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首页> 外文期刊>Journal of Biotechnology >Analysis of cyclic-stretching responses using cell-adhesion-patterned cells
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Analysis of cyclic-stretching responses using cell-adhesion-patterned cells

机译:使用细胞粘附模式的细胞分析循环拉伸反应

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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 phosphorylation 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 2mum fibronectin dots (10 linesx10 columns in a 38mum square) before seeding them with human umbilical vein endothelial cells (HUVEC). We found that most HUVEC attached to the patterned dots after 2h 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.
机译:人血管内皮细胞形成血流和血管壁之间的界面,并不断受到机械刺激。当内皮细胞沿一个轴循环拉伸时,它们垂直于拉伸轴排列。我们以前报道过,向细胞施加循环的单轴应变会引起粘着斑激酶的酪氨酸磷酸化并刺激丝裂原活化的蛋白激酶。然而,由于这些细胞随机形成粘着斑,因此难以量化和分析这些细胞中酪氨酸磷酸化的空间分布。在这项研究中,我们开发了一个系统来解决此问题,方法是准备单个,均匀,有图案的细胞,该细胞可以循环和单轴拉伸。我们构建了聚二甲基硅氧烷拉伸腔室,并使用微接触印刷技术在将其植入人脐静脉内皮细胞(HUVEC)之前,将2mum纤连蛋白点的图案(38平方毫米中的10行x 10列)压印出来。我们发现,基于相差显微镜,大多数HUVEC在2小时后附着在图案点上,并且大小和形态相似。在这个系统中,当我们使用循环显微镜观察到环状单轴应变时,我们能够统计分析这些图案化细胞中的酪氨酸磷酸化和肌动蛋白聚合。

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