The role of VASP in regulation of cAMP- and Rac 1-mediated endothelial barrier stabilization
The role of VASP in regulation of cAMP- and Rac 1-mediated endothelial barrier stabilization
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DOI：
10.1152/ajpcell.00273.2007
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发表时间：
2008-01-01
影响因子：
5.5
通讯作者：
Waschke, J.
中科院分区：
文献类型：
--
作者：
Schlegel, N.;Burger, S.;Waschke, J.
肌动蛋白动力学的调节是内皮屏障功能的关键。我们提供的证据表明，肌动蛋白结合蛋白血管扩张剂刺激的磷酸蛋白(VASP)是维持内皮屏障所必需的。Vasp缺陷(Vasp(-/-))微血管心肌内皮细胞(MyEnd)的基线通透性显著增加，而黏附和紧密连接的免疫染色没有明显变化。Forskolin和Rolipram(F/R)升高cAMP和细胞毒性坏死性因子1(CNF-1)激活RAC1对Vasp(-/-)和野生型(Wt)细胞屏障功能的稳定作用相同。在wt细胞中，Vasp在F/R反应中被磷酸化，但不定位于细胞间连接。相反，CNF-1和固有活性RAC-1的表达诱导Vasp移位到wt细胞的细胞边界，在那里它与活性Rac-1共定位。在Vasp(-/-)细胞中，Rac-1活性在对照组降低到wt水平的0.4倍，在CNF-1刺激下增加20倍，与wt细胞的7倍激活相似。此外，与wt细胞相比，致死毒素使Rac 1失活导致通透性增加。这些数据表明Vasp参与了Rac-1活性的调节。综合这些发现，我们的研究表明，VASP至少在一定程度上通过控制RhoFamily GTP酶来稳定内皮屏障功能。
Regulation of actin dynamics is critical for endothelial barrier functions. We provide evidence that the actin-binding protein vasodilator-stimulated phosphoprotein (VASP) is required for endothelial barrier maintenance. Baseline permeability was significantly increased in VASP-deficient (VASP(-/-)) microvascular myocardial endothelial cells (MyEnd) in the absence of discernible alterations of immunostaining for adherens and tight junctions. We tested whether VASP is involved in the endothelium-stabilizing effects of cAMP or Rac 1. Forskolin and rolipram (F/R) to increase cAMP and cytotoxic necrotizing factor 1 (CNF-1) to activate Rac 1 were equally efficient to stabilize barrier functions in VASP(-/-) and wild-type (wt) cells. In wt cells, VASP was phosphorylated in response to F/R but did not localize to intercellular junctions. In contrast, CNF-1 and expression of constitutively active Rac 1 induced translocation of VASP to cell borders in wt cells, where it colocalized with active Rac 1. In VASP(-/-) cells, Rac 1 activity was reduced to 0.4 of wt levels in controls and increased similar to 20-fold in response to CNF-1 compared with 7-fold activation in wt cells. Moreover, inactivation of Rac 1 by lethal toxin led to a greater increase of permeability compared with wt cells. All these data suggest that VASP is involved in the regulation of Rac 1 activity. Taking these findings together, our study indicates that VASP at least in part stabilizes endothelial barrier functions by control of Rhofamily GTPases.