Quantification of stretch-induced cytoskeletal remodeling in vascular endothelial cells by image processing

被引:72
|
作者
Yoshigi, M
Clark, EB
Yost, HJ
机构
[1] Univ Utah, Dept Pediat, Salt Lake City, UT 84132 USA
[2] Univ Utah, Primary Childrens Med Ctr, Salt Lake City, UT USA
[3] Univ Utah, Huntsman Canc Inst, Dept Oncol Sci, Salt Lake City, UT USA
来源
CYTOMETRY PART A | 2003年 / 55A卷 / 02期
关键词
cyclic stretching; cytoskeletal remodeling; image processing; mechanotransduction;
D O I
10.1002/cyto.a.10076
中图分类号
Q5 [生物化学];
学科分类号
071010 ; 081704 ;
摘要
Background: Reorientation of the cell axis induced by cyclic stretching is an early response to mechanical forces in vitro. However, quantitative assay for this phenomenon has been difficult due to lack of robust methods. We hypothesized that cell orientation may be redefined by the orientation of actin fibers. We developed image processing methods to quantitate the orientation and density of actin fibers. Methods: A convolution filter using Sobel kernels was adapted to determine the orientation and density of actin fibers in human endothelial cells. Unidirectional stretching (10%, 0.5 Hz) was applied to induce cytoskeletal remodeling by varying the duration of stimulation (control, 0.5, 1, 2, 5, 10, and 20 h). Actin fibers were visualized by fluorescent phalloidin. The image processing method was compared with the manual method for reproducibility. Both confluent and subconfluent cells were tested to assess the efficacy of the methods. Results: Cyclic stretch-induced dense and uninterrupted actin cabling formed across the cell body and, later, the actin fibers became aligned perpendicular to the stretch direction. The variance of actin fiber orientation became smaller after 2 h of stretch (F < 0.01). The actin fiber density index, a derived parameter related to the density of actin fibers, increased as early as 30 min of stretching (P < 0.05) and decreased after 10 h of stretching. Reproducibility of our method was extremely good. Applicability of the method was not compromised by cell density. Conclusions: Our method is reliable for quantifying cytoskeletal remodeling induced by mechanical force. (C) 2003 Wiley-Liss, Inc.
引用
收藏
页码:109 / 118
页数:10
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