記述言語：英語   出版者・発行元：Bba Advances  

All eukaryotic cell surfaces are coated with various types of glycans, which are essential molecules in biological events. In this review, we summarize recent integrated glycomics studies using various biological samples. We introduce an improved sialic acid linkage-specific alkylamidation (SALSA) method for sialylated glycan analysis and an automated glycosphingolipid-glycan preparation system for large-scale glycomic analysis of human plasma/serum. Finally, we explain the importance of integrated glycomics of glycoconjugates through total glycomic analysis of human serum and mouse brain tissue, and discuss prospects for exploring glycans as effective biomarkers of biological phenomena.

DOI： 10.1016/j.bbadva.2025.100144

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PubMed

記述言語：英語   出版者・発行元：Antiviral Research  

Zoonotic avian influenza viruses have continued to infect people on occasion. During treatment, antiviral resistant viruses have occasionally emerged, highlighting the need for a novel strategy for treating human illness. After pancreatin treatment, edible bird's nest (EBN), swiftlet saliva consumed for health purposes, possesses anti-avian viral activity by inhibiting receptor-binding hemagglutinin (HA) activity. Glycan analysis revealed an abundance of α2,3Neu5Ac decoy receptors in pancreatin-treated EBN. Fucosylated tri-α2,3Neu5Ac tri-antennary N-glycans (N-35) and di-α2,3Neu5Ac core 2 O-glycans (O-15) are predominant, accounting for 53.46% and 44.66% of total N- and O-glycan amounts, respectively. Isobologram analysis revealed that the treated EBN had a strong synergistic effect with either oseltamivir carboxylate or zanamivir, a competitive inhibitor of receptor-destroying neuraminidases (NAs), against the avian H5N1 virus. Taken together, EBN has the potential to be developed as a food-derived avian viral trap to prevent and decrease avian virus infection as well as in combination with a viral releasing-NA inhibitor to increase therapeutic potency, reduce toxicity, delay resistance development, and potentially prevent pandemic onset.

DOI： 10.1016/j.antiviral.2024.106040

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記述言語：英語   出版者・発行元：Frontiers in Immunology  

Background: Cartilage repair is a significant clinical challenge because of the limited intrinsic healing capacity. Current therapeutic strategies, such as cell transplantation therapy, aim to overcome this challenge by replacing damaged tissue with healthy cells. However, the long-term survival and functionality of transplanted cells remain major hurdles. Objective: This study investigated the impact of chondrocyte passaging on glycan profiles and their antigenic properties. We hypothesized that alterations in glycan composition due to passaging may contribute to the enhanced ability to activate macrophages, thereby affecting the outcome of cell transplantation therapy. Methods: Peritoneal macrophages and primary articular chondrocytes were isolated from C57BL/6 mice to establish direct and indirect coculture models. Macrophage activation was assessed by measuring the concentrations of IL-6 and nitric oxide in the culture supernatants or their gene expression. Glycome analysis of various glycoconjugates was performed by glycoblotting methods combined with the SALSA procedure for N-glycans and GSLs and the BEP method for O-glycans. Results: Our results revealed that direct coculture of macrophages with passaged chondrocytes increased the production of proinflammatory cytokines, including IL-6 and NO, as the number of passages increased. With increasing passage number, the expression of GD3 substantially decreased, and the expression of GM3, especially GD1a, significantly increased. Coculturing passaged GM3S knockout chondrocytes with macrophages significantly suppressed IL-6 expression, implying reduced macrophage activation. Conclusion: The observed activation of macrophages due to alterations in the glycan profile of chondrocytes provides a possible explanation for the antigenicity and immune rejection of transplanted cells.

