Ohtani-Kaneko Rsituko, Sato Kenjiro, Tsutiya Atsuhiro, Nakagawa Yuka, Hashizume Kazutoshi, Tazawa Hidekatsu
Department of Life Sciences, Toyo University, 1-1-1 Itakura, Oura, Gunma, 374-0193, Japan.
Department of Biochemistry, St. Marianna University School of Medicine, 2-16-1 Sugao, Miyamae, Kawasaki, 216-8511, Japan.
Biomed Microdevices. 2017 Oct 9;19(4):91. doi: 10.1007/s10544-017-0229-5.
Induced pluripotent stem cell-derived endothelial cells (iPSC-ECs) can contribute to elucidating the pathogenesis of heart and vascular diseases and developing their treatments. Their precise characteristics in fluid flow however remain unclear. Therefore, the aim of the present study is to characterise these features. We cultured three types of ECs in a microfluidic culture system: commercially available human iPS-ECs, human umbilical vein endothelial cells (HUVECs) and human umbilical artery endothelial cells (HUAECs). We then examined the mRNA expression levels of endothelial marker gene cluster of differentiation 31 (CD31), fit-related receptor tyrosine kinase (Flk-1), and the smooth muscle marker gene smooth muscle alpha-actin, and investigated changes in plasminogen activator inhibitor-1 (PAI-1) secretion and intracellular F-actin arrangement following heat stress. We also compared expressions of the arterial and venous marker genes ephrinB2 and EphB4, and the endothelial gap junction genes connexin (Cx) 37, 40, and 43 under fluidic shear stress to determine their arterial or venous characteristics. We found that iPS-ECs had similar endothelial marker gene expressions and exhibited similar increases in PAI-1 secretion under heat stress as HUVECs and HUAECs. In addition, F-actin arrangement in iPSC-ECs also responded to heat stress, as previously reported. However, they had different expression patterns of arterial and venous marker genes and Cx genes under different fluidic shear stress levels, showing that iPSC-ECs exhibit different characteristics from arterial and venous ECs. This microfluidic culture system equipped with variable shear stress control will provide an easy-to-use assay tool to examine characteristics of iPS-ECs generated by different protocols in various laboratories and contribute to basic and applied biomedical researches on iPS-ECs.
诱导多能干细胞衍生的内皮细胞（iPSC-ECs）有助于阐明心脏和血管疾病的发病机制并开发相应治疗方法。然而，它们在流体流动中的精确特征仍不清楚。因此，本研究的目的是表征这些特征。我们在微流控培养系统中培养了三种类型的内皮细胞：市售的人iPS-ECs、人脐静脉内皮细胞（HUVECs）和人脐动脉内皮细胞（HUAECs）。然后，我们检测了内皮标记基因分化簇31（CD31）、fit相关受体酪氨酸激酶（Flk-1）以及平滑肌标记基因平滑肌α-肌动蛋白的mRNA表达水平，并研究了热应激后纤溶酶原激活物抑制剂-1（PAI-1）分泌和细胞内F-肌动蛋白排列的变化。我们还比较了流体剪切应力下动脉和静脉标记基因ephrinB2和EphB4以及内皮间隙连接基因连接蛋白（Cx）37、40和43的表达，以确定它们的动脉或静脉特征。我们发现，iPS-ECs具有与HUVECs和HUAECs相似的内皮标记基因表达，并且在热应激下PAI-1分泌也有类似的增加。此外，iPSC-ECs中的F-肌动蛋白排列也如先前报道的那样对热应激有反应。然而，在不同的流体剪切应力水平下，它们的动脉和静脉标记基因以及Cx基因具有不同的表达模式，表明iPSC-ECs表现出与动脉和静脉内皮细胞不同的特征。这种配备可变剪切应力控制的微流控培养系统将提供一种易于使用的检测工具，用于检测不同实验室通过不同方案生成的iPS-ECs的特征，并有助于iPS-ECs的基础和应用生物医学研究。
