Activation of RhoA by thrombin in endothelial hyperpermeability - Role of Rho kinase and protein tyrosine kinases
Activation of RhoA by thrombin in endothelial hyperpermeability - Role of Rho kinase and protein tyrosine kinases
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DOI：
10.1161/01.res.87.4.335
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发表时间：
2000-08-18
影响因子：
20.1
通讯作者：
van Hinsbergh, VWM
中科院分区：
文献类型：
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作者：
Amerongen, GPV;van Delft, S;van Hinsbergh, VWM
内皮细胞（EC）主动调节血液成分的外渗。在血管活性剂和凝血酶的刺激下，内皮细胞改变其细胞骨架结构，相邻细胞之间形成小间隙。这些变化部分取决于 [Ca2+](i) 的增加和 Ca2+/钙调蛋白依赖性肌球蛋白轻链激酶的激活。在本研究中，进一步研究了促进凝血酶增强内皮通透性的机制。我们提供了直接证据表明凝血酶在人脐静脉 EC 中诱导 RhoA 快速且短暂的激活。在相同条件下，相关蛋白 Rac 的活性不受影响，这伴随着肌球蛋白轻链磷酸化的增加、F-肌动蛋白应激纤维的生成以及内皮通透性的长期增加，用特异性抑制剂 Y-27632 抑制 RhoA 靶点 Rho 激酶可显着降低所有这些影响。在 Y-27632 存在的情况下，BAPTA 螯合 [Ca2+](i) 进一步降低了凝血酶增强的通透性。这些数据表明，RhoA/Rho 激酶和 Ca2+ 代表作用于内皮通透性的 2 条途径。此外，蛋白酪氨酸激酶抑制剂金雀异黄素可降低凝血酶诱导的内皮通透性，而不影响凝血酶对 RhoA 的激活。我们的数据支持凝血酶诱导的内皮通透性模型，该通透性受 3 种细胞信号转导途径调节。
Endothelial cells (ECs) actively regulate the extravasation of blood constituents. On stimulation by vasoactive agents and thrombin, ECs change their cytoskeletal architecture and small gaps are formed between neighboring cells. These changes partly depend on a rise in [Ca2+](i) and activation of the Ca2+/calmodulin-dependent myosin light chain kinase, In this study, mechanisms that contribute to the thrombin-enhanced endothelial permeability were further investigated. We provide direct evidence that thrombin induces a rapid and transient activation of RhoA in human umbilical vein ECs. Under the same conditions, the activity of the related protein Rac was not affected, This was accompanied by an increase in myosin light chain phosphorylation, the generation of F-actin stress fibers, and a prolonged increase in endothelial permeability, Inhibition of the RhoA target Rho kinase with the specific inhibitor Y-27632 reduced all of these effects markedly. In the presence of Y-27632, the thrombin-enhanced permeability was additionally reduced by chelation of [Ca2+](i) by BAPTA. These data indicate that RhoA/Rho kinase and Ca2+ represent 2 pathways that act on endothelial permeability. In addition, the protein tyrosine kinase inhibitor genistein reduced thrombin-induced endothelial permeability without affecting activation of RhoA by thrombin. Our data support a model of thrombin-induced endothelial permeability that is regulated by 3 cellular signal transduction pathways.