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Cryopreservation of human pluripotent stem cell-derived cardiomyocytes is not detrimental to their molecular and functional properties.

ABSTRACT: Human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) have emerged as a powerful platform for in vitro modelling of cardiac diseases, safety pharmacology and drug screening. All these applications require large quantities of well-characterised and standardised batches of hiPSC-CMs. Cryopreservation of hiPSC-CMs without affecting their biochemical or biophysical phenotype is essential for facilitating this, but ideally requires the cells being unchanged by the freeze-thaw procedure. We therefore compared the in vitro functional and molecular characteristics of fresh and cryopreserved hiPSC-CMs generated from multiple independent hiPSC lines. While the frozen hiPSC-CMs exhibited poorer replating than their freshly-derived counterparts, there was no difference in the proportion of cardiomyocytes retrieved from the mixed population when this was factored in, although for several lines a higher percentage of ventricular-like hiPSC-CMs were recovered following cryopreservation. Furthermore, cryopreserved hiPSC-CMs from one line exhibited longer action potential durations. These results provide evidence that cryopreservation does not compromise the in vitro molecular, physiological and mechanical properties of hiPSC-CMs, though can lead to an enrichment in ventricular myocytes. It also validates this procedure for storing hiPSC-CMs, thereby allowing the same batch of hiPSC-CMs to be used for multiple applications and evaluations.

SUBMITTER: van den Brink L

PROVIDER: S-EPMC7611364 | biostudies-literature | 2020 Mar

REPOSITORIES: biostudies-literature

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Cryopreservation of human pluripotent stem cell-derived cardiomyocytes is not detrimental to their molecular and functional properties.

van den Brink Lettine L Brandão Karina O KO Yiangou Loukia L Mol Mervyn P H MPH Grandela Catarina C Mummery Christine L CL Verkerk Arie O AO Davis Richard P RP

Stem cell research 20200107

Human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) have emerged as a powerful platform for in vitro modelling of cardiac diseases, safety pharmacology and drug screening. All these applications require large quantities of well-characterised and standardised batches of hiPSC-CMs. Cryopreservation of hiPSC-CMs without affecting their biochemical or biophysical phenotype is essential for facilitating this, but ideally requires the cells being unchanged by the freeze-thaw procedu ...[more]

PMID: 31945612

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Project description:Despite preclinical studies demonstrating the functional benefit of transplanting human pluripotent stem cell-derived cardiomyocytes (PSC-CMs) into damaged myocardium, the ability of these immature cells to adopt a more adult-like cardiomyocyte (CM) phenotype remains uncertain. To address this issue, we tested the hypothesis that prolonged in vitro culture of human embryonic stem cell (hESC)- and human induced pluripotent stem cell (hiPSC)-derived CMs would result in the maturation of their structural and contractile properties to a more adult-like phenotype. Compared to their early-stage counterparts (PSC-CMs after 20-40 days of in vitro differentiation and culture), late-stage hESC-CMs and hiPSC-CMs (80-120 days) showed dramatic differences in morphology, including increased cell size and anisotropy, greater myofibril density and alignment, sarcomeres visible by bright-field microscopy, and a 10-fold increase in the fraction of multinucleated CMs. Ultrastructural analysis confirmed improvements in the myofibrillar density, alignment, and morphology. We measured the contractile performance of late-stage hESC-CMs and hiPSC-CMs and noted a doubling in shortening magnitude with slowed contraction kinetics compared to the early-stage cells. We then examined changes in the calcium-handling properties of these matured CMs and found an increase in calcium release and reuptake rates with no change in the maximum amplitude. Finally, we performed electrophysiological assessments in hESC-CMs and found that late-stage myocytes have hyperpolarized maximum diastolic potentials, increased action potential amplitudes, and faster upstroke velocities. To correlate these functional changes with gene expression, we performed qPCR and found a robust induction of the key cardiac structural markers, including ?-myosin heavy chain and connexin-43, in late-stage hESC-CMs and hiPSC-CMs. These findings suggest that PSC-CMs are capable of slowly maturing to more closely resemble the phenotype of adult CMs and may eventually possess the potential to regenerate the lost myocardium with robust de novo force-producing tissue.

Dataset Information

Cryopreservation of human pluripotent stem cell-derived cardiomyocytes is not detrimental to their molecular and functional properties.

Publications

Cryopreservation of human pluripotent stem cell-derived cardiomyocytes is not detrimental to their molecular and functional properties.

Similar Datasets

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