Abstract
BackgroundLong-QT syndrome type 2 (LQT2) is a common malignant hereditary arrhythmia. Due to the lack of suitable animal and human models, the pathogenesis of LQT2 caused by human ether-a-go-go-related gene (hERG) deficiency is still unclear. In this study, we generated an hERG-deficient human cardiomyocyte (CM) model that simulates ‘human homozygous hERG mutations’ to explore the underlying impact of hERG dysfunction and the genotype–phenotype relationship of hERG deficiency.MethodsThe KCNH2 was knocked out in the human embryonic stem cell (hESC) H9 line using the CRISPR/Cas9 system. Using a chemically defined differentiation protocol, we obtained and verified hERG-deficient CMs. Subsequently, high-throughput microelectrode array (MEA) assays and drug interventions were performed to characterise the electrophysiological signatures of hERG-deficient cell lines.ResultsOur results showed that KCNH2 knockout did not affect the pluripotency or differentiation efficiency of H9 cells. Using high-throughput MEA assays, we found that the electric field potential duration and action potential duration of hERG-deficient CMs were significantly longer than those of normal CMs. The hERG-deficient lines also exhibited irregular rhythm and some early afterdepolarisations. Moreover, we used the hERG-deficient human CM model to evaluate the potency of agents (nifedipine and magnesium chloride) that may ameliorate the phenotype.ConclusionsWe established an hERG-deficient human CM model that exhibited QT prolongation, irregular rhythm and sensitivity to other ion channel blockers. This model serves as an important tool that can aid in understanding the fundamental impact of hERG dysfunction, elucidate the genotype–phenotype relationship of hERG deficiency and facilitate drug development.
Highlights
Long-QT syndrome type 2 (LQT2) is a common malignant hereditary arrhythmia
HESC-H9 cells were subjected to electroporation with a plasmid containing stranded guide RNA (sgRNA) and Cas9, followed by puromycin screening
To determine whether KCNH2 knockout played the same role in different stem cell lines, we established a KCNH2−/−human pluripotent stem cells (hPSCs) cell line using the same method
Summary
Long-QT syndrome type 2 (LQT2) is a common malignant hereditary arrhythmia. Due to the lack of suitable animal and human models, the pathogenesis of LQT2 caused by human ether-a-go-go-related gene (hERG) deficiency is still unclear. The hERG channel constitutes a rapidly activated delayed rectifier K+ channel (IKr) in the heart [2] and is responsible for myocardial cell repolarisation when the cardiac action potential ends [3]. Previous evidence shows that KCNH2 mutations are associated with hereditary long-QT syndrome type 2 (LQT2) [4]. LQT2 has the characteristics of a prolonged QT interval and action potential duration (APD), which predispose patients to torsade de pointes, a type of polymorphic ventricular tachycardia and sudden cardiac death [6,7,8]. The need to establish an hERG-deficient model to determine the pathological mechanism of this disease is urgent
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