DOI： 10.3389/fimmu.2024.1475473

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記述言語：英語   出版者・発行元：Analytica Chimica Acta  

Background: In sharp contrast with analysis of N-glycan that can be prepared by PNGase F, O-glycan analysis remains challenging due to a lack of versatile and simple procedures, especially those mediating cleavage of O-glycans from proteins. Most N-glycans and O-glycans are modified with sialic acids at the non-reducing end and their glycosidic linkages are labile, making it difficult to measure glycans by mass spectrometric analysis. In addition, sialic acid residues present on glycan chains via α2,3-, α2,6-, and α2,8-linkages as structural isomers. Results: In this study, we firstly established a direct and linkage-specific derivatization method for sialylated O-glycans on proteins via linkage-specific lactone-opening aminolysis. In this procedure, labile sialylated glycans were not only stabilized, but also allowed distinguishing between sialyl linkages. Furthermore, we revealed that general reductive β-elimination was not useful for O-glycan cleavages with undesirable degradations of resulting methyl amides. Using β-elimination in the presence of pyrazolone (PMP), with low pH despite alkali base concentration, SALSA-derivatized O-glycans could be cleaved with minimal degradations. Cleaved and PMP-labeled O-glycans could be efficiently prepared in an open reaction system at high temperature (evaporative BEP reaction) and detected by simple liquid-phase extraction. Moreover, in the evaporative BEP reaction by changing the alkali solution with LiOH, the lithiated O-glycans could be observed and provided a lot of fragment information reflecting the complex structure of the O-glycans. Significance: Direct sialic acid linkage-specific derivatization of O-glycans on glycoproteins is simple protocol containing in-solution aminolysis-SALSA and acetonitrile precipitation for removal of excess reagents. Evaporative β-elimination with pyrazolone makes possible intact O-linked glycan analysis just by liquid-phase extraction. These analytical methods established by the appropriate combination of direct-SALSA and evaporative β-elimination will facilitate O-glycomic studies in various biological samples.

DOI： 10.1016/j.aca.2024.342945

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記述言語：日本語   出版者・発行元：生化学工業株式会社  

DOI： 10.32285/glycoforum.27a9j

CiNii Research

記述言語：英語   出版者・発行元：SEIKAGAKU CORPORATION  

DOI： 10.32285/glycoforum.27a9

CiNii Research

記述言語：英語   出版者・発行元：eLife  

This study aimed to investigate the glycan structural changes that occur before histological degeneration in osteoarthritis (OA) and to determine the mechanism by which these glycan conformational changes affect cartilage degeneration. An OA model was established in rabbits using mannosidase injection, which reduced high-mannose type N-glycans and led to cartilage degeneration. Further analysis of glycome in human OA cartilage identified specific corefucosylated N-glycan expression patterns. Inhibition of N-glycan corefucosylation in mice resulted in unrecoverable cartilage degeneration, while cartilage-specific blocking of corefucosylation led to accelerated development of aging-associated and instability-induced OA models. We conclude that α1,6 fucosyltransferase is required postnatally to prevent preosteoarthritic deterioration of articular cartilage. These findings provide a novel definition of early OA and identify glyco-phenotypes of OA cartilage, which may distinguish individuals at higher risk of progression.

DOI： 10.7554/eLife.92275

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記述言語：英語   出版者・発行元：Cell Death and Disease  

Chronic metabolic stress paradoxically elicits pro-tumorigenic signals that facilitate cancer stem cell (CSC) development. Therefore, elucidating the metabolic sensing and signaling mechanisms governing cancer cell stemness can provide insights into ameliorating cancer relapse and therapeutic resistance. Here, we provide convincing evidence that chronic metabolic stress triggered by hyaluronan production augments CSC-like traits and chemoresistance by partially impairing nucleotide sugar metabolism, dolichol lipid-linked oligosaccharide (LLO) biosynthesis and N-glycan assembly. Notably, preconditioning with either low-dose tunicamycin or 2-deoxy-D-glucose, which partially interferes with LLO biosynthesis, reproduced the promoting effects of hyaluronan production on CSCs. Multi-omics revealed characteristic changes in N-glycan profiles and Notch signaling activation in cancer cells exposed to mild glycometabolic stress. Restoration of N-glycan assembly with glucosamine and mannose supplementation and Notch signaling blockade attenuated CSC-like properties and further enhanced the therapeutic efficacy of cisplatin. Therefore, our findings uncover a novel mechanism by which tolerable glycometabolic stress boosts cancer cell resilience through altered N-glycosylation and Notch signaling activation.

