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Yamanoi K, Fujii C, Nakayama A, Matsuura N, Takatori Y, Kato M, Yahagi N, Nakayama J. Decreased expression of TFF2 and decreased αGlcNAc glycosylation are malignant biomarkers of pyloric gland adenoma of the duodenum. Sci Rep 2023; 13:21641. [PMID: 38062108 PMCID: PMC10703765 DOI: 10.1038/s41598-023-49040-1] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/29/2023] [Accepted: 12/03/2023] [Indexed: 12/18/2023] Open
Abstract
Pyloric gland adenoma (PGA) is a duodenal neoplasm expressing MUC6 and is often associated with high-grade dysplasia and adenocarcinoma. MUC6 secreted from the pyloric gland cells carries unique O-glycans exhibiting terminal α1,4-linked N-acetylglucosamine residues (αGlcNAc). The small peptide trefoil factor 2 (TFF2) is also secreted from pyloric gland cells and binds to αGlcNAc. We recently demonstrated that αGlcNAc serves as a tumor suppressor for gastric neoplasm including PGA, but the significance of TFF2 expression remains unknown. We examined 20 lesions representing low- and high-grade PGA in 22 cases by immunohistochemistry for αGlcNAc, TFF2, MUC6, MUC5AC, MUC2 and p53. αGlcNAc, TFF2 and MUC6 were co-expressed on the cell surface and a dot-like pattern in the cytosol in low-grade PGA lesions. High-grade PGA also expressed MUC6, but reduced αGlcNAc and TFF2 expression. The ratios of αGlcNAc or TFF2 to MUC6 score in high-grade PGA were significantly lower than low-grade PGA (P < 0.001). Co-expression of αGlcNAc-glycosylated MUC6 and TFF2 in PGA suggests the existence of αGlcNAc/TFF2 form complex in PGA cells, a finding consistent with our observations in non-neoplastic Brunner's gland cells. The decreased αGlcNAc and TFF2 expression are associated with high grade atypical cells, indicative of the malignant potential of PGA.
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Affiliation(s)
- Kazuhiro Yamanoi
- Department of Pathology, School of Medicine, Keio University, 35 Shinanomachi, Shinjuku, Tokyo, 160-8582, Japan.
- Department of Molecular Pathology, Shinshu University School of Medicine, Matsumoto, Japan.
| | - Chifumi Fujii
- Department of Molecular Pathology, Shinshu University School of Medicine, Matsumoto, Japan
- Department of Biotechnology, Institute for Biomedical Sciences, Interdisciplinary Cluster for Cutting Edge Research, Shinshu University, Matsumoto, Japan
- Center for Medical Education and Clinical Training, Shinshu University School of Medicine, Matsumoto, Japan
| | - Atsushi Nakayama
- Division of Research and Development for Minimally Invasive Treatment, Cancer Center, Keio University School of Medicine, Tokyo, Japan
| | - Noriko Matsuura
- Division of Research and Development for Minimally Invasive Treatment, Cancer Center, Keio University School of Medicine, Tokyo, Japan
| | - Yusaku Takatori
- Division of Research and Development for Minimally Invasive Treatment, Cancer Center, Keio University School of Medicine, Tokyo, Japan
| | - Motohiko Kato
- Division of Research and Development for Minimally Invasive Treatment, Cancer Center, Keio University School of Medicine, Tokyo, Japan
| | - Naohisa Yahagi
- Division of Research and Development for Minimally Invasive Treatment, Cancer Center, Keio University School of Medicine, Tokyo, Japan
| | - Jun Nakayama
- Department of Molecular Pathology, Shinshu University School of Medicine, Matsumoto, Japan
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2
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Yang H, Wang L, Zhang M, Hu B. The Role of Adhesion in Helicobacter pylori Persistent Colonization. Curr Microbiol 2023; 80:185. [PMID: 37071212 DOI: 10.1007/s00284-023-03264-6] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/12/2022] [Accepted: 03/10/2023] [Indexed: 04/19/2023]
Abstract
Helicobacter pylori (H. pylori) has coevolved with its human host for more than 100 000 years. It can safely colonize around the epithelium of gastric glands via their specific microstructures and proteins. Unless patients receive eradication treatment, H. pylori infection is always lifelong. However, few studies have discussed the reasons. This review will focus on the adhesion of H. pylori from the oral cavity to gastric mucosa and summarize the possible binding and translocation characteristics. Adhesion is the first step for persistent colonization after the directional motility, and factors related to adhesion are necessary. Outer membrane proteins, such as the blood group antigen binding adhesin (BabA) and the sialic acid binding adhesin (SabA), play pivotal roles in binding to human mucins and cellular surfaces. And this may offer different perspectives on eradication.
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Affiliation(s)
- Hang Yang
- Department of Gastroenterology, West China Hospital, Sichuan University, No.37, Guo Xue Alley, Wu Hou District, Chengdu City, 610041, Sichuan Province, China
| | - Lixia Wang
- The Second Department of Gastroenterology, The First Affiliated Hospital of Dalian Medical University, Dalian, Liaoning, China
| | - Miao Zhang
- Department of Gastroenterology, West China Hospital, Sichuan University, No.37, Guo Xue Alley, Wu Hou District, Chengdu City, 610041, Sichuan Province, China
| | - Bing Hu
- Department of Gastroenterology, West China Hospital, Sichuan University, No.37, Guo Xue Alley, Wu Hou District, Chengdu City, 610041, Sichuan Province, China.
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3
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Fass D, Thornton DJ. Mucin networks: Dynamic structural assemblies controlling mucus function. Curr Opin Struct Biol 2023; 79:102524. [PMID: 36753925 DOI: 10.1016/j.sbi.2022.102524] [Citation(s) in RCA: 22] [Impact Index Per Article: 11.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/28/2022] [Revised: 12/01/2022] [Accepted: 12/11/2022] [Indexed: 02/08/2023]
Abstract
Contrary to first appearances, mucus structural biology is not an oxymoron. Though mucus hydrogels derive their characteristics largely from intrinsically disordered, heavily glycosylated polypeptide segments, the secreted mucin glycoproteins that constitute mucus undergo an orderly assembly process controlled by folded domains at their termini. Recent structural studies revealed how mucin complexes promote disulphide-mediated polymerization to produce the mucus gel scaffold. Additional protein-protein and protein-glycan interactions likely tune the mesoscale properties, stability, and activities of mucins. Evidence is emerging that even intrinsically disordered glycosylated segments have specific structural roles in the production and properties of mucus. Though soft-matter biophysical approaches to understanding mucus remain highly relevant, high-resolution structural studies of mucins and other mucus components are providing new perspectives on these vital, protective hydrogels.
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Affiliation(s)
- Deborah Fass
- Department of Chemical and Structural Biology, Weizmann Institute of Science, Rehovot 7610001, Israel
| | - David J Thornton
- Wellcome Centre for Cell-Matrix Research and the Lydia Becker Institute of Immunology and Inflammation, Faculty of Biology, Medicine and Health, University of Manchester, Manchester, UK.
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4
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Morris KM, Mishra A, Raut AA, Gaunt ER, Borowska D, Kuo RI, Wang B, Vijayakumar P, Chingtham S, Dutta R, Baillie K, Digard P, Vervelde L, Burt DW, Smith J. The molecular basis of differential host responses to avian influenza viruses in avian species with differing susceptibility. Front Cell Infect Microbiol 2023; 13:1067993. [PMID: 36926515 PMCID: PMC10011077 DOI: 10.3389/fcimb.2023.1067993] [Citation(s) in RCA: 4] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/12/2022] [Accepted: 02/09/2023] [Indexed: 03/04/2023] Open
Abstract
Introduction Highly pathogenic avian influenza (HPAI) viruses, such as H5N1, continue to pose a serious threat to animal agriculture, wildlife and to public health. Controlling and mitigating this disease in domestic birds requires a better understanding of what makes some species highly susceptible (such as turkey and chicken) while others are highly resistant (such as pigeon and goose). Susceptibility to H5N1 varies both with species and strain; for example, species that are tolerant of most H5N1 strains, such as crows and ducks, have shown high mortality to emerging strains in recent years. Therefore, in this study we aimed to examine and compare the response of these six species, to low pathogenic avian influenza (H9N2) and two strains of H5N1 with differing virulence (clade 2.2 and clade 2.3.2.1) to determine how susceptible and tolerant species respond to HPAI challenge. Methods Birds were challenged in infection trials and samples (brain, ileum and lung) were collected at three time points post infection. The transcriptomic response of birds was examined using a comparative approach, revealing several important discoveries. Results We found that susceptible birds had high viral loads and strong neuro-inflammatory response in the brain, which may explain the neurological symptoms and high mortality rates exhibited following H5N1 infection. We discovered differential regulation of genes associated with nerve function in the lung and ileum, with stronger differential regulation in resistant species. This has intriguing implications for the transmission of the virus to the central nervous system (CNS) and may also indicate neuro-immune involvement at the mucosal surfaces. Additionally, we identified delayed timing of the immune response in ducks and crows following infection with the more deadly H5N1 strain, which may account for the higher mortality in these species caused by this strain. Lastly, we identified candidate genes with potential roles in susceptibility/resistance which provide excellent targets for future research. Discussion This study has helped elucidate the responses underlying susceptibility to H5N1 influenza in avian species, which will be critical in developing sustainable strategies for future control of HPAI in domestic poultry.
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Affiliation(s)
- Katrina M. Morris
- The Roslin Institute and R(D)SVS, The University of Edinburgh, Edinburgh, United Kingdom
- *Correspondence: Katrina M. Morris, ;
| | - Anamika Mishra
- National Institute of High Security Animal Diseases, Indian Council of Agricultural Research, Bhopal, India
| | - Ashwin A. Raut
- National Institute of High Security Animal Diseases, Indian Council of Agricultural Research, Bhopal, India
| | - Eleanor R. Gaunt
- The Roslin Institute and R(D)SVS, The University of Edinburgh, Edinburgh, United Kingdom
| | - Dominika Borowska
- The Roslin Institute and R(D)SVS, The University of Edinburgh, Edinburgh, United Kingdom
| | - Richard I. Kuo
- The Roslin Institute and R(D)SVS, The University of Edinburgh, Edinburgh, United Kingdom
| | - Bo Wang
- The Roslin Institute and R(D)SVS, The University of Edinburgh, Edinburgh, United Kingdom
| | - Periyasamy Vijayakumar
- National Institute of High Security Animal Diseases, Indian Council of Agricultural Research, Bhopal, India
| | - Santhalembi Chingtham
- National Institute of High Security Animal Diseases, Indian Council of Agricultural Research, Bhopal, India
| | - Rupam Dutta
- National Institute of High Security Animal Diseases, Indian Council of Agricultural Research, Bhopal, India
| | - Kenneth Baillie
- The Roslin Institute and R(D)SVS, The University of Edinburgh, Edinburgh, United Kingdom
| | - Paul Digard
- The Roslin Institute and R(D)SVS, The University of Edinburgh, Edinburgh, United Kingdom
| | - Lonneke Vervelde
- The Roslin Institute and R(D)SVS, The University of Edinburgh, Edinburgh, United Kingdom
| | - David W. Burt
- The Roslin Institute and R(D)SVS, The University of Edinburgh, Edinburgh, United Kingdom
| | - Jacqueline Smith
- The Roslin Institute and R(D)SVS, The University of Edinburgh, Edinburgh, United Kingdom
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5
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Fujii C, Harumiya S, Sato Y, Kawakubo M, Matoba H, Nakayama J. α1,4-linked N-acetylglucosamine suppresses gastric cancer development by inhibiting MUC1-mediated signaling. Cancer Sci 2022; 113:3852-3863. [PMID: 35959971 PMCID: PMC9633294 DOI: 10.1111/cas.15530] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/02/2022] [Revised: 07/28/2022] [Accepted: 08/06/2022] [Indexed: 11/29/2022] Open
Abstract
Gastric cancer is the second leading cause of cancer deaths worldwide, and more understanding of its molecular basis is urgently needed. Gastric gland mucin secreted from pyloric gland cells, mucous neck cells, and cardiac gland cells of the gastric mucosa harbors unique O‐glycans carrying terminal α1,4‐linked N‐acetylglucosamine (αGlcNAc) residues. We previously reported that αGlcNAc loss correlated positively with poor outcomes for patients with differentiated‐type gastric cancer. However, the molecular mechanisms underlying these outcomes remained poorly understood. Here, we examined the effects of upregulated αGlcNAc expression on malignant phenotypes of the differentiated‐type gastric cancer cell lines, AGS and MKN7. Upregulation of αGlcNAc following ectopic expression of its biosynthetic enzyme attenuated cell proliferation, motility, and invasiveness of AGS and MKN7 cells in vitro. Moreover, AGS cell tumorigenicity was significantly suppressed by αGlcNAc overexpression in a xenograft model. To define the molecular mechanisms underlying these phenotypes, we investigated αGlcNAc binding proteins in AGS cells and identified Mucin‐1 (MUC1) and podocalyxin. Both proteins were colocalized with αGlcNAc on human gastric cancer cells. We also found that αGlcNAc was bound to MUC1 in murine normal gastric mucosa. When we assessed the effects of αGlcNAc binding to MUC1, we found that αGlcNAc blocked galectin‐3 binding to MUC1, phosphorylation of the MUC1 C‐terminus, and recruitment of Src and β‐catenin to that C‐terminus. These results suggest that αGlcNAc regulates cancer cell phenotypes by dampening MUC1 signal transduction.
