利用液相色谱-四极杆-轨道阱串联质谱分析 MIN6 小鼠胰腺 beta 细胞膜 N-糖的结构和定量特征。

IF 3.1 3区 医学 Q2 CHEMISTRY, ANALYTICAL Journal of pharmaceutical and biomedical analysis Pub Date : 2024-10-01 DOI:10.1016/j.jpba.2024.116494
Ji Yeon Jang, Chulmin Moon, Kyuran Kim, Chi Soo Park, Leeseul Jang, Chang Myeong Jeong, Han Seul Lee, Haeun Byeon, Ha Hyung Kim
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引用次数: 0

摘要

MIN6是一种小鼠胰腺β细胞系,被用于糖尿病研究,细胞膜上的N-糖蛋白对调节胰岛素分泌的新陈代谢非常重要。然而,MIN6细胞中N-糖的特性尚未完全阐明。本研究采用液相色谱-电喷雾离子化-高能碰撞解离-串联质谱法分析了N-糖的结构。同时还获得了每种 N-聚糖相对于 N-聚糖总量(100%)的丰度(%)。共获得 50 个 N-聚糖(每个 N-聚糖的相对丰度大于 0.5%),其中 22 个为双链 N-乙酰葡糖胺(GlcNAc;与细胞粘附和生长有关;每个 N-聚糖的相对丰度总和为 27.1%),21 个为核心 N-聚糖(GlcNAc;与细胞粘附和生长有关;每个 N-聚糖的相对丰度总和为 27.1%):27.1%)、21 个核心-岩藻糖基化(与葡萄糖传感和胰岛素分泌调节有关;28.3%)和 16 个ialylated(N-乙酰神经氨酸;与葡萄糖转运体的表达和糖尿病有关;15.5%)N-聚糖。与裂解液(细胞内容物)相比,膜含有较高的双链GlcNAc和核心岩藻糖基化,类似的硅氨酰化,但较少的高甘露糖基化。值得注意的是,所有双ecting GlcNAc N-聚糖都被分为具有(16.6%)或不具有(10.5%)核心-岩藻糖基化和具有(6.9%)或不具有(20.2%)硅烷基化的结构。在人类胰岛中没有发现二叉GlcNAc结构;此外,硅氨酰化水平是人类胰岛的6.9倍。N-聚糖的这些结构特征通过β细胞之间的同源相互作用影响其细胞粘附和分布,从而提高胰岛素分泌效率。这项研究首次确定了MIN6细胞膜中50种N-聚糖的结构和数量,这些N-聚糖可能在调节胰岛β细胞功能方面发挥重要作用。
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Structural and quantitative characterization of membrane N-glycans from MIN6 mouse pancreatic beta cells using liquid chromatography-quadrupole-Orbitrap tandem mass spectrometry
MIN6, a mouse pancreatic beta cell line, is used in diabetes research, and the cellular N-glycoproteins in membrane are important in regulating the metabolism of insulin secretion. However, the identities of N-glycans in MIN6 cells are yet to be fully elucidated. In this study, the structures of N-glycans were analyzed using liquid chromatography-electrospray ionization-higher energy collisional dissociation-tandem mass spectrometry. The abundances (%) of each N-glycan relative to the total N-glycans (100 %) were also obtained. Fifty N-glycans (with relative abundance of each > 0.5 %) were obtained, revealing 22 bisecting N-acetylglucosamine (GlcNAc; associated with cell adhesion and growth; sum of relative abundance of each: 27.1 %), 21 core-fucosylated (associated with glucose sensing and insulin secretion regulation; 28.3 %), and 16 sialylated (N-acetylneuraminic acid; related to the expression of glucose transporters and diabetes;15.5 %) N-glycans. Membranes contained higher bisecting GlcNAc and core-fucosylation, similar sialylation, but less high-mannosylation than the lysate (the cellular contents). Notably, all bisecting GlcNAc N-glycans were categorized into structures with (16.6 %) or without (10.5 %) core-fucosylation and with (6.9 %) or without (20.2 %) sialylation. The bisecting GlcNAc structures were not found in human islets; moreover, sialylation levels were 6.9 times higher than for human islets. These structural characteristics of N-glycans affect their cell adhesion and distribution through homologous interactions between beta cells, leading to increased insulin secretion efficiency. This study is the first to identify the structures and quantities of 50 N-glycans in MIN6 cell membranes that may play an important role in regulating the functions of pancreatic beta cells.
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CiteScore
6.70
自引率
5.90%
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审稿时长
37 days
期刊介绍: This journal is an international medium directed towards the needs of academic, clinical, government and industrial analysis by publishing original research reports and critical reviews on pharmaceutical and biomedical analysis. It covers the interdisciplinary aspects of analysis in the pharmaceutical, biomedical and clinical sciences, including developments in analytical methodology, instrumentation, computation and interpretation. Submissions on novel applications focusing on drug purity and stability studies, pharmacokinetics, therapeutic monitoring, metabolic profiling; drug-related aspects of analytical biochemistry and forensic toxicology; quality assurance in the pharmaceutical industry are also welcome. Studies from areas of well established and poorly selective methods, such as UV-VIS spectrophotometry (including derivative and multi-wavelength measurements), basic electroanalytical (potentiometric, polarographic and voltammetric) methods, fluorimetry, flow-injection analysis, etc. are accepted for publication in exceptional cases only, if a unique and substantial advantage over presently known systems is demonstrated. The same applies to the assay of simple drug formulations by any kind of methods and the determination of drugs in biological samples based merely on spiked samples. Drug purity/stability studies should contain information on the structure elucidation of the impurities/degradants.
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