Microparticle-associated endothelial protein C receptor and the induction of cytoprotective and anti-inflammatory effects
Microparticle-associated endothelial protein C receptor and the induction of cytoprotective and anti-inflammatory effects
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DOI：
10.3324/haematol.13547
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发表时间：
2009-03
期刊：
影响因子：
10.1
通讯作者：
M. Pérez-Casal;C. Downey;Beatriz Cutillas-Moreno;M. Zuzel;K. Fukudome;C. Toh
中科院分区：
文献类型：
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作者：
M. Pérez-Casal;C. Downey;Beatriz Cutillas-Moreno;M. Zuzel;K. Fukudome;C. Toh
内皮蛋白C受体在蛋白C途径中调节凝血和炎症中起重要作用。本研究提供了活化蛋白C导致内皮细胞释放微颗粒形式的内皮蛋白C受体的机制信息。参见第313页的相关观点文章。背景内皮细胞蛋白C受体在蛋白C通路中起重要作用，调节凝血和炎症反应。最近，我们描述了内皮蛋白C受体可以通过外源性活化蛋白C以微粒形式从原代内皮细胞中释放。与内皮蛋白C受体结合的活化蛋白C保留了抗凝活性，我们假设这种微粒内皮蛋白C受体活化蛋白C复合物也可以切割内皮蛋白酶活化受体1，以调节炎症并增加细胞存活。因此，我们的主要目的是研究微粒相关内皮蛋白C受体激活蛋白C对内皮功能的影响。设计与方法采用微阵列技术，以内皮细胞经微粒相关蛋白C受体激活蛋白C处理后获得的cDNA为探针，通过真实的时间聚合酶链反应证实结果。通过内皮细胞凋亡和通透性测定，在存在和不存在内皮蛋白C受体、蛋白C和蛋白酶激活受体1的特异性阻断的情况下，进一步分析了基因水平变化的功能相关性。结果40 nmol/L活化蛋白C微粒刺激内皮细胞后，内皮细胞抗凋亡和炎症通路的基因谱发生明显变化。这是伴随着蛋白酶激活受体1依赖性抗凋亡和屏障保护作用，后者还涉及鞘氨醇1-磷酸受体和血管内皮生长因子受体-2/激酶插入结构域受体。蛋白C阻断逆转了这些作用，显示了对微粒上活化蛋白C的特异性。此外，共聚焦显微镜和酶联免疫吸附试验从脓毒症患者重组活化蛋白C治疗过程中获得的血浆显示其存在于体内的证据。结论活化蛋白C对微粒相关内皮细胞蛋白C受体释放的影响可诱导蛋白酶活化受体1依赖的内皮效应。屏障保护的机制涉及1-磷酸鞘氨醇受体和激酶插入结构域受体。
The endothelial protein C receptor plays an important role within the protein C pathway in regulating coagulation and inflammation. This study provides information on the mechanisms by which activated protein C causes the release of the endothelial protein C receptor in microparticulate form from endothelial cells. See related perspective article on page 313. Background The endothelial protein C receptor plays an important role within the protein C pathway in regulating coagulation and inflammation. Recently, we described that endothelial protein C receptor can be released in vitro in microparticulate form from primary endothelial cells by exogenous activated protein C. Activated protein C bound to this endothelial protein C receptor retains anticoagulant activity and we hypothesize that this microparticulate endothelial protein C receptor-activated protein C complex can also cleave endothelial protease-activated receptor 1 to modulate inflammation and increase cell survival. Our main objective was, therefore, to study the effect that microparticle-associated endothelial protein C receptor-activated protein C has on endothelial function. Design and Methods Mini-arrays were used and probed with cDNA obtained from endothelial cells after treatment with microparticle-associated endothelial protein C receptor-activated protein C and results were confirmed by real time polymerase chain reaction. The functional relevance of changes at gene level were further analyzed by endothelial apoptosis and permeability assays, in the presence and absence of specific blockade of endothelial protein C receptor, protein C and protease-activated receptor 1. Results Gene profiling of endothelial cells stimulated by 40 nmol/L activated protein C on microparticles showed significant changes in anti-apoptotic and inflammatory pathways. This was accompanied by protease-activated receptor 1-dependent anti-apoptotic and barrier protective effects, the latter of which also involved sphingosine 1-phosphate receptor and vascular endothelial growth factor receptor-2/ kinase insert domain receptor. Protein C blockade reversed these effects showing specificity for activated protein C on microparticles. Furthermore, confocal microscopy and enzyme-linked immunosorbent assay of plasma obtained from septic patients during recombinant activated protein C treatment showed evidence of their presence in vivo. Conclusions Activated protein C on microparticle-associated endothelial protein C receptor release can induce protease-activated receptor 1-dependent endothelial effects. The mechanisms underlying barrier protection involve sphingosine 1-phosphate receptor and kinase insert domain receptor.