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Affiliation(s)
- Chifumi Fujii
- Department of Molecular Pathology, Shinshu University School of Medicine, 3-1-1 Asahi, Matsumoto 390-8621, Japan.,Department of Biotechnology, Institute for Biomedical Sciences, Interdisciplinary Cluster for Cutting Edge Research, Shinshu University, 390-8621, Matsumoto, Japan
| | - Satoru Harumiya
- Department of Molecular Pathology, Shinshu University School of Medicine, 3-1-1 Asahi, Matsumoto 390-8621, Japan
| | - Yoshiko Sato
- Department of Molecular Pathology, Shinshu University School of Medicine, 3-1-1 Asahi, Matsumoto 390-8621, Japan
| | - Masatomo Kawakubo
- Department of Molecular Pathology, Shinshu University School of Medicine, 3-1-1 Asahi, Matsumoto 390-8621, Japan
| | - Hisanori Matoba
- Department of Molecular Pathology, Shinshu University School of Medicine, 3-1-1 Asahi, Matsumoto 390-8621, Japan
| | - Jun Nakayama
- Department of Molecular Pathology, Shinshu University School of Medicine, 3-1-1 Asahi, Matsumoto 390-8621, Japan
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6
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MUC6 expression is a preferable prognostic marker for invasive mucinous adenocarcinoma of the lung. Histochem Cell Biol 2022; 157:671-684. [PMID: 35353213 DOI: 10.1007/s00418-022-02093-1] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Accepted: 02/15/2022] [Indexed: 11/04/2022]
Abstract
Gastric gland mucin consists of core protein MUC6 with residues heavily glycosylated by unique O-glycans carrying α1,4-linked N-acetylglucosamine (αGlcNAc). αGlcNAc-glycosylated MUC6 protein is seen in normal gastric and duodenal glands. Decreased αGlcNAc glycosylation on MUC6-positive tumor cells is often observed in premalignant lesions of the stomach, pancreas, and bile duct, and decreased MUC6 expression is seen in invasive cancer of these organs. Lung cancer (LC) is the most common cause of cancer death worldwide. Recently, the adenocarcinoma subtype has become the most common histological subtype of LC, and one of its invasive forms is invasive mucinous adenocarcinoma (IMA). Currently, prognostic markers of LC IMA are unknown. Here, we analyzed MUC5AC, MUC6, and αGlcNAc expression in 54 IMA LC cases. MUC5AC was positively expressed in 50 (93%), MUC6 in 38 (70%), and αGlcNAc in 19 (35%). Each expression level was scored from 0 to 3. The αGlcNAc expression score was significantly decreased relative to MUC6 (P < 0.001). Interestingly, disease-free survival was significantly higher in MUC6-positive than MUC6-negative cases based on the log-rank test (P = 0.021). For in vitro analysis, we ectopically expressed MUC6 in A549 cells, derived from LC and harboring a KRAS mutation. MUC6-expressing A549 cells showed significantly lower proliferation, motility, and invasiveness than control cells. Finally, F-actin staining in MUC6-expressing cells revealed a decrease or loss of filopodia associated with decreased levels of FSCN transcripts, which encodes an actin-bundling protein fascin1 necessary for cell migration. We conclude that MUC6 expression is a preferable prognostic biomarker in IMA LC.
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7
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Zhang Y, Wang L, Ocansey DKW, Wang B, Wang L, Xu Z. Mucin-Type O-Glycans: Barrier, Microbiota, and Immune Anchors in Inflammatory Bowel Disease. J Inflamm Res 2021; 14:5939-5953. [PMID: 34803391 PMCID: PMC8598207 DOI: 10.2147/jir.s327609] [Citation(s) in RCA: 20] [Impact Index Per Article: 5.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/02/2021] [Accepted: 09/19/2021] [Indexed: 12/21/2022] Open
Abstract
Inflammatory bowel disease (IBD), which affects about 7 million people globally, is a chronic inflammatory condition of the gastrointestinal tract caused by gut microbiota alterations, immune dysregulation, and genetic and environmental factors. The association of microbial and immune molecules with mucin-type O-glycans has been increasingly noticed by researchers. Mucin is the main component of mucus, which forms a protective barrier between the microbiota and immune cells in the colon. Mucin-type O-glycans alter the diversity of gastrointestinal microorganisms, which in turn increases the level of O-glycosylation of host intestinal proteins via the utilization of glycans. Additionally, alterations in mucin-type O-glycans not only increase the activity and stability of immune cells but are also involved in the maintenance of intestinal mucosal immune tolerance. Although there is accumulating evidence indicating that mucin-type O-glycans play an important role in IBD, there is limited literature that integrates available data to present a complete picture of exactly how O-glycans affect IBD. This review emphasizes the roles of the mucin-type O-glycans in IBD. This seeks to provide a better understanding and encourages future studies on IBD glycosylation and the design of novel glycan-inspired therapies for IBD.
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Affiliation(s)
- Yaqin Zhang
- Key Laboratory of Medical Science and Laboratory Medicine of Jiangsu Province, School of Medicine, Jiangsu University, Zhenjiang, Jiangsu, 212013, People's Republic of China
| | - Lan Wang
- Danyang Blood Station, Zhenjiang, Jiangsu, 212300, People's Republic of China
| | - Dickson Kofi Wiredu Ocansey
- Key Laboratory of Medical Science and Laboratory Medicine of Jiangsu Province, School of Medicine, Jiangsu University, Zhenjiang, Jiangsu, 212013, People's Republic of China.,Directorate of University Health Services, University of Cape Coast, PMB, Cape Coast, Ghana
| | - Bo Wang
- Key Laboratory of Medical Science and Laboratory Medicine of Jiangsu Province, School of Medicine, Jiangsu University, Zhenjiang, Jiangsu, 212013, People's Republic of China
| | - Li Wang
- Huai'an Maternity and Children Hospital, Huaian, Jiangsu, 223002, People's Republic of China
| | - Zhiwei Xu
- Key Laboratory of Medical Science and Laboratory Medicine of Jiangsu Province, School of Medicine, Jiangsu University, Zhenjiang, Jiangsu, 212013, People's Republic of China
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8
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Yuki A, Fujii C, Yamanoi K, Matoba H, Harumiya S, Kawakubo M, Nakayama J. Glycosylation of MUC6 by α1,4-linked N-acetylglucosamine enhances suppression of pancreatic cancer malignancy. Cancer Sci 2021; 113:576-586. [PMID: 34808019 PMCID: PMC8819301 DOI: 10.1111/cas.15209] [Citation(s) in RCA: 11] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/01/2021] [Revised: 11/04/2021] [Accepted: 11/08/2021] [Indexed: 11/27/2022] Open
Abstract
Biomarkers for early diagnosis of pancreatic cancer are greatly needed, as the high fatality of this cancer is in part due to delayed detection. α1,4‐linked N‐acetylglucosamine (αGlcNAc), a unique O‐glycan specific to gastric gland mucus, is biosynthesized by α1,4‐N‐acetylglucosaminyltransferase (α4GnT) and primarily bound at the terminal glycosylated residue to scaffold protein MUC6. We previously reported that αGlcNAc expression decreases at early stages of neoplastic pancreatic lesions, followed by decreased MUC6 expression, although functional effects of these outcomes were unknown. Here, we ectopically expressed α4GnT, the αGlcNAc biosynthetic enzyme, together with MUC6 in the human pancreatic cancer cell lines MIA PaCa‐2 and PANC‐1, neither of which expresses α4GnT and MUC6. We observed significantly suppressed proliferation in both lines following coexpression of α4GnT and MUC6. Moreover, cellular motility decreased following MUC6 ectopic expression, an effect enhanced by cotransduction with α4GnT. MUC6 expression also attenuated invasiveness of both lines relative to controls, and this effect was also enhanced by additional α4GnT expression. We found αGlcNAc‐bound MUC6 formed a complex with trefoil factor 2. Furthermore, analysis of survival curves of patients with pancreatic ductal adenocarcinoma using a gene expression database showed that samples marked by higher A4GNT or MUC6 mRNA levels were associated with relatively favorable prognosis. These results strongly suggest that αGlcNAc and MUC6 function as tumor suppressors in pancreatic cancer and that decreased expression of both may serve as a biomarker of tumor progression to pancreatic cancer.
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Affiliation(s)
- Atsuko Yuki
- Department of Molecular Pathology, Shinshu University School of Medicine, Matsumoto, Japan
| | - Chifumi Fujii
- Department of Molecular Pathology, Shinshu University School of Medicine, Matsumoto, Japan.,Department of Biotechnology, Interdisciplinary Cluster for Cutting Edge Research, Institute for Biomedical Sciences, Shinshu University, Matsumoto, Japan
| | - Kazuhiro Yamanoi
- Department of Molecular Pathology, Shinshu University School of Medicine, Matsumoto, Japan.,Department of Pathology, Keio University School of Medicine, Tokyo, Japan
| | - Hisanori Matoba
- Department of Molecular Pathology, Shinshu University School of Medicine, Matsumoto, Japan
| | - Satoru Harumiya
- Department of Molecular Pathology, Shinshu University School of Medicine, Matsumoto, Japan
| | - Masatomo Kawakubo
- Department of Molecular Pathology, Shinshu University School of Medicine, Matsumoto, Japan
| | - Jun Nakayama
- Department of Molecular Pathology, Shinshu University School of Medicine, Matsumoto, Japan
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9
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Ohya A, Matoba H, Fujinaga Y, Nakayama J. Decreased Gastric Gland Mucin-specific O-glycans Are Involved in the Progression of Ovarian Primary Mucinous Tumours. Acta Histochem Cytochem 2021; 54:115-122. [PMID: 34511650 PMCID: PMC8424251 DOI: 10.1267/ahc.21-00032] [Citation(s) in RCA: 4] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/22/2021] [Accepted: 05/31/2021] [Indexed: 01/22/2023] Open
Abstract
Ovarian primary mucinous tumours (OPMTs) show an adenoma–borderline–carcinoma sequence with gastrointestinal metaplasia. Gastric gland mucin-specific O-glycans are unique with an α1,4-linked N-acetylglucosamine (αGlcNAc) residue attached to mucin 6 (MUC6). Although αGlcNAc is expected to be expressed in OPMTs, the relationship between αGlcNAc expression and OPMT progression remains unknown. Here, we analysed 104 areas of benign mucinous tumours (benign), 55 areas of borderline mucinous tumours (borderline), and 18 areas of malignant mucinous tumours (malignant) to investigate the expression patterns of αGlcNAc, mucin 2 (MUC2), mucin 5AC (MUC5AC), and MUC6 during the progression of OPMT from benign to malignant. MUC5AC expression was observed in all areas. The frequencies of MUC6- and αGlcNAc-positive areas were decreased with tumour progression. In particular, the decrease in αGlcNAc-positive areas was remarkable. Furthermore, αGlcNAc expression was lower than MUC6 expression at all grades (benign, p < 0.0001; borderline, p = 0.0014; malignant, p = 0.0039). Conversely, there was no difference in the expression frequency or level of MUC2 among the three grades. These results suggest that decreased expression of αGlcNAc relative to MUC6 occurs early in tumour development and marks the initiation of OPMT progression.
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Affiliation(s)
- Ayumi Ohya
- Department of Radiology, Shinshu University School of Medicine
| | - Hisanori Matoba
- Department of Molecular Pathology, Shinshu University School of Medicine
| | | | - Jun Nakayama
- Department of Molecular Pathology, Shinshu University School of Medicine
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10
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Effects of orally administered Euglena gracilis and its reserve polysaccharide, paramylon, on gastric dysplasia in A4gnt knockout mice. Sci Rep 2021; 11:13640. [PMID: 34210998 PMCID: PMC8249615 DOI: 10.1038/s41598-021-92013-5] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/29/2020] [Accepted: 05/24/2021] [Indexed: 12/28/2022] Open
Abstract
Euglena gracilis is widely utilized as food or supplement to promote human and animal health, as it contains rich nutrients. In this study, we administered spray-dried powder of E. gracilis and paramylon, β-glucan stored in E. gracilis cells, to A4gnt knockout (KO) mice. A4gnt KO mice are a mutant mouse model that spontaneously develops gastric cancer through hyperplasia-dysplasia-adenocarcinoma sequence in the antrum of the stomach, and we observed the effects of E. gracilis and paramylon on the early involvements of A4gnt KO mice. Male and female 10-week-old A4gnt KO mice and their age-matched wildtype C57BL/6J mice were orally administered with 50 mg of E. gracilis or paramylon suspended in saline or saline as a control. After 3-week administration, animals were euthanatized and the stomach was examined histopathologically and immunohistochemically. Gene expression patterns of the stomach, which have been reported to be altered with A4gnt KO, and IgA concentration in small intestine were also analyzed with real-time PCR and ELISA, respectively. Administration of Euglena significantly reduced the number of stimulated CD3-positive T-lymphocytes in pyloric mucosa of A4gnt KO mice and tend to reduce polymorphonuclear leukocytes infiltration. Euglena administration further downregulated the expression of Il11 and Cxcl1 of A4gnt KO mice. Euglena administration also affected IgA concentration in small intestinal contents of A4gnt KO mice. Paramylon administration reduced the number of CD3-positive lymphocytes in pyloric mucosa of A4gnt KO mice, and downregulated the expressions of Il11 and Ccl2 of A4gnt KO mice. Although we found no significant effects on gross and microscopic signs of gastric dysplasia and cell proliferation, the present study suggests that the administration of Euglena and paramylon may ameliorate the early involvements of A4gnt mice through the effects on inflammatory reactions in the gastric mucosa. The cancer-preventing effects should be studied with long-term experiments until actual gastric cancer formation.
