Skog Maria S, Nystedt Johanna, Korhonen Matti, Anderson Heidi, Lehti Timo A, Pajunen Maria I, Finne Jukka
Biochemistry and Biotechnology, Department of Biosciences, University of Helsinki, P.O. Box 56, FI-00014, Helsinki, Finland.
Cell Therapy Services, Finnish Red Cross Blood Service, Kivihaantie 7, FI-00310, Helsinki, Finland.
Stem Cell Res Ther. 2016 Aug 15;7(1):113. doi: 10.1186/s13287-016-0373-5.
In order to develop novel clinical applications and to gain insights into possible therapeutic mechanisms, detailed molecular characterization of human bone marrow-derived mesenchymal stromal cells (hBM-MSCs) is needed. Neural cell adhesion molecule (NCAM, CD56) is a transmembrane glycoprotein modulating cell-cell and cell-matrix interactions. An additional post-translational modification of NCAM is the α2,8-linked polysialic acid (polySia). Because of its background, NCAM is often considered a marker of neural lineage commitment. Generally, hBM-MSCs are considered to be devoid of NCAM expression, but more rigorous characterization is needed.
We have studied NCAM and polySia expression in five hBM-MSC lines at mRNA and protein levels. Cell surface localization was confirmed by immunofluorescence staining and expression frequency in the donor-specific lines by flow cytometry. For the detection of poorly immunogenic polySia, a fluorochrome-tagged catalytically defective enzyme was employed.
All five known NCAM isoforms are expressed in these cells at mRNA level and the three main isoforms are present at protein level. Both polysialyltransferases, generally responsible for NCAM polysialylation, are expressed at mRNA level, but only very few cells express polySia at the cell surface.
Our results underline the need for a careful control of methods and conditions in the characterization of MSCs. This study shows that, against the generally held view, clinical-grade hBM-MSCs do express NCAM. In contrast, although both polysialyltransferase genes are transcribed in these cells, very few express polySia at the cell surface. NCAM and polySia represent new candidate molecules for influencing MSC interactions.
为开发新的临床应用并深入了解可能的治疗机制，需要对人骨髓间充质基质细胞（hBM-MSCs）进行详细的分子特征分析。神经细胞黏附分子（NCAM，CD56）是一种调节细胞间和细胞与基质相互作用的跨膜糖蛋白。NCAM的另一种翻译后修饰是α2,8-连接的多唾液酸（polySia）。由于其背景，NCAM常被视为神经谱系定向的标志物。一般来说，hBM-MSCs被认为不表达NCAM，但需要更严格的特征分析。
我们在mRNA和蛋白质水平研究了5种hBM-MSC系中NCAM和polySia的表达。通过免疫荧光染色确认细胞表面定位，并通过流式细胞术检测供体特异性系中的表达频率。为检测免疫原性较弱的polySia，使用了荧光染料标记的催化缺陷酶。
所有5种已知的NCAM异构体在这些细胞中均以mRNA水平表达，3种主要异构体以蛋白质水平存在。通常负责NCAM多唾液酸化的两种多唾液酸转移酶均以mRNA水平表达，但只有极少数细胞在细胞表面表达polySia。
我们的结果强调了在间充质干细胞特征分析中仔细控制方法和条件的必要性。这项研究表明，与普遍观点相反，临床级hBM-MSCs确实表达NCAM。相比之下，尽管这两种多唾液酸转移酶基因在这些细胞中均有转录，但只有极少数细胞在细胞表面表达polySia。NCAM和polySia是影响间充质干细胞相互作用的新候选分子。
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