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Defined Engineered Human Myocardium With Advanced Maturation for Applications in Heart Failure Modeling and Repair.

ABSTRACT: BACKGROUND:Advancing structural and functional maturation of stem cell-derived cardiomyocytes remains a key challenge for applications in disease modeling, drug screening, and heart repair. Here, we sought to advance cardiomyocyte maturation in engineered human myocardium (EHM) toward an adult phenotype under defined conditions. METHODS:We systematically investigated cell composition, matrix, and media conditions to generate EHM from embryonic and induced pluripotent stem cell-derived cardiomyocytes and fibroblasts with organotypic functionality under serum-free conditions. We used morphological, functional, and transcriptome analyses to benchmark maturation of EHM. RESULTS:EHM demonstrated important structural and functional properties of postnatal myocardium, including: (1) rod-shaped cardiomyocytes with M bands assembled as a functional syncytium; (2) systolic twitch forces at a similar level as observed in bona fide postnatal myocardium; (3) a positive force-frequency response; (4) inotropic responses to ?-adrenergic stimulation mediated via canonical ?1- and ?2-adrenoceptor signaling pathways; and (5) evidence for advanced molecular maturation by transcriptome profiling. EHM responded to chronic catecholamine toxicity with contractile dysfunction, cardiomyocyte hypertrophy, cardiomyocyte death, and N-terminal pro B-type natriuretic peptide release; all are classical hallmarks of heart failure. In addition, we demonstrate the scalability of EHM according to anticipated clinical demands for cardiac repair. CONCLUSIONS:We provide proof-of-concept for a universally applicable technology for the engineering of macroscale human myocardium for disease modeling and heart repair from embryonic and induced pluripotent stem cell-derived cardiomyocytes under defined, serum-free conditions.

SUBMITTER: Tiburcy M

PROVIDER: S-EPMC5501412 | biostudies-literature | 2017 May

REPOSITORIES: biostudies-literature

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Defined Engineered Human Myocardium With Advanced Maturation for Applications in Heart Failure Modeling and Repair.

Tiburcy Malte M Hudson James E JE Balfanz Paul P Schlick Susanne S Meyer Tim T Chang Liao Mei-Ling ML Levent Elif E Raad Farah F Zeidler Sebastian S Wingender Edgar E Riegler Johannes J Wang Mouer M Gold Joseph D JD Kehat Izhak I Wettwer Erich E Ravens Ursula U Dierickx Pieterjan P van Laake Linda W LW Goumans Marie Jose MJ Khadjeh Sara S Toischer Karl K Hasenfuss Gerd G Couture Larry A LA Unger Andreas A Linke Wolfgang A WA Araki Toshiyuki T Neel Benjamin B Keller Gordon G Gepstein Lior L Wu Joseph C JC Zimmermann Wolfram-Hubertus WH

Circulation 20170206 19

<h4>Background</h4>Advancing structural and functional maturation of stem cell-derived cardiomyocytes remains a key challenge for applications in disease modeling, drug screening, and heart repair. Here, we sought to advance cardiomyocyte maturation in engineered human myocardium (EHM) toward an adult phenotype under defined conditions.<h4>Methods</h4>We systematically investigated cell composition, matrix, and media conditions to generate EHM from embryonic and induced pluripotent stem cell-der ...[more]

PMID: 28167635

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Project description:The ability to differentiate human pluripotent stem cells (hPSCs) into cardiomyocytes (CMs) makes them an attractive source for repairing injured myocardium, disease modeling, and drug testing. Although current differentiation protocols yield hPSC-CMs to >90% efficiency, hPSC-CMs exhibit immature characteristics. With the goal of overcoming this limitation, we tested the effects of varying passive stretch on engineered heart muscle (EHM) structural and functional maturation, guided by computational modeling. Human embryonic stem cells (hESCs, H7 line) or human induced pluripotent stem cells (IMR-90 line) were differentiated to hPSC-derived cardiomyocytes (hPSC-CMs) in vitro using a small molecule based protocol. hPSC-CMs were characterized by troponin+ flow cytometry as well as electrophysiological measurements. Afterwards, 1.2 × 106 hPSC-CMs were mixed with 0.4 × 106 human fibroblasts (IMR-90 line) (3:1 ratio) and type-I collagen. The blend was cast into custom-made 12-mm long polydimethylsiloxane reservoirs to vary nominal passive stretch of EHMs to 5, 7, or 9 mm. EHM characteristics were monitored for up to 50 days, with EHMs having a passive stretch of 7 mm giving the most consistent formation. Based on our initial macroscopic observations of EHM formation, we created a computational model that predicts the stress distribution throughout EHMs, which is a function of cellular composition, cellular ratio, and geometry. Based on this predictive modeling, we show cell alignment by immunohistochemistry and coordinated calcium waves by calcium imaging. Furthermore, coordinated calcium waves and mechanical contractions were apparent throughout entire EHMs. The stiffness and active forces of hPSC-derived EHMs are comparable with rat neonatal cardiomyocyte-derived EHMs. Three-dimensional EHMs display increased expression of mature cardiomyocyte genes including sarcomeric protein troponin-T, calcium and potassium ion channels, ?-adrenergic receptors, and t-tubule protein caveolin-3. Passive stretch affects the structural and functional maturation of EHMs. Based on our predictive computational modeling, we show how to optimize cell alignment and calcium dynamics within EHMs. These findings provide a basis for the rational design of EHMs, which enables future scale-up productions for clinical use in cardiovascular tissue engineering. Stem Cells 2018;36:265-277.

Dataset Information

Defined Engineered Human Myocardium With Advanced Maturation for Applications in Heart Failure Modeling and Repair.

Publications

Defined Engineered Human Myocardium With Advanced Maturation for Applications in Heart Failure Modeling and Repair.

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