Abstract
Myocardial infarction (MI) can lead to irreversible loss of cardiomyocytes (CMs), primarily localized to the left ventricle (LV) of the heart. The CMs of the LV are predominantly derived from first heart field (FHF) progenitors, whereas the majority of CMs within the right ventricle originate from the second heart field (SHF) during early cardiogenesis. Human embryonic stem cells (hESCs) serve as a valuable source of CMs for understanding early cardiac development and lineage commitment of CMs within these two heart fields that ultimately enable theĀ development of more effective candidates for cell therapy. An ideal candidate may be FHF CMs that share the same ontogeny with the LV CMs that die after MI. We previously generated a double reporter hESC line that utilizes two important cardiac transcription factors, TBX5 and NKX2-5. TBX5 marks FHF progenitors and CMs, while NKX2-5 is expressed in nearly all myocytes of the developing heart. Here, we describe a step-by-step approach to efficiently generate FHF and SHF CMs using this double reporter hESC line. In addition, this approach can be applied to any non-genetically modified hESC lines to enrich FHF and SHF CMs. Obtaining enriched populations of these two CM subtypes provides a platform for downstream comparative analyses and in vitro studies to facilitate a deeper understanding of cardiovascular lineage commitment and theĀ development of more effective candidates for cell therapy to treat diseases or defects that affect specific regions of the heart.
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
Ilic D, Ogilvie C (2017) Concise review: human embryonic stem cells-what have we done? What are we doing? Where are we going? Stem Cells 35(1):17ā25. https://doi.org/10.1002/stem.2450
Romagnuolo R, Masoudpour H, Porta-SĆ”nchez A et al (2019) Human embryonic stem cell-derived cardiomyocytes regenerate the infarcted pig heart but induce ventricular tachyarrhythmias. Stem Cell Reports 12(5):967ā981. https://doi.org/10.1016/j.stemcr.2019.04.005
Kim D, Kim SB, Ryu JL et al (2020) Human embryonic stem cell-derived Wilson's disease model for screening drug efficacy. Cells 9(4):872. https://doi.org/10.3390/cells9040872
Crespo M, Vilar E, Tsai SY et al (2017) Colonic organoids derived from human induced pluripotent stem cells for modeling colorectal cancer and drug testing. Nat Med 23(7):878ā884. https://doi.org/10.1038/nm.4355
DeLaughter DM, Bick AG, Wakimoto H et al (2016) Single-cell resolution of temporal gene expression during heart development. Dev Cell 39(4):480ā490. https://doi.org/10.1016/j.devcel.2016.10.001
Kattman SJ, Witty AD, Gagliardi M et al (2011) Stage-specific optimization of activin/nodal and BMP signaling promotes cardiac differentiation of mouse and human pluripotent stem cell lines. Cell Stem Cell 8(2):228ā240. https://doi.org/10.1016/j.stem.2010.12.008
Lian X, Zhang J, Azarin SM et al (2013) Directed cardiomyocyte differentiation from human pluripotent stem cells by modulating Wnt/Ī²-catenin signaling under fully defined conditions. Nat Protoc 8(1):162ā175. https://doi.org/10.1038/nprot.2012.150
Yang L, Soonpaa MH, Adler ED et al (2008) Human cardiovascular progenitor cells develop from a KDR+ embryonic-stem-cell-derived population. Nature 453(7194):524ā528. https://doi.org/10.1038/nature06894
Liu X, Yagi H, Saeed S et al (2017) The complex genetics of hypoplastic left heart syndrome. Nat Genet 49(7):1152ā1159. https://doi.org/10.1038/ng.3870
Corrado D, Link MS, Calkins H (2017) Arrhythmogenic right ventricular cardiomyopathy. N Engl J Med 376(15):1489ā1490. https://doi.org/10.1056/NEJMc1701400
Pezhouman A, Engel JL, Nguyen NB et al (2021) Isolation and characterization of hESC-derived heart field-specific cardiomyocytes unravels new insights into their transcriptional and electrophysiological profiles. Cardiovasc Res. https://doi.org/10.1093/cvr/cvab102
Y-27632 dihydrochloride. https://www.tocris.com/products/y-27632-dihydrochloride_1254?gclid=CjwKCAjwm_P5BRAhEiwAwRzSO4qVIcMe6q8JJZPJ981Ad_mG07y4RMghUf5GSx5AaXzSRBqKHGHODhoCcmoQAvD_BwE
Acknowledgments
This work was supported in part by grants from the Eli & Edythe Broad Center of Regenerative Medicine and Stem Cell Research at UCLA Postdoctoral Fellowship (A.P.), Department of Defense Discovery Award (W81XWH-19-1-0244) (A.P.), Ruth L. Kirschstein Predoctoral Fellowship (HL144057) (N.B.N), California Institute for Regenerative Medicine (CIRM) (RN3-06378) (R.A.) and Eli & Edythe Broad Center of Regenerative Medicine and Stem Cell Research at UCLA Research Award (R.A.).
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
Ā© 2022 Springer Science+Business Media, LLC, part of Springer Nature
About this protocol
Cite this protocol
Pezhouman, A., Nguyen, N.B., Shevtsov, A., Qiao, R., Ardehali, R. (2022). In Vitro Generation of Heart Field Specific Cardiomyocytes. In: Kannan, N., Beer, P. (eds) Stem Cell Assays. Methods in Molecular Biology, vol 2429. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-1979-7_17
Download citation
DOI: https://doi.org/10.1007/978-1-0716-1979-7_17
Published:
Publisher Name: Humana, New York, NY
Print ISBN: 978-1-0716-1978-0
Online ISBN: 978-1-0716-1979-7
eBook Packages: Springer Protocols