Efficient and reproducible generation of human iPSC-derived cardiomyocytes using a stirred bioreactor
Abstract In the last decade human iPSC-derived cardiomyocytes (hiPSC-CMs) proved to be valuable for cardiac disease modeling and cardiac regeneration, yet challenges with scale, quality, inter-batch consistency, and cryopreservation remain, reducing experimental reproducibility and limiting clinical translation. Here, we report a robust cardiac differentiation protocol that uses Wnt modulation and a stirred suspension bioreactor to produce on average 124 million hiPSC-CMs with >90% purity using a variety of hiPSC lines (19 differentiations; 10 iPSC lines). After controlled freeze and thaw, bioreactor-derived CMs (bCMs) showed high viability (>90%), interbatch reproducibility in cellular morphology, function, drug response and ventricular identity, which was further supported by single cell transcriptomes. bCMs on microcontact printed substrates revealed a higher degree of sarcomere maturation and viability during long-term culture compared to monolayer-derived CMs (mCMs). Moreover, functional investigation of bCMs in 3D engineered heart tissues showed earlier and stronger force production during long-term culture, and robust pacing capture up to 4 Hz when compared to mCMs. bCMs derived from this differentiation protocol will expand the applications of hiPSC-CMs by providing a reproducible, scalable, and resource efficient method to generate cardiac cells with well-characterized structural and functional properties superior to standard mCMs..
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Preprint| Erscheinungsjahr: |
2024 |
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| Erschienen: |
2024 |
| Enthalten in: |
bioRxiv.org - (2024) vom: 01. März Zur Gesamtaufnahme - year:2024 |
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| Sprache: |
Englisch |
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| Beteiligte Personen: |
Prondzynski, Maksymilian [VerfasserIn] |
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| Links: |
Volltext [kostenfrei] |
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| doi: |
10.1101/2024.02.24.581789 |
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| PPN (Katalog-ID): |
XBI042663067 |
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| 520 | |a Abstract In the last decade human iPSC-derived cardiomyocytes (hiPSC-CMs) proved to be valuable for cardiac disease modeling and cardiac regeneration, yet challenges with scale, quality, inter-batch consistency, and cryopreservation remain, reducing experimental reproducibility and limiting clinical translation. Here, we report a robust cardiac differentiation protocol that uses Wnt modulation and a stirred suspension bioreactor to produce on average 124 million hiPSC-CMs with >90% purity using a variety of hiPSC lines (19 differentiations; 10 iPSC lines). After controlled freeze and thaw, bioreactor-derived CMs (bCMs) showed high viability (>90%), interbatch reproducibility in cellular morphology, function, drug response and ventricular identity, which was further supported by single cell transcriptomes. bCMs on microcontact printed substrates revealed a higher degree of sarcomere maturation and viability during long-term culture compared to monolayer-derived CMs (mCMs). Moreover, functional investigation of bCMs in 3D engineered heart tissues showed earlier and stronger force production during long-term culture, and robust pacing capture up to 4 Hz when compared to mCMs. bCMs derived from this differentiation protocol will expand the applications of hiPSC-CMs by providing a reproducible, scalable, and resource efficient method to generate cardiac cells with well-characterized structural and functional properties superior to standard mCMs. | ||
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| 700 | 1 | |a Bortolin, Raul H. |4 aut | |
| 700 | 1 | |a Berkson, Paul |4 aut | |
| 700 | 1 | |a Trembley, Michael A. |4 aut | |
| 700 | 1 | |a Shani, Kevin |4 aut | |
| 700 | 1 | |a Sweat, Mason E. |4 aut | |
| 700 | 1 | |a Mayourian, Joshua |4 aut | |
| 700 | 1 | |a Cordoves, Albert M. |4 aut | |
| 700 | 1 | |a Anyanwu, Nnaemeka J. |4 aut | |
| 700 | 1 | |a Tharani, Yashasvi |4 aut | |
| 700 | 1 | |a Cotton, Justin |4 aut | |
| 700 | 1 | |a Milosh, Joseph B. |4 aut | |
| 700 | 1 | |a Walker, David |4 aut | |
| 700 | 1 | |a Zhang, Yan |4 aut | |
| 700 | 1 | |a Liu, Fujian |4 aut | |
| 700 | 1 | |a Liu, Xujie |4 aut | |
| 700 | 1 | |a Parker, Kevin K. |4 aut | |
| 700 | 1 | |a Bezzerides, Vassilios J. |4 aut | |
| 700 | 1 | |a Pu, William T. |4 aut | |
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