表达细胞间粘附分子的内皮细胞外囊泡反映了内皮渗透性和败血症的严重程度

目前，无法获得评估败血症内皮内皮通透性的临床指标。内皮衍生的细胞外囊泡（EDEV）是内皮损伤的生物标志物。血小板内皮细胞粘附分子（PECAM）和血管内皮（VE） - 钙粘蛋白是组成型表达的内皮内皮间粘附分子，可调节细胞间粘附和渗透率。在此，我们研究了表达细胞间粘附分子（PECAM或VE-cadherin Edevs）的EDEV与内皮渗透性和脓毒症的严重程度之间的可能关联。直接与肿瘤坏死因子（TNF-α）刺激了渗透性的脐带静脉内皮细胞（HUVEC）。血管生成素-1，前列环素或血管内皮生长因子（VEGF）改变TNF-α诱导的内皮过度过度性。使用葡聚糖测定法或跨内皮电阻测量内皮渗透性。此外，还进行了前瞻性横断面观测研究，以分析败血症患者的EDEV水平。使用流式细胞仪，在HUVEC培养上清液或患者血浆中检查EDEV在HUVEC培养上清液或患者血浆（nonsepsis，n = 30; sepsis，n = 30; spic shock，n = 42）中检查。 Wilcoxon等级测试用于2组之间的比较。使用Steel-Dwass测试进行了3个或更多组之间的比较。 Spearman的测试用于相关分析。统计显着性设置为p <.05.TNF-α的HUVEC刺激显着提高EDEV释放和内皮渗透性。用血管生成素-1或前列环素预处理抑制了TNF-α诱导的内皮渗透性增加，并抑制了PECAM和VE-Cadherin Edevs的释放。相反，用VEGF进行预处理增加了TNF-α诱导的内皮渗透性以及PECAM和VE-Cadherin Edevs的释放。然而，用渗透率修饰试剂进行预处理不会影响表达炎症性刺激诱导的内皮粘附分子（例如E-选择蛋白），细胞内粘附分子-1或血管细胞粘附分子1的EDEV的释放。败血症组（232 [124，590]/μL）入院的PECAM EDEV数量明显高于脓毒症组（138 [77,267]/μL）的数量（p = .043），平均治疗效果为98/μl（95％置信区间[CI），and 2-2-270 ver，and 2-267/μl）在败血症组中（173 [76,339]/μL）也明显高于败血症组（81 [42,159]/μL），平均治疗效果（ATE）为79/μL（95％CI，19-171/μL）；这些EDEV水平一直升高到第5天。表达细胞间粘附分子（PECAM或VE-Cadherin Edevs）的EDEV可能反映了内皮通透性的增加，并且可能是脓毒症的有价值的诊断和预后标记。

Currently, clinical indicators for evaluating endothelial permeability in sepsis are unavailable. Endothelium-derived extracellular vesicles (EDEVs) are emerging as biomarkers of endothelial injury. Platelet endothelial cell adhesion molecule (PECAM) and vascular endothelial (VE)-cadherin are constitutively expressed endothelial intercellular adhesion molecules that regulate intercellular adhesion and permeability. Herein, we investigated the possible association between EDEVs expressing intercellular adhesion molecules (PECAM+ or VE-cadherin+ EDEVs) and endothelial permeability and sepsis severity.Human umbilical vein endothelial cells (HUVECs) were stimulated with tumor necrosis factor alpha (TNF-α) directly or after pretreatment with permeability-modifying reagents such as angiopoietin-1, prostacyclin, or vascular endothelial growth factor (VEGF) to alter TNF-α-induced endothelial hyperpermeability. Endothelial permeability was measured using the dextran assay or transendothelial electrical resistance. Additionally, a prospective cross-sectional observational study was conducted to analyze circulating EDEV levels in patients with sepsis. EDEVs were examined in HUVEC culture supernatants or patient plasma (nonsepsis, n = 30; sepsis, n = 30; septic shock, n = 42) using flow cytometry. The Wilcoxon rank-sum test was used for comparisons between 2 groups. Comparisons among 3 or more groups were performed using the Steel-Dwass test. Spearman's test was used for correlation analysis. Statistical significance was set at P < .05.TNF-α stimulation of HUVECs significantly increased EDEV release and endothelial permeability. Pretreatment with angiopoietin-1 or prostacyclin suppressed the TNF-α-induced increase in endothelial permeability and inhibited the release of PECAM+ and VE-cadherin+ EDEVs. In contrast, pretreatment with VEGF increased TNF-α-induced endothelial permeability and the release of PECAM+ and VE-cadherin+ EDEVs. However, pretreatment with permeability-modifying reagents did not affect the release of EDEVs expressing inflammatory stimulus-inducible endothelial adhesion molecules such as E-selectin, intracellular adhesion molecule-1, or vascular cell adhesion molecule-1. The number of PECAM+ EDEVs on admission in the septic-shock group (232 [124, 590]/μL) was significantly higher (P = .043) than that in the sepsis group (138 [77,267]/μL), with an average treatment effect of 98/μL (95% confidence interval [CI], 2-270/μL), and the number of VE-cadherin+ EDEVs in the septic-shock group (173 [76,339]/μL) was also significantly higher (P = .004) than that in the sepsis group (81 [42,159]/μL), with an average treatment effect (ATE) of 79/μL (95% CI, 19-171/μL); these EDEV levels remained elevated until day 5.EDEVs expressing intercellular adhesion molecules (PECAM+ or VE-cadherin+ EDEVs) may reflect increased endothelial permeability and could be valuable diagnostic and prognostic markers for sepsis.