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。

基本信息

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

项目摘要

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）的分子特性和激活机制仍然知之甚少，我们发现大鼠脑动脉肌细胞（CAM）中的TRPM4样阳离子通道被膜拉伸激活，即负压激活单通道。在分离的CAM中，这些通道可渗透Na ^+ 和Cs ^+ 并被Gd ^ 3+ 和DIDS抑制。膜切除消除了负压的影响，但随后应用了膜。膜的细胞内侧的Ca 2+ (>100nM)恢复了与SAC无法区分的单通道活性。HEK293细胞中hTRPM4B的过度表达导致了被负压和Ca 2+ 激活的阳离子通道的出现。 <2+> 与 CAM 中的 SAC 相比具有非常相似的生物物理和药理学特性。这些结果表明 CAM 中的 TRPM4 样通道可以被膜激活。拉伸，这可能有助于机械刺激后完整脑动脉的去极化和伴随的血管收缩 2) 低渗应激 (HTS) 和溶血磷脂酸 (LPA) 通过连续激活肌动蛋白诱导 ATP 释放和肌动蛋白短暂重组。人脐带静脉中的 RhoA/Rho 激酶和粘着斑激酶 F-肌动蛋白 (FAK)/桩蛋白已知 LPA 可通过其特定受体诱导 Rho A 的激活，但我们尚未发现抗整合素 α5β1 抗体 (Ab)，但 HTS 启动这些细胞内信号的机制尚不清楚。整合素 α2、α6、αv 或 β4 抗体，抑制 HTS 诱导的 RhoA 易位、FAK 和桩蛋白的酪氨酸磷酸化、ATP 释放，以及然而，LPA 诱导的 ATP 释放和肌动蛋白重组不受任何这些抗整合素抗体的抑制，表明整合素 α5 β1 在 HTS 诱导的反应中发挥关键作用，但在 LPA 诱导的反应中不起关键作用。因此，可以合理地假设这种特定的整合素亚型参与了 HUVEC 中机械刺激诱导的反应的启动。