Pharmacological study on the mechano-sensitive molecules involved in vascular smooth muscle and endothelial cells.

参与血管平滑肌和内皮细胞的力敏感分子的药理学研究。

• 批准号: 17390067
• 负责人: ITO Yushi
• 金额: $ 9.34万
• 依托单位: KYUSHU UNIVERSITY
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (B)
• 财政年份: 2005
• 资助国家: 日本
• 起止时间: 2005 至 2006
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-17390067/
• 关键词: stretch; TRPM4; cerebral artery; smooth muscle; endothelial cells; Rho A; integrin; actin-cytoskeleton; 大脳動脈; 平滑筋細胞; 末梢抵抗血管; Ca^<2+>動員機構; 機械刺激感受性チャネル; TRP蛋白質; 血管平滑筋細胞; 脂質メディエーター; リアノジン受容体; stretch; TRPM4; cerebral artery; smooth muscle; endothelial cells; Rho A; integrin; actin-cytoskeleton; 大脳動脈; 平滑筋細胞; 末梢抵抗血管; Ca^<2+>動員機構; 機械刺激感受性チャネル; TRP蛋白質; 血管平滑筋細胞; 脂質メディエーター; リアノジン受容体

1)The molecular identity and activation mechanisms of Stretch-activated cation channels (SACs) remains poorly understood. We found that TRPM4-like cation channels are activated by membrane stretch in rat cerebral artery myocytes (CAMs). Namely, negative pressure activated single channels in isolated CAMs. These channels were permeable to Na^+ and Cs^+ and inhibited by Gd^<3+> and DIDS. The effect of negative pressure was abolished by membrane excision, but subsequent application of Ca^<2+> (>100nM) to the intracellular side of the membrane restored single channel activity that was indistinguishable from SACs. Overexpression of hTRPM4B in HEK293 cells resulted in the appearance of cation channels which were activated by both negative pressure and Ca^<2+> and which had very similar biophysical and pharmacological properties as compared with SACs in CAMs. These results indicate that TRPM4-like channels in CAMs can be activated by membrane stretch, and this may contribute to the depolariza … More tion and concomitant vasoconstriction of intact cerebral arteries following mechanical stimulation. 2)Both hypotonic stress (HTS) and lysophosphatidic acid (LPA) induce ATP release and a transient reorganization of actin through sequential activation of RhoA/Rho-kinase and focal adhesion kinase F-actin (FAK)/paxillin in human umbilical cord vein endothelial cells (HUVECs). LPA is known to induce the activation of Rho A via its specific receptors, but the mechanisms by which HTS initiates these intracellular signals are not known. We found anti-integrin α5β1 antibody(Ab), but not anti-integrin α2,α6,αv, or β4 antibodies, inhibited HTS-induced RhoA translocation, tyrosine phosphorylation of FAK and paxillin, ATP release, and actin reorganization. However, the LPA-induced ATP release and actin reorganization were not inhibited by any of these anti-integrin antibodies, indicating the integrin α5 β1 plays a pivotal role in the HTS-induced but not in the LPA-induced responses. It is therefore reasonable to assume that this particular subtype of integrin is involved in the initiation of the responses induced by mechanical stimuli in HUVECs. Less

1）拉伸激活的阳离子通道（SAC）的分子身份和激活机制知之甚少。我们发现，大鼠脑动脉心肌细胞（CAM）中的膜拉伸激活TRPM4样阳离子通道。也就是说，负压激活了孤立的凸轮中的单个通道。这些通道可渗透到na^+和cs^+，并被gd^<3+>和dids抑制。膜惊喜消除了负压的影响，但随后将Ca^<2+>（> 100nm）应用于膜的细胞内侧，恢复了与SACS无法区分的单个通道活性。 HEK293细胞中HTRPM4B的过表达导致阳离子通道的出现，与CAM中的SAC相比，与SAC相比，其生物物理和药物特性非常相似。这些结果表明，CAM中的TRPM4样通道可以通过膜拉伸激活，这可能有助于depolariza……在机械模拟后，更多的和随之而来的完整脑动脉的血管收缩。 2）通过RhoA/Rho-激酶和局灶性粘附激酶F-肌动蛋白（FAK）/Paxillin人类脐带菌丝索素静脉静脉静脉静脉静脉细胞（Huveecs）（Huveecs）（Huveecs）（Huveecs），通过顺序激活RhoA/Rho-激酶和局灶性粘附激酶F-肌动蛋白（FAK）诱导肌动蛋白的短暂释放和肌动蛋白的短暂重组。已知LPA通过其特定受体诱导Rho A的激活，但是HTS启动这些细胞内信号的机制尚不清楚。我们发现抗整合蛋白α5β1抗体（AB），但没有抑制HTS诱导的RhoA易位，FAK和PAXILLIN，ATP释放以及Actin Reorlanization抗HTS诱导的RhoA易位，抗凝集素α2，α6，αV或β4抗体。然而，这些抗积蛋白抗体中的任何一种均未抑制LPA诱导的ATP释放和肌动蛋白的重组，表明整联蛋白α5β1在HTS诱导的HTS诱导中起着关键作用，但在LPA诱导的反应中均未发挥作用。因此，合理地假设整合素的这种特定亚型参与了HUVEC中机械刺激引起的反应的倡议。