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11
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Okumura M, Yamanoi K, Uehara T, Nakayama J. Decreased alpha-1,4-linked N-acetylglucosamine glycosylation in biliary tract cancer progression from biliary intraepithelial neoplasia to invasive adenocarcinoma. Cancer Sci 2020; 111:4629-4635. [PMID: 33020993 PMCID: PMC7734011 DOI: 10.1111/cas.14677] [Citation(s) in RCA: 11] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/09/2020] [Revised: 09/21/2020] [Accepted: 09/30/2020] [Indexed: 12/14/2022] Open
Abstract
Biliary tract cancer (BTC) is typically lethal due to the difficulty of early stage diagnosis. Thus, novel biomarkers of BTC precursors are necessary. Biliary intraepithelial neoplasia (BilIN) is a major precursor of BTC and is classified as low or high grade based on cell atypia. In normal gastric mucosa, gastric gland mucin‐specific O‐glycans are unique in having α1,4‐linked N‐acetylglucosamine (αGlcNAc) attached to MUC6. Previously, we reported that αGlcNAc functions as a tumor suppressor of differentiated‐type gastric adenocarcinoma and that decreased αGlcNAc glycosylation on MUC6 in gastric, pancreatic, and uterine cervical neoplasms occurs in cancer as well as in their precursor lesions. However, αGlcNAc and MUC6 expression patterns in biliary tract neoplasms have remained unclear. Here, we analyzed MUC5AC, MUC6, and αGlcNAc expression status in 51 BTC cases and compared the expression of each with progression from low‐grade BilIN to invasive adenocarcinoma (IAC). The frequency of αGlcNAc‐positive and MUC6‐positive lesions decreased with tumor progression. When we compared each marker’s expression level with tumor progression, we found that the MUC6 expression score in IAC was significantly lower than in low‐grade or high‐grade BilIN (P < 0.001 or P < 0.01, respectively). However, the αGlcNAc expression score was low irrespective of histological grade, and also lower than that of MUC6 across all histological grades (P < 0.001 for low‐grade and high‐grade BilIN, and P < 0.01 for IAC). These results suggest that decreased expression of αGlcNAc relative to MUC6 marks the initiation of BTC progression.
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Affiliation(s)
- Motohiro Okumura
- Department of Molecular Pathology, Shinshu University School of Medicine, Matsumoto, Japan.,Department of Surgery, Shinshu University School of Medicine, Matsumoto, Japan
| | - Kazuhiro Yamanoi
- Department of Molecular Pathology, Shinshu University School of Medicine, Matsumoto, Japan.,Department of Pathology, Keio University School of Medicine, Tokyo, Japan
| | - Takeshi Uehara
- Department of Laboratory Medicine, Shinshu University Hospital, Matsumoto, Japan
| | - Jun Nakayama
- Department of Molecular Pathology, Shinshu University School of Medicine, Matsumoto, Japan
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12
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Kudelka MR, Stowell SR, Cummings RD, Neish AS. Intestinal epithelial glycosylation in homeostasis and gut microbiota interactions in IBD. Nat Rev Gastroenterol Hepatol 2020; 17:597-617. [PMID: 32710014 PMCID: PMC8211394 DOI: 10.1038/s41575-020-0331-7] [Citation(s) in RCA: 198] [Impact Index Per Article: 39.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Accepted: 06/05/2020] [Indexed: 02/08/2023]
Abstract
Inflammatory bowel disease (IBD) affects 6.8 million people globally. A variety of factors have been implicated in IBD pathogenesis, including host genetics, immune dysregulation and gut microbiota alterations. Emerging evidence implicates intestinal epithelial glycosylation as an underappreciated process that interfaces with these three factors. IBD is associated with increased expression of truncated O-glycans as well as altered expression of terminal glycan structures. IBD genes, glycosyltransferase mislocalization, altered glycosyltransferase and glycosidase expression and dysbiosis drive changes in the glycome. These glycan changes disrupt the mucus layer, glycan-lectin interactions, host-microorganism interactions and mucosal immunity, and ultimately contribute to IBD pathogenesis. Epithelial glycans are especially critical in regulating the gut microbiota through providing bacterial ligands and nutrients and ultimately determining the spatial organization of the gut microbiota. In this Review, we discuss the regulation of intestinal epithelial glycosylation, altered epithelial glycosylation in IBD and mechanisms for how these alterations contribute to disease pathobiology. We hope that this Review provides a foundation for future studies on IBD glycosylation and the emergence of glycan-inspired therapies for IBD.
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Affiliation(s)
- Matthew R Kudelka
- Medical Scientist Training Program, Emory University School of Medicine, Atlanta, GA, USA
- Department of Internal Medicine, Weill Cornell Medicine, New York, NY, USA
| | - Sean R Stowell
- Department of Pathology and Laboratory Medicine, Emory University School of Medicine, Atlanta, GA, USA
| | - Richard D Cummings
- Department of Surgery, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA, USA
| | - Andrew S Neish
- Department of Pathology and Laboratory Medicine, Emory University School of Medicine, Atlanta, GA, USA.
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13
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Yamada S, Yamanoi K, Sato Y, Nakayama J. Diffuse MIST1 expression and decreased α1,4-linked N-acetylglucosamine (αGlcNAc) glycosylation on MUC6 are distinct hallmarks for gastric neoplasms showing oxyntic gland differentiation. Histopathology 2020; 77:413-422. [PMID: 32502322 DOI: 10.1111/his.14165] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/04/2020] [Revised: 05/24/2020] [Accepted: 05/29/2020] [Indexed: 12/12/2022]
Abstract
AIMS Gastric neoplasms showing oxyntic gland differentiation (GAOGs) constitute a gastric neoplasm subtype that shows low atypia, thus similar to non-neoplastic gastric oxyntic glands. Therefore, their diagnosis in biopsy specimens is difficult. GAOGs were first described in 2007, and introduced in the latest World Health Organization classification book as gastric adenocarcinoma of the fundic gland type (GA-FG) and oxyntic gland adenoma. Previously, we assessed α1,4-linked N-acetylglucosamine (αGlcNAc) residues attached to the MUC6 scaffold in gastric neoplasms, and observed decreased αGlcNAc glycosylation in both differentiated-type gastric cancer and high-grade pyloric gland adenoma (PGA), a gastric cancer precursor. GA-FG and PGA often harbour the same mutations. However, the αGlcNAc status in GAOGs remained unknown. To elucidate αGlcNAc expression in GAOGs, we performed the study. METHODS AND RESULTS We assessed the expression of αGlcNAc; the mucin markers MUC6, MUC5AC, and MUC2; the gastric gland cell markers MIST1, pepsinogen 1 (PG1), H/K-ATPase and chromogranin-A (CGA); and the proliferation marker Ki67 in 13 GAOG lesions. All 13 (100%) were MUC6-positive, whereas 10 (76.2%) were αGlcNAc-negative. Moreover, all 13 (100%) were MIST1- and PG1-positive, three (23.1%) were MUC5AC-positive, four (30.8%) were H/K-ATPase-positive, and one (7.7%) was CGA-positive. CONCLUSIONS GAOGs frequently lost αGlcNAc residues on MUC6, but expressed the gastric gland progenitor marker MIST1 and aberrantly expressed various types of gastric gland cell lineage marker, suggestive of immature differentiation to gastric gland cells. Thus, diffuse MIST1 positivity and decreased αGlcNAc glycosylation on MUC6-positive cells could serve as important biomarkers for the histopathological diagnosis of GAOG.
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Affiliation(s)
- Shigenori Yamada
- Department of Molecular Pathology, Shinshu University School of Medicine, Matsumoto, Japan.,Department of 2nd Internal Medicine, Shinshu University School of Medicine, Matsumoto, Japan.,Division of Gastroenterology, Iiyama Red Cross Hospital, Iiyama, Japan
| | - Kazuhiro Yamanoi
- Department of Molecular Pathology, Shinshu University School of Medicine, Matsumoto, Japan.,Department of Pathology, Keio University School of Medicine, Tokyo, Japan
| | - Yoshiko Sato
- Department of Molecular Pathology, Shinshu University School of Medicine, Matsumoto, Japan
| | - Jun Nakayama
- Department of Molecular Pathology, Shinshu University School of Medicine, Matsumoto, Japan
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14
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Ida K, Yamanoi K, Asaka S, Takeuchi H, Miyamoto T, Shiozawa T, Nakayama J. αGlcNAc and its catalyst α4GnT are diagnostic and prognostic markers in uterine cervical tumor, gastric type. Sci Rep 2019; 9:13043. [PMID: 31506488 PMCID: PMC6737144 DOI: 10.1038/s41598-019-49376-7] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/11/2019] [Accepted: 08/20/2019] [Indexed: 02/01/2023] Open
Abstract
Cervical adenocarcinoma, gastric type (GAS) is not associated with human papilloma virus (HPV) infection. GAS patients prognoses are significantly worse compared with cervical adenocarcinoma associated with HPV infection, as their tumors exhibit resistance to conventional chemotherapy and radiotherapy. GAS is often associated with lobular endocervical glandular hyperplasia (LEGH), which is regarded as a precursor to GAS in the latest WHO classification. Recently, we reported that a decrease in expression of terminal α1,4-linked N-acetylglucosamine (αGlcNAc) relative to that of MUC6 was already apparent in atypical LEGH in the LEGH-GAS sequence. Here, we analyzed expression of α1,4-N-acetylglucosaminyltransferase (α4GnT), the sole enzyme catalyzing αGlcNAc biosynthesis, and that of αGlcNAc and MUC6 in cases representing non-neoplastic endocervical gland (NNEG) (11 cases), LEGH (26 cases) and GAS (12 cases). α4GnT protein was detected in a "dot-like" pattern, indicating localization in the Golgi apparatus in all 26 LEGH cases and 5 of 12 GAS cases. α4GnT- and αGlcNAc-positive cells largely overlapped, suggesting that α4GnT gene expression regulates αGlcNAc biosynthesis. Interestingly, all NNEG cases were negative for α4GnT and αGlcNAc expression, but 7 of 11 NNEG and all LEGH cases were MUC6-positive. In GAS cases, patients whose tumors were α4GnT- and αGlcNAc-positive had more favorable prognosis than others. Multivariate analysis revealed that positive expressions of α4GnT and αGlcNAc were independent prognostic indicators. These results indicate that α4GnT and αGlcNAc could serve as useful markers not only to distinguish LEGH from NNEG but to evaluate prognoses of GAS patients.
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Affiliation(s)
- Koichi Ida
- Department of Obstetrics and Gynecology, Shinshu University School of Medicine, Matsumoto, 390-8621, Japan
| | - Kazuhiro Yamanoi
- Department of Molecular Pathology, Shinshu University School of Medicine, Matsumoto, 390-8621, Japan.
- Institute for Biomedical Sciences, Interdisciplinary Cluster for Cutting Edge Research, Shinshu University, Matsumoto, 390-8621, Japan.
