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Gelatin-based hydrogels combined with electrical stimulation to modulate neonatal rat cardiomyocyte beating and promote maturation

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Abstract

Cardiovascular diseases are the leading cause of morbidity and mortality throughout the world underlining the importance of efficient treatments including disease modeling and drug discovery by cardiac tissue engineering. However, the predictive power of these applications is currently limited by the immature state of the cardiomyocytes. Here, we developed gelatin hydrogels chemically crosslinked by genipin, a biocompatible crosslinker, as cell culture scaffolds. Neonatal rat cardiomyocytes appear synchronous beating within 2 days after seeding on hydrogels. Furthermore, we applied the electrical stimulation as a conditioning treatment to promote the maturation of cardiomyocytes cultured on the hydrogels. Our results show that electrical stimulation improves the organization of sarcomeres, establishment of gap junctions, calcium-handling capacity and propagation of pacing signals, thereby, increase the beating velocity of cardiomyocytes and responsiveness to external pacing. The above system can be applied in promoting physiological function maturation of engineered cardiac tissues, exhibiting promising applications in cardiac tissue engineering and drug screening.

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Acknowledgements

This work was financially supported by the National Natural Science Foundation of China (Grant No. 31871017), the Natural Science Foundation of Jiangsu Province, China (Grant No. BK20171352), the Southeast University-Nanjing Medical University Cooperative research project (2242019K3DN05), the Medical Science and Technology Development Foundation, Jiangsu Provincial Commission of Health and Family Planning, China (ZDRCA2016073), and the “111” Project (B17011, Ministry of Education of China).

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Authors and Affiliations

  1. State Key Laboratory of Bioelectronics, School of Biological Science and Medical Engineering, Southeast University, Si Pai Lou 2#, Nanjing, 210096, China

    Feng Zhang, Kaiyun Qu, Xiaopei Li, Chaoming Liu, Lazarus S. Ortiz & Ningping Huang

  2. Cardiovascular Center, Children’s Hospital of Nanjing Medical University, Nanjing, 210008, China

    Kaihong Wu

  3. Department of Cardiovascular Surgery, The First Affiliated Hospital of Nanjing Medical University, Nanjing, 210029, China

    Xiaowei Wang

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  1. Feng Zhang
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Contributions

FZ participated in the experimental research, data analysis, writing and editing of the manuscript. KYQ and XPL performed cardiomyocyte harvest and culture. CML and LSO established the experimental setup for electrical stimulation and developed softwares for data analysis. KHW and XWW provided guidance on cell study and helped to revise the manuscript. NPH conducted the design of the work as well as the deep review, editing, guidance, and supervision. All authors have read and approved the article for publication.

Corresponding author

Correspondence to Ningping Huang.

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Conflict of interest

The authors declare that there is no conflict of interest.

Ethical approval

Cardiomyocytes were extracted from neonatal rat ventricles of two-day-old Sprague–Dawley rats. All related procedures were carried out as approved by the Institutional Animal Care and Use Committee (IACUC) of Southeast University.

Electronic supplementary material

Below is the link to the electronic supplementary material.

Supporting movie S1: Synchronously beating behavior and calcium transient of cardiomyocytes on the gelatin-based hydrogels at day 2 after cell seeding. (AVI 15053 kb)

Supporting movie S2: Cardiomyocytes cultured on the culture dishes over time. (AVI 14868 kb)

Supporting movie S3: Beating behavior of cardiomyocytes cultured on different substrates. (AVI 19331 kb)

Supporting movie S4: The electrical stimulation can modulate the beating behavior of cardiomyocytes. (AVI 26793 kb)

Supporting movie S5: Electrical stimulation promotes the elongation and alignment of cardiomyocytes. (AVI 34992 kb)

42242_2020_100_MOESM6_ESM.docx

The following files are available free of charge: Characterization of gelatin-based scaffolds and electrical stimulation device; Immunofluorescent staining of cardiac-specific proteins (Cx43: in red; α-actinin: in green) on cardiomyocytes and vimentin (in blue) on non-muscle cells including fibroblasts and endothelial cells, with and without electrical stimulation. (DOCX 2130 kb)

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Zhang, F., Qu, K., Li, X. et al. Gelatin-based hydrogels combined with electrical stimulation to modulate neonatal rat cardiomyocyte beating and promote maturation. Bio-des. Manuf. 4, 100–110 (2021). https://doi.org/10.1007/s42242-020-00100-9

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  • DOI: https://doi.org/10.1007/s42242-020-00100-9

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