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基于亚型特异性启动子的动作电位成像技术,用于在人诱导多能干细胞衍生的心肌细胞中进行精确的疾病建模和药物测试。

Subtype-specific promoter-driven action potential imaging for precise disease modelling and drug testing in hiPSC-derived cardiomyocytes.

机构信息

I. Department of Medicine (Cardiology), Klinikum rechts der Isar, Technische Universität München, Munich 81675, Germany.

Institute for Molecular Cell Biology, Medical Faculty, University Homburg/Saar, Universität des Saarlandes, Homburg/Saar 66421, Germany.

出版信息

Eur Heart J. 2017 Jan 21;38(4):292-301. doi: 10.1093/eurheartj/ehw189.


DOI:10.1093/eurheartj/ehw189
PMID:28182242
原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC5381588/
Abstract

AIMS: Cardiomyocytes (CMs) generated from human induced pluripotent stem cells (hiPSCs) are increasingly used in disease modelling and drug evaluation. However, they are typically a heterogeneous mix of ventricular-, atrial-, and nodal-like cells based on action potentials (APs) and gene expression. This heterogeneity and the paucity of methods for high-throughput functional phenotyping hinder the full exploitation of their potential. We aimed at developing a method for rapid, sequential, and subtype-specific phenotyping of hiPSC-CMs with respect to AP morphology and single-cell arrhythmias. METHODS AND RESULTS: We used cardiac lineage-specific promoters to drive the expression of a voltage-sensitive fluorescent protein (VSFP-CR) in hiPSC-CMs, enabling subtype-specific optical AP recordings. In a patient-specific hiPSC model of long-QT syndrome type 1, AP prolongation and frequent early afterdepolarizations were evident in mutant ventricular- and atrial like, but not in nodal-like hiPSC-CMs compared with their isogenic controls, consistent with the selective expression of the disease-causing gene. Furthermore, we demonstrate the feasibility of sequentially probing a cell over several days to investigate genetic rescue of the disease phenotype and to discern CM subtype-specific drug effects. CONCLUSION: By combining a genetically encoded membrane voltage sensor with promoters that drive its expression in the major subtypes of hiPSC-CMs, we developed a convenient system for disease modelling and drug evaluation in the relevant cell type, which has the potential to advance the emerging utility of hiPSCs in cardiovascular medicine.

摘要

目的:源自人诱导多能干细胞(hiPSC)的心肌细胞(CMs)越来越多地用于疾病建模和药物评估。然而,基于动作电位(APs)和基因表达,它们通常是心室、心房和结样细胞的异质混合物。这种异质性以及高通量功能表型分析方法的缺乏阻碍了其潜力的充分发挥。我们旨在开发一种用于快速、连续和亚型特异性表型分析 hiPSC-CMs 的方法,以研究其 AP 形态和单细胞心律失常。

方法和结果:我们使用心脏谱系特异性启动子驱动 hiPSC-CMs 中电压敏感荧光蛋白(VSFP-CR)的表达,从而实现亚型特异性的光学 AP 记录。在 1 型长 QT 综合征的患者特异性 hiPSC 模型中,与同基因对照相比,突变型心室和心房样 hiPSC-CMs 中可见 AP 延长和频繁的早期后除极,而结样 hiPSC-CMs 中则没有,这与致病基因的选择性表达一致。此外,我们还证明了在几天内连续探测一个细胞以研究疾病表型的基因矫正并辨别 CM 亚型特异性药物作用的可行性。

结论:通过将基因编码的膜电压传感器与在 hiPSC-CMs 的主要亚型中驱动其表达的启动子相结合,我们开发了一种方便的疾病建模和药物评估系统,该系统有可能推进 hiPSC 在心血管医学中的新兴应用。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/201b/5381588/ac4cfa0df476/ehw18905.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/201b/5381588/6bf9f84d00ab/ehw18901.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/201b/5381588/5073015cb569/ehw18902.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/201b/5381588/5e3770297e23/ehw18903.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/201b/5381588/f28d3e1c7260/ehw18904.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/201b/5381588/ac4cfa0df476/ehw18905.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/201b/5381588/6bf9f84d00ab/ehw18901.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/201b/5381588/5073015cb569/ehw18902.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/201b/5381588/5e3770297e23/ehw18903.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/201b/5381588/f28d3e1c7260/ehw18904.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/201b/5381588/ac4cfa0df476/ehw18905.jpg

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Subtype-specific promoter-driven action potential imaging for precise disease modelling and drug testing in hiPSC-derived cardiomyocytes.

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Recent advancements of human iPSC derived cardiomyocytes in drug screening and tissue regeneration.

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[2]
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[3]
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[4]
Role of Cx43 in iPSC-CM Damage Induced by Microwave Radiation.

Int J Mol Sci. 2023-8-8

[5]
Human-Induced Pluripotent Stem Cell-Based Differentiation of Cardiomyocyte Subtypes for Drug Discovery and Cell Therapy.

Handb Exp Pharmacol. 2023

[6]
Recapitulating porcine cardiac development : from expanded potential stem cell to embryo culture models.

Front Cell Dev Biol. 2023-5-15

[7]
Cardiomyocyte electrophysiology and its modulation: current views and future prospects.

Philos Trans R Soc Lond B Biol Sci. 2023-6-19

[8]
High-throughput optical action potential recordings in hiPSC-derived cardiomyocytes with a genetically encoded voltage indicator in the locus.

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[9]
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[10]
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本文引用的文献

[1]
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Stem Cell Reports. 2015-9-12

[2]
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Int J Mol Sci. 2015-9-8

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Nat Biotechnol. 2015-7-20

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Circ Res. 2015-6-15

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Stem Cells. 2015-9

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Characterization of a novel KCNQ1 mutation for type 1 long QT syndrome and assessment of the therapeutic potential of a novel IKs activator using patient-specific induced pluripotent stem cell-derived cardiomyocytes.

Stem Cell Res Ther. 2015-3-19

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Stem Cells Transl Med. 2015-5

[8]
Antisense-mediated exon skipping: a therapeutic strategy for titin-based dilated cardiomyopathy.

EMBO Mol Med. 2015-5

[9]
Atrial-like cardiomyocytes from human pluripotent stem cells are a robust preclinical model for assessing atrial-selective pharmacology.

EMBO Mol Med. 2015-4

[10]
Mechanism of automaticity in cardiomyocytes derived from human induced pluripotent stem cells.

J Mol Cell Cardiol. 2015-4

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