Microparticle-Induced Activation of the Vascular Endothelium Requires Caveolin-1/Caveolae

Microparticles (MPs) are small membrane fragments shed from normal as well as activated, apoptotic or injured cells. Emerging evidence implicates MPs as a causal and/or contributing factor in altering normal vascular cell phenotype through initiation of proinflammatory signal transduction events and paracrine delivery of proteins, mRNA and miRNA. However, little is known regarding the mechanism by which MPs influence these events. Caveolae are important membrane microdomains that function as centers of signal transduction and endocytosis. Here, we tested the concept that the MP-induced pro-inflammatory phenotype shift in endothelial cells (ECs) depends on caveolae. Consistent with previous reports, MP challenge activated ECs as evidenced by upregulation of intracellular adhesion molecule-1 (ICAM-1) expression. ICAM-1 upregulation was mediated by activation of NF-κB, Poly [ADP-ribose] polymerase 1 (PARP-1) and the epidermal growth factor receptor (EGFR). This response was absent in ECs lacking caveolin-1/caveolae. To test whether caveolae-mediated endocytosis, a dynamin-2 dependent process, is a feature of the proinflammatory response, EC’s were pretreated with the dynamin-2 inhibitor dynasore. Similar to observations in cells lacking caveolin-1, inhibition of endocytosis significantly attenuated MPs effects including, EGFR phosphorylation, activation of NF-κB and upregulation of ICAM-1 expression. Thus, our results indicate that caveolae play a role in mediating the pro-inflammatory signaling pathways which lead to EC activation in response to MPs.微颗粒（Microparticles, MPs）是从正常、活化、凋亡或损伤细胞中脱落的小型膜片段。越来越多的研究证据表明，微颗粒可通过启动促炎信号转导事件、旁分泌递送蛋白质、mRNA及microRNA（miRNA），参与调控正常血管细胞表型的改变，是该过程的致病诱因或辅助因素。然而，目前关于微颗粒调控此类过程的具体分子机制仍尚未明确。胞膜窖（Caveolae）是一类重要的膜微结构域，兼具信号转导枢纽与内吞作用位点的功能。本研究旨在验证如下假说：微颗粒诱导内皮细胞（endothelial cells, ECs）发生促炎表型转化的过程，依赖于胞膜窖的介导。与既往研究结果一致，微颗粒刺激可激活内皮细胞，该效应可通过细胞间黏附分子-1（intracellular adhesion molecule-1, ICAM-1）的表达上调得到证实。ICAM-1的表达上调，由核因子κB（NF-κB）、多聚ADP核糖聚合酶1（Poly [ADP-ribose] polymerase 1, PARP-1）以及表皮生长因子受体（epidermal growth factor receptor, EGFR）的活化所介导。在缺失窖蛋白-1（caveolin-1）或胞膜窖的内皮细胞中，此类促炎反应并未出现。为验证胞膜窖介导的内吞作用——一种依赖动力蛋白2（dynamin-2）的生物学过程——是否为促炎反应的核心特征，我们使用动力蛋白2抑制剂dynasore对内皮细胞进行预处理。与缺失窖蛋白-1的内皮细胞中的实验结果一致，抑制内吞作用可显著削弱微颗粒介导的各项效应，包括EGFR磷酸化、NF-κB活化以及ICAM-1的表达上调。综上，本研究结果表明，胞膜窖在介导促炎信号通路中发挥关键作用，该通路可促使内皮细胞在微颗粒刺激下发生活化。