Hypoxia-reoxygenation-induced endothelial barrier failure: role of RhoA, Rac1 and myosin light chain kinase
Hypoxia-reoxygenation-induced endothelial barrier failure: role of RhoA, Rac1 and myosin light chain kinase
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DOI：
10.1113/jphysiol.2012.237834
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发表时间：
2013-01-01
影响因子：
5.5
通讯作者：
Guenduez, Dursun
中科院分区：
文献类型：
--
作者：
Aslam, Muhammad;Schluter, Klaus-Dieter;Guenduez, Dursun
缺氧-再氧合导致内皮屏障功能丧失和水肿形成，这是器官恢复的主要障碍。内皮屏障的完整性高度依赖于其收缩机制和肌动蛋白动力学，这是由Rho GTPases精确调节的。这些rho - gtpase在缺氧-再氧作用下的活性紊乱会导致肌动蛋白细胞骨架的紊乱，从而可能影响内皮屏障的完整性。本研究的目的是分析这些gtpase在培养的猪主动脉内皮细胞和离体灌注大鼠心脏缺氧-再氧化过程中调节内皮屏障功能的作用。缺氧-再氧化诱导内皮单层白蛋白通透性增加，并伴有内皮收缩机制的激活、肌动蛋白细胞骨架的紊乱和细胞连接处ve -钙粘蛋白的丢失。ML-7对收缩激活的抑制部分保护了缺氧再氧诱导的高通透性。同样，再氧化引起RhoA增加和Rac1活性降低，并伴有应力纤维形成增强和外周肌动蛋白丢失。RhoA或Rock抑制剂抑制RhoA/rho激酶（Rock）信号可导致肌动蛋白细胞骨架完全解聚和紊乱，并加重缺氧-再氧诱导的高通透性。使用cAMP类似物8-CPT-O'-Me-cAMP激活Rac1，特异性激活Epac/Rap1信号，恢复细胞连接处肌动蛋白和ve -钙粘蛋白的外周定位，并消除再氧诱导的高通透性。在离体盐水灌注的大鼠心脏中也出现了类似的结果。这些数据表明，激活Rac1而非抑制RhoA可保护内皮完整性，防止再氧化诱导的屏障功能丧失。
Hypoxia-reoxygenation induces loss of endothelial barrier function and oedema formation, which presents a major impediment for recovery of the organ. The integrity of the endothelial barrier is highly dependent on its contractile machinery and actin dynamics, which are precisely regulated by Rho GTPases. Perturbed activities of these Rho-GTPases under hypoxia-reoxygenation lead to derangement of the actin cytoskeleton and therefore may affect the integrity of the endothelial barrier. The aim of the present study was to analyse the role of these GTPases in regulating endothelial barrier function during hypoxia-reoxygenation in cultured porcine aortic endothelial cells and isolated perfused rat hearts. Hypoxia-reoxygenation induced an increase in albumin permeability of endothelial monolayers accompanied by an activation of the endothelial contractile machinery, derangement of the actin cytoskeleton and loss of VE-cadherin from cellular junctions. Inhibition of contractile activation with ML-7 partially protected against hypoxia-reoxygenation-induced hyperpermeability. Likewise, reoxygenation caused an increase in RhoA and a reduction in Rac1 activity accompanied by enhanced stress fibre formation and loss of peripheral actin. Inhibition of RhoA/rho kinase (Rock) signalling with RhoA or Rock inhibitors led to a complete depolymerisation and derangement of the actin cytoskeleton and worsened hypoxia-reoxygenation-induced hyperpermeability. Activation of Rac1 using a cAMP analogue, 8-CPT-O'-Me-cAMP, which specifically activates Epac/Rap1 signalling, restored peripheral localisation of actin and VE-cadherin at cellular junctions and abrogated reoxygenation-induced hyperpermeability. Similar results were reproduced in isolated saline-perfused rat hearts. These data show that activation of Rac1 but not the inhibition of RhoA preserves endothelial integrity against reoxygenation-induced loss of barrier function.