Assays for in vitro monitoring of human airway smooth muscle (ASM) and human pulmonary arterial vascular smooth muscle (VSM) cell migration
Assays for in vitro monitoring of human airway smooth muscle (ASM) and human pulmonary arterial vascular smooth muscle (VSM) cell migration
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DOI：
10.1038/nprot.2006.434
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发表时间：
2006-01-01
期刊：
影响因子：
14.8
通讯作者：
Krymskaya, Vera P.
中科院分区：
文献类型：
--
作者：
Goncharova, Elena A.;Goncharov, Dmitry A.;Krymskaya, Vera P.
人肺血管平滑肌（VSM）细胞的迁移有助于肺动脉高压和动脉粥样硬化的血管重塑。证据还表明，气道平滑肌（ASM）细胞的迁移可能部分导致与哮喘相关的气道重塑。在这里，我们描述了VSM和ASM细胞在体外迁移使用Transwell或Boyden室测定。由于解剖信号机制调节细胞迁移需要分子的方法，我们的协议也描述了如何评估转染的VSM和ASM细胞的迁移。Transwell或Boyden室试验可在约8小时内完成，包括将去除血清的VSM或ASM细胞悬液接种在预涂有胶原蛋白的膜上，细胞向趋化梯度迁移，以及膜的目视（Transwell）或数字（Boyden室）分析。虽然Transwell检测很容易，但Boyden室检测需要实践经验;然而，这两种检测都是可靠的基于细胞的方法，提供了有关趋化和炎症因子如何调节VSM和ASM迁移的有价值的信息。
Migration of human pulmonary vascular smooth muscle (VSM) cells contributes to vascular remodeling in pulmonary arterial hypertension and atherosclerosis. Evidence also indicates that, in part, migration of airway smooth muscle (ASM) cells may contribute to airway remodeling associated with asthma. Here we describe migration of VSM and ASM cells in vitro using Transwell or Boyden chamber assays. Because dissecting signaling mechanisms regulating cell migration requires molecular approaches, our protocol also describes how to assess migration of transfected VSM and ASM cells. Transwell or Boyden chamber assays can be completed in approximately 8 h and include plating of serum-deprived VSM or ASM cell suspension on membrane precoated with collagen, migration of cells toward chemotactic gradient and visual (Transwell) or digital (Boyden chamber) analysis of membrane. Although the Transwell assay is easy, the Boyden chamber assay requires hands-on experience; however, both assays are reliable cell-based approaches providing valuable information on how chemotactic and inflammatory factors modulate VSM and ASM migration.