- Department of Pathology, Keio University School of Medicine, Tokyo, 160-8582, Japan.
| | - Shiho Asaka
- Department of Clinical Laboratory, Shinshu University Hospital, Matsumoto, 390-8621, Japan
| | - Hodaka Takeuchi
- Department of Obstetrics and Gynecology, Shinshu University School of Medicine, Matsumoto, 390-8621, Japan
| | - Tsutomu Miyamoto
- Department of Obstetrics and Gynecology, Shinshu University School of Medicine, Matsumoto, 390-8621, Japan
| | - Tanri Shiozawa
- Department of Obstetrics and Gynecology, Shinshu University School of Medicine, Matsumoto, 390-8621, Japan
| | - Jun Nakayama
- Department of Molecular Pathology, Shinshu University School of Medicine, Matsumoto, 390-8621, Japan
- Institute for Biomedical Sciences, Interdisciplinary Cluster for Cutting Edge Research, Shinshu University, Matsumoto, 390-8621, Japan
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15
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Stowell CP, Stowell SR. Biologic roles of the ABH and Lewis histo-blood group antigens Part I: infection and immunity. Vox Sang 2019; 114:426-442. [PMID: 31070258 DOI: 10.1111/vox.12787] [Citation(s) in RCA: 46] [Impact Index Per Article: 7.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/30/2018] [Revised: 04/08/2019] [Accepted: 04/10/2019] [Indexed: 12/22/2022]
Abstract
The ABH and Lewis antigens were among the first of the human red blood cell polymorphisms to be identified and, in the case of the former, play a dominant role in transfusion and transplantation. But these two therapies are largely twentieth century innovations, and the ABH and related carbohydrate antigens are not only expressed on a very wide range of human tissues, but were present in primates long before modern humans evolved. Although we have learned a great deal about the biochemistry and genetics of these structures, the biological roles that they play in human health and disease are incompletely understood. This review and its companion, to appear in a later issue of Vox Sanguinis, will focus on a few of the biologic and pathologic processes which appear to be affected by histo-blood group phenotype. The first of the two reviews will explore the interactions of two bacteria with the ABH and Lewis glycoconjugates of their human host cells, and describe the possible connections between the immune response of the human host to infection and the development of the AB-isoagglutinins. The second review will describe the relationship between ABO phenotype and thromboembolic disease, cardio-vascular disease states, and general metabolism.
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Affiliation(s)
- Christopher P Stowell
- Blood Transfusion Service, Massachusetts General Hospital, Boston, MA, USA.,Department of Pathology, Harvard Medical School, Boston, MA, USA
| | - Sean R Stowell
- Center for Apheresis, Center for Transfusion and Cellular Therapies, Emory Hospital, Emory University School of Medicine, Atlanta, GA, USA.,Department of Pathology, Emory University School of Medicine, Atlanta, GA, USA
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16
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Desamero MJ, Kakuta S, Chambers JK, Uchida K, Hachimura S, Takamoto M, Nakayama J, Nakayama H, Kyuwa S. Orally administered brown seaweed-derived β-glucan effectively restrained development of gastric dysplasia in A4gnt KO mice that spontaneously develop gastric adenocarcinoma. Int Immunopharmacol 2018; 60:211-220. [PMID: 29763881 DOI: 10.1016/j.intimp.2018.05.002] [Citation(s) in RCA: 15] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/07/2018] [Revised: 04/22/2018] [Accepted: 05/06/2018] [Indexed: 02/06/2023]
Abstract
β-Glucan refers to a heterogeneous group of chemically defined storage polysaccharides containing β-(1,3)-d-linked glucose polymers with branches connected by either β-(1,4) or β-(1,6) glycosidic linkage. To date, an extensive amount of scientific evidence supports their multifunctional biological activities, but their potential involvement in the progression of premalignant lesions remains to be clarified. A4gnt KO mice that lack α1,4-N-acetylglucosamine-capped O-glycans in gastric gland mucin are a unique animal model for gastric cancer because the mutant mice spontaneously develop gastric cancer through hyperplasia-dysplasia-adenocarcinoma sequence. In particular, A4gnt KO mice show gastric dysplasia during 10-20 weeks of age. Here we investigated the putative gastro-protective activity of brown seaweed-derived β-glucan (Laminaran) against development of gastric dysplasia, precancerous lesion for gastric cancer in A4gnt KO mice. The mutant mice at 12 weeks of age were randomly assigned into three treatment groups namely, wildtype control + distilled water (normal control), A4gnt KO mice + distilled water (untreated control), and A4gnt KO mice + 100 mg/kg Laminaran. After 3 weeks, the stomach was removed and examined for morphology and gene expression patterns. In contrast to the untreated control group, administration of Laminaran substantially attenuated gastric dysplasia development and counterbalanced the increased induction in cell proliferation and angiogenesis. Furthermore, Laminaran treatment effectively overcame the A4gnt KO-induced alteration in the gene expression profile of selected cytokines as revealed by real-time PCR analysis. Collectively, our present findings indicate that β-glucan can potentially restrain the development of gastric dysplasia to mediate their tissue-preserving activity.
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Affiliation(s)
- Mark Joseph Desamero
- Laboratory of Biomedical Science, Graduate School of Agricultural and Life Sciences, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-8657, Japan; Department of Basic Veterinary Sciences, College of Veterinary Medicine, University of the Philippines Los Baños, Laguna 4031, Philippines
| | - Shigeru Kakuta
- Laboratory of Biomedical Science, Graduate School of Agricultural and Life Sciences, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-8657, Japan.
| | - James Kenn Chambers
- Laboratory of Veterinary Pathology, Graduate School of Agricultural and Life Sciences, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-8657, Japan
| | - Kazuyuki Uchida
- Laboratory of Veterinary Pathology, Graduate School of Agricultural and Life Sciences, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-8657, Japan
| | - Satoshi Hachimura
- Research Center for Food Safety, Graduate School of Agricultural and Life Sciences, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-8657, Japan
| | - Masaya Takamoto
- Department of Infection and Host Defense, Shinshu University School of Medicine, 3-1-1 Asahi, Matsumoto, Nagano 390-8621, Japan
| | - Jun Nakayama
- Department of Molecular Pathology, Shinshu University School of Medicine, 3-1-1 Asahi, Matsumoto, Nagano 390-8621, Japan
| | - Hiroyuki Nakayama
- Laboratory of Veterinary Pathology, Graduate School of Agricultural and Life Sciences, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-8657, Japan
| | - Shigeru Kyuwa
- Laboratory of Biomedical Science, Graduate School of Agricultural and Life Sciences, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-8657, Japan
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17
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Yamanoi K, Ishii K, Tsukamoto M, Asaka S, Nakayama J. Gastric gland mucin-specific O-glycan expression decreases as tumor cells progress from lobular endocervical gland hyperplasia to cervical mucinous carcinoma, gastric type. Virchows Arch 2018; 473:305-311. [PMID: 29845361 DOI: 10.1007/s00428-018-2381-6] [Citation(s) in RCA: 18] [Impact Index Per Article: 2.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/07/2018] [Revised: 05/09/2018] [Accepted: 05/16/2018] [Indexed: 01/27/2023]
Abstract
Gastric gland mucin-specific O-glycans are unique in having α1,4-linked N-acetylglucosamine (αGlcNAc) attached to MUC6. We previously reported decreased expression of αGlcNAc relative to MUC6 in gastric and pancreatic neoplasms, but its significance in cervical glandular lesions remained unclear. Here, we analyzed MUC5AC, MUC6, αGlcNAc, and p16 expression in 9 lesions of mucinous carcinoma, gastric type with minimal deviation adenocarcinoma (GAS-MDA), 5 of GAS with nonMDA (GAS-nonMDA), 14 of typical lobular endocervical gland hyperplasia (LEGH), and 5 of atypical LEGH (33 total lesions). All 33 were MUC5AC-positive. Moreover, all 14 typical LEGH, 5 atypical LEGH, 8 of 9 GAS-MDA, and 3 of 5 GAS-nonMDA were MUC6-positive. All 14 typical LEGH, 2 of 5 atypical LEGH, 3 of 9 GAS-MDA, and 1 of 5 GAS-nonMDA were αGlcNAc-positive. The proportion of αGlcNAc-positive atypical LEGH or GAS-MDA or GAS-nonMDA lesions was significantly smaller than that seen in typical LEGH lesions (P < 0.001 and P < 0.01, respectively). Of 33 lesions, 32 were p16-negative. Furthermore, when we evaluated MUC6 and αGlcNAc immunoreactivity semi-quantitatively in all 33 lesions, in typical LEGH and GAS-MDA, the immunohistochemical score for αGlcNAc was significantly lower than that for MUC6 (P < 0.01). We did not observe significantly decreased αGlcNAc expression relative to MUC6 in typical LEGH lesions. These studies suggest that αGlcNAc expression decreases as typical LEGH progresses to GAS. Given the difficulty in distinguishing MDA and atypical LEGH from typical LEGH in H.E. staining, we propose that immunohistochemical analysis of αGlcNAc and MUC6 could be useful.
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Affiliation(s)
- Kazuhiro Yamanoi
- Department of Molecular Pathology, Shinshu University School of Medicine, 3-1-1 Asahi, Matsumoto, 390-8621, Japan. .,Institute for Biomedical Sciences, Interdisciplinary Cluster for Cutting Edge Research, Shinshu University, Matsumoto, 390-8621, Japan.
| | - Keiko Ishii
- Department of Laboratory Medicine, Shinshu University Hospital, Matsumoto, 390-8621, Japan.,Department of Pathology, Okaya City Hospital, Okaya, 394-8512, Japan
| | - Michihiko Tsukamoto
- Department of Molecular Pathology, Shinshu University School of Medicine, 3-1-1 Asahi, Matsumoto, 390-8621, Japan
| | - Shiho Asaka
- Department of Laboratory Medicine, Shinshu University Hospital, Matsumoto, 390-8621, Japan
| | - Jun Nakayama
- Department of Molecular Pathology, Shinshu University School of Medicine, 3-1-1 Asahi, Matsumoto, 390-8621, Japan.,Institute for Biomedical Sciences, Interdisciplinary Cluster for Cutting Edge Research, Shinshu University, Matsumoto, 390-8621, Japan
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18
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Binding of Helicobacter pylori to Human Gastric Mucins Correlates with Binding of TFF1. Microorganisms 2018; 6:microorganisms6020044. [PMID: 29783620 PMCID: PMC6027488 DOI: 10.3390/microorganisms6020044] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/27/2018] [Revised: 04/23/2018] [Accepted: 05/01/2018] [Indexed: 02/06/2023] Open
Abstract
Helicobacter pylori binds to the gastric mucin, MUC5AC, and to trefoil factor, TFF1, which has been shown to interact with gastric mucin. We examined the interactions of TFF1 and H. pylori with purified gastrointestinal mucins from different animal species and from humans printed on a microarray platform to investigate whether TFF1 may play a role in locating H. pylori in gastric mucus. TFF1 bound almost exclusively to human gastric mucins and did not interact with human colonic mucins. There was a strong correlation between binding of TFF1 and H. pylori to human gastric mucins, and between binding of both TFF1 and H. pylori to gastric mucins with that of Griffonia simplicifolia lectin-II, which is specific for terminal non-reducing α- or β-linked N-acetyl-d-glucosamine. These results suggest that TFF1 may help to locate H. pylori in a discrete layer of gastric mucus and hence restrain their interactions with epithelial cells.
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19
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Reduced αGlcNAc glycosylation on gastric gland mucin is a biomarker of malignant potential for gastric cancer, Barrett’s adenocarcinoma, and pancreatic cancer. Histochem Cell Biol 2018; 149:569-575. [DOI: 10.1007/s00418-018-1667-8] [Citation(s) in RCA: 14] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Accepted: 04/10/2018] [Indexed: 01/22/2023]
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20
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Ohya A, Yamanoi K, Shimojo H, Fujii C, Nakayama J. Gastric gland mucin-specific O-glycan expression decreases with tumor progression from precursor lesions to pancreatic cancer. Cancer Sci 2017; 108:1897-1902. [PMID: 28685935 PMCID: PMC5581519 DOI: 10.1111/cas.13317] [Citation(s) in RCA: 24] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/30/2017] [Revised: 06/20/2017] [Accepted: 07/01/2017] [Indexed: 01/06/2023] Open
Abstract
Pancreatic cancer is lethal, as it is often detected late. Thus, novel biomarkers of precursor lesions are needed to devise timely therapies. Pancreatic intraepithelial neoplasia (PanIN) and intraductal papillary mucinous neoplasm (IPMN) are major precursors of pancreatic cancer. In normal gastric mucosa, gastric gland mucin‐specific O‐glycans are unique in having α1,4‐linked N‐acetylglucosamine (αGlcNAc) residues attached to MUC6. Recently we reported that αGlcNAc functions as a tumor suppressor for differentiated‐type gastric adenocarcinoma (Karasawa et al., J Clin Invest 122, 923, 2012). MUC6 is also expressed in pancreatic neoplasms, including PanIN and IPMN, but the role of αGlcNAc expression in pancreatic neoplasms remains unknown. Here, we analyze expression patterns of αGlcNAc, MUC6 and MUC5AC in pancreatic neoplasms and compare them with progression from PanIN to invasive ductal adenocarcinoma (IDAC) (the PanIN‐IDAC sequence; 20 cases) and from IPMN to IPMN with associated invasive carcinoma (IPMNAIC) (the IPMN‐IPMNAIC sequence; 20 cases). At both sequences, the frequency of MUC6‐positive and αGlcNAc‐positive lesions decreased with tumor progression. We then compared expression levels of αGlcNAc and MUC6 at each step of the progression. At the PanIN‐IDAC sequence, αGlcNAc expression significantly decreased relative to MUC6 in low‐grade PanIN (P = 0.021), high‐grade PanIN/intraductal spread of IDAC (P = 0.031) and IDAC (P = 0.013). At the IPMN‐IPMNAIC sequence, decreased αGlcNAc expression was also observed in low‐grade IPMN exhibiting gastric‐type morphology (P = 0.020). These results suggest that decreased expression of αGlcNAc relative to MUC6 occurs early and marks the initiation of tumor progression to pancreatic cancer.