DOI： 10.1038/s41419-024-06432-z

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記述言語：英語   出版者・発行元：Biochimica et Biophysica Acta - General Subjects  

This study determined the effect of brefeldin A (BFA) on the free N-glycomic profile of HepG2 cells to better understand the effect of blocking intracellular vesicle formation and transport of proteins from the endoplasmic reticulum to the Golgi apparatus. A series of exoglycosidase- and endoglycosidase-assisted analyses clarified the complex nature of altered glycomic profiles. A key feature of BFA-mediated alterations in Gn2-type glycans was the expression of unusual hybrid-, monoantennary- and complex-type free N-glycans (FNGs). BFA-mediated alterations in Gn1-type glycans were characterized by the expression of unusual hybrid- and monoantennary-FNGs, without significant expression of complex-type FNGs. A time course analysis revealed that sialylated hybrid- and complex-type Gn2-type FNGs were generated later than asialo-Gn2-type FNGs, and the expression profiles of Gn2-type FNGs and N-glycans were found to be similar, suggesting that the metabolic flux of FNGs is the same as that of protein-bound N-glycans. Subcellular glycomic analysis revealed that almost all FNGs were detected in the cytoplasmic extracts. Our data suggest that hybrid-, monoantennary- and complex-type Gn2-type FNGs were cleaved from glycoproteins in the cytosol by cytosolic PNGase, and subsequently digested by cytosolic endo-β-N-acetylglucosaminidase (ENGase) to generate Gn1-type FNGs. The substrate specificity of ENGase explains the limited expression of complex Gn1 type FNGs.

DOI： 10.1016/j.bbagen.2023.130331

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記述言語：英語   出版者・発行元：Glycoconjugate Journal  

Characterization of O-glycans linked to serine or threonine residues in glycoproteins has mostly been achieved using chemical reaction approaches because there are no known O-glycan-specific endoglycosidases. Most O-glycans are modified with sialic acid residues at the non-reducing termini through various linkages. In this study, we developed a novel approach for sialic acid linkage-specific O-linked glycan analysis through lactone-driven ester-to-amide derivatization combined with non-reductive β-elimination in the presence of hydroxylamine. O-glycans released by non-reductive β-elimination were efficiently purified using glycoblotting via chemoselective ligation between carbohydrates and a hydrazide-functionalized polymer, followed by modification of methyl or ethyl ester groups of sialic acid residues on solid-phase. In-solution lactone-driven ester-to-amide derivatization of ethyl-esterified O-glycans was performed, and the resulting sialylated glycan isomers were discriminated by mass spectrometry. In combination with PNGase F digestion, we carried out simultaneous, quantitative, and sialic acid linkage-specific N- and O-linked glycan analyses of a model glycoprotein and human cartilage tissue. This novel glycomic approach will facilitate detailed characterization of biologically relevant sialylated N- and O-glycans on glycoproteins.

DOI： 10.1007/s10719-023-10109-8

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記述言語：英語   出版者・発行元：Journal of Chromatography A  

Glycosaminoglycans (GAGs), which are one of the major components of proteoglycans, play a pivotal role in physiological processes such as signal transduction, cell adhesion, growth, and differentiation. Characterization of GAGs is challenging due to the tremendous structural diversity of heteropolysaccharides with numerous sulfate or carboxyl groups. In this present study, we examined the analysis of 2-aminobenzamide (2-AB) labeled GAG disaccharides by high-performance liquid chromatography (HPLC) using a reverse-phase (RP)-column with adamantyl groups. Under the analytical conditions, 17 types of 2-AB labeled GAG disaccharides derived from heparan sulfate, chondroitin/dermatan sulfates, and hyaluronan were sequentially separated in a single analysis. The analysis time was fast with high retention time reproducibility. Moreover, the RP-HPLC column with adamantyl groups allowed the quantification of GAGs in various biological samples, such as serum, cultured cells, and culture medium.

DOI： 10.1016/j.chroma.2022.463748

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