Mechanism-based facilitated maturation of human pluripotent stem cell-derived cardiomyocytes.
Mechanism-based facilitated maturation of human pluripotent stem cell-derived cardiomyocytes.
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DOI：
10.1161/circep.111.973420
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发表时间：
2013-02
期刊：
影响因子：
--
通讯作者：
Li RA
中科院分区：
文献类型：
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作者：
Lieu DK;Fu JD;Chiamvimonvat N;Tung KC;McNerney GP;Huser T;Keller G;Kong CW;Li RA
人胚胎干细胞（hESC）可以使用阶段特异性诱导方案有效且可重复地定向为心肌细胞（CM）。然而，其功能特性和临床和其他应用的适用性尚未得到评价。在这里，我们发现来自多种多能人类干细胞系的CM（hESC：H1、HES 2）和类型（诱导多能干细胞或iPSC），使用不同体外分化方案（胚状体形成、内胚层诱导、定向分化）通常显示出不成熟的促胚性动作电位（AP）特性，如高度自律性、去极化静息膜电位（RMP）、第4相-去极化和延迟后去极化（DAD）。在研究的肌膜离子电流（INa+/ICaL 2 +/IKr+/INCX+/If+/Ito+/IK 1-/IKs-）中，我们指出Kir2.1编码的内向整流K+电流（IK 1）的缺乏是hESC-CM中观察到的不成熟电生理特性的单一机制贡献者。Kir2.1在hESC-CM中的强制表达导致了Ba 2+敏感性IK 1的稳健表达，更重要的是，完全消除了所有的促细胞凋亡AP特征，使得电生理表型与成人对应物无法区分。这些结果提供了一个复杂的发育停滞表型的主要效应基因的第一个链接，重要的是，进一步引导我们开发一种仿生培养策略，以提高成熟。通过提供传统培养方法中缺少的环境线索，这种方法不需要任何遗传或药理干预。我们的研究结果可以通过提高衍生CM的产量、安全性和有效性来促进临床应用、药物发现和心脏毒性筛选。
Human embryonic stem cells (hESCs) can be efficiently and reproducibly directed into cardiomyocytes (CMs) using stage-specific induction protocols. However, their functional properties and suitability for clinical and other applications have not been evaluated. Here we showed that CMs derived from multiple pluripotent human stem cell lines (hESC: H1, HES2) and types (induced pluripotent stem cell or iPSC) using different in vitro differentiation protocols (embryoid body formation, endodermal induction, directed differentiation) commonly displayed immature, pro-arrhythmic action potential (AP) properties such as high-degree of automaticity, depolarized resting membrane potential (RMP), Phase 4- depolarization and delayed after-depolarization (DAD). Among the panoply of sarcolemmal ionic currents investigated (INa+/ICaL2+/IKr+/INCX+/If+/Ito+/IK1-/IKs-), we pinpointed the lack of the Kir2.1-encoded inwardly rectifying K+ current (IK1) as the single mechanistic contributor to the observed immature electrophysiological properties in hESC-CMs. Forced expression of Kir2.1 in hESC-CMs led to robust expression of Ba2+-sensitive IK1 and more importantly, completely ablated all the pro-arrhythmic AP traits, rendering the electrophysiological phenotype indistinguishable from the adult counterparts. These results provided the first link of a complex developmentally arrested phenotype to a major effector gene, and importantly, further led us to develop a biomimetic culturing strategy for enhancing maturation. By providing the environmental cues that are missing in conventional culturing method, this approach did not require any genetic or pharmacological interventions. Our findings can facilitate clinical applications, drug discovery and cardiotoxicity screening by improving the yield, safety and efficacy of derived CMs.