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

内皮细胞呈纺锤形，其经度与血管运行平行。这种特殊的形状和排列对于防止细胞被血流剥离具有重要意义。然而，培养的内皮细胞没有显示出这样的形状和排列。众所周知，对细胞的机械应力，如周期性的血液循环拉伸，足以诱导内皮细胞的这种形态。也有人认为细胞骨架和粘附分子参与了这种形态的形成。该项目的最终目标是阐明在培养内皮细胞拉伸诱导的形态发生中的细胞内信号级联，并了解细胞中二维极性形成的分子机制。利用Ca- γ - D12+ γ - D1显像、膜片钳、生物化学和分子生物学技术，我们可以确定主要的信号级联：<单轴周期性拉伸…More→SA通道激活→细胞内Ca动员→酪氨酸激酶活化，src，→黏附蛋白酪氨酸磷酸化→应力纤维重组和形式黏附→形态改变>。在本研究中，我们重点研究了细胞极性的起源，得到了以下结果。: 1)随着拉伸激活细胞内CaィイD12 +ィエD1增加空间均匀,这可能不是细胞极性的真正原因。2)拉伸依赖性cAMP增加可能与形态学变化无直接关系。3)另一方面，应力纤维在形态变化之前趋向于垂直于拉伸轴。因此，可能有另一种信号机制从整合素流动，机械力直接施加到细胞骨架上。5)为了研究整合素的作用，我们开发了一个近场显微镜系统，通过它我们可以实时观察整合素分子的动态。6)整合素分子的易位速度比我们预期的要快。我们推测通过整合素的信号机制和Ca γ γ γ - D12+ γ γ - D1信号机制可能参与了细胞极性的形成。我们研究的下一个目标应该是证明这一假设。少

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。

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

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

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。