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Affiliation(s)
- Ayumi Ohya
- Department of Molecular Pathology, Shinshu University Graduate School of Medicine, Matsumoto, Japan.,Department of Radiology, Shinshu University School of Medicine, Matsumoto, Japan
| | - Kazuhiro Yamanoi
- Department of Molecular Pathology, Shinshu University Graduate School of Medicine, Matsumoto, Japan.,Institute for Biomedical Sciences, Interdisciplinary Cluster for Cutting Edge Research, Shinshu University, Matsumoto, Japan
| | - Hisashi Shimojo
- Department of Pathology, Shinshu University School of Medicine, Matsumoto, Japan
| | - Chifumi Fujii
- Department of Molecular Pathology, Shinshu University Graduate School of Medicine, Matsumoto, Japan.,Institute for Biomedical Sciences, Interdisciplinary Cluster for Cutting Edge Research, Shinshu University, Matsumoto, Japan
| | - Jun Nakayama
- Department of Molecular Pathology, Shinshu University Graduate School of Medicine, Matsumoto, Japan.,Institute for Biomedical Sciences, Interdisciplinary Cluster for Cutting Edge Research, Shinshu University, Matsumoto, Japan
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21
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Wang SY, Laborda P, Lu AM, Wang M, Duan XC, Liu L, Voglmeir J. Chemo-enzymatic approach to access diastereopure α-substituted GlcNAc derivatives. J Carbohydr Chem 2017. [DOI: 10.1080/07328303.2017.1321116] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 10/19/2022]
Affiliation(s)
- Su-Yan Wang
- Glycomics and Glycan Bioengineering Research Center (GGBRC), College of Food Science and Technology, Nanjing Agricultural University, Nanjing, People's Republic of China
| | - Pedro Laborda
- Glycomics and Glycan Bioengineering Research Center (GGBRC), College of Food Science and Technology, Nanjing Agricultural University, Nanjing, People's Republic of China
| | - Ai-Min Lu
- College of Sciences, Nanjing Agricultural University, Nanjing, People's Republic of China
| | - Meng Wang
- Glycomics and Glycan Bioengineering Research Center (GGBRC), College of Food Science and Technology, Nanjing Agricultural University, Nanjing, People's Republic of China
| | - Xu-Chu Duan
- Glycomics and Glycan Bioengineering Research Center (GGBRC), College of Food Science and Technology, Nanjing Agricultural University, Nanjing, People's Republic of China
| | - Li Liu
- Glycomics and Glycan Bioengineering Research Center (GGBRC), College of Food Science and Technology, Nanjing Agricultural University, Nanjing, People's Republic of China
- Qlyco Ltd., Nanjing, People's Republic of China
| | - Josef Voglmeir
- Glycomics and Glycan Bioengineering Research Center (GGBRC), College of Food Science and Technology, Nanjing Agricultural University, Nanjing, People's Republic of China
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22
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Protein glycosylation in gastric and colorectal cancers: Toward cancer detection and targeted therapeutics. Cancer Lett 2017; 387:32-45. [DOI: 10.1016/j.canlet.2016.01.044] [Citation(s) in RCA: 45] [Impact Index Per Article: 5.6] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/16/2015] [Revised: 01/20/2016] [Accepted: 01/22/2016] [Indexed: 12/25/2022]
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23
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Corfield A. Eukaryotic protein glycosylation: a primer for histochemists and cell biologists. Histochem Cell Biol 2017; 147:119-147. [PMID: 28012131 PMCID: PMC5306191 DOI: 10.1007/s00418-016-1526-4] [Citation(s) in RCA: 94] [Impact Index Per Article: 11.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Accepted: 11/25/2016] [Indexed: 12/21/2022]
Abstract
Proteins undergo co- and posttranslational modifications, and their glycosylation is the most frequent and structurally variegated type. Histochemically, the detection of glycan presence has first been performed by stains. The availability of carbohydrate-specific tools (lectins, monoclonal antibodies) has revolutionized glycophenotyping, allowing monitoring of distinct structures. The different types of protein glycosylation in Eukaryotes are described. Following this educational survey, examples where known biological function is related to the glycan structures carried by proteins are given. In particular, mucins and their glycosylation patterns are considered as instructive proof-of-principle case. The tissue and cellular location of glycoprotein biosynthesis and metabolism is reviewed, with attention to new findings in goblet cells. Finally, protein glycosylation in disease is documented, with selected examples, where aberrant glycan expression impacts on normal function to let disease pathology become manifest. The histological applications adopted in these studies are emphasized throughout the text.
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Affiliation(s)
- Anthony Corfield
- Mucin Research Group, School of Clinical Sciences, Bristol Royal Infirmary, University of Bristol, Bristol, BS2 8HW, UK.
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24
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Sinha J, Cao Z, Dai J, Tang H, Partyka K, Hostetter G, Simeone DM, Feng Z, Allen PJ, Brand RE, Haab BB. A Gastric Glycoform of MUC5AC Is a Biomarker of Mucinous Cysts of the Pancreas. PLoS One 2016; 11:e0167070. [PMID: 27992432 PMCID: PMC5167232 DOI: 10.1371/journal.pone.0167070] [Citation(s) in RCA: 20] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/24/2016] [Accepted: 10/22/2016] [Indexed: 02/07/2023] Open
Abstract
Molecular indicators to specify the risk posed by a pancreatic cyst would benefit patients. Previously we showed that most cancer-precursor cysts, termed mucinous cysts, produce abnormal glycoforms of the proteins MUC5AC and endorepellin. Here we sought to validate the glycoforms as a biomarker of mucinous cysts and to specify the oligosaccharide linkages that characterize MUC5AC. We hypothesized that mucinous cysts secrete MUC5AC displaying terminal N-acetylglucosamine (GlcNAc) in either alpha or beta linkage. We used antibody-lectin sandwich assays to detect glycoforms of MUC5AC and endorepellin in cyst fluid samples from three independent cohorts of 49, 32, and 66 patients, and we used monoclonal antibodies to test for terminal, alpha-linked GlcNAc and the enzyme that produces it. A biomarker panel comprising the previously-identified glycoforms of MUC5AC and endorepellin gave 96%, 96%, and 87% accuracy for identifying mucinous cysts in the three cohorts with an average sensitivity of 92% and an average specificity of 94%. Glycan analysis showed that MUC5AC produced by a subset of mucinous cysts displays terminal alpha-GlcNAc, a motif expressed in stomach glands. The alpha-linked glycoform of MUC5AC was unique to intraductal papillary mucinous neoplasms (IPMN), whereas terminal beta-linked GlcNAc was increased in both IPMNs and mucinous cystic neoplasms (MCN). The enzyme that synthesizes alpha-GlcNAc, A4GNT, was expressed in the epithelia of mucinous cysts that expressed alpha-GlcNAc, especially in regions with high-grade dysplasia. Thus IPMNs secrete a gastric glycoform of MUC5AC that displays terminal alpha-GlcNAc, and the combined alpha-GlcNAc and beta-GlcNAc glycoforms form an accurate biomarker of mucinous cysts.
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Affiliation(s)
- Jessica Sinha
- Van Andel Research Institute, Grand Rapids, MI, United States of America
| | - Zheng Cao
- Van Andel Research Institute, Grand Rapids, MI, United States of America
| | - Jianliang Dai
- MD Anderson Cancer Center, Houston, TX, United States of America
| | - Huiyuan Tang
- Van Andel Research Institute, Grand Rapids, MI, United States of America
| | - Katie Partyka
- Van Andel Research Institute, Grand Rapids, MI, United States of America
| | - Galen Hostetter
- Van Andel Research Institute, Grand Rapids, MI, United States of America
| | - Diane M. Simeone
- University of Michigan School of Medicine, Ann Arbor, MI, United States of America
| | - Ziding Feng
- MD Anderson Cancer Center, Houston, TX, United States of America
| | - Peter J. Allen
- Memorial Sloan Kettering Cancer Center, New York, NY, United States of America
| | - Randall E. Brand
- University of Pittsburgh Medical Center, Pittsburgh, PA, United States of America
| | - Brian B. Haab
- Van Andel Research Institute, Grand Rapids, MI, United States of America
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25
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Gabius HJ, Manning JC, Kopitz J, André S, Kaltner H. Sweet complementarity: the functional pairing of glycans with lectins. Cell Mol Life Sci 2016; 73:1989-2016. [PMID: 26956894 PMCID: PMC11108359 DOI: 10.1007/s00018-016-2163-8] [Citation(s) in RCA: 62] [Impact Index Per Article: 6.9] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/10/2015] [Revised: 02/08/2016] [Accepted: 02/09/2016] [Indexed: 02/07/2023]
Abstract
Carbohydrates establish the third alphabet of life. As part of cellular glycoconjugates, the glycans generate a multitude of signals in a minimum of space. The presence of distinct glycotopes and the glycome diversity are mapped by sugar receptors (antibodies and lectins). Endogenous (tissue) lectins can read the sugar-encoded information and translate it into functional aspects of cell sociology. Illustrated by instructive examples, each glycan has its own ligand properties. Lectins with different folds can converge to target the same epitope, while intrafamily diversification enables functional cooperation and antagonism. The emerging evidence for the concept of a network calls for a detailed fingerprinting. Due to the high degree of plasticity and dynamics of the display of genes for lectins the validity of extrapolations between different organisms of the phylogenetic tree yet is inevitably limited.
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Affiliation(s)
- H-J Gabius
- Institute of Physiological Chemistry, Faculty of Veterinary Medicine, Ludwig-Maximilians-University Munich, Veterinärstr. 13, 80539, Munich, Germany.
| | - J C Manning
- Institute of Physiological Chemistry, Faculty of Veterinary Medicine, Ludwig-Maximilians-University Munich, Veterinärstr. 13, 80539, Munich, Germany
| | - J Kopitz
- Institute of Pathology, Department of Applied Tumor Biology, Ruprecht-Karls-University Heidelberg, Im Neuenheimer Feld 224, 69120, Heidelberg, Germany
| | - S André
- Institute of Physiological Chemistry, Faculty of Veterinary Medicine, Ludwig-Maximilians-University Munich, Veterinärstr. 13, 80539, Munich, Germany
| | - H Kaltner
- Institute of Physiological Chemistry, Faculty of Veterinary Medicine, Ludwig-Maximilians-University Munich, Veterinärstr. 13, 80539, Munich, Germany
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26
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Merging carbohydrate chemistry with lectin histochemistry to study inhibition of lectin binding by glycoclusters in the natural tissue context. Histochem Cell Biol 2015; 145:185-99. [PMID: 26553286 DOI: 10.1007/s00418-015-1383-6] [Citation(s) in RCA: 15] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Accepted: 10/28/2015] [Indexed: 01/22/2023]
Abstract
Recognition of glycans by lectins leads to cell adhesion and growth regulation. The specificity and selectivity of this process are determined by carbohydrate structure (sequence and shape) and topology of its presentation. The synthesis of (neo)glycoconjugates with bi- to oligo-valency (glycoclusters) affords tools to delineate structure-activity relationships by blocking lectin binding to an artificial matrix, often a glycoprotein, or cultured cell lines. The drawback of these assays is that glycan presentation is different from that in tissues. In order to approach the natural context, we here introduce lectin histochemistry on fixed tissue sections to determine the susceptibility of binding of two plant lectins, i.e., GSA-II and WGA, to a series of 10 glycoclusters. Besides valency, this panel covers changes in the anomeric position (α/β) and the atom at the glycosidic linkage (O/S). Flanked by cell and solid-phase assays with human tumor lines and two mucins, respectively, staining (intensity and profile) was analyzed in sections of murine jejunum, stomach and epididymis as a function of glycocluster presence. The marked and differential sensitivity of signal generation to structural aspects of the glycoclusters proves the applicability of this method. This enables comparisons between data sets obtained by using (neo)glycoconjugates, cells and the tissue context as platforms. The special advantage of processing tissue sections is the monitoring of interference with lectin association at sites that are relevant for functionality. Testing glycoclusters in lectin histochemistry will especially be attractive in cases of multi-target recognition (glycans, proteins and lipids) by a tissue lectin.
