Analysis of cyclic-stretching responses using cell-adhesion-patterned cells
Analysis of cyclic-stretching responses using cell-adhesion-patterned cells
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DOI：
10.1016/j.jbiotec.2007.09.017
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发表时间：
2008-01-01
影响因子：
4.1
通讯作者：
Naruse, Keiji
中科院分区：
文献类型：
--
作者：
Katanosaka, Yuki;Bao, Jin-Hua;Naruse, Keiji
人血管内皮细胞形成血流和血管壁之间的界面，并持续受到机械刺激。当内皮细胞沿沿着一个轴循环拉伸时，它们垂直于拉伸轴排列。我们以前报道，应用一个周期性的，单轴应变细胞诱导酪氨酸磷酸化的粘着斑激酶和刺激丝裂原活化蛋白激酶。然而，这是很难量化和分析的酪氨酸磷酸化在这些细胞中的空间分布，因为它们形成局灶性粘连随机。在这项研究中，我们开发了一个系统来克服这个问题，通过制备单独的，均匀的，图案化的细胞，可以周期性和单轴拉伸。我们构建了聚二甲基硅氧烷拉伸室，并使用微接触印刷技术压印2 μ m的纤连蛋白点（10行× 10列在38 μ m的正方形）的图案，然后将它们与人脐静脉内皮细胞（HUVEC）接种。我们发现，大多数HUVEC连接到图案化的点后2小时，在大小和形态相似，基于相差显微镜。在这个系统中，我们能够统计分析酪氨酸磷酸化和肌动蛋白聚合在这些图案化的细胞，当受到一个循环，单轴应变，使用荧光显微镜。(C)2007 Elsevier B.V.保留所有权利。
Human vascular endothelial cells form the interface between the bloodstream and vessel walls and are continuously subjected to mechanical stimulation. When endothelial cells are stretched cyclically, along one axis, they align perpendicular to the axis of stretch. We previously reported that applying a cyclic, uni-axial strain to cells induced tyrosine phospborylation of focal adhesion kinase and stimulated mitogen-activated protein kinase. However, it is difficult to quantify and analyze the spatial distribution of tyrosine phosphorylation in these cells, as they form focal adhesions randomly. In this study, we developed a system to overcome this problem by preparing individual, uniform, patterned cells that could be stretched cyclically and uni-axially. We constructed polydimethylsiloxane stretch chambers and used microcontact printing technology to imprint a pattern of 2 mu m fibronectin dots (10 lines x 10 columns in a 38 mu m square) before seeding them with human umbilical vein endothelial cells (HUVEC). We found that most HUVEC attached to the patterned dots after 2 h and were similar in size and morphology, based on phase-contrast microscopy. In this system we were able to statistically analyze tyrosine phosphorylation and actin polymerization in these patterned cells, when subjected to a cyclic, uni-axial strain, using fluorescent microscopy. (C) 2007 Elsevier B.V. All rights reserved.