从乳腺癌患者淋巴液渗出物中分离的细胞外囊泡表面标志物的特征。
Characterization of surface markers on extracellular vesicles isolated from lymphatic exudate from patients with breast cancer.
机构信息
Sahlgrenska Center for Cancer Research and Wallenberg Centre for Molecular and Translational Medicine, Department of Surgery, Institute of Clinical Sciences, Sahlgrenska Academy, University of Gothenburg, Gothenburg, Sweden.
Department of Surgery, Sahlgrenska University Hospital, Gothenburg, Sweden.
出版信息
BMC Cancer. 2022 Jan 10;22(1):50. doi: 10.1186/s12885-021-08870-w.
BACKGROUND
Breast cancer is the most common cancer, and the leading cause of cancer-related deaths, among females world-wide. Recent research suggests that extracellular vesicles (EVs) play a major role in the development of breast cancer metastasis. Axillary lymph node dissection (ALND) is a procedure in patients with known lymph node metastases, and after surgery large amounts of serous fluid are produced from the axilla. The overall aim was to isolate and characterize EVs from axillary serous fluid, and more specifically to determine if potential breast cancer biomarkers could be identified.
METHODS
Lymphatic drain fluid was collected from 7 patients with breast cancer the day after ALND. EVs were isolated using size exclusion chromatography, quantified and detected by nanoparticle tracking analysis, electron microscopy, nano flow cytometry and western blot. The expression of 37 EV surface proteins was evaluated by flow cytometry using the MACSPlex Exosome kit.
RESULTS
Lymphatic drainage exudate retrieved after surgery from all 7 patients contained EVs. The isolated EVs were positive for the typical EV markers CD9, CD63, CD81 and Flotillin-1 while albumin was absent, indicating low contamination from blood proteins. In total, 24 different EV surface proteins were detected. Eleven of those proteins were detected in all patients, including the common EV markers CD9, CD63 and CD81, cancer-related markers CD24, CD29, CD44 and CD146, platelet markers CD41b, CD42a and CD62p as well as HLA-DR/DP/DQ. Furthermore, CD29 and CD146 were enriched in Her2+ patients compared to patients with Her2- tumors.
CONCLUSIONS
Lymphatic drainage exudate retrieved from breast cancer patients after surgery contains EVs that can be isolated using SEC isolation. The EVs have several cancer-related markers including CD24, CD29, CD44 and CD146, proteins of potential interest as biomarkers as well as to increase the understanding of the mechanisms of cancer biology.
背景
乳腺癌是全球女性中最常见的癌症，也是癌症相关死亡的主要原因。最近的研究表明，细胞外囊泡（EVs）在乳腺癌转移的发展中起主要作用。腋窝淋巴结清扫术（ALND）是已知淋巴结转移患者的一种手术程序，手术后腋窝会产生大量浆液性液体。总体目标是从腋窝浆液中分离和表征 EVs，更具体地说，是确定是否可以识别出潜在的乳腺癌生物标志物。
方法
在 ALND 后一天，从 7 名乳腺癌患者中收集淋巴引流液。使用排阻色谱法分离 EVs，通过纳米颗粒跟踪分析、电子显微镜、纳米流式细胞术和 Western blot 进行定量和检测。使用 MACSPlex Exosome 试剂盒通过流式细胞术评估 37 种 EV 表面蛋白的表达。
结果
从所有 7 名患者手术后的淋巴引流渗出物中均提取到 EVs。分离的 EVs 对典型的 EV 标志物 CD9、CD63、CD81 和 Flotillin-1 呈阳性，而白蛋白呈阴性，表明血液蛋白污染较低。总共检测到 24 种不同的 EV 表面蛋白。其中 11 种蛋白在所有患者中均被检测到，包括常见的 EV 标志物 CD9、CD63 和 CD81、与癌症相关的标志物 CD24、CD29、CD44 和 CD146、血小板标志物 CD41b、CD42a 和 CD62p 以及 HLA-DR/DP/DQ。此外，与 Her2-肿瘤患者相比，CD29 和 CD146 在 Her2+患者中更为丰富。
结论
手术后从乳腺癌患者中提取的淋巴引流渗出物含有可通过 SEC 分离的 EVs。这些 EVs 具有多种与癌症相关的标志物，包括 CD24、CD29、CD44 和 CD146，这些蛋白可能作为生物标志物，也可能有助于增加对癌症生物学机制的理解。
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