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Rat-Induced Pluripotent Stem Cells-Derived Cardiac Myocytes in a Cell Culture Dish

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Embryonic Stem Cell Protocols

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2520))

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Abstract

Induced pluripotent stem (iPS) cells are genetically reprogrammed somatic cells that exhibit embryonic stem cell-like characteristics such as self-renewal and pluripotency. These cells have broad differentiation capability to convert into diverse cell types that make up the primary germ layers during embryonic development. iPS cells can spontaneously differentiate and form cell aggregates termed embryoid bodies (EBs) in the absence of differentiation inhibitory factors. Unlike other methods used to generate EBs, “the hanging drop” method offers reproducibility and homogeneity from a set number of iPS cells. As such, we describe the differentiation of rat-induced pluripotent stem cells into cardiac myocytes in vitro using the hanging drop method. Both the confirmation and identification of the cardiac myocytes are done using immunocytochemistry, RT-PCR, Western Blot, and Flow Cytometry. Briefly, a specific number of iPS cells are placed in droplets on the lid of culture dishes and incubated for 2 days, yielding embryoid bodies, which are suspended and plated. Spontaneous beating of cardiomyocytes can be seen 7–14 days after the plating of EBs and specific cardiac markers can be observed through identification assays.

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Acknowledgments

This study was supported in part by National Institutes of Health grant 1R01DK120866-01 and 5R01CA221813-04 to D.K. Singla. Dr. Singla is also the holder of the Advent Health Endowed Chair in Cardiovascular Sciences and research is in part supported by this award. Dr. P.K. Singal is the holder of the Dr. Naranjan S. Dhalla Chair in Cardiovascular Research supported by the St. Boniface Hospital Foundation.

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Authors and Affiliations

  1. Division of Metabolic and Cardiovascular Sciences, Burnett School of Biomedical Sciences, College of Medicine, University of Central Florida, Orlando, FL, USA

    Fatima Bianca A. Dessouki & Dinender K. Singla

  2. Institute of Cardiovascular Sciences, St. Boniface Hospital Albrechtsen Research Centre, University of Manitoba, Winnipeg, MB, Canada

    Pawan K. Singal

Authors
  1. Fatima Bianca A. Dessouki
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  2. Pawan K. Singal
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  3. Dinender K. Singla
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Corresponding author

Correspondence to Dinender K. Singla .

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Editors and Affiliations

  1. Ottawa, ON, Canada

    Kursad Turksen

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Dessouki, F.B.A., Singal, P.K., Singla, D.K. (2021). Rat-Induced Pluripotent Stem Cells-Derived Cardiac Myocytes in a Cell Culture Dish. In: Turksen, K. (eds) Embryonic Stem Cell Protocols . Methods in Molecular Biology, vol 2520. Humana, New York, NY. https://doi.org/10.1007/7651_2021_406

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  • DOI: https://doi.org/10.1007/7651_2021_406

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-2436-4

  • Online ISBN: 978-1-0716-2437-1

  • eBook Packages: Springer Protocols

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