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Electrophysiological Analysis of human Pluripotent Stem Cell-derived Cardiomyocytes hPSC-CMs Using Multi-electrode Arrays MEAs
Published on: May 12, 2017
Exploring SCN5A variants associated with atrial fibrillation in atrial cardiomyocytes derived from human induced
Marion Pierre1, Mohammed Djemai1, Valérie Pouliot1
1CERVO Brain Research Centre, Quebec City, Quebec, Canada.
Insights
Investigating atrial fibrillation (AF) linked SCN5A variants using human induced pluripotent stem cells (hiPSCs) reveals distinct gain and loss-of-function effects impacting cardiac electrical activity and arrhythmogenesis.
Area of Science:
- Cardiovascular Genetics
- Stem Cell Biology
- Electrophysiology
Background:
- Atrial fibrillation (AF) is a significant risk factor for heart failure, myocardial infarction, and stroke.
- While SCN5A variants are implicated in AF, their precise mechanisms remain elusive.
- Human induced pluripotent stem cells (hiPSCs) offer a powerful model for studying AF-related SCN5A variants.
Purpose of the Study:
- To evaluate the electrophysiological consequences of three AF-associated SCN5A variants (K1493R, M1875T, N1986K).
- To elucidate the functional impact of these variants on cardiac sodium channel NaV1.5 activity.
Main Methods:
- Generation of a NaV1.5 knockout hiPSC line using CRISPR-Cas9.
- Differentiation into atrial cardiomyocytes and introduction of wild-type (WT) or variant SCN5A channels.
- Analysis via molecular biology, optical mapping, and electrophysiology.
Main Results:
- NaV1.5 knockout altered cardiac gene expression, conduction velocity, action potential parameters, and calcium handling.
- WT channel reintroduction restored normal electrophysiological function.
- One variant (N1986K) caused loss-of-function; two variants caused gain-of-function in NaV1.5 channel activity.
- AF variants induced cellular excitability changes and early afterdepolarizations.
Conclusions:
- Specific alterations in NaV1.5 channel function contribute to atrial excitability defects and arrhythmogenesis in AF.
- The developed knockout model provides a novel platform for studying SCN5A variants in a human cardiac context.
Background:
Atrial fibrillation (AF) poses a major risk for heart failure, myocardial infarction, and stroke. Several studies have linked SCN5A variants to AF, but their precise mechanistic contribution remains unclear. Human induced pluripotent stem cells (hiPSCs) provide a promising platform for modeling AF-linked SCN5A variants and their functional alterations.
Objective:
The purpose of this study was to assess the electrophysiological impact of 3 AF-linked SCN5A variants (K1493R, M1875T, N1986K) identified in 3 unrelated individuals.
Methods:
CRISPR-Cas9 was used to generate a new hiPSC line in which NaV1.5 was knocked out. Following differentiation into specific atrial cardiomyocyte by using retinoic acid, the adult wild-type (WT) and 3 AF variants were introduced into the NaV1.5 knockout (KO) line through transfection. Subsequent analysis including molecular biology, optical mapping, and electrophysiology were performed.
Results:
The absence of NaV1.5 channels altered the expression of key cardiac genes. NaV1.5 KO atrial-like cardiomyocytes derived from human induced pluripotent stem cells displayed slower conduction velocities, altered action potential (AP) parameters, and impaired calcium transient propagation. The transfection of the WT channel restored sodium current density, AP characteristics and the expression of several cardiac genes. Among the AF variants, 1 induced a loss of function (N1986K) while the other 2 induced a gain of function in NaV1.5 channel activity. Cellular excitability alterations and early afterdepolarizations were observed in AF variants.
Conclusion:
Our findings suggest that distinct alterations in NaV1.5 channel properties may trigger altered atrial excitability and arrhythmogenic activity in AF. Our KO model offers an innovative approach for investigating SCN5A variants in an adult human cardiac environment.
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