内皮细胞衍生的细胞外囊泡表达细胞间粘附分子，反映内皮通透性与败血症严重程度

目前，尚无临床指标用于评估败血症中的内皮通透性。内皮细胞外囊泡（EDEVs）正逐渐成为内皮损伤的潜在生物标志物。血小板内皮细胞粘附分子（PECAM）和血管内皮（VE）钙粘蛋白是内皮细胞中普遍表达的细胞间粘附分子，调控细胞间粘附与通透性。本研究探讨表达细胞间粘附分子的EDEVs（PECAM+或VE-钙粘蛋白+ EDEVs）与内皮通透性及败血症严重程度之间的潜在关联。用肿瘤坏死因子α（TNF-α）刺激人脐静脉内皮细胞（HUVECs），或在刺激前用调节通透性的试剂如血管生成素-1、前列环素或血管内皮生长因子（VEGF）处理，以改变TNF-α诱导的内皮高通透性。通过葡聚糖实验或跨内皮电阻测定内皮通透性。还进行一项前瞻性横断面观察研究，分析败血症患者循环中EDEV水平。利用流式细胞术在HUVEC培养上清液或患者血浆（非败血症组30例，败血症组30例，败血症性休克组42例）中检测EDEVs。使用Wilcoxon秩和检验进行组间比较，Steel-Dwass检验用于三组或多组比较，Spearman相关分析评估相关性，统计显著性设定为P < .05。TNF-α刺激显著增加EDEV释放和内皮通透性。血管生成素-1或前列环素预处理抑制TNF-α引起的内皮通透性升高，并减少PECAM+和VE-钙粘蛋白+ EDEVs的释放。而VEGF预处理则增强TNF-α引起的内皮通透性及EDEVs的释放。预处理并不影响表达炎症诱导性内皮粘附分子的EDEVs（如E-selectin、细胞内粘附分子-1或血管细胞粘附分子-1）。败血症性休克组（232 [124, 590]/μL）在入院时PECAM+ EDEVs的数量显著高于败血症组（138 [77,267]/μL）（P = .043），平均治疗效应为98/μL（95% CI，2-270/μL）；VE-钙粘蛋白+ EDEVs的数量（173 [76,339]/μL）也显著高于败血症组（81 [42,159]/μL）（P = .004），平均治疗效应为79/μL（95% CI，19-171/μL）；这些EDEV水平在第5天仍然升高。表达细胞间粘附分子的EDEVs（PECAM+或VE-钙粘蛋白+ EDEVs）可能反映内皮通透性增加，具有作为败血症诊断和预后标志物的潜力。

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.