Cardiomyocyte differentiation

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Cardiomyocyte differentiation timecourse

Time course ID: human_HES3-GFP_Embryonic_Stem_cells
Sample provider:  Christine Mummery and Robert Passier

Introduction

Human H3 embryonic stem cells differentiated to cardiomyocytes

Human pluripotent stem cells have the potential to differentiate to all cell types of the human body, including cardiomyocytes. Since the adult human heart does not have the capacity to regenerate, loss of cardiomyocytes after myocardial infarction eventually may lead to heart failure[1]. Cardiomyocytes derived from human pluripotent stem cells may represent a source for future cell replacement in patients with cardiac injury. Furthermore, human cardiomyocytes may also be used for toxicity screening, drug discovery, and for studying mechanisms related to cardiac disease and development. It is important to study the regulatory molecular networks during cardiomyocyte differentiation in order to get a better understanding of processes such as specification, maturation and proliferation of cardiomyocytes. This will lead most likely to novel insights regarding endogenous cardiac regeneration, tissue engineering and cardiac disease.

Samples

For the differentiation of human pluripotent stem cells we used the human embryonic stem cell line HES3-GFP, ubiquitously expressing GFP. Previously, we have shown that co-culture of human embryonic stem cells with a mouse endoderm cell line, END-2, lead to beating cardiomyocytes within 12 days. END-2 cells were treated with mitocmycin C to block proliferation. In general, using this differentiation procedure (in the absence of serum) beating clusters contained 25 % cardiomyocytes[2], [3]. END-2 cells are cultured as a monolayer, whereas differentiated stem cells are grown on top as three-dimensional structures, which allows separation of human embryonic stem cell-derived populations. From undifferentiated human embryonic stem cells and every day during cardiomyocyte differentiation until day 12 samples were collected and used for RNA isolation (n=3).

Quality control

Expression of key markers for this differentiation course is based on previous experiments[4]. For undifferentiated stem cells, OCT4, NANOG, SOX2 are expressed at high levels. At day 3 we expect higher expression levels of MESP1, WNT3A, BrachyuryT. At day MEF2C, TBX5, NKX2.5 should be expressed at higher levels. Finally, at day 12, ACTN2, TNNT, PLN are expressed at high levels.

Human HES3-GFP Embryonic Stem cells.png

Figure 1: CAGE expression of marker genes in TPM.

References

[1] C. L. Mummery, J. Zhang, E. S. Ng, D. A. Elliott, A. G. Elefanty, and T. J. Kamp, “Differentiation of human embryonic stem cells and induced pluripotent stem cells to cardiomyocytes: a methods overview.,” Circ. Res., vol. 111, no. 3, pp. 344–358, Jul. 2012.

[2] R. Passier, D. W.-V. Oostwaard, J. Snapper, J. Kloots, R. J. Hassink, E. Kuijk, B. Roelen, A. B. de la Riviere, and C. Mummery, “Increased cardiomyocyte differentiation from human embryonic stem cells in serum-free cultures.,” Stem Cells, vol. 23, no. 6, pp. 772–780, Jun. 2005.

[3] C. Mummery, D. Ward-van Oostwaard, P. Doevendans, R. Spijker, S. van den Brink, R. Hassink, M. van der Heyden, T. Opthof, M. Pera, A. B. de la Riviere, R. Passier, and L. Tertoolen, “Differentiation of human embryonic stem cells to cardiomyocytes: role of coculture with visceral endoderm-like cells.,” Circulation, vol. 107, no. 21, pp. 2733–2740, Jun. 2003.

[4] A. Beqqali, J. Kloots, D. Ward-van Oostwaard, C. Mummery, and R. Passier, “Genome-wide transcriptional profiling of human embryonic stem cells differentiating to cardiomyocytes.,” Stem Cells, vol. 24, no. 8, pp. 1956–1967, Aug. 2006.






















































































Beginning of non-public section

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Zenbu configurations and status


Expression profiles

MARA based network results

Related samples

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References

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Quality control

https://fantom5-collaboration.gsc.riken.jp/webdav/home/arner/timecourse/time_course_main_paper_freeze_feb2013/qc_release_130226/human_HES3-GFP_Embryonic_Stem_cells/

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ISMARA analysis results

All samples: http://ismara.unibas.ch/timecourses/cardiomyocyte/ismara_report/index.html

Replicate averaged: http://ismara.unibas.ch/timecourses/cardiomyocyte-avgd/averaged_report/index.html


For more information, see ISMARA.

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