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使用凝集素芯片和液相色谱-电喷雾串联质谱法鉴定区分大鼠内皮细胞和成纤维细胞的细胞表面标志物。

Identification of cell surface markers to differentiate rat endothelial and fibroblast cells using lectin arrays and LC-ESI-MS/MS.

作者信息

Lee Ji Eun, Mirza Shama P, Didier Daniela N, Scalf Mark, Olivier Michael, Greene Andrew S, Smith Lloyd M

机构信息

Department of Chemistry, University of Wisconsin-Madison, 1101 University Avenue, Madison, Wisconsin 53706, USA.

出版信息

Anal Chem. 2008 Nov 1;80(21):8269-75. doi: 10.1021/ac801390b. Epub 2008 Sep 27.

Abstract

Vascular endothelial cells located at the inner surface of blood vessels are a key component in angiogenesis and are employed as a primary cell type in the study of angiogenesis. These endothelial cells are, however, easily contaminated with fibroblast cells, which are located in proximity to the endothelial cells, during their isolation from tissue. It is thus important to find markers to distinguish the two cell types. In the present work, lectin arrays were prepared using aldehyde-terminated self-assembled monolayers (SAMs) and utilized to explore cell surface carbohydrate expression patterns on endothelial and fibroblast cells. It was found that the lectins Griffonia simplicifolia II (GS II) and Ulex europaeus agglutinin I (UEA I) selectively bind to rat fibroblast cells and not to rat endothelial cells. GS II-binding glycoproteins on fibroblast cells, which are potential cell surface markers to differentiate endothelial and fibroblast cells, were captured on a GS II lectin column and analyzed by LC-ESI-MS/MS. Six candidate cell surface glycoproteins were identified. Differential expression was confirmed by Western blot analysis for two of these proteins, lysosome-associated membrane glycoprotein-1 and transmembrane glycoprotein NMB.

摘要

位于血管内表面的血管内皮细胞是血管生成的关键组成部分,并且在血管生成研究中作为主要细胞类型被采用。然而,这些内皮细胞在从组织中分离过程中很容易被位于其附近的成纤维细胞污染。因此,找到区分这两种细胞类型的标志物很重要。在本研究中,使用醛基终止的自组装单分子层(SAMs)制备凝集素阵列,并用于探索内皮细胞和成纤维细胞的细胞表面碳水化合物表达模式。研究发现,凝集素西非单叶豆凝集素II(GS II)和荆豆凝集素I(UEA I)选择性地结合大鼠成纤维细胞,而不结合大鼠内皮细胞。成纤维细胞上与GS II结合的糖蛋白是区分内皮细胞和成纤维细胞的潜在细胞表面标志物,将其在GS II凝集素柱上捕获并用液相色谱-电喷雾串联质谱(LC-ESI-MS/MS)进行分析。鉴定出六种候选细胞表面糖蛋白。通过蛋白质印迹分析证实了其中两种蛋白,即溶酶体相关膜糖蛋白-1和跨膜糖蛋白NMB的差异表达。

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