Department of Anesthesiology, Pain and Perioperative Medicine, The First Affiliated Hospital of Zhengzhou University, Zhengzhou, China.
Institutes of Biomedical Sciences, Fudan University, Shanghai, China.
Front Immunol. 2021 Sep 28;12:732209. doi: 10.3389/fimmu.2021.732209. eCollection 2021.
Perioperative hypersensitivity reaction (HR) is an IgE-FcϵRI-mediated hypersensitivity reaction with degranulation and activation of mast cells and basophils. Several studies have focused on assessing the degranulation and activation of mast cells and basophils to diagnose and predict the prognosis of drug induced HR. However, it is challenging to isolate sufficiently pure mast cells and basophils from human sources to investigate. Effective biomarkers to assess mast cells and basophils activation could potentially have high diagnostic and prognostic values. In the present study, we investigated EVs pelleted from serum in patients with succinylated gelatin induced HR.
Extracellular vesicles (EVs) were isolated using a total exosome isolation kit and ultracentrifugation, characterized by Western blot, transmission electron microscopy, and nanoparticle tracking analysis. Basophils were isolated from fresh peripheral blood by negative selection using Basophil Isolation Kit II. Human mast cell line was stimulated with IL4. The expression levels of proteins related to the hypersensitive response were evaluated by Western blotting and flow Cytometer. Histamine and tryptase levels were tested using a commercial ELISA kit, and gene expression of inflammatory mediators was evaluated by qRT-PCR. The receiver operating characteristic (ROC) curve was used to evaluate the specificity and sensitivity of biomarker in predicting HR.
The concentration of EVs and protein expression level of CD63, FcϵRI, CD203c and tryptase were significantly (< 0.05) increased in HR samples. The expression level of mast cell/basophil specific CD203c were significantly increased in EVs derived from serum and basophils of HR patients, and the CD203c-EVs production in mast cells is dramatically increased in the presence of IL4, which positively correlated with histamine, tryptase and inflammatory mediators. Moreover, the ROC curve of EVs concentration and CD203c expression indicated that CD203c-EVs had a strong diagnostic ability for HR.
Serum CD203c+-EVs serves as a novel diagnostic and prognostic biomarker for HR.
围手术期过敏反应 (HR) 是一种 IgE-FcϵRI 介导的过敏反应，伴有脱颗粒和肥大细胞和嗜碱性粒细胞的激活。几项研究集中在评估肥大细胞和嗜碱性粒细胞的脱颗粒和激活，以诊断和预测药物诱导的 HR 的预后。然而，从人体来源分离足够纯的肥大细胞和嗜碱性粒细胞进行研究具有挑战性。有效的生物标志物来评估肥大细胞和嗜碱性粒细胞的激活，可能具有较高的诊断和预后价值。在本研究中，我们研究了在琥珀酰明胶诱导的 HR 患者血清中沉淀的 EVs。
使用总外泌体分离试剂盒和超速离心法分离细胞外囊泡 (EVs)，通过 Western blot、透射电子显微镜和纳米颗粒跟踪分析进行表征。使用 Basophil Isolation Kit II 通过阴性选择从新鲜外周血中分离嗜碱性粒细胞。用人肥大细胞系用 IL4 刺激。通过 Western blot 和流式细胞仪评估与过敏反应相关的蛋白质的表达水平。使用商业 ELISA 试剂盒测试组胺和类胰蛋白酶水平，并通过 qRT-PCR 评估炎症介质的基因表达。使用接收者操作特征 (ROC) 曲线评估生物标志物预测 HR 的特异性和敏感性。
HR 样本中 EVs 的浓度和 CD63、FcϵRI、CD203c 和类胰蛋白酶的蛋白表达水平显著升高(<0.05)。HR 患者血清和嗜碱性粒细胞衍生的 EVs 中肥大细胞/嗜碱性粒细胞特异性 CD203c 的表达水平显著增加，IL4 存在时，CD203c-EVs 的产生明显增加，与组胺、类胰蛋白酶和炎症介质呈正相关。此外，EVs 浓度和 CD203c 表达的 ROC 曲线表明，CD203c-EVs 对 HR 具有很强的诊断能力。
血清 CD203c+-EVs 可作为 HR 的新型诊断和预后生物标志物。
