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  Subtype-specific differentiation of cardiac pacemaker cell clusters from human induced pluripotent stem cells

Schweizer, P. A., Darche, F. F., Ullrich, N. D., Geschwill, P., Greber, B., Rivinius, R., et al. (2017). Subtype-specific differentiation of cardiac pacemaker cell clusters from human induced pluripotent stem cells. Stem Cell Research, 8(229), 1-15. doi:10.1186/s13287-017-0681-4.

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Item Permalink: http://hdl.handle.net/11858/00-001M-0000-002E-1048-2 Version Permalink: http://hdl.handle.net/21.11116/0000-0000-299A-5
Genre: Journal Article

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 Creators:
Schweizer, Patrick A., Author
Darche, Fabrice F., Author
Ullrich, Nina D., Author
Geschwill, Pascal, Author
Greber, Boris, Author
Rivinius, Rasmus, Author
Seyler, Claudia, Author
Müller-Decker, Karin, Author
Draguhn, Andreas, Author
Utikal, Jochen, Author
Koenen, Michael1, Author              
Katus, Hugo A., Author
Thomas, Dierk, Author
Affiliations:
1Department of Cell Physiology, Max Planck Institute for Medical Research, Max Planck Society, ou_1497701              

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Free keywords: Pacemaker, Pluripotent stem cells, Automaticity, Differentiation
 Abstract: Background Human induced pluripotent stem cells (hiPSC) harbor the potential to differentiate into diverse cardiac cell types. Previous experimental efforts were primarily directed at the generation of hiPSC-derived cells with ventricular cardiomyocyte characteristics. Aiming at a straightforward approach for pacemaker cell modeling and replacement, we sought to selectively differentiate cells with nodal-type properties. Methods hiPSC were differentiated into spontaneously beating clusters by co-culturing with visceral endoderm-like cells in a serum-free medium. Subsequent culturing in a specified fetal bovine serum (FBS)-enriched cell medium produced a pacemaker-type phenotype that was studied in detail using quantitative real-time polymerase chain reaction (qRT-PCR), immunocytochemistry, and patch-clamp electrophysiology. Further investigations comprised pharmacological stimulations and co-culturing with neonatal cardiomyocytes. Results hiPSC co-cultured in a serum-free medium with the visceral endoderm-like cell line END-2 produced spontaneously beating clusters after 10–12 days of culture. The pacemaker-specific genes HCN4, TBX3, and TBX18 were abundantly expressed at this early developmental stage, while levels of sarcomeric gene products remained low. We observed that working-type cardiomyogenic differentiation can be suppressed by transfer of early clusters into a FBS-enriched cell medium immediately after beating onset. After 6 weeks under these conditions, sinoatrial node (SAN) hallmark genes remained at high levels, while working-type myocardial transcripts (NKX2.5, TBX5) were low. Clusters were characterized by regular activity and robust beating rates (70–90 beats/min) and were triggered by spontaneous Ca2+ transients recapitulating calcium clock properties of genuine pacemaker cells. They were responsive to adrenergic/cholinergic stimulation and able to pace neonatal rat ventricular myocytes in co-culture experiments. Action potential (AP) measurements of cells individualized from clusters exhibited nodal-type (63.4%) and atrial-type (36.6%) AP morphologies, while ventricular AP configurations were not observed. Conclusion We provide a novel culture media-based, transgene-free approach for targeted generation of hiPSC-derived pacemaker-type cells that grow in clusters and offer the potential for disease modeling, drug testing, and individualized cell-based replacement therapy of the SAN.

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Language(s): eng - English
 Dates: 2017-05-062017-09-252017-10-16
 Publication Status: Published online
 Pages: 15
 Publishing info: -
 Table of Contents: -
 Rev. Method: Peer
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Title: Stem Cell Research
Source Genre: Journal
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Publ. Info: Amsterdam : Elsevier
Pages: - Volume / Issue: 8 (229) Sequence Number: - Start / End Page: 1 - 15 Identifier: ISSN: 1873-5061
CoNE: https://pure.mpg.de/cone/journals/resource/1873-5061