Cellular repressor of E1A stimulated genes enhances endothelial monolayer integrity
Cellular repressor of E1A stimulated genes enhances endothelial monolayer integrity
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DOI：
10.1007/s11033-012-2373-6
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发表时间：
2013-04
影响因子：
2.8
通讯作者：
Y. Duan;Shaowei Liu;J. Tao;Yang You;Gui-tang Yang;Chenghui Yan;Yaling Han
中科院分区：
文献类型：
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作者：
Y. Duan;Shaowei Liu;J. Tao;Yang You;Gui-tang Yang;Chenghui Yan;Yaling Han
E1 A刺激基因的细胞阻遏物（CREG）是一种新型的维持血管细胞稳态的调节因子。本研究旨在探讨CREG对肿瘤坏死因子（TNF-α）介导的血管内皮细胞炎性损伤的影响。培养人脐静脉内皮细胞（HUVECs），用TNF-α攻击CREG过表达（VC）、敲低（VS）和假转染（VE）的HUVECs。通过罗丹明-鬼笔环肽染色，我们证明TNF-α促进了细胞间丝状肌动蛋白（F-actin）应激纤维的形成。Transwell法和罗丹明B异硫氰酸盐-葡聚糖染色显示TNF-α可诱导HUVEC单层细胞间通透性增高。这些作用在具有强制CREG过表达的VC细胞中减弱，但在具有CREG沉默的VS细胞中显著增强。TNF-α刺激后，VE细胞IL-6和IL-8分泌明显增加，诱导型一氧化氮合酶（iNOS）表达显著增加，而VC细胞上述作用明显减弱。相反，在VS细胞中，炎症标志物的增加显著增强。免疫荧光染色显示，TNF-α刺激VC细胞后，核因子κB（NF-κB）可缓慢、短暂地移位入细胞核。然而，一个更迅速和持续的核转位，观察VS相比，VE细胞。还观察到其蛋白表达模式的相应变化。这些结果表明，CREG可抑制NF-κB的活化、TNF-α诱导的炎症反应和内皮细胞的通透性增高，因此可能成为治疗病理性血管损伤的潜在靶点。
Cellular repressor of E1A stimulated genes (CREG) is a novel modulator that maintains the homeostasis of vascular cells. The present study aimed to investigate the effects of CREG on tumor necrosis factor (TNF)-α-mediated inflammatory injury of vascular endothelial cells. Human umbilical vein endothelial cells (HUVECs) were cultured and CREG overexpressing (VC), knockdown (VS) and mock-transfected (VE) HUVECs were challenged with TNF-α. We demonstrated that TNF-α prompted robust intercellular filamentous actin (F-actin) stress fiber formation as examined by rhodamin-phalloidin staining. Transwell assay and rhodamine B isothiocyanate–dextran staining indicated that TNF-α induced intercellular hyperpermeability of the HUVEC monolayers. These effects were attenuated in VC cells with forced CREG overexpression but significantly potentiated in VS cells with CREG silencing. After TNF-α stimulation, interleukin (IL)-6 and IL-8 secretions in VE cells were markedly increased and inducible nitric oxidase (iNOS) expression substantially elevated, whereas these effects were pronouncedly damped in VC cells. Conversely, in VS cells, the increase in inflammatory markers was substantially potentiated. Immunofluorescence staining demonstrated that nuclear factor κB (NF-κB) slowly and transiently translocated into the nuclei of VC cells upon TNF-α stimulation. However, a more swift and sustained nuclear translocation was observed in VS as compared to VE cells. Corresponding changes in the pattern of its protein expression was also observed. These data suggested that CREG can inhibit NF-κB activation, TNF-α-induced inflammatory responses and the hyperpermeability of endothelial cells, and may therefore represent a potential therapeutic target for pathological vascular injury.