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Gene editing reverses arrhythmia susceptibility in humanized PLN-R14del mice: modeling a European cardiomyopathy with global impact

Title
Gene editing reverses arrhythmia susceptibility in humanized PLN-R14del mice: modeling a European cardiomyopathy with global impact
Author
정동탁
Keywords
Humanized mouse; Phospholamban R14del mutation; CRISPR; Cas9; Ventricular tachycardia; Gene therapy
Issue Date
2022-02
Publisher
OXFORD UNIV PRESS
Citation
CARDIOVASCULAR RESEARCH; FEB 22 2022, 11p.
Abstract
Aims A mutation in the phospholamban (PLN) gene, leading to deletion of Arg14 (R14del), has been associated with malignant arrhythmias and ventricular dilation. Identifying pre-symptomatic carriers with vulnerable myocardium is crucial because arrhythmia can result in sudden cardiac death, especially in young adults with PLN-R14del mutation. This study aimed at assessing the efficiency and efficacy of in vivo genome editing, using CRISPR/Cas9 and a cardiotropic adeno-associated virus-9 (AAV9), in improving cardiac function in young adult mice expressing the human PLN-R14del. Methods and results Humanized mice were generated expressing human wild-type (hPLN-WT) or mutant (hPLN-R14del) PLN in the heterozygous state, mimicking human carriers. Cardiac magnetic resonance imaging at 12 weeks of age showed bi-ventricular dilation and increased stroke volume in mutant vs. WT mice, with no deficit in ejection fraction or cardiac output. Challenge of ex vivo hearts with isoproterenol and rapid pacing unmasked higher propensity for sustained ventricular tachycardia (VT) in hPLN-R14del relative to hPLN-WT. Specifically, the VT threshold was significantly reduced (20.3 ± 1.2 Hz in hPLN-R14del vs. 25.7 ± 1.3 Hz in WT, P < 0.01) reflecting higher arrhythmia burden. To inactivate the R14del allele, mice were tail-vein-injected with AAV9.CRISPR/Cas9/gRNA or AAV9 empty capsid (controls). CRISPR-Cas9 efficiency was evaluated by droplet digital polymerase chain reaction and NGS-based amplicon sequencing. In vivo gene editing significantly reduced end-diastolic and stroke volumes in hPLN-R14del CRISPR-treated mice compared to controls. Susceptibility to VT was also reduced, as the VT threshold was significantly increased relative to controls (30.9 ± 2.3 Hz vs. 21.3 ± 1.5 Hz; P < 0.01). Conclusions This study is the first to show that disruption of hPLN-R14del allele by AAV9-CRISPR/Cas9 improves cardiac function and reduces VT susceptibility in humanized PLN-R14del mice, offering preclinical evidence for translatable approaches to therapeutically suppress the arrhythmogenic phenotype in human patients with PLN-R14del disease.
URI
https://academic.oup.com/cardiovascres/advance-article/doi/10.1093/cvr/cvac021/6534329?login=truehttps://repository.hanyang.ac.kr/handle/20.500.11754/170953
ISSN
00086363; 17553245
DOI
10.1093/cvr/cvac021
Appears in Collections:
COLLEGE OF SCIENCE AND CONVERGENCE TECHNOLOGY[E](과학기술융합대학) > MOLECULAR AND LIFE SCIENCE(분자생명과학과) > Articles
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