology Information
building	NCBI PubMed
collection	Online Access
language	English
format	Online
author	Kempf, Henning Olmer, Ruth Haase, Alexandra Franke, Annika Bolesani, Emiliano Schwanke, Kristin Robles-Diaz, Diana Coffee, Michelle Göhring, Gudrun Dräger, Gerald Pötz, Oliver Joos, Thomas Martinez-Hackert, Erik Haverich, Axel Buettner, Falk F. R. Martin, Ulrich Zweigerdt, Robert
spellingShingle	Kempf, Henning Olmer, Ruth Haase, Alexandra Franke, Annika Bolesani, Emiliano Schwanke, Kristin Robles-Diaz, Diana Coffee, Michelle Göhring, Gudrun Dräger, Gerald Pötz, Oliver Joos, Thomas Martinez-Hackert, Erik Haverich, Axel Buettner, Falk F. R. Martin, Ulrich Zweigerdt, Robert Bulk cell density and Wnt/TGFbeta signalling regulate mesendodermal patterning of human pluripotent stem cells
author_facet	Kempf, Henning Olmer, Ruth Haase, Alexandra Franke, Annika Bolesani, Emiliano Schwanke, Kristin Robles-Diaz, Diana Coffee, Michelle Göhring, Gudrun Dräger, Gerald Pötz, Oliver Joos, Thomas Martinez-Hackert, Erik Haverich, Axel Buettner, Falk F. R. Martin, Ulrich Zweigerdt, Robert
author_sort	Kempf, Henning
title	Bulk cell density and Wnt/TGFbeta signalling regulate mesendodermal patterning of human pluripotent stem cells
title_short	Bulk cell density and Wnt/TGFbeta signalling regulate mesendodermal patterning of human pluripotent stem cells
title_full	Bulk cell density and Wnt/TGFbeta signalling regulate mesendodermal patterning of human pluripotent stem cells
title_fullStr	Bulk cell density and Wnt/TGFbeta signalling regulate mesendodermal patterning of human pluripotent stem cells
title_full_unstemmed	Bulk cell density and Wnt/TGFbeta signalling regulate mesendodermal patterning of human pluripotent stem cells
title_sort	bulk cell density and wnt/tgfbeta signalling regulate mesendodermal patterning of human pluripotent stem cells
description	In vitro differentiation of human pluripotent stem cells (hPSCs) recapitulates early aspects of human embryogenesis, but the underlying processes are poorly understood and controlled. Here we show that modulating the bulk cell density (BCD: cell number per culture volume) deterministically alters anteroposterior patterning of primitive streak (PS)-like priming. The BCD in conjunction with the chemical WNT pathway activator CHIR99021 results in distinct paracrine microenvironments codifying hPSCs towards definitive endoderm, precardiac or presomitic mesoderm within the first 24 h of differentiation, respectively. Global gene expression and secretome analysis reveals that TGFß superfamily members, antagonist of Nodal signalling LEFTY1 and CER1, are paracrine determinants restricting PS progression. These data result in a tangible model disclosing how hPSC-released factors deflect CHIR99021-induced lineage commitment over time. By demonstrating a decisive, functional role of the BCD, we show its utility as a method to control lineage-specific differentiation. Furthermore, these findings have profound consequences for inter-experimental comparability, reproducibility, bioprocess optimization and scale-up.
publisher	Nature Publishing Group
publishDate	2016
url	https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5155150/
_version_	1613779191346495488

Bulk cell density and Wnt/TGFbeta signalling regulate mesendodermal patterning of human pluripotent stem cells

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