Division of Pathology, Department of Laboratory Medicine, Karolinska Institutet, 141 52 Stockholm, Sweden.
Division of BCM, Department of Laboratory Medicine, Karolinska Institutet, 141 52 Stockholm, Sweden.
Biomolecules. 2021 Oct 29;11(11):1606. doi: 10.3390/biom11111606.
Extracellular vesicles (EVs), comprising exosomes, microvesicles, and apoptotic bodies, are released by all cells into the extracellular matrix and body fluids, where they play important roles in intercellular communication and matrix remodeling in various pathological conditions. Malignant pleural mesothelioma (MPM) is a primary tumor of mesothelial origin, predominantly related to asbestos exposure. The detection of MPM at an early stage and distinguishing it from benign conditions and metastatic adenocarcinomas (AD) is sometimes challenging. Pleural effusion is often the first available biological material and an ideal source for characterizing diagnostic and prognostic factors. Specific proteins have previously been identified as diagnostic markers in effusion, but it is not currently known whether these are associated with vesicles or released in soluble form. Here, we study and characterize tumor heterogeneity and extracellular vesicle diversity in pleural effusion as diagnostic or prognostic markers for MPM. We analyzed extracellular vesicles and soluble proteins from 27 pleural effusions, which were collected and processed at the department of pathology and cytology at Karolinska University Hospital, representing three different patient groups, MPM ( = 9), benign ( = 6), and AD ( = 12). The vesicles were fractionated into apoptotic bodies, microvesicles, and exosomes by differential centrifugation and characterized by nanoparticle tracking analysis and Western blotting. Multiplex bead-based flow cytometry analysis showed that exosomal markers were expressed differently on EVs present in different fractions. Further characterization of exosomes by a multiplex immunoassay (Luminex) showed that all soluble proteins studied were also present in exosomes, though the ratio of protein concentration present in supernatant versus exosomes varied. The proportion of Angiopoietin-1 present in exosomes was generally higher in benign compared to malignant samples. The corresponding ratios of Mesothelin, Galectin-1, Osteopontin, and VEGF were higher in MPM effusions compared to those in the benign group. These findings demonstrate that relevant diagnostic markers can be recovered from exosomes.
细胞外囊泡（EVs）包括外泌体、微囊泡和凋亡小体，由所有细胞释放到细胞外基质和体液中，在各种病理条件下发挥着重要的细胞间通讯和基质重塑作用。恶性胸膜间皮瘤（MPM）是一种主要来源于间皮细胞的原发性肿瘤，主要与石棉暴露有关。在早期发现 MPM 并将其与良性病变和转移性腺癌（AD）区分开来有时具有挑战性。胸腔积液通常是第一个可用的生物材料，也是用于表征诊断和预后因素的理想来源。先前已经确定了胸腔积液中的特定蛋白质作为诊断标志物，但目前尚不清楚这些标志物是否与囊泡有关或以可溶性形式释放。在这里，我们研究并表征了胸腔积液中肿瘤异质性和细胞外囊泡多样性作为 MPM 的诊断或预后标志物。我们分析了来自 27 份胸腔积液的细胞外囊泡和可溶性蛋白，这些胸腔积液是在卡罗林斯卡大学医院的病理学和细胞学系收集和处理的，代表了三个不同的患者群体，MPM（=9）、良性（=6）和 AD（=12）。通过差速离心将囊泡分为凋亡小体、微囊泡和外泌体，并通过纳米颗粒跟踪分析和 Western blot 进行表征。多聚体珠基流式细胞术分析显示，不同分数中存在的 EV 上表达了不同的外泌体标志物。通过多聚体免疫测定（Luminex）进一步对外泌体进行表征表明，所有研究的可溶性蛋白都存在于外泌体中，尽管上清液与外泌体中存在的蛋白浓度比有所不同。与良性样本相比，外泌体中 Angiopoietin-1 的比例通常更高。与良性组相比，MPM 积液中外泌体中 Mesothelin、Galectin-1、Osteopontin 和 VEGF 的相应比例更高。这些发现表明，可以从外泌体中回收相关的诊断标志物。