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27
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Yamada S, Okamura T, Kobayashi S, Tanaka E, Nakayama J. Reduced gland mucin-specific O-glycan in gastric atrophy: A possible risk factor for differentiated-type adenocarcinoma of the stomach. J Gastroenterol Hepatol 2015; 30:1478-84. [PMID: 25967588 DOI: 10.1111/jgh.13000] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Accepted: 04/13/2015] [Indexed: 12/25/2022]
Abstract
BACKGROUND AND AIMS O-glycans exhibiting terminal α1,4-linked N-acetylglucosamine (αGlcNAc) are attached to MUC6 in gastric gland mucins and serve as a tumor suppressor for gastric adenocarcinoma. Gastric atrophy is associated with risk for gastric cancer. However, the significance of αGlcNAc expression in pyloric glands of chronic atrophic gastritis remains unknown. Here, we asked whether reduced αGlcNAc expression in chronic atrophic gastritis is associated with risk for gastric cancer. METHODS We quantitatively analyzed expression of αGlcNAc relative to MUC6 in pyloric glands by immunohistochemistry in 67 patients with normal mucosa, 70 with chronic atrophic gastritis, 68 with intramucosal differentiated-type adenocarcinoma, and 11 with intramucosal undifferentiated-type adenocarcinoma. We also compared the Ki-67 labeling index in gastric epithelial cells between chronic atrophic gastritis and normal gastric mucosa with respect to αGlcNAc reduction. RESULTS In normal pyloric mucosa, αGlcNAc was co-expressed with MUC6. By contrast, in chronic atrophic gastritis, pyloric gland αGlcNAc expression was significantly reduced relative to MUC6. In intramucosal gastric cancer, αGlcNAc expression in pyloric glands found just beneath differentiated-type adenocarcinoma was also reduced relative to MUC6. However, pyloric glands present beneath undifferentiated-type adenocarcinoma exhibited no αGlcNAc decrease. The Ki-67 labeling index in chronic atrophic gastritis showing αGlcNAc reduction was significantly increased relative to that in normal gastric mucosa. CONCLUSIONS Because αGlcNAc prevents the gastric cancer development, reduced αGlcNAc expression in chronic atrophic gastritis is a possible risk factor for differentiated-type adenocarcinoma of the stomach.
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Affiliation(s)
- Shigenori Yamada
- Department of Molecular Pathology, Shinshu University Graduate School of Medicine, Matsumoto, Japan.,Department of Gastroenterology, Shinshu University School of Medicine, Matsumoto, Japan
| | - Takuma Okamura
- Department of Molecular Pathology, Shinshu University Graduate School of Medicine, Matsumoto, Japan.,Department of Gastroenterology, Shinshu University School of Medicine, Matsumoto, Japan
| | - Satoshi Kobayashi
- Department of Molecular Pathology, Shinshu University Graduate School of Medicine, Matsumoto, Japan.,Department of Gastroenterology, Shinshu University School of Medicine, Matsumoto, Japan.,Department of Gastroenterology, Iiyama Red Cross Hospital, Iiyama, Japan
| | - Eiji Tanaka
- Department of Gastroenterology, Shinshu University School of Medicine, Matsumoto, Japan
| | - Jun Nakayama
- Department of Molecular Pathology, Shinshu University Graduate School of Medicine, Matsumoto, Japan
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28
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Hoffmann W. TFF2, a MUC6-binding lectin stabilizing the gastric mucus barrier and more (Review). Int J Oncol 2015. [PMID: 26201258 DOI: 10.3892/ijo.2015.3090] [Citation(s) in RCA: 64] [Impact Index Per Article: 6.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/06/2022] Open
Abstract
The peptide TFF2 (formerly 'spasmolytic polypeptide'), a member of the trefoil factor family (TFF) containing two TFF domains, is mainly expressed together with the mucin MUC6 in the gastric epithelium and duodenal Brunner's glands. Pathologically, TFF2 expression is observed ectopically during stone diseases, chronic inflammatory conditions and in several metaplastic and neoplastic epithelia; most prominent being the 'spasmolytic polypeptide-expressing metaplasia' (SPEM), which is an established gastric precancerous lesion. TFF2 plays a critical role in maintaining gastric mucosal integrity and appears to restrain tumorigenesis in the stomach. Recently, porcine TFF2 has been shown to interact with the gastric mucin MUC6 and thus stabilize the gastric mucus barrier. On the one hand, TFF2 binds to MUC6 via non-covalent lectin interactions with the glycotope GlcNAcα1→4Galβ1→R. On the other hand, TFF2 is probably also covalently bound to MUC6 via disulfide bridges. Thus, implications for the complex multimeric assembly, cross-linking, and packaging of MUC6 as well as the rheology of gastric mucus are discussed in detail in this review. Furthermore, TFF2 is also expressed in minor amounts in the immune and nervous systems. Thus, similar to galectins, its lectin activity would perfectly enable TFF2 to form multivalent complexes and cross-linked lattices with a plethora of transmembrane glycoproteins and thus modulate different signal transduction processes. This could explain the multiple and diverse biological effects of TFF2 [e.g., motogenic, (anti)apoptotic, and angiogenic effects]. Finally, a function during fertilization is also possible for TFF domains because they occur as shuffled modules in certain zona pellucida proteins.
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Affiliation(s)
- Werner Hoffmann
- Institute of Molecular Biology and Medicinal Chemistry, Otto-von-Guericke-University Magdeburg, D-39120 Magdeburg, Germany
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29
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Yamanoi K, Sekine S, Higuchi K, Kushima R, Nakayama J. Decreased expression of gastric gland mucin-specific glycan α1,4-linked N-acetylglucosamine on its scaffold mucin 6 is associated with malignant potential of pyloric gland adenoma of the stomach. Histopathology 2015; 67:898-904. [PMID: 25929994 DOI: 10.1111/his.12728] [Citation(s) in RCA: 13] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/17/2015] [Accepted: 04/25/2015] [Indexed: 12/22/2022]
Abstract
AIMS Pyloric gland adenoma (PGA) is a unique gastric neoplasm expressing mucin 6 (MUC6), and is often associated with high-grade dysplasia and/or adenocarcinoma. MUC6 secreted from the gastric gland mucous cells, such as pyloric gland cells, carries unique O-glycans with terminal α1,4-linked N-acetylglucosamine (αGlcNAc) residues on its molecule. As we recently demonstrated that αGlcNAc serves as a tumour suppressor for gastric adenocarcinoma, this study aimed to investigate the significance of αGlcNAc expression in PGA. METHODS AND RESULTS Eighteen patients with PGA were examined with immunohistochemistry for αGlcNAc and MUC6. αGlcNAc and MUC6 were coexpressed in 12 of 18 PGAs. However, reduced αGlcNAc expression relative to MUC6 expression was observed in six cases. When the MIB-1 labelling index (LI) of tumour cells was examined with respect to reduced αGlcNAc expression, the MIB-1 LI was significantly higher in PGAs showing decreased αGlcNAc expression relative to MUC6 expression than in PGAs with unchanged αGlcNAc expression (P = 0.023). CONCLUSIONS The present study indicates that coexpression of αGlcNAc and MUC6 in PGA suggests the presence of fully glycosylated MUC6 on tumour cells, consistent with pyloric gland differentiation. However, the decreased glycosylation of αGlcNAc on MUC6 is associated with high mitotic activity of tumour cells, indicative of malignant potential of PGA.
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Affiliation(s)
- Kazuhiro Yamanoi
- Department of Pathology, Aizawa Hospital, Matsumoto, Japan.,Department of Pathology, School of Medicine, Keio University, Tokyo, Japan
| | - Shigeki Sekine
- Pathology and Clinical Laboratory Division, National Cancer Centre Hospital, Tokyo, Japan
| | - Kayoko Higuchi
- Department of Pathology, Aizawa Hospital, Matsumoto, Japan
| | - Ryoji Kushima
- Pathology and Clinical Laboratory Division, National Cancer Centre Hospital, Tokyo, Japan.,Division of Diagnostic Pathology, Department of Clinical Laboratory Medicine, Shiga University of Medical Science, Shiga, Japan
| | - Jun Nakayama
- Department of Molecular Pathology, Shinshu University Graduate School of Medicine, Matsumoto, Japan
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30
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Joncquel Chevalier Curt M, Lecointe K, Mihalache A, Rossez Y, Gosset P, Léonard R, Robbe-Masselot C. Alteration or adaptation, the two roads for human gastric mucin glycosylation infected by Helicobacter pylori. Glycobiology 2015; 25:617-31. [DOI: 10.1093/glycob/cwv004] [Citation(s) in RCA: 16] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/19/2014] [Accepted: 01/06/2015] [Indexed: 12/24/2022] Open
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31
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Hanisch FG, Bonar D, Schloerer N, Schroten H. Human trefoil factor 2 is a lectin that binds α-GlcNAc-capped mucin glycans with antibiotic activity against Helicobacter pylori. J Biol Chem 2014; 289:27363-75. [PMID: 25124036 DOI: 10.1074/jbc.m114.597757] [Citation(s) in RCA: 46] [Impact Index Per Article: 4.2] [Reference Citation Analysis] [Abstract] [Key Words] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/16/2022] Open
Abstract
Helicobacter pylori infection is the major cause of gastric cancer and remains an important health care challenge. The trefoil factor peptides are a family of small highly conserved proteins that are claimed to play essential roles in cytoprotection and epithelial repair within the gastrointestinal tract. H. pylori colocalizes with MUC5AC at the gastric surface epithelium, but not with MUC6 secreted in concert with TFF2 by deep gastric glands. Both components of the gastric gland secretome associate non-covalently and show increased expression upon H. pylori infection. Although blood group active O-glycans of the Lewis-type form the basis of H. pylori adhesion to the surface mucin layer and to epithelial cells, α1,4-GlcNAc-capped O-glycans on gastric mucins were proposed to inhibit H. pylori growth as a natural antibiotic. We show here that the gastric glycoform of TFF2 is a calcium-independent lectin, which binds with high specificity to O-linked α1,4-GlcNAc-capped hexasaccharides on human and porcine stomach mucin. The structural assignments of two hexasaccharide isomers and the binding active glycotope were based on mass spectrometry, linkage analysis, (1)H nuclear magnetic resonance spectroscopy, glycan inhibition, and lectin competition of TFF2-mucin binding. Neoglycolipids derived from the C3/C6-linked branches of the two isomers revealed highly specific TFF2 binding to the 6-linked trisaccharide in GlcNAcα1-4Galβ1-4GlcNAcβ1-6(Fucα1-2Galβ1-3)GalNAc-ol(Structure 1). Supposedly, lectin TFF2 is involved in protection of gastric epithelia via a functional relationship to defense against H. pylori launched by antibiotic α1,4-GlcNAc-capped mucin glycans. Lectin-carbohydrate interaction may have also an impact on more general functional aspects of TFF members by mediating their binding to cell signaling receptors.
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Affiliation(s)
- Franz-Georg Hanisch
- From the Institute of Biochemistry II, Medical Faculty, University of Cologne, Joseph-Stelzmann-Str. 52, 50931 Köln, the Center for Molecular Medicine Cologne, University of Cologne, Robert-Koch-Str. 21, 50931 Köln,
| | - David Bonar
- From the Institute of Biochemistry II, Medical Faculty, University of Cologne, Joseph-Stelzmann-Str. 52, 50931 Köln
| | - Nils Schloerer
- the Institute of Organic Chemistry, University of Cologne, Greinstr. 4, 50939 Köln, and
| | - Horst Schroten
- the University Children's Hospital, Mannheim, Heidelberg University, Theodor-Kutzer-Ufer 1-3, 68167 Mannheim, Germany
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32
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Molecular Mechanism of Gastric Carcinogenesis in Helicobacter pylori-Infected Rodent Models. Diseases 2014. [DOI: 10.3390/diseases2020168] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/05/2023] Open
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33
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Dunne C, Dolan B, Clyne M. Factors that mediate colonization of the human stomach by Helicobacter pylori. World J Gastroenterol 2014; 20:5610-24. [PMID: 24914320 PMCID: PMC4024769 DOI: 10.3748/wjg.v20.i19.5610] [Citation(s) in RCA: 83] [Impact Index Per Article: 7.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 10/25/2013] [Revised: 12/04/2013] [Accepted: 01/19/2014] [Indexed: 02/06/2023] Open
Abstract
Helicobacter pylori (H. pylori) colonizes the stomach of humans and causes chronic infection. The majority of bacteria live in the mucus layer overlying the gastric epithelial cells and only a small proportion of bacteria are found interacting with the epithelial cells. The bacteria living in the gastric mucus may act as a reservoir of infection for the underlying cells which is essential for the development of disease. Colonization of gastric mucus is likely to be key to the establishment of chronic infection. How H. pylori manages to colonise and survive in the hostile environment of the human stomach and avoid removal by mucus flow and killing by gastric acid is the subject of this review. We also discuss how bacterial and host factors may together go some way to explaining the susceptibility to colonization and the outcome of infection in different individuals. H. pylori infection of the gastric mucosa has become a paradigm for chronic infection. Understanding of why H. pylori is such a successful pathogen may help us understand how other bacterial species colonise mucosal surfaces and cause disease.
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34
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Nakayama J. Dual Roles of Gastric Gland Mucin-specific O-glycans in Prevention of Gastric Cancer. Acta Histochem Cytochem 2014; 47:1-9. [PMID: 24761044 PMCID: PMC3972424 DOI: 10.1267/ahc.13034] [Citation(s) in RCA: 27] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/15/2013] [Accepted: 12/24/2013] [Indexed: 11/22/2022] Open
Abstract
Gastric gland mucin is secreted from gland mucous cells, including pyloric gland cells and mucous neck cells located in the lower layer of the gastric mucosa. These mucins typically contain O-glycans carrying terminal α1,4-linked N-acetylglucosamine residues (αGlcNAc) attached to the scaffold protein MUC6, and biosynthesis of the O-glycans is catalyzed by the glycosyltransferase, α1,4-N-acetylglucosaminyltransferase (α4GnT). We previously used expression cloning to isolate cDNA encoding α4GnT, and then demonstrated that αGlcNAc functions as natural antibiotic against Helicobacter pylori, a microbe causing various gastric diseases including gastric cancer. More recently, it was shown that αGlcNAc serves as a tumor suppressor for differentiated-type adenocarcinoma. This review summarizes these findings and identifies dual roles for αGlcNAc in gastric cancer.
