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Collagen scaffolds with or without the addition of RGD peptides support cardiomyogenesis after aggregation of mouse embryonic stem cells

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Abstract

Embryonic stem (ES) cell-based cardiac muscle repair using tissue-engineered scaffolds is an attractive prospective treatment option for patients suffering from heart disease. In this study, our aim was to characterize mouse ES cell-derived cardiomyocytes growing on collagen I/III scaffolds, modified with the adhesion peptides arginine-glycine-aspartic acid (RGD). Mouse ES-derived embryoid bodies (EBs) differentiated efficiently into beating cardiomyocytes on the collagen scaffolds. QPCR analysis and immunofluorescent staining showed that cardiomyocytes expressed cardiac muscle-related transcripts and proteins. Analysis of cardiomyocytes by electron microscopy identified muscle fiber bundles and Z bands, typical of ES-derived cardiomyocytes. No differences were detected between the collagen + RGD and collagen control scaffolds. ES cells that were not differentiated as EBs prior to seeding on the scaffold, did not differentiate into cardiomyocytes. These results indicate that a collagen I/III scaffold supports cardiac muscle development and function after EB formation, and that this scaffold appears suitable for future in vivo testing. The addition of the RGD domain to the collagen scaffold did not improve cardiomyocyte development or viability, indicating that RGD signaling to integrins was not a rate-limiting event for cardiomyogenesis from EBs seeded on a collagen scaffold.

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Acknowledgments

This work is supported by Canadian Institute of Health Research (CIHR) to IS and MR (Grant 53277), and Ontario Graduate Scholarship (OGS) to JD. We thank Peter Rippstein and Junhui Tan at the Core Pathology Laboratory and Division of Cardiology, University of Ottawa Heart Institute, for performing the electron microscopic imaging and sample processing. We thank Dr. deBold at the University of Ottawa Heart Institute for his advice on the EM images.

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Authors and Affiliations

  1. Department of Cellular and Molecular Medicine, University of Ottawa, 451 Smyth Road, Ottawa, ON, Canada, K1H 8M5

    Jennifer Dawson, Olivier Schussler & Marc Ruel

  2. Department of Biochemistry, Microbiology, and Immunology, University of Ottawa, 451 Smyth Road, Ottawa, ON, Canada, K1H 8M5

    Jennifer Dawson, Ashraf Al-Madhoun & Ilona S. Skerjanc

  3. Division of Cardiac Surgery, University of Ottawa Heart Institute, 40 Ruskin Street, Ottawa, ON, Canada, K1Y 4W7

    Olivier Schussler & Marc Ruel

  4. Centre de Recherche de Biochimie Macromoleculaire, CNRS UMR5237, 1919 Route de Mende, 34293, Montpellier, France

    Claudine Menard

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  1. Jennifer Dawson
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Correspondence to Ilona S. Skerjanc.

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Supplementary Figure 1

Five-day-old EBs grown until day 10 on the collagen (+RGD) and control scaffolds, and gelatin substrates differentiated more efficiently into beating cardiomyocytes than 3-day-old EBs or unaggregated cells. Higher levels of MHC, cardiac a actin, and Nkx2.5 transcripts were observed in cardiomyocytes aggregated for 5 d, when compared to unaggregated cells and 3-day-old mEBs grown on the substrates until day 10. Results are expressed relative to undifferentiated mES cells grown in monolayer, n = 3. (JPEG 73 kb)

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Dawson, J., Schussler, O., Al-Madhoun, A. et al. Collagen scaffolds with or without the addition of RGD peptides support cardiomyogenesis after aggregation of mouse embryonic stem cells. In Vitro Cell.Dev.Biol.-Animal 47, 653–664 (2011). https://doi.org/10.1007/s11626-011-9453-0

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