Generation and Maturation of Human IPSC-Derived Myocardium in a Closed and Automated Bioreactor System

 

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Objectives: The acceptance of iPSC-derived myocardium as invaluable tool for in vitro disease modeling and drug development is steadily growing. To increase usability and broaden its spectrum of applications, novel strategies for scalable production, cultivation, and analysis are needed.

Methods: We developed a closed microscopically accessible bioreactor system for the formation, automated cultivation and characterization of miniaturized rod-shaped 3D bioartificial cardiac tissue constructs (BCTs). Each collagen type I-based BCT (starting volume: 330 μL) comprised 10⁶ human wild type cardiomyocytes and 10⁵ irradiated fibroblasts. The liquid cell–matrix mixture was injected into a closed vessel via a perfusion inlet. After autonomous matrix solidification, resulting in a semi-solid tissue precursor, the chamber was automatically perfused with 0.2 mL/h of culture medium at 37°C. Physiological characterization was performed in a custom-made multimodal bioreactor setup. Field potentials (FPs) and action potentials (APs) were recorded using a multi electrode array and a sharp-electrode impaling set-up.

Result: Spontaneous contraction of BCTs started from day two of cultivation and substantial compaction occurred between day two and 14 (2.85 ± 0.65 mm²; 0.45 ± 0.11 mm²). Further tissue maturation between day 14 and 21 was indicated by a significant reduction in beating frequency (1.33 ± 0.29 Hz; 0.64 ± 0.13 Hz) and an increase in maximum contractile tension (2.13 ± 0.63 kPa; 5.23 ± 0.4 kPa). Although a Frank-Starling mechanism was already detectable on day 14, it was significantly more pronounced on day 21. FP recordings on day 21 of BCTs exposed to cardio-active substances showed typical physiological responses. APs acquired on day 21 also showed near physiological values, with a resting membrane potential of −46.11 ± 1.9 mV, an action potential amplitude of 69.59 ± 2.2 mV and an action potential duration of 280.9 ± 13.7 milliseconds at 90% repolarization (APD₉₀). Histology on day 21 revealed an even distribution of CMs within BCTs along the axis of strain with well-pronounced sarcomeres.

Conclusion: In this proof-of-concept study we show that BCTs with advanced maturity can be generated in a closed and automated system without interim manipulation. Physiological characteristics and histological appearance of resulting tissue constructs resemble native myocardium to a large extent. Therefore, our results may serve as a base for future applications at industry-compatible scales.

Publication History

Article published online:
19 February 2021

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