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Affiliation(s)
- Jun Nakayama
- Department of Molecular Pathology, Shinshu University Graduate School of Medicine
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35
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Shiratsu K, Higuchi K, Nakayama J. Loss of gastric gland mucin-specific O-glycan is associated with progression of differentiated-type adenocarcinoma of the stomach. Cancer Sci 2014; 105:126-33. [PMID: 24138592 PMCID: PMC4317868 DOI: 10.1111/cas.12305] [Citation(s) in RCA: 29] [Impact Index Per Article: 2.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/14/2013] [Revised: 10/08/2013] [Accepted: 10/16/2013] [Indexed: 12/25/2022] Open
Abstract
Gastric gland mucin secreted from the lower portion of the gastric mucosa contains unique O-linked oligosaccharides having terminal α1,4-linked N-acetylglucosamine (αGlcNAc) residues largely attached to a MUC6 scaffold. Previously, we generated A4gnt-deficient mice, which totally lack αGlcNAc, and showed that αGlcNAc functions as a tumor suppressor for gastric cancer. Here, to determine the clinicopathological significance of αGlcNAc in gastric carcinomas, we examined immunohistochemical expression of αGlcNAc and mucin phenotypic markers including MUC5AC, MUC6, MUC2, and CD10 in 214 gastric adenocarcinomas and compared those expression patterns with clinicopathological parameters and cancer-specific survival. The αGlcNAc loss was evaluated in MUC6-positive gastric carcinoma. Thirty-three (61.1%) of 54 differentiated-type gastric adenocarcinomas exhibiting MUC6 in cancer cells lacked αGlcNAc expression. Loss of αGlcNAc was significantly correlated with depth of invasion, stage, and venous invasion by differentiated-type adenocarcinoma. Loss of αGlcNAc was also significantly associated with poorer patient prognosis in MUC6-positive differentiated-type adenocarcinoma. By contrast, no significant correlation between αGlcNAc loss and any clinicopathologic variable was observed in undifferentiated-type adenocarcinoma. Expression of MUC6 was also significantly correlated with several clinicopathological variables in differentiated-type adenocarcinoma. However, unlike the case with αGlcNAc, its expression showed no correlation with cancer-specific survival in patients. In undifferentiated-type adenocarcinoma, we observed no significant correlation between mucin phenotypic marker expression, including MUC6, and any clinicopathologic variable. These results together indicate that loss of αGlcNAc in MUC6-positive cancer cells is associated with progression and poor prognosis in differentiated, but not undifferentiated, types of gastric adenocarcinoma.
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Affiliation(s)
- Kazuo Shiratsu
- Department of Molecular Pathology, Shinshu University Graduate School of MedicineMatsumoto, Japan
- Department of Gastroenterology, Aizawa HospitalMatsumoto, Japan
| | - Kayoko Higuchi
- Department of Pathology, Aizawa HospitalMatsumoto, Japan
| | - Jun Nakayama
- Department of Molecular Pathology, Shinshu University Graduate School of MedicineMatsumoto, Japan
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36
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Ito Y, Vela JL, Matsumura F, Hoshino H, Tyznik A, Lee H, Girardi E, Zajonc DM, Liddington R, Kobayashi M, Bao X, Bugaytsova J, Borén T, Jin R, Zong Y, Seeberger PH, Nakayama J, Kronenberg M, Fukuda M. Helicobacter pylori cholesteryl α-glucosides contribute to its pathogenicity and immune response by natural killer T cells. PLoS One 2013; 8:e78191. [PMID: 24312443 PMCID: PMC3846475 DOI: 10.1371/journal.pone.0078191] [Citation(s) in RCA: 49] [Impact Index Per Article: 4.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/28/2013] [Accepted: 09/09/2013] [Indexed: 12/31/2022] Open
Abstract
Approximately 10–15% of individuals infected with Helicobacter pylori will develop ulcer disease (gastric or duodenal ulcer), while most people infected with H. pylori will be asymptomatic. The majority of infected individuals remain asymptomatic partly due to the inhibition of synthesis of cholesteryl α-glucosides in H. pylori cell wall by α1,4-GlcNAc-capped mucin O-glycans, which are expressed in the deeper portion of gastric mucosa. However, it has not been determined how cholesteryl α-glucosyltransferase (αCgT), which forms cholesteryl α-glucosides, functions in the pathogenesis of H. pylori infection. Here, we show that the activity of αCgT from H. pylori clinical isolates is highly correlated with the degree of gastric atrophy. We investigated the role of cholesteryl α-glucosides in various aspects of the immune response. Phagocytosis and activation of dendritic cells were observed at similar degrees in the presence of wild-type H. pylori or variants harboring mutant forms of αCgT showing a range of enzymatic activity. However, cholesteryl α-glucosides were recognized by invariant natural killer T (iNKT) cells, eliciting an immune response in vitro and in vivo. Following inoculation of H. pylori harboring highly active αCgT into iNKT cell-deficient (Jα18−/−) or wild-type mice, bacterial recovery significantly increased in Jα18−/− compared to wild-type mice. Moreover, cytokine production characteristic of Th1 and Th2 cells dramatically decreased in Jα18−/− compared to wild-type mice. These findings demonstrate that cholesteryl α-glucosides play critical roles in H. pylori-mediated gastric inflammation and precancerous atrophic gastritis.
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Affiliation(s)
- Yuki Ito
- Cancer Center, Sanford-Burnham Medical Research Institute, La Jolla, California, United States of America
| | - Jose Luis Vela
- La Jolla Institute for Allergy & Immunology, La Jolla, California, United States of America
| | - Fumiko Matsumura
- Cancer Center, Sanford-Burnham Medical Research Institute, La Jolla, California, United States of America
| | - Hitomi Hoshino
- Department of Molecular Pathology, Shinshu University Graduate School of Medicine, Matsumoto, Nagano, Japan
| | - Aaron Tyznik
- La Jolla Institute for Allergy & Immunology, La Jolla, California, United States of America
| | - Heeseob Lee
- Cancer Center, Sanford-Burnham Medical Research Institute, La Jolla, California, United States of America
| | - Enrico Girardi
- La Jolla Institute for Allergy & Immunology, La Jolla, California, United States of America
| | - Dirk M. Zajonc
- La Jolla Institute for Allergy & Immunology, La Jolla, California, United States of America
| | - Robert Liddington
- Infectious and Inflammatory Disease Center, Sanford-Burnham Medical Research Institute, La Jolla, California, United States of America
| | - Motohiro Kobayashi
- Department of Molecular Pathology, Shinshu University Graduate School of Medicine, Matsumoto, Nagano, Japan
| | - Xingfeng Bao
- Cancer Center, Sanford-Burnham Medical Research Institute, La Jolla, California, United States of America
| | - Jeanna Bugaytsova
- Department of Medical Biochemistry and Biophysics, Umeå University, Umeå, Sweden
| | - Thomas Borén
- Department of Medical Biochemistry and Biophysics, Umeå University, Umeå, Sweden
| | - Rongsheng Jin
- Del E. Webb Neuroscience, Aging and Stem Cell Research Center, Sanford-Burnham Medical Research Institute, La Jolla, California, United States of America
| | - Yinong Zong
- Del E. Webb Neuroscience, Aging and Stem Cell Research Center, Sanford-Burnham Medical Research Institute, La Jolla, California, United States of America
| | - Peter H. Seeberger
- Cancer Center, Sanford-Burnham Medical Research Institute, La Jolla, California, United States of America
| | - Jun Nakayama
- Department of Molecular Pathology, Shinshu University Graduate School of Medicine, Matsumoto, Nagano, Japan
| | - Mitchell Kronenberg
- La Jolla Institute for Allergy & Immunology, La Jolla, California, United States of America
| | - Minoru Fukuda
- Cancer Center, Sanford-Burnham Medical Research Institute, La Jolla, California, United States of America
- * E-mail:
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Cheng PF, Snovida S, Ho MY, Cheng CW, Wu AM, Khoo KH. Increasing the depth of mass spectrometry-based glycomic coverage by additional dimensions of sulfoglycomics and target analysis of permethylated glycans. Anal Bioanal Chem 2013; 405:6683-95. [PMID: 23797909 DOI: 10.1007/s00216-013-7128-2] [Citation(s) in RCA: 26] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/02/2013] [Revised: 05/30/2013] [Accepted: 06/06/2013] [Indexed: 01/08/2023]
Abstract
Hog or porcine gastric mucin resembles the human source in carrying not only blood group antigens but also the rather rare α4-GlcNAc-capped terminal epitope functionally implicated in protection against Helicobacter pylori infection. Being more readily available and reasonably well characterized, it serves as a good reagent for immunobiological studies, as well as a standard for analytical methodology developments. Current approaches in mass spectrometry (MS)-based glycomic mapping remain vastly inadequate in revealing the full complexity of glycosylation, particularly for cases such as the extremely heterogeneous O-glycosylation of mucosal mucins that can be further sulfated. We demonstrate here a novel concerted workflow that extends the conventional matrix-assisted laser desorption/ionization–mass spectrometry (MALDI-MS) mapping of permethylated glycans in positive ion mode to include a further step of sulfoglycomic analysis in negative ion mode. This was facilitated by introducing a mixed-mode solid-phase extraction step, which allows direct cleanup and simultaneous fractionation of the permethylated glycans into separate non-sulfated and sulfated pools in one single step. By distinct MALDI-MS/MS fragmentation patterns, all previously known structural features of porcine gastric mucin including the terminal epitopes and location of sulfates could be readily defined. We additionally showed that both arms of the core 2 structures could be extended via 6-O-sulfated GlcNAc to yield a series of disulfated O-glycans not previously reported, thus expanding its current glycomic coverage. However, a targeted LC-MSn analysis was required and best suited to dig even deeper into validating the occurrence of very minor structural isomers carrying the Lewis Y epitope implicated by positive antibody binding.
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Affiliation(s)
- Ping-Fu Cheng
- Institute of Biochemical Sciences, National Taiwan University, Roosevelt Road, PO Box 23-106, Taipei, 10617, Taiwan
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Biochemical studies on sphingolipids of Artemia franciscana: complex neutral glycosphingolipids. Glycoconj J 2012; 30:257-68. [PMID: 22890904 PMCID: PMC3606520 DOI: 10.1007/s10719-012-9436-8] [Citation(s) in RCA: 10] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/31/2012] [Revised: 07/18/2012] [Accepted: 07/20/2012] [Indexed: 01/03/2023]
Abstract
Brine shrimp are primitive crustacean arthropodal model organisms, second to daphnia, which can survive in high-salinity environments. Their oviposited cysts, cuticle-covered diapausing eggs, are highly resistant to dryness. To elucidate specialties of brine shrimp, this study characterized glycosphingolipids, which are signal transduction-associated material. A group of novel and complex fucosyl glycosphingolipids were separated and identified from cysts of the brine shrimp Artemia franciscana by repeated lipid extraction, alkaline methanolysis, acid treatment, successive column chromatography, and post-source decay measurements by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. Structures of the glycosphingolipids were elucidated by conventional structural characterization and mass spectrometry, and the compounds were identified as GlcNAcβ1-3GalNAcβ1-4(GlcNAcα1-2Fucα1-3)GlcNAcβ1-3Manβ1-4Glcβ1-Cer, GalNAcβ1-4(Fucα1-3)GlcNAcβ1-3GalNAcβ1-4(GlcNAcα1-2Fucα1-3)GlcNAcβ1-3Manβ1-4Glcβ1-Cer, and GalNAcβ1-4(GlcNAcα1-2Fucα1-3)GlcNAcβ1-3GalNAcβ1-4(GlcNAcα1-2Fucα1-3)GlcNAcβ1-3Manβ1-4Glcβ1-Cer. These compounds also contained a branching, non-arthro-series disaccharide with an α-GlcNAc terminus, similar to that found in a previously reported ceramide hexasaccharide (III3(GlcNAcα2Fucα)-At4Cer). The glycans within these complex GSLs are longer than reported glycans of the animal kingdom containing α-GlcNAc terminus. These complex GSLs as well as the longest GSL with ten sugar residues, ceramide decasaccharide (CDeS), contain the fucosylated LacdiNAc sequence reported to associate with parasitism/immunosuppression and the α-GlcNAc terminus reported to show a certain antibacterial effect in other reports. CDeS, the longest GSL of this species, was found in the highest amount, which indicates that CDeS may be functionally important.
