Reactive oxygen species activate focal adhesion kinase, paxillin and P130CAS tyrosine phosphorylation in endothelial cells
Reactive oxygen species activate focal adhesion kinase, paxillin and P130CAS tyrosine phosphorylation in endothelial cells
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DOI：
10.1016/s0891-5849(98)00134-8
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发表时间：
1998-12-01
影响因子：
7.4
通讯作者：
Pasquier, C
中科院分区：
文献类型：
--
作者：
Gozin, A;Franzini, E;Pasquier, C
活性氧（ROS），特别是羟基自由基（HO），增加中性粒细胞对培养的次黄嘌呤-黄嘌呤氧化酶（HX-XO）处理的人脐静脉内皮细胞（HUVEC）的粘附。酪氨酸激酶抑制剂染料木素（30 μ M）和herbimycin A （0.9 μ M）抑制了这种粘附，表明酪氨酸激酶参与其中。几种HUVEC蛋白在120-130和70 kDa范围内的磷酸化被发现取决于XO浓度和刺激时间。抗氧化剂二甲基硫脲（DMTU, 0.75 ~ 7.5 mM）和己酮茶碱（Ptx, 0.1 mM）以及铁螯合剂去铁胺（DF, 1 mM）和羟苄乙二胺（HBED, 0.5 mM）抑制了磷酸化，表明HO参与了磷酸化过程。分离得到3种酪氨酸磷酸化蛋白：黏着斑激酶（p125(FAK)）、paxillin （PAX）和p130cas，并通过免疫沉淀和western blotting对其进行表征。抗氧化剂和铁螯合剂降低了它们的磷酸化。ROS处理15 min后，HUVEC出现肌动蛋白应激纤维形成。细胞松弛素D （5 μ M）抑制酪氨酸磷酸化和PMN-HUVEC粘附，表明细胞骨架完整性在这两种功能中的重要性。总之，HO。它参与了PMN-HUVEC粘附的增加，也增加了三种主要细胞骨架蛋白上的酪氨酸磷酸化，这些蛋白似乎在这种粘附中起作用。(C) 1998爱思唯尔科学有限公司
Reactive oxygen species (ROS), particularly hydroxyl radical (HO.), increase neutrophil adherence to hypoxanthine-xanthine oxidase (HX-XO)-treated human umbilical vein endothelial cells (HUVEC) in culture. This adherence is inhibited by the tyrosine kinase inhibitors genistein (30 mu M) and herbimycin A (0.9 mu M), suggesting the involvement of tyrosine kinase. Phosphorylation of several HUVEC proteins in the range of 120-130 and 70 kDa was found to depend on the XO concentration and stimulation time. This phosphorylation was inhibited by the antioxidants dimethylthiourea (DMTU, 0.75 to 7.5 mM) and pentoxifylline (Ptx, 0.1 mM), and by the iron chelators desferrioxamine (DF, 1 mM) and hydroxybenzyl ethylene diamine (HBED, 0.5 mM), suggesting the involvement of HO.. Three tyrosine-phosphorylated proteins, focal adhesion kinase (p125(FAK)), paxillin (PAX) and p130cas were isolated and characterized by immunoprecipitation and western blotting. Antioxidants and iron chelators reduced their phosphorylation. HUVEC treated with ROS for 15 min showed actin stress fiber formation. Cytochalasin D (5 mu M) inhibited tyrosine phosphorylation and PMN-HUVEC adherence, showing the importance of cytoskeleton integrity in these two functions. In conclusion, HO., which is involved in increased PMN-HUVEC adhesion, also increases tyrosine phosphorylation on three major cytoskeleton proteins which seem to play a role in this adhesion. (C) 1998 Elsevier Science Inc.