CELL REMODELING BY MECHANICAL STIMULI : ROLE OF FOVAL ADHESION MOLECULES IN POLARITY FORMATION

机械刺激的细胞重塑：卵泡粘附分子在极性形成中的作用

• 批准号: 10480180
• 负责人: SOKABE Masahiro
• 金额: $ 7.17万
• 依托单位: Nagoya University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (B)
• 财政年份: 1998
• 资助国家: 日本
• 起止时间: 1998 至 1999
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-10480180/
• 关键词: Endothelial cells; Stretch stimuli; SA channel; Intracellular calcium; Morphological response; Cell polarity; Integrin; Near field microscopy; 形態変化; ライブイメージング; 機械刺激; 細胞内Ca^<2+>; 接着斑

Endothelial cells exhibit spindle like shape aligning their longitude parallel with vessel running. This peculiar shape and alignment is significant to prevent the cells from being pealed off by blood flow. However, cultured endothelial cells do not show such a shape and alignment. It is known that mechanical stresses onto the cells, like periodic circumferential stretch by pulsative blood flow, is enough to induce such a morphology of endothelial cells. It has been also suggested that cytoskeletons and adhesion molecules contribute to this morphogenesis. The altimate goal of this project is to elucidate the intracellular signaling cascade in the stretch induced morphogenesis in cultured endothelial cells and to understand the molecular mechanism underlying the formation of two dimensional polarity in the cell. Using Ca ィイD12+ィエD1 imaging, patch clamp, biochemical and molecular biological techniques, we could identify the major signaling cascade as follows : < uniaxial periodic stretch … More → activation of SA channels→ intracellular Ca-mobilization→activation of tyrosine kinase, src,→tyrosine phosphorylation of adhesion proteins→ reorganization of stress fibers and forcal adhesion→ morphological change>. In the present study, we focused on the origin of the cell polarity and obtained following results. : 1) As the stretch activated intracellular CaィイD12+ィエD1 increase was spatially uniform, this may not be true cause of the cell polarity. 2) Stretch dependent cAMP increase may not directly relate to the morphological change. 3) On the other hand, stress fibres tend to align perpendiculary to the stretch axis prior to the morphological change. 4) Therefore, there may be another signaling mechanism flowing from the integrin, upon which mechanical forces are directly imposed, to cytoskeletons. 5) To investigate the role of the integrin, we developed a near field microscope system by which we can visualize the dynamics of integrin molecules in real time. 6) Integrin molecules were found to be translocated rapidly than we expected. We speculate that the signaling mechanism via integrins along with the CaィイD12+ィエD1 signaling mechanism may contribute to the formation of the cell polarity. Next aim of our study should be to prove this hypothesis. Less

内皮细胞表现出纺锤体的形状对齐，其经度与容器的运行平行。这种特殊的形状和​​比对非常重要，可防止细胞被血流渗出。但是，培养的内皮细胞没有显示出这种形状和比对。众所周知，在细胞上的机械应力，例如脉动血流周期性的圆周拉伸，足以诱导内皮细胞的形态。还建议细胞骨架和粘附分子有助于这种形态发生。该项目的最终目标是阐明拉伸中细胞内信号传导级联反应在培养的内皮细胞中诱导的形态发生，并了解细胞中二维极性形成的基础机制。使用CA D12+IE D1成像，斑块夹，生化和分子生物学技术，我们可以确定主要的信号传导级联反应如下：<单轴周期性伸展…更多→更多→激活SA通道→细胞内Ca-Mobilization→激活酪氨酸激酶的激活酪氨酸激酶，SRC，SRC的激活，→酪蛋白激酶的蛋白质→腐败的刺激性→酪蛋白的刺激性，腐败的腐败量，腐气化的腐蚀性是腐败的。纤维和锻造粘合剂→形态变化>。在本研究中，我们专注于细胞极性的起源，并获得了以下结果。 ：1）随着拉伸激活的细胞内CAIY D12+IE D1在空间上均匀，这可能不是细胞极性的真正原因。 2）伸展依赖的营地增加可能与形态变化直接相关。 3）另一方面，应力纤维倾向于在形态变化之前垂直于伸展轴。 4）因此，可能有一种从整合素流动的信号机制，直接施加了机械力，直接施加到细胞骨架上。 5）为了研究整联蛋白的作用，我们开发了一个近场显微镜系统，通过该系统可以实时可视化整联蛋白分子的动力学。 6）发现整联蛋白分子的翻译比我们预期的迅速。我们推测通过整合素的信号传导机制以及CAIY D12+IE D1信号传导机制可能有助于细胞极性的形成。我们研究的下一个目标应该是证明这一假设。较少的

项目成果

Naruse K, Sai, X, Yokohama N, Sokabe M.: "Uniaxial cyclic stretch indeces c-src activation and translocation in human endothelial cells via SA channel activation"FEBS Lett. 441. 111-115 (1998)

Naruse K、Sai、X、Yokohama N、Sokabe M.：“单轴循环拉伸通过 SA 通道激活导致人内皮细胞中的 c-src 激活和易位”FEBS Lett。

曽我部正博: "イオンシステム ―神経興奮と筋収縮の分子機構― In「わかりやすい分子生物学」(榊、菊池、村松編)"丸善. 277 (1999)

Masahiro Sogabe：“离子系统 - 神经兴奋和肌肉收缩的分子机制 - 在“易于理解的分子生物学”（Sakaki，Kikuchi，Muramatsu 编辑）”Maruzen 277（1999）。

Kanzaki M, Nagasawa M, Kojima I, Sato C, Naruse K, Sokabe M, Iida H.: "Molecular identification of a eukaryotic, stretch-activated nonselective cation channel."Science. 285. 882-886 (1999)

Kanzaki M、Nagasawa M、Kojima I、Sato C、Naruse K、Sokabe M、Iida H.：“真核拉伸激活非选择性阳离子通道的分子鉴定。”科学。

Kanzaki M, Nagasawa M, Kojima I, Sato C, Naruse K, Sokabe M, Iida H.: "Molecular identification of a eukaryotic, stretch-activated nonselective cation channel"Science. 285. 882-886 (1999)

Kanzaki M、Nagasawa M、Kojima I、Sato C、Naruse K、Sokabe M、Iida H.：“真核拉伸激活非选择性阳离子通道的分子鉴定”科学。

Sokabe M, Usukura J.: "Bioimaging"kyoritu Shuppan, Tokyo. 266 (1998)

Sokabe M，Usukura J.：“生物成像”kyoritu Shuppan，东京。