Table_1_Proteomic Profile of Procoagulant Extracellular Vesicles Reflects Complement System Activation and Platelet Hyperreactivity of Patients with Severe COVID-19.xlsx

BackgroundExtracellular vesicles (EVs) are a valuable source of biomarkers and display the pathophysiological status of various diseases. In COVID-19, EVs have been explored in several studies for their ability to reflect molecular changes caused by SARS-CoV-2. Here we provide insights into the roles of EVs in pathological processes associated with the progression and severity of COVID-19.
MethodsIn this study, we used a label-free shotgun proteomic approach to identify and quantify alterations in EV protein abundance in severe COVID-19 patients. We isolated plasma extracellular vesicles from healthy donors and patients with severe COVID-19 by size exclusion chromatography (SEC). Then, flow cytometry was performed to assess the origin of EVs and to investigate the presence of circulating procoagulant EVs in COVID-19 patients. A total protein extraction was performed, and samples were analyzed by nLC-MS/MS in a Q-Exactive HF-X. Finally, computational analysis was applied to signify biological processes related to disease pathogenesis.
ResultsWe report significant changes in the proteome of EVs from patients with severe COVID-19. Flow cytometry experiments indicated an increase in total circulating EVs and with tissue factor (TF) dependent procoagulant activity. Differentially expressed proteins in the disease groups were associated with complement and coagulation cascades, platelet degranulation, and acute inflammatory response.
ConclusionsThe proteomic data reinforce the changes in the proteome of extracellular vesicles from patients infected with SARS-CoV-2 and suggest a role for EVs in severe COVID-19.

【背景】细胞外囊泡（Extracellular vesicles, EVs）是极具价值的生物标志物来源，可反映多种疾病的病理生理状态。在新型冠状病毒肺炎（Corona Virus Disease 2019, COVID-19）领域，多项研究已探索了EVs反映严重急性呼吸综合征冠状病毒2（Severe Acute Respiratory Syndrome Coronavirus 2, SARS-CoV-2）诱导的分子变化的能力。本研究旨在阐明EVs在与COVID-19进展及病情严重程度相关的病理过程中的作用。
【方法】本研究采用无标记鸟枪法蛋白质组学方法，对重型COVID-19患者EVs的蛋白质丰度变化进行鉴定与定量分析。研究通过尺寸排阻色谱法（Size Exclusion Chromatography, SEC）从健康志愿者及重型COVID-19患者的血浆中分离细胞外囊泡。随后通过流式细胞术（Flow Cytometry）评估EVs的来源，并探究COVID-19患者体内循环促凝EVs的存在情况。对样本进行总蛋白提取后，采用Q-Exactive HF-X质谱仪结合纳流液相色谱-串联质谱法（nLC-MS/MS）完成样本分析。最终通过生物信息学分析明确与疾病发病机制相关的生物学过程。
【结果】本研究发现重型COVID-19患者EVs的蛋白质组存在显著变化。流式细胞术实验显示，患者体内循环总EVs水平升高，且伴随组织因子（Tissue Factor, TF）依赖性促凝活性增强。疾病组中差异表达的蛋白质与补体与凝血级联反应、血小板脱颗粒以及急性炎症应答密切相关。
【结论】本蛋白质组学数据验证了SARS-CoV-2感染患者EVs蛋白质组的变化，并提示EVs在重型COVID-19发病过程中发挥特定作用。