Activation of Rac-1 and Cdc42 stabilizes the microvascular endothelial barrier
Activation of Rac-1 and Cdc42 stabilizes the microvascular endothelial barrier
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DOI：
10.1007/s00418-005-0080-2
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发表时间：
2006-04-01
影响因子：
2.3
通讯作者：
Adamson, RH
中科院分区：
文献类型：
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作者：
Waschke, J;Burger, S;Adamson, RH
我们之前已经证明Rho家族GTPase Rac-1是维持小鼠微血管心肌内皮细胞（MyEnd）和大鼠肠系膜微血管内皮屏障功能所必需的。在本研究中，我们验证了特异性激活Rac-1可以稳定微血管内皮屏障功能的假设。为此，我们使用大肠杆菌细胞毒性坏死因子（CNF-1）在300 ng/ml， 120 min的条件下，在MyEnd细胞中强烈激活Rac-1和Cdc42，但不激活Rho A。在这些条件下，CNF-1诱导肌动蛋白结合蛋白皮质蛋白和血管扩张剂刺激磷酸化蛋白（VASP）向细胞连接处易位，增加了连接相关的肌动蛋白丝带，降低了单层通透性。我们还通过单次灌注肠系膜微血管测试了CNF-1对内皮屏障特性的影响。与培养的微血管单层相比，CNF-1不会降低基线屏障功能，如水力传导率（Lp）。然而，在预处理120分钟后，CNF-1显著降低了血小板活化因子（PAF, 10 nM）对Lp的响应峰值，从单独使用PAF的46.2 +/- 10x10(-7) cm/(s cmH(2)O)降至12.6 +/- 4 × 10(-7) cm/(s cmH(2)O。这些实验表明，激活Rac-1和Cdc42在体外和体内稳定微血管内皮屏障功能，可能是通过增加连接相关的肌动蛋白细胞骨架。
We have demonstrated previously that the Rho family GTPase Rac-1 is required for maintenance of endothelial barrier functions in mouse microvascular myocardial endothelial (MyEnd) cells in vitro as well as in rat mesenteric microvessels in vivo. In this study, we tested the hypothesis that specific activation of Rac-1 would stabilize microvascular endothelial barrier functions. For this purpose we used Escherichia coli Cytotoxic necrotizing factor (CNF-1) under conditions (300 ng/ml, 120 min) where it strongly activated Rac-1 and Cdc42 but not Rho A in MyEnd cells. Under these conditions, CNF-1 induced translocation of the actin-binding proteins cortactin and vasodilator-stimulated phosphoprotein (VASP) to cell junctions, increased the junction-associated actin filament belt, and reduced monolayer permeability. We also tested the effect of CNF-1 on endothelial barrier properties in vivo using single-perfused mesenteric microvessels. In contrast to cultured microvascular monolayers, CNF-1 did not reduce baseline barrier functions assayed as hydraulic conductivity (Lp). However, following 120 min pretreatment, CNF-1 significantly attenuated the peak Lp increase in response to platelet-activating factor (PAF, 10 nM) to 12.6 +/- 4x10(-7) cm/(s cmH(2)O) compared to 46.2 +/- 10x10(-7) cm/(s cmH(2)O) in experiments using PAF alone. These experiments indicate that activation of Rac-1 and Cdc42 stabilizes microvascular endothelial barrier functions in vitro and in vivo, likely by increasing the junction-associated actin cytoskeleton.