Derivation of highly purified cardiomyocytes from human induced pluripotent stem cells using small molecule-modulated differentiation and subsequent glucose starvation.
Derivation of highly purified cardiomyocytes from human induced pluripotent stem cells using small molecule-modulated differentiation and subsequent glucose starvation.
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DOI：
10.3791/52628
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发表时间：
2015-03-18
期刊：
影响因子：
--
通讯作者：
Wu SM
中科院分区：
文献类型：
--
作者：
Sharma A;Li G;Rajarajan K;Hamaguchi R;Burridge PW;Wu SM
在这里，我们描述了一个强大的协议，人类心肌细胞的衍生相结合的小分子调节的心脏分化和葡萄糖剥夺介导的心肌细胞纯化，使生产纯化的心肌细胞的心血管疾病建模和药物筛选的目的。人诱导多能干细胞衍生的心肌细胞（hiPSC-CM）已成为解决可用于基础研究和转化应用的原代心肌细胞缺乏的重要细胞来源。为了使hiPSC分化成心肌细胞，已经开发了包括基于胚状体（EB）的分化和生长因子诱导的各种方案。然而，这些方案是低效的，并且在它们产生纯化的心肌细胞的能力方面高度可变。最近，一种基于小分子的方案利用Wnt/β-Catenin信号转导的调节被证明可以高效地促进心脏分化。使用该方案，始终观察到超过50-60%的分化细胞是心肌肌钙蛋白阳性心肌细胞。为了进一步提高心肌细胞纯度，使分化的细胞经受葡萄糖饥饿以基于心肌细胞和非心肌细胞之间的代谢差异特异性地消除非心肌细胞。使用这种选择策略，我们一致地获得了分化细胞群体中心肌细胞与非心肌细胞的比率增加大于30%。这些高度纯化的心肌细胞应该增强来自基于人iPSC的体外疾病建模研究和药物筛选测定的结果的可靠性。
Here, we describe a robust protocol for human cardiomyocyte derivation that combines small molecule-modulated cardiac differentiation and glucose deprivation-mediated cardiomyocyte purification, enabling production of purified cardiomyocytes for the purposes of cardiovascular disease modeling and drug screening. Human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) have become an important cell source to address the lack of primary cardiomyocytes available for basic research and translational applications. To differentiate hiPSCs into cardiomyocytes, various protocols including embryoid body (EB)-based differentiation and growth factor induction have been developed. However, these protocols are inefficient and highly variable in their ability to generate purified cardiomyocytes. Recently, a small molecule-based protocol utilizing modulation of Wnt/β-Catenin signaling was shown to promote cardiac differentiation with high efficiency. With this protocol, greater than 50–60% of differentiated cells were cardiac troponin-positive cardiomyocytes were consistently observed. To further increase cardiomyocyte purity, the differentiated cells were subjected to glucose starvation to specifically eliminate non-cardiomyocytes based on the metabolic differences between cardiomyocytes and non-cardiomyocytes. Using this selection strategy, we consistently obtained a greater than 30% increase in the ratio of cardiomyocytes to non-cardiomyocytes in a population of differentiated cells. These highly purified cardiomyocytes should enhance the reliability of results from human iPSC-based in vitro disease modeling studies and drug screening assays.
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影响因子：
23.9
作者：
Burridge, Paul W.;Keller, Gordon;Gold, Joseph D.;Wu, Joseph C.
通讯作者：
Wu, Joseph C.
影响因子：
20.1
作者：
Zhang J;Wilson GF;Soerens AG;Koonce CH;Yu J;Palecek SP;Thomson JA;Kamp TJ
通讯作者：
Kamp TJ
影响因子：
48
作者：
Burridge, Paul W.;Matsa, Elena;Shukla, Praveen;Lin, Ziliang C.;Churko, Jared M.;Ebert, Antje D.;Lan, Feng;Diecke, Sebastian;Huber, Bruno;Mordwinkin, Nicholas M.;Plews, Jordan R.;Abilez, Oscar J.;Cui, Bianxiao;Gold, Joseph D.;Wu, Joseph C.
通讯作者：
Wu, Joseph C.
影响因子：
20.1
作者：
Sharma A;Marceau C;Hamaguchi R;Burridge PW;Rajarajan K;Churko JM;Wu H;Sallam KI;Matsa E;Sturzu AC;Che Y;Ebert A;Diecke S;Liang P;Red-Horse K;Carette JE;Wu SM;Wu JC
通讯作者：
Wu JC
影响因子：
15.9
作者：
Kehat, I;Kenyagin-Karsenti, D;Gepstein, L
通讯作者：
Gepstein, L