Evaluation of Residual Human-Induced Pluripotent Stem Cells in Human Chondrocytes by Cell Type-Specific Glycosphingolipid Glycome Analysis Based on the Aminolysis-SALSA Technique
Evaluation of Residual Human-Induced Pluripotent Stem Cells in Human Chondrocytes by Cell Type-Specific Glycosphingolipid Glycome Analysis Based on the Aminolysis-SALSA Technique
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DOI：
10.3390/ijms21010231
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发表时间：
2020-01-01
影响因子：
5.6
通讯作者：
Iwasaki, Norimasa
中科院分区：
文献类型：
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作者：
Miyazaki, Takuji;Hanamatsu, Hisatoshi;Iwasaki, Norimasa
软骨损伤可能最终导致骨关节炎，因为它很难修复。人诱导多能干细胞（iPSC）衍生的软骨细胞可能用于治疗软骨损伤，但iPSC的致瘤性是其在再生医学应用中的主要问题。许多糖缀合物作为干细胞标记物，鞘糖脂（GSLs）包括H型抗原（Fuc α 1- 2gal β 1- 3glcnac）已在iPSCs表面表达。本研究的目的是探讨gsl - glysugar分析是否有助于软骨细胞中残留iPSCs的质量控制。我们对软骨细胞中未分化的ipsc进行了gsl -糖聚糖分析，通过结合糖印迹和氨基水解-唾液酸连接特异性烷基酰胺化（SALSA）方法，能够从5 × 10(4)个细胞中检测到少量ipsc特异性gsl -聚糖。此外，我们使用R-17F抗体估计iPSCs的残留量，该抗体对依赖于GSL的Lacto-N-fucopentaose I （LNFP I）的iPSCs具有细胞毒活性。此外，我们在间充质干细胞（mesenchymal stem cells, MSCs）向iPSCs分化的过程中检测到少量的LNFP I。这是首次证明gsl - glysugar分析可用于检测未分化的iPSCs，从而支持安全的再生医学。
Cartilage damage may eventually lead to osteoarthritis because it is difficult to repair. Human-induced pluripotent stem cell (iPSC)-derived chondrocytes may potentially be used to treat cartilage damage, but the tumorigenicity of iPSCs is a major concern for their application in regenerative medicine. Many glycoconjugates serve as stem cell markers, and glycosphingolipids (GSLs) including H type 1 antigen (Fuc alpha 1-2Gal beta 1-3GlcNAc) have been expressed on the surface of iPSCs. The purpose of the present study was to investigate whether GSL-glycome analysis is useful for quality control of residual iPSCs in chondrocytes. We performed GSL-glycome analysis of undifferentiated iPSCs in chondrocytes by combining glycoblotting and aminolysis-sialic acid linkage-specific alkylamidation (SALSA) method, enabling the detection of small quantities of iPSC-specific GSL-glycans from 5 x 10(4) cells. Furthermore, we estimated the residual amount of iPSCs using R-17F antibody, which possesses cytotoxic activity toward iPSCs that is dependent on the Lacto-N-fucopentaose I (LNFP I) of GSL. Moreover, we could detect a small number of LNFP I during mesenchymal stem cells (MSCs) differentiation from iPSCs. This is the first demonstration that GSL-glycome analysis is useful for detecting undifferentiated iPSCs, and can thereby support safe regenerative medicine.