Jeziorowska Dorota, Fontaine Vincent, Jouve Charlène, Villard Eric, Dussaud Sébastien, Akbar David, Letang Valérie, Cervello Pauline, Itier Jean-Michiel, Pruniaux Marie-Pierre, Hulot Jean-Sébastien
Sorbonne Universités, UPMC Univ Paris 06, AP-HP, INSERM, Pitié-Salpêtrière Hospital, F-75013 Paris, France.
Institute of Cardiometabolism and Nutrition (ICAN), F-75013 Paris, France.
Int J Mol Sci. 2017 Jun 1;18(6):1173. doi: 10.3390/ijms18061173.
Human induced pluripotent stem cells (iPSCs) represent a powerful human model to study cardiac disease in vitro, notably channelopathies and sarcomeric cardiomyopathies. Different protocols for cardiac differentiation of iPSCs have been proposed either based on embroid body formation (3D) or, more recently, on monolayer culture (2D). We performed a direct comparison of the characteristics of the derived cardiomyocytes (iPSC-CMs) on day 27 ± 2 of differentiation between 3D and 2D differentiation protocols with two different Wnt-inhibitors were compared: IWR1 (inhibitor of Wnt response) or IWP2 (inhibitor of Wnt production). We firstly found that the level of Troponin T () expression measured by FACS was significantly higher for both 2D protocols as compared to the 3D protocol. In the three methods, iPSC-CM show sarcomeric structures. However, iPSC-CM generated in 2D protocols constantly displayed larger sarcomere lengths as compared to the 3D protocol. In addition, mRNA and protein analyses reveal higher cTNi to ssTNi ratios in the 2D protocol using IWP2 as compared to both other protocols, indicating a higher sarcomeric maturation. Differentiation of cardiac myocytes with 2D monolayer-based protocols and the use of IWP2 allows the production of higher yield of cardiac myocytes that have more suitable characteristics to study sarcomeric cardiomyopathies.
人诱导多能干细胞（iPSC）是一种强大的人体模型，可用于体外研究心脏病，尤其是离子通道病和肌节性心肌病。已经提出了基于胚状体形成（3D）或最近基于单层培养（2D）的不同iPSC心脏分化方案。我们对3D和2D分化方案在分化第27±2天获得的心肌细胞（iPSC-CM）的特征进行了直接比较，比较了两种不同的Wnt抑制剂：IWR1（Wnt反应抑制剂）或IWP2（Wnt产生抑制剂）。我们首先发现，与3D方案相比，两种2D方案通过流式细胞术测量的肌钙蛋白T（）表达水平均显著更高。在这三种方法中，iPSC-CM均显示出肌节结构。然而，与3D方案相比，2D方案生成的iPSC-CM的肌节长度始终更长。此外，mRNA和蛋白质分析显示，与其他两种方案相比，使用IWP2的2D方案中cTNi与ssTNi的比率更高，表明肌节成熟度更高。基于2D单层的方案分化心肌细胞并使用IWP2能够产生更高产量的心肌细胞，这些心肌细胞具有更适合研究肌节性心肌病 的特征。
