NMMHC IIA inhibition impedes tissue factor expression and venous thrombosis via Akt/GSK3β-NF-κB signalling pathways in the endothelium.
NMMHC IIA inhibition impedes tissue factor expression and venous thrombosis via Akt/GSK3β-NF-κB signalling pathways in the endothelium.
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DOI：
10.1160/th14-10-0880
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发表时间：
2015-07
影响因子：
6.7
通讯作者：
Yu B
中科院分区：
文献类型：
--
作者：
Zhai K;Tang Y;Zhang Y;Li F;Wang Y;Cao Z;Yu J;Kou J;Yu B
非肌球蛋白重链IIA （NMMHC IIA）已被证明参与内皮细胞血栓形成和炎症微粒释放。然而，NMMHC IIA在调节组织因子（TF）表达和深静脉血栓形成中的作用仍有待阐明。本研究用肿瘤坏死因子-α (TNF-α)刺激内皮细胞诱导TF表达。用NMMHC II抑制剂blebbistatin预处理后，TF的mRNA和蛋白表达以及促凝活性均呈剂量依赖性。Blebbistatin增强Akt和GSK3β磷酸化，抑制NF-κB p65核易位和i -κB□降解。这些观察结果与GSK3β抑制剂CHIR99021的效果相似。blebbistatin对TF的下调可被PI3K抑制剂wortmannin拮抗。此外，siRNA敲低NMMHC IIA，而不敲低IIB或IIC，可抑制TF表达，激活Akt/GSK3β，抑制NF-κB信号通路，而NMMHC IIA过表达可增加TF表达。在TNF-α刺激的内皮细胞中，NMMHC IIA和TNF受体2的结合介导信号内化。重要的是，在小鼠深静脉血栓形成模型中，blebbistatin降低内皮细胞NMMHC IIA和TF的表达，通过诱导其磷酸化使GSK3β失活，抑制p65核易位，并抑制血栓形成。我们的研究结果提供了确凿的证据，证明抑制NMMHC II，最有可能是NMMHC IIA，通过Akt/GSK3β-NF-κB信号通路在体外和体内内皮中阻碍TF表达和静脉血栓形成。NMMHC IIA可能是治疗血栓性疾病的潜在新靶点。
Non-muscle myosin heavy chain IIA (NMMHC IIA) has been shown to be involved in thrombus formation and inflammatory microparticle release in endothelial cells. However, the role of NMMHC IIA in regulating the expression of tissue factor (TF) and deep venous thrombosis remains to be elucidated. In the present study, endothelial cells were stimulated with tumour necrosis factor-α (TNF-α) to induce TF expression. Pretreatment with the NMMHC II inhibitor blebbistatin suppressed the mRNA and protein expressions as well as the procoagulant activity of TF in a dose-dependent manner. Blebbistatin enhanced Akt and GSK3β phosphorylation and inhibited NF-κB p65 nuclear translocation and IκB□ degradation. These observations were similar to the effect of CHIR99021, a GSK3β inhibitor. TF downregulation by blebbistatin was antagonised by the PI3K inhibitor, wortmannin. Furthermore, siRNA knockdown of NMMHC IIA, but not IIB or IIC, inhibited TF expression, activated Akt/GSK3β and suppressed NF-κB signalling pathways, whereas the overexpression of NMMHC IIA increased TF expression. The binding of NMMHC IIA and TNF receptor 2 mediated signal internalisation in TNF-α-stimulated endothelial cells. Importantly, blebbistatin decreased endothelium NMMHC IIA and TF expression, deactivated GSK3β by inducing its phosphorylation, suppressed p65 nuclear translocation, and inhibited thrombus formation in a mouse deep venous thrombosis model. Our findings provide solid evidence that inhibition of NMMHC II, most likely NMMHC IIA, impedes TF expression and venous thrombosis via Akt/GSK3β-NF-κB signalling pathways in the endothelium both in vitro and in vivo. NMMHC IIA might be a potential novel target for the treatment of thrombotic disorders.