Unraveling the Inconsistencies of Cardiac Differentiation Efficiency Induced by the GSK3β Inhibitor CHIR99021 in Human Pluripotent Stem Cells.
Unraveling the Inconsistencies of Cardiac Differentiation Efficiency Induced by the GSK3β Inhibitor CHIR99021 in Human Pluripotent Stem Cells.
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DOI：
10.1016/j.stemcr.2018.03.023
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发表时间：
2018-06-05
影响因子：
5.9
通讯作者：
Oh S
中科院分区：
文献类型：
--
作者：
Laco F;Woo TL;Zhong Q;Szmyd R;Ting S;Khan FJ;Chai CLL;Reuveny S;Chen A;Oh S
心脏分化效率受到不一致性和低再现性的阻碍。我们分析了多种人多能干细胞（hPSC）系在不同细胞培养条件下响应于动态GSK 3 β抑制的分化过程。hPSC在不同的培养物汇合度下显示出细胞周期谱的强烈差异。具有较高百分比的细胞处于细胞周期的G1期的hPSC表现出细胞死亡，并且需要较低剂量的GSK 3 β抑制剂来诱导心脏分化。GSK 3 β抑制通过细胞周期蛋白D1启动细胞周期进程，并调节Wnt信号传导和转录因子（TCF）水平，导致中胚层分化加速或延迟。TCF水平是使用CHIR 99021的hPSC分化期间的关键调节因子。我们的结果解释了hPSC细胞系和培养条件的差异如何影响细胞死亡和心脏分化。通过分析细胞周期，我们能够选择高心源性hPSC系，并通过预测分化结果增加实验重现性。谱系多样性和细胞培养密度影响hPSC中的细胞周期CHIR 99021对hPSC具有细胞毒性，S/G2/M细胞周期阶段减少心脏分化的可重复性取决于hPSC中细胞周期的一致性细胞周期和TCF蛋白水平调节CHIR 99021诱导的分化在这篇文章中，Laco和同事表明，人类多能干细胞的特征在于单个细胞周期谱。当应用GSK 3 β抑制剂时，细胞周期谱的变化（主要由细胞培养密度引起）影响Wnt途径诱导的心脏分化和细胞死亡。我们表明，细胞周期验证是干细胞质量和实验重现性的关键。
Cardiac differentiation efficiency is hampered by inconsistencies and low reproducibility. We analyzed the differentiation process of multiple human pluripotent stem cell (hPSC) lines in response to dynamic GSK3β inhibition under varying cell culture conditions. hPSCs showed strong differences in cell-cycle profiles with varying culture confluency. hPSCs with a higher percentage of cells in the G1 phase of the cell cycle exhibited cell death and required lower doses of GSK3β inhibitors to induce cardiac differentiation. GSK3β inhibition initiated cell-cycle progression via cyclin D1 and modulated both Wnt signaling and the transcription factor (TCF) levels, resulting in accelerated or delayed mesoderm differentiation. The TCF levels were key regulators during hPSC differentiation with CHIR99021. Our results explain how differences in hPSC lines and culture conditions impact cell death and cardiac differentiation. By analyzing the cell cycle, we were able to select for highly cardiogenic hPSC lines and increase the experimental reproducibility by predicting differentiation outcomes. Lineage variety and cell culture density affect the cell cycle in hPSCs CHIR99021 is cytotoxic to hPSCs with reduced S/G2/M cell-cycle phases Cardiac differentiation reproducibility depends on cell-cycle consistency in hPSCs Cell cycle and TCF protein levels modulate CHIR99021-induced differentiation In this article, Laco and colleagues show that human pluripotent stem cells are characterized by individual cell-cycle profiles. Changes in cell-cycle profiles, mainly due to cell culture density, affect Wnt pathway-induced cardiac differentiation and cell death when GSK3β inhibitors are applied. We show that cell-cycle validation is crucial for stem cell quality and experimental reproducibility.
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