Skip to main content

Research Repository

Advanced Search

CRISPR/Cas9 editing in human pluripotent stem cell-cardiomyocytes highlights arrhythmias, hypocontractility, and energy depletion as potential therapeutic targets for hypertrophic cardiomyopathy

Mosqueira, Diogo; Mannhardt, Ingra; Bhagwan, Jamie R.; Lis-Slimak, Katarzyna; Katili, Puspita; Scott, Elizabeth; Hassan, Mustafa; Prondzynski, Maksymilian; Harmer, Stephen C.; Tinker, Andrew; Smith, James G.W.; Carrier, Lucie; Williams, Philip M.; Gaffney, Daniel; Eschenhagen, Thomas; Hansen, Arne; Denning, Chris

CRISPR/Cas9 editing in human pluripotent stem cell-cardiomyocytes highlights arrhythmias, hypocontractility, and energy depletion as potential therapeutic targets for hypertrophic cardiomyopathy Thumbnail


Authors

Diogo Mosqueira

Ingra Mannhardt

Jamie R. Bhagwan

Katarzyna Lis-Slimak

Puspita Katili

Elizabeth Scott

Mustafa Hassan

Maksymilian Prondzynski

Stephen C. Harmer

Andrew Tinker

James G.W. Smith

Lucie Carrier

PHIL WILLIAMS PHIL.WILLIAMS@NOTTINGHAM.AC.UK
Professor of Biophysics

Daniel Gaffney

Thomas Eschenhagen

Arne Hansen

CHRIS DENNING chris.denning@nottingham.ac.uk
Professor of Stem Cell Biology



Abstract

Aims: Sarcomeric gene mutations frequently underlie hypertrophic cardiomyopathy (HCM), a prevalent and complex condition leading to left ventricle thickening and heart dysfunction. We evaluated isogenic genome-edited human pluripotent stem cell-cardiomyocytes (hPSC-CM) for their validity to model, and add clarity to, HCM. Methods and results: CRISPR/Cas9 editing produced 11 variants of the HCM-causing mutation c.C9123T-MYH7 [(p.R453C-β-myosin heavy chain (MHC)] in 3 independent hPSC lines. Isogenic sets were differentiated to hPSC-CMs for high-throughput, non-subjective molecular and functional assessment using 12 approaches in 2D monolayers and/or 3D engineered heart tissues. Although immature, edited hPSC-CMs exhibited the main hallmarks of HCM (hypertrophy, multi-nucleation, hypertrophic marker expression, sarcomeric disarray). Functional evaluation supported the energy depletion model due to higher metabolic respiration activity, accompanied by abnormalities in calcium handling, arrhythmias, and contraction force. Partial phenotypic rescue was achieved with ranolazine but not omecamtiv mecarbil, while RNAseq highlighted potentially novel molecular targets. Conclusion: Our holistic and comprehensive approach showed that energy depletion affected core cardiomyocyte functionality. The engineered R453C-βMHC-mutation triggered compensatory responses in hPSC-CMs, causing increased ATP production and αMHC to energy-efficient βMHC switching. We showed that pharmacological rescue of arrhythmias was possible, while MHY7: MYH6 and mutant: wild-type MYH7 ratios may be diagnostic, and previously undescribed lncRNAs and gene modifiers are suggestive of new mechanisms.

Citation

Mosqueira, D., Mannhardt, I., Bhagwan, J. R., Lis-Slimak, K., Katili, P., Scott, E., …Denning, C. (2018). CRISPR/Cas9 editing in human pluripotent stem cell-cardiomyocytes highlights arrhythmias, hypocontractility, and energy depletion as potential therapeutic targets for hypertrophic cardiomyopathy. European Heart Journal, 39(43), 3879-3892. https://doi.org/10.1093/eurheartj/ehy249

Journal Article Type Article
Acceptance Date Apr 11, 2018
Online Publication Date May 8, 2018
Publication Date Nov 14, 2018
Deposit Date Apr 12, 2018
Publicly Available Date May 8, 2018
Journal European heart journal
Print ISSN 0195-668X
Electronic ISSN 1522-9645
Publisher Oxford University Press
Peer Reviewed Peer Reviewed
Volume 39
Issue 43
Pages 3879-3892
DOI https://doi.org/10.1093/eurheartj/ehy249
Keywords Hypertrophic cardiomyopathy; Disease modelling; CRISPR/Cas9; Genome-edited human pluripotent stem cell-cardiomyocytes; R453C-βMHC
Public URL https://nottingham-repository.worktribe.com/output/931354
Publisher URL https://academic.oup.com/eurheartj/advance-article/doi/10.1093/eurheartj/ehy249/4993867

Files




You might also like



Downloadable Citations