Abstract
In drug discovery, there is an increasing demand for more physiological in vitro models that recapitulate the disease situation in patients. Human induced pluripotent stem (hiPS) cell-derived model cells could serve this purpose. To date, several directed differentiation approaches have been described to generate definitive endoderm (DE) from hiPS cells, but protocols suitable for drug development and high-throughput screening (HTS) have not been reported yet. In this work, a large-scale expansion of hiPS cells for high-throughput adaption is presented and an optimized stepwise differentiation of hiPS cells into DE cells is described. The produced DE cells were demonstrated to express classical DE markers on the gene expression and protein level. The here described DE cells are multipotent progenitors and act as starting points for a broad spectrum of endodermal model cells in HTS and other areas of drug discovery.
This is a preview of subscription content, log in via an institution to check access.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Takahashi K, Yamanaka S (2006) Induction of pluripotent stem cells from mouse embryonic and adult fibroblast cultures by defined factors. Cell 126:663–676
Traub S, Stahl H, Rosenbrock H et al (2017) Upscaling of hiPS cell-derived neurons for high-throughput screening. SLAS Discov 22:274–286
Heilker R, Traub S, Reinhardt P et al (2014) iPS cell derived neuronal cells for drug discovery. Trends Pharmacol Sci 35:510–519
Kim PT, Ong CJ (2012) Differentiation of definitive endoderm from mouse embryonic stem cells. Results Probl Cell Differ 55:303–319
Zorn AM, Wells JM (2009) Vertebrate endoderm development and organ formation. Annu Rev Cell Dev Biol 25:221–251
Wells JM, Melton DA (1999) Vertebrate endoderm development. Annu Rev Cell Dev Biol 15:393–410
Kraus MR, Grapin-Botton A (2012) Patterning and shaping the endoderm in vivo and in culture. Curr Opin Genet Dev 22:347–353
Kubo A, Shinozaki K, Shannon JM et al (2004) Development of definitive endoderm from embryonic stem cells in culture. Development 131:1651–1662
Wong AP, Chin S, Xia S et al (2015) Efficient generation of functional CFTR-expressing airway epithelial cells from human pluripotent stem cells. Nat Protoc 10:363–381
Huang SX, Green MD, De Carvalho AT et al (2015) The in vitro generation of lung and airway progenitor cells from human pluripotent stem cells. Nat Protoc 10:413–425
D’amour KA, Agulnick AD, Eliazer S et al (2005) Efficient differentiation of human embryonic stem cells to definitive endoderm. Nat Biotechnol 23:1534–1541
Gotoh S, Ito I, Nagasaki T et al (2014) Generation of alveolar epithelial spheroids via isolated progenitor cells from human pluripotent stem cells. Stem Cell Reports 3:394–403
Ghaedi M, Mendez JJ, Bove PF et al (2014) Alveolar epithelial differentiation of human induced pluripotent stem cells in a rotating bioreactor. Biomaterials 35:699–710
Wang P, Mcknight KD, Wong DJ et al (2012) A molecular signature for purified definitive endoderm guides differentiation and isolation of endoderm from mouse and human embryonic stem cells. Stem Cells Dev 21:2273–2287
Ng VY, Ang SN, Chan JX et al (2010) Characterization of epithelial cell adhesion molecule as a surface marker on undifferentiated human embryonic stem cells. Stem Cells 28:29–35
Grapin-Botton A (2008) Endoderm specification. In: StemBook, Cambridge MA
Green MD, Chen A, Nostro MC et al (2011) Generation of anterior foregut endoderm from human embryonic and induced pluripotent stem cells. Nat Biotechnol 29:267–272
Acknowledgments
The research leading to these results has received support from the Innovative Medicines Initiative Joint Undertaking under (grant no. 115439), resources of which are composed of financial contribution from the European Union’s Seventh Framework Programme (FP7/2007-2013) and EFPIA companies. This publication reflects only the author’s views and neither the IMI JU nor EFPIA nor the European Commission are liable for any use that may be made of the information contained therein. We sincerely thank Marcel Leist and Dirk Stenkamp for their excellent advice as well as Michael Traub for his technical assistance.
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2019 Springer Science+Business Media, LLC, part of Springer Nature
About this protocol
Cite this protocol
Bluhmki, T. et al. (2019). Differentiation of hiPS Cells into Definitive Endoderm for High-Throughput Screening. In: Mandenius, CF., Ross, J. (eds) Cell-Based Assays Using iPSCs for Drug Development and Testing. Methods in Molecular Biology, vol 1994. Humana, New York, NY. https://doi.org/10.1007/978-1-4939-9477-9_9
Download citation
DOI: https://doi.org/10.1007/978-1-4939-9477-9_9
Published:
Publisher Name: Humana, New York, NY
Print ISBN: 978-1-4939-9476-2
Online ISBN: 978-1-4939-9477-9
eBook Packages: Springer Protocols