Actin Cytoskeleton Regulates Stretch-Activated Ca2+Influx in Human Pulmonary Microvascular Endothelial Cells
Actin Cytoskeleton Regulates Stretch-Activated Ca2+Influx in Human Pulmonary Microvascular Endothelial Cells
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DOI：
10.1165/rcmb.2009-0073oc
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发表时间：
2010-07-01
影响因子：
6.4
通讯作者：
Sokabe, Masahiro
中科院分区：
文献类型：
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作者：
Ito, Satoru;Suki, Bela;Sokabe, Masahiro
在急性肺损伤（ALI）/急性呼吸窘迫综合征（ARDS）患者的高潮气量机械通气期间，肺部区域受到过度拉伸，导致炎症反应和进一步的肺部损伤。本研究在人肺微血管内皮细胞 (HPMVEC) 中研究了机械拉伸对调节多种内皮特性的细胞内 Ca2+ 浓度 ([Ca2+](i)) 的影响。使用细胞拉伸装置对在纤连蛋白包被的硅室上生长的 HPMVEC 进行单轴拉伸。拉伸和随后卸载后，通过 fura-2 荧光测量，[Ca2+](i) 以应变幅度依赖性方式瞬时增加。拉伸引起的 [Ca2+](i) 升高在无 Ca2+ 溶液中并不明显，并且被 Gd3+（一种拉伸激活通道抑制剂）或钌红（一种瞬时受体电位香草醛抑制剂）阻断。用细胞松弛素 D 破坏肌动蛋白聚合可抑制拉伸诱导的 [Ca2+](i) 升高。相反，毒胡萝卜素或凝血酶诱导的 [Ca2+](i) 增加不受细胞松弛素 D 的影响。1-磷酸鞘氨醇或茉莉酮内酯增加肌动蛋白聚合可增强拉伸诱导的 [Ca2+](i) 升高。还开发了一个简单的细胞骨架网络模型，以支持肌动蛋白应力纤维需要有效的力传递以打开拉伸激活的 Ca2+ 通道的概念。总之，机械拉伸通过拉伸激活通道激活 Ca2+ 流入，该通道受到肌动蛋白细胞骨架的严格调节，与 HPMVEC 中受体操作和储存操作的 Ca2+ 进入等其他 Ca2+ 流入途径不同。这些结果表明，机械通气期间过度机械牵拉导致的异常 Ca2+ 稳态可能在 ALI/ARDS 的进展中发挥作用。
During high tidal volume mechanical ventilation in patients with acute lung injury (ALI)/acute respiratory distress syndrome (ARDS), regions of the lung are exposed to excessive stretch, causing inflammatory responses and further lung damage. In this study, the effects of mechanical stretch on intracellular Ca2+ concentration ([Ca2+](i)), which regulates a variety of endothelial properties, were investigated in human pulmonary microvascular endothelial cells (HPMVECs). HPMVECs grown on fibronectin-coated silicon chambers were exposed to uniaxial stretching, using a cell-stretching apparatus. After stretching and subsequent unloading, [Ca2+](i), as measured by fura-2 fluorescence, was transiently increased in a strain amplitude-dependent manner. The elevation of [Ca2+](i) induced by stretch was not evident in the Ca2+-free solution and was blocked by Gd3+, a stretch-activated channel inhibitor, or ruthenium red, a transient receptor potential vanilloid inhibitor. The disruption of actin polymerization with cytochalasin D inhibited the stretch-induced elevation of [Ca2+](i). In contrast, increases in [Ca2+](i) induced by thapsigargin or thrombin were not affected by cytochalasin D. Increased actin polymerization with sphingosine-1-phosphate or jasplakinolide enhanced the stretch-induced elevation of [Ca2+](i). A simple network model of the cytoskeleton was also developed in support of the notion that actin stress fibers are required for efficient force transmission to open stretch-activated Ca2+ channels. In conclusion, mechanical stretch activates Ca2+ influx via stretch-activated channels which are tightly regulated by the actin cytoskeleton different from other Ca2+ influx pathways such as receptor-operated and store-operated Ca2+ entries in HPMVECs. These results suggest that abnormal Ca2+ homeostasis because of excessive mechanical stretch during mechanical ventilation may play a role in the progression of ALI/ARDS.