PDF(Pubmed)
Abstract：
BACKGROUND: Cardiomyocytes (CMs) derived from human induced pluripotent stem cells (hiPSCs) by traditional methods are a mix of atrial and ventricular CMs and many other non-cardiomyocyte cells. Retinoic acid (RA) plays an important role in regulation of the spatiotemporal development of the embryonic heart.
METHODS: CMs were derived from hiPSC (hi-PCS-CM) using different concentrations of RA (Control without RA, LRA with 0.05μM and HRA with 0.1 μM) between day 3-6 of the differentiation process. Engineered heart tissues (EHTs) were generated by assembling hiPSC-CM at high cell density in a low collagen hydrogel.
RESULTS: In the HRA group, hiPSC-CMs exhibited highest expression of contractile proteins MYH6, MYH7 and cTnT. The expression of TBX5, NKX2.5 and CORIN, which are marker genes for left ventricular CMs, was also the highest in the HRA group. In terms of EHT, the HRA group displayed the highest contraction force, the lowest beating frequency, and the highest sensitivity to hypoxia and isoprenaline, which means it was functionally more similar to the left ventricle. RNAsequencing revealed that the heightened contractility of EHT within the HRA group can be attributed to the promotion of augmented extracellular matrix strength by RA.
CONCLUSIONS: By interfering with the differentiation process of hiPSC with a specific concentration of RA at a specific time, we were able to successfully induce CMs and EHTs with a phenotype similar to that of the left ventricle or right ventricle.
METHODS: CMs were derived from hiPSC (hi-PCS-CM) using different concentrations of RA (Control without RA, LRA with 0.05μM and HRA with 0.1 μM) between day 3-6 of the differentiation process. Engineered heart tissues (EHTs) were generated by assembling hiPSC-CM at high cell density in a low collagen hydrogel.
RESULTS: In the HRA group, hiPSC-CMs exhibited highest expression of contractile proteins MYH6, MYH7 and cTnT. The expression of TBX5, NKX2.5 and CORIN, which are marker genes for left ventricular CMs, was also the highest in the HRA group. In terms of EHT, the HRA group displayed the highest contraction force, the lowest beating frequency, and the highest sensitivity to hypoxia and isoprenaline, which means it was functionally more similar to the left ventricle. RNAsequencing revealed that the heightened contractility of EHT within the HRA group can be attributed to the promotion of augmented extracellular matrix strength by RA.
CONCLUSIONS: By interfering with the differentiation process of hiPSC with a specific concentration of RA at a specific time, we were able to successfully induce CMs and EHTs with a phenotype similar to that of the left ventricle or right ventricle.
摘要：
背景：通过传统方法衍生自人诱导多能干细胞（hiPSC）的心肌细胞（CM）是心房和心室CM以及许多其他非心肌细胞的混合物。维甲酸（RA）在调节胚胎心脏的时空发育中起着重要作用。
方法：使用不同浓度的RA从hiPSC（hi-PCS-CM）衍生出CM（无RA的对照，在分化过程的第3-6天之间，LRA为0.05μM，HRA为0.1μM）。通过在低胶原水凝胶中以高细胞密度组装hiPSC-CM来产生工程化心脏组织(EHT)。
结果：在HRA组中，hiPSC-CM表现出最高的收缩蛋白MYH6，MYH7和cTnT表达。TBX5、NKX2.5和CORIN的表达,它们是左心室CMs的标记基因，在HRA组中也是最高的。就EHT而言,HRA组表现出最高的收缩力，最低的跳动频率，对缺氧和异丙肾上腺素的敏感性最高，这意味着它在功能上更类似于左心室。RNA测序表明，HRA组中EHT的收缩性增强可归因于RA促进了细胞外基质强度的增强。
结论：通过在特定时间用特定浓度的RA干扰hiPSC的分化过程，我们成功诱导出表型与左心室或右心室相似的CM和EHT.
方法：使用不同浓度的RA从hiPSC（hi-PCS-CM）衍生出CM（无RA的对照，在分化过程的第3-6天之间，LRA为0.05μM，HRA为0.1μM）。通过在低胶原水凝胶中以高细胞密度组装hiPSC-CM来产生工程化心脏组织(EHT)。
结果：在HRA组中，hiPSC-CM表现出最高的收缩蛋白MYH6，MYH7和cTnT表达。TBX5、NKX2.5和CORIN的表达,它们是左心室CMs的标记基因，在HRA组中也是最高的。就EHT而言,HRA组表现出最高的收缩力，最低的跳动频率，对缺氧和异丙肾上腺素的敏感性最高，这意味着它在功能上更类似于左心室。RNA测序表明，HRA组中EHT的收缩性增强可归因于RA促进了细胞外基质强度的增强。
结论：通过在特定时间用特定浓度的RA干扰hiPSC的分化过程，我们成功诱导出表型与左心室或右心室相似的CM和EHT.
