Department of Anatomy and Embryology, Leiden University Medical Center, Einthovenweg 20, 2333 ZC Leiden, The Netherlands.
Department of Anatomy and Embryology, Leiden University Medical Center, Einthovenweg 20, 2333 ZC Leiden, The Netherlands.
Stem Cell Reports. 2019 Apr 9;12(4):647-656. doi: 10.1016/j.stemcr.2019.02.003. Epub 2019 Mar 7.
Vascular smooth muscle cells (vSMCs) are highly heterogeneous across different vascular beds. This is partly dictated by their developmental origin but also their position in the vascular tree, reflected in their differential responses to vasoactive agonists depending on which arteriolar or venular segment they are located. Functional assays are necessary to capture this heterogeneity in vitro since there are no markers that distinguish subtypes. Here we describe methods for determining real-time intracellular Ca release and contraction in vSMCs of neural crest origin differentiated from human induced pluripotent stem cells using multiple protocols, and compare these with primary human brain vascular pericytes and smooth muscle cells. Open-source software was adapted for automated high-density analysis of Ca-release kinetics and contraction by tracking individual cells. Simultaneous measurements on hundreds of cells revealed heterogeneity in responses to vasoconstrictors that would likely be overlooked using manual low-throughput assays or marker expression.
血管平滑肌细胞（vascular smooth muscle cells，vSMCs）在不同的血管床中具有高度的异质性。这部分是由它们的发育起源决定的，但也反映在它们在血管树中的位置上，这影响了它们对血管活性激动剂的不同反应，取决于它们所在的小动脉或小静脉段。由于没有区分亚型的标志物，因此需要功能测定来在体外捕获这种异质性。在这里，我们描述了使用多种方案从人诱导多能干细胞分化出神经嵴来源的 vSMCs 来确定实时细胞内 Ca 释放和收缩的方法，并将其与原代人脑血管周细胞和平滑肌细胞进行了比较。我们改编了开源软件，用于通过跟踪单个细胞自动进行高密度 Ca 释放动力学和收缩的分析。对数百个细胞的同时测量揭示了对血管收缩剂反应的异质性，如果使用手动低通量测定或标志物表达，这种异质性很可能会被忽略。
