Morphological Study of Fibroblasts Treated with Cytochalasin D and Colchicine Using a Confocal Laser Scanning Microscopy
Morphological Study of Fibroblasts Treated with Cytochalasin D and Colchicine Using a Confocal Laser Scanning Microscopy
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DOI：
10.2170/physiolsci.rp007708
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发表时间：
2008-12-01
影响因子：
2.3
通讯作者：
Wada, Shigeo
中科院分区：
文献类型：
--
作者：
Ujihara, Yoshihiro;Miyazaki, Hiroshi;Wada, Shigeo
利用共聚焦激光扫描显微镜和基于区域生长方法的图像分析研究了肌动蛋白丝和微管在三维细胞形态中的作用。用细胞松弛素D或秋水仙素分别处理成纤维细胞以破坏肌动蛋白丝或微管，并使用共聚焦激光扫描显微镜观察这些细胞骨架丝的结构和分布。从细胞骨架的3D重建荧光图像中，确定了单个细胞的形态学参数，如体积、粘附面积、高度和体积比。体积比定义为每10%高度的部分体积与总单元体积的比率。肌动蛋白丝和微管断裂后，细胞体积略有下降，但变化不显著。细胞粘附面积显着减少后，破坏的肌动蛋白丝和微管，并显着小于肌动蛋白破坏细胞比微管破坏细胞。细胞高度增加后，肌动蛋白丝中断，而它几乎保持不变后，微管中断。体积比的分析表明，细胞的形状从一个圆锥体改变成一个半球后，破坏肌动蛋白丝和微管破坏后，稍微朝半球状的形状。这些结果表明，肌动蛋白丝的主要组成部分，负责维持全球细胞的形状和微管的全球细胞形态的贡献是远远低于肌动蛋白丝。
The role of actin filaments and microtubules in 3D cell morphology was investigated using confocal laser scanning microscopy and image analysis based on a region-growing method. Fibroblasts were treated with cytochalasin D or colchicine to disrupt the actin filaments or microtubules, respectively, and the structure and distribution of these cytoskeletal filaments were observed using a confocal laser scanning microscope. From the 3D reconstructed fluorescence images of the cytoskeleton, morphological parameters such as volume, adhesion area, height, and volume ratio of individual cells were determined. The volume ratio was defined as the ratio of the partial volume for every 10% of the height to the total cell volume. The cell volume decreased slightly after the disruption of actin filaments and microtubules, but the change was not significant. The cell adhesion area was significantly decreased after the disruption of actin filaments and microtubules, and was significantly smaller in actin filament-disrupted cells than in microtubule-disrupted cells. Cell height increased significantly after actin filament disruption, whereas it remained almost unchanged after microtubule disruption. Analysis of the volume ratio revealed that the cell shape changed from a cone to a hemisphere after disruption of actin filaments and slightly shifted toward a hemisphere-like shape after microtubule disruption. These results suggest that actin filaments are the major component responsible for the maintenance of global cell shape and that the contribution of microtubules to global cell morphology is much less than that of actin filaments.