Comparative Glycomic Analysis of Exosome Subpopulations Derived from Pancreatic Cancer Cell Lines
Comparative Glycomic Analysis of Exosome Subpopulations Derived from Pancreatic Cancer Cell Lines
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DOI：
10.1021/acs.jproteome.0c00200
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发表时间：
2020-06-05
影响因子：
4.4
通讯作者：
Narimatsu, Hisashi
中科院分区：
文献类型：
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作者：
Matsuda, Atsushi;Kuno, Atsushi;Narimatsu, Hisashi
胞外小泡，如外周小体，通常被一系列由宿主细胞糖合成机制控制的多糖所覆盖，类似于分泌型和膜型糖蛋白。几个外体亚群根据它们的Tetraspanin表达进行分类，在疾病的背景下进行了研究。然而，从来没有人尝试过对他们的血糖进行比较分析。在这里，我们报告了一种比较分析胰腺癌细胞系外体亚群的方法。从8个细胞系获得了细胞外小泡、分泌糖蛋白和膜糖蛋白的糖蛋白糖谱。统计分析显示，囊泡中有大量的PHA-L结合蛋白。胞外囊泡表面用Cy3标记，并用抗Tetraspanins抗体(CD9、CD63和CD81)的磁珠捕获。共沉淀的囊泡被裂解，并接受凝集素微阵列分析。使用19个糖链图谱进行的系统聚类分析证实，除了CD81阳性的外切体外，大多数亚群都可以根据宿主细胞的种类来区分。主成分分析和随后对完整外切体的凝集素亲和捕获表明，CD81阳性的外切体在其表面优先表达的不是PHA-L-而是LEL结合蛋白。这些数据表明，胞外糖组分依赖于宿主细胞类型和亚群。
Extracellular vesicles such as exosomes are generally covered with an array of glycans, which are controlled by the hostcell glyco-synthetic machinery, similar to secreted and membrane glycoproteins. Several exosome subpopulations classified by their tetraspanin expression have been investigated in the context of diseases. However, a comparative analysis of their glycomics has never been attempted. Herein, we report a method for the comparative glycomic analysis of exosome subpopulations among pancreatic cancer cell lines. Glycomic profiles were obtained for extracellular vesicles, secreted glycoproteins, and membrane glycoproteins from eight cell lines. Statistical analyses revealed high populations of PHA-L-binding proteins in the vesicles. The surfaces of extracellular vesicles were labeled with Cy3 and captured by magnetic beads with antibodies against tetraspanins (CD9, CD63, and CD81). The coprecipitated vesicles were lysed and subjected to a lectin microarray analysis. A hierarchical clustering analysis using 19 glycomic profiles confirmed that most subpopulations, except CD81-positive exosomes, could be distinguished according to the host-cell species. Principal component analysis and subsequent lectin-affinity capturing of intact exosomes highlighted that CD81-positive exosomes preferentially expressed not PHA-L- but LEL-binding proteins on their surfaces. These data suggested that exosomal glycomics depended on the host-cell type and subpopulation.