Chen Chu-Yen, Chou Fu-Sheng, Wang Pei-Shan
Department of Pediatrics, University of Kansas Medical Center, Kansas City, KS, USA.
Department of Pediatrics, University of Missouri-Kansas City, Kansas City, MO, USA.
Bio Protoc. 2019 Jun 20;9(12):e3275. doi: 10.21769/BioProtoc.3275.
Cell motility has been extensively studied in models using fibroblasts and keratocytes, but the cell type-specific mechanisms underlying migration of lineage- or disease-specific cells, such as neural and glial progenitor cells, remain an active field for investigation. The migrating neural and glial progenitor cells contribute to the development, tissue repair and tumor invasion in the central nervous system (CNS). Cell migration is a highly dynamic process which relies on membranous protrusions to assemble, extend, disassemble and retract. In the CNS, the motility of neural and glial progenitor cells is affected by various cell-autonomous and non-cell-autonomous mechanisms such as signaling molecules, actin and microtubule interactions, and environmental cues. Here, we described a live-cell migration assay for use in the assessment of neural and glial progenitor cell migration. We first will demonstrate the procedures for isolating and culturing neural and glial progenitor cells. Next, we will demonstrate the acquisition of time-lapse images using phase contrast microscopy, the methods for quantification and the analyses of various motility parameters including speed, velocity, straightness and leading-edge dynamics. This method allows researchers to dissect the mechanisms of cell motility in response to different environmental cues, such as chemoattractive and repulsive signals, matrix adhesiveness and stiffness. This assay also allows researchers to study migration of pharmacologically and genetically manipulated cells.
细胞运动性已在使用成纤维细胞和角膜细胞的模型中得到广泛研究，但谱系特异性或疾病特异性细胞（如神经和神经胶质祖细胞）迁移背后的细胞类型特异性机制仍是一个活跃的研究领域。迁移的神经和神经胶质祖细胞有助于中枢神经系统（CNS）的发育、组织修复和肿瘤侵袭。细胞迁移是一个高度动态的过程，依赖于膜状突起的组装、延伸、解体和回缩。在中枢神经系统中，神经和神经胶质祖细胞的运动性受多种细胞自主和非细胞自主机制的影响，如信号分子、肌动蛋白和微管相互作用以及环境线索。在此，我们描述了一种用于评估神经和神经胶质祖细胞迁移的活细胞迁移测定法。我们首先将演示分离和培养神经和神经胶质祖细胞的步骤。接下来，我们将演示使用相差显微镜获取延时图像的方法、定量方法以及对各种运动参数（包括速度、速率、直线度和前沿动态）的分析。该方法使研究人员能够剖析细胞对不同环境线索（如趋化性和排斥信号、基质粘附性和硬度）作出反应的运动机制。该测定法还使研究人员能够研究经药理学和基因操作的细胞的迁移。