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Cholleti SR, Agravat S, Morris T, Saltz JH, Song X, Cummings RD, Smith DF. Automated motif discovery from glycan array data. OMICS-A JOURNAL OF INTEGRATIVE BIOLOGY 2012; 16:497-512. [PMID: 22877213 DOI: 10.1089/omi.2012.0013] [Citation(s) in RCA: 35] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 02/02/2023]
Abstract
Assessing interactions of a glycan-binding protein (GBP) or lectin with glycans on a microarray generates large datasets, making it difficult to identify a glycan structural motif or determinant associated with the highest apparent binding strength of the GBP. We have developed a computational method, termed GlycanMotifMiner, that uses the relative binding of a GBP with glycans within a glycan microarray to automatically reveal the glycan structural motifs recognized by a GBP. We implemented the software with a web-based graphical interface for users to explore and visualize the discovered motifs. The utility of GlycanMotifMiner was determined using five plant lectins, SNA, HPA, PNA, Con A, and UEA-I. Data from the analyses of the lectins at different protein concentrations were processed to rank the glycans based on their relative binding strengths. The motifs, defined as glycan substructures that exist in a large number of the bound glycans and few non-bound glycans, were then discovered by our algorithm and displayed in a web-based graphical user interface ( http://glycanmotifminer.emory.edu ). The information is used in defining the glycan-binding specificity of GBPs. The results were compared to the known glycan specificities of these lectins generated by manual methods. A more complex analysis was also carried out using glycan microarray data obtained for a recombinant form of human galectin-8. Results for all of these lectins show that GlycanMotifMiner identified the major motifs known in the literature along with some unexpected novel binding motifs.
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Affiliation(s)
- Sharath R Cholleti
- Center for Comprehensive Informatics, Emory University, Atlanta, Georgia, USA
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Ficko-Blean E, Stuart CP, Suits MD, Cid M, Tessier M, Woods RJ, Boraston AB. Carbohydrate recognition by an architecturally complex α-N-acetylglucosaminidase from Clostridium perfringens. PLoS One 2012; 7:e33524. [PMID: 22479408 PMCID: PMC3313936 DOI: 10.1371/journal.pone.0033524] [Citation(s) in RCA: 42] [Impact Index Per Article: 3.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/08/2011] [Accepted: 02/14/2012] [Indexed: 11/19/2022] Open
Abstract
CpGH89 is a large multimodular enzyme produced by the human and animal pathogen Clostridium perfringens. The catalytic activity of this exo-α-D-N-acetylglucosaminidase is directed towards a rare carbohydrate motif, N-acetyl-β-D-glucosamine-α-1,4-D-galactose, which is displayed on the class III mucins deep within the gastric mucosa. In addition to the family 89 glycoside hydrolase catalytic module this enzyme has six modules that share sequence similarity to the family 32 carbohydrate-binding modules (CBM32s), suggesting the enzyme has considerable capacity to adhere to carbohydrates. Here we suggest that two of the modules, CBM32-1 and CBM32-6, are not functional as carbohydrate-binding modules (CBMs) and demonstrate that three of the CBMs, CBM32-3, CBM32-4, and CBM32-5, are indeed capable of binding carbohydrates. CBM32-3 and CBM32-4 have a novel binding specificity for N-acetyl-β-D-glucosamine-α-1,4-D-galactose, which thus complements the specificity of the catalytic module. The X-ray crystal structure of CBM32-4 in complex with this disaccharide reveals a mode of recognition that is based primarily on accommodation of the unique bent shape of this sugar. In contrast, as revealed by a series of X-ray crystal structures and quantitative binding studies, CBM32-5 displays the structural and functional features of galactose binding that is commonly associated with CBM family 32. The functional CBM32s that CpGH89 contains suggest the possibility for multivalent binding events and the partitioning of this enzyme to highly specific regions within the gastrointestinal tract.
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Affiliation(s)
- Elizabeth Ficko-Blean
- Biochemistry and Microbiology, University of Victoria, Victoria, British Columbia, Canada
| | - Christopher P. Stuart
- Biochemistry and Microbiology, University of Victoria, Victoria, British Columbia, Canada
| | - Michael D. Suits
- Biochemistry and Microbiology, University of Victoria, Victoria, British Columbia, Canada
| | - Melissa Cid
- Biochemistry and Microbiology, University of Victoria, Victoria, British Columbia, Canada
| | - Matthew Tessier
- Complex Carbohydrate Research Center, University of Georgia, Athens, Georgia, United States of America
| | - Robert J. Woods
- Complex Carbohydrate Research Center, University of Georgia, Athens, Georgia, United States of America
- School of Chemistry, National University of Ireland, Galway, Ireland
| | - Alisdair B. Boraston
- Biochemistry and Microbiology, University of Victoria, Victoria, British Columbia, Canada
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Karasawa F, Shiota A, Goso Y, Kobayashi M, Sato Y, Masumoto J, Fujiwara M, Yokosawa S, Muraki T, Miyagawa S, Ueda M, Fukuda MN, Fukuda M, Ishihara K, Nakayama J. Essential role of gastric gland mucin in preventing gastric cancer in mice. J Clin Invest 2012; 122:923-34. [PMID: 22307328 DOI: 10.1172/jci59087] [Citation(s) in RCA: 83] [Impact Index Per Article: 6.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/19/2011] [Accepted: 12/21/2011] [Indexed: 02/06/2023] Open
Abstract
Gastric gland mucin secreted from the lower portion of the gastric mucosa contains unique O-linked oligosaccharides (O-glycans) having terminal α1,4-linked N-acetylglucosamine residues (αGlcNAc). Previously, we identified human α1,4-N-acetylglucosaminyltransferase (α4GnT), which is responsible for the O-glycan biosynthesis and characterized αGlcNAc function in suppressing Helicobacter pylori in vitro. In the present study, we engineered A4gnt(-/-) mice to better understand its role in vivo. A4gnt(-/-) mice showed complete lack of αGlcNAc expression in gastric gland mucin. Surprisingly, all the mutant mice developed gastric adenocarcinoma through a hyperplasia-dysplasia-carcinoma sequence in the absence of H. pylori infection. Microarray and quantitative RT-PCR analysis revealed upregulation of genes encoding inflammatory chemokine ligands, proinflammatory cytokines, and growth factors, such as Ccl2, Il-11, and Hgf in the gastric mucosa of A4gnt(-/-) mice. Further supporting an important role for this O-glycan in cancer progression, we also observed significantly reduced αGlcNAc in human gastric adenocarcinoma and adenoma. Our results demonstrate that the absence of αGlcNAc triggers gastric tumorigenesis through inflammation-associated pathways in vivo. Thus, αGlcNAc-terminated gastric mucin plays dual roles in preventing gastric cancer by inhibiting H. pylori infection and also suppressing tumor-promoting inflammation.
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Affiliation(s)
- Fumitoshi Karasawa
- Department of Molecular Pathology, Shinshu University Graduate School of Medicine, Matsumoto, Japan
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Kojima H, Shimizu T, Sugita M, Itonori S, Fujita N, Ito M. Biochemical studies on sphingolipids of Artemia franciscana: novel neutral glycosphingolipids. J Lipid Res 2011; 52:308-17. [PMID: 21062954 PMCID: PMC3023551 DOI: 10.1194/jlr.m010173] [Citation(s) in RCA: 10] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/30/2010] [Revised: 10/15/2010] [Indexed: 12/27/2022] Open
Abstract
Neutral glycosphingolipids containing one to six sugars in their oligosaccharide chains have been isolated from cysts of the brine shrimp Artemia franciscana. The structures of these glycolipids were identified by methylation analysis, partial acid hydrolysis, gas-liquid chromatography, combined gas-liquid chromatography-mass spectrometry, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry, and proton nuclear magnetic resonance spectroscopy to be Glcβ1-Cer, Manβ1-4Glcβ1-Cer, Fucα1-3Manβ1-4Glcβ1-Cer, GlcNAcβ1-3Manβ1-4Glcβ1-Cer, GlcNAcα1-2Fucα1-3Manβ1-4Glcβ1-Cer, GalNAcβ1-4GlcNAcβ1-3Manβ1-4Glcβ1-Cer, GalNAcβ1-4(Fucα1-3)GlcNAcβ1-3Manβ1-4Glcβ1-Cer (CPS), and GalNAcβ1-4(GlcNAcα1-2Fucα1-3)GlcNAcβ1-3Manβ1-4Glcβ1-Cer (CHS). Two glycosphingolipids, CPS and CHS, were characterized as novel structures. Because Artemia contains a certain series of glycosphingolipids (-Fucα3Manβ4GlcβCer), which differ from the core sugar sequences reported thus far, we tentatively designated the glycosphingolipids characterized as nonarthro-series ones. Furthermore, CHS exhibited a hybrid structure of arthro-series and nonarthro-series sugar chain. Two novel glycosphingolipids were characterized from the brine shrimp Artemia franciscana; one was composed of arthrotetraose and a branching fucose attached to N-acetylglucosamine residue, and the other was composed of CPS with an additional N-acetylglucosamine residue attached to the branching fucose.
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Affiliation(s)
- Hisao Kojima
- Department of Bioinformatics, Faculty of Pharmacy, Ritsumeikan University, 1-1-1 Nojihigashi, Kusatsu, Shiga 525-8577, Japan
- Department of Chemistry, Faculty of Liberal Arts and Education, Shiga University, 2-5-1 Hiratsu, Otsu, Shiga 520-0862, Japan
| | - Takemasa Shimizu
- Department of Chemistry, Faculty of Liberal Arts and Education, Shiga University, 2-5-1 Hiratsu, Otsu, Shiga 520-0862, Japan
| | - Mutsumi Sugita
- Department of Chemistry, Faculty of Liberal Arts and Education, Shiga University, 2-5-1 Hiratsu, Otsu, Shiga 520-0862, Japan
| | - Saki Itonori
- Department of Chemistry, Faculty of Liberal Arts and Education, Shiga University, 2-5-1 Hiratsu, Otsu, Shiga 520-0862, Japan
| | - Norihisa Fujita
- Institute of Science and Engineering, and Laboratory of Pharmcoinformatics, Faculty of Pharmacy, Ritsumeikan University, 1-1-1 Nojihigashi, Kusatsu, Shiga 525-8577, Japan
| | - Masahiro Ito
- Department of Bioinformatics, Faculty of Pharmacy, Ritsumeikan University, 1-1-1 Nojihigashi, Kusatsu, Shiga 525-8577, Japan
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Ismail MN, Stone EL, Panico M, Lee SH, Luu Y, Ramirez K, Ho SB, Fukuda M, Marth JD, Haslam SM, Dell A. High-sensitivity O-glycomic analysis of mice deficient in core 2 {beta}1,6-N-acetylglucosaminyltransferases. Glycobiology 2011; 21:82-98. [PMID: 20855471 PMCID: PMC2998984 DOI: 10.1093/glycob/cwq134] [Citation(s) in RCA: 38] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/16/2010] [Revised: 08/25/2010] [Accepted: 08/26/2010] [Indexed: 11/14/2022] Open
Abstract
Core 2 β1,6-N-acetylglucosaminyltransferase (C2GnT), which exists in three isoforms, C2GnT1, C2GnT2 and C2GnT3, is one of the key enzymes in the O-glycan biosynthetic pathway. These isoenzymes produce core 2 O-glycans and have been correlated with the biosynthesis of core 4 O-glycans and I-branches. Previously, we have reported mice with single and multiple deficiencies of C2GnT isoenzyme(s) and have evaluated the biological and structural consequences of the loss of core 2 function. We now present more comprehensive O-glycomic analyses of neutral and sialylated glycans expressed in the colon, small intestine, stomach, kidney, thyroid/trachea and thymus of wild-type, C2GnT2 and C2GnT3 single knockouts and the C2GnT1-3 triple knockout mice. Very high-quality data have emerged from our mass spectrometry techniques with the capability of detecting O-glycans up to at least 3500 Da. We were able to unambiguously elucidate the types of O-glycan core, branching location and residue linkages, which allowed us to exhaustively characterize structural changes in the knockout tissues. The C2GnT2 knockout mice suffered a major loss of core 2 O-glycans as well as glycans with I-branches on core 1 antennae especially in the stomach and the colon. In contrast, core 2 O-glycans still dominated the O-glycomic profile of most tissues in the C2GnT3 knockout mice. Analysis of the C2GnT triple knockout mice revealed a complete loss of both core 2 O-glycans and branched core 1 antennae, confirming that the three known isoenzymes are entirely responsible for producing these structures. Unexpectedly, O-linked mannosyl glycans are upregulated in the triple deficient stomach. In addition, our studies have revealed an interesting terminal structure detected on O-glycans of the colon tissues that is similar to the RM2 antigen from glycolipids.
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Affiliation(s)
- Mohd Nazri Ismail
- Division of Molecular Biosciences, Faculty of Natural Sciences, Imperial College London, London SW7 2AZ, UK
| | - Erica L Stone
- Department of Cellular and Molecular Medicine, University of California-San Diego, La Jolla, CA 92093, USA
| | - Maria Panico
- Division of Molecular Biosciences, Faculty of Natural Sciences, Imperial College London, London SW7 2AZ, UK
| | - Seung Ho Lee
- Burnham Institute for Medical Research, La Jolla, CA 92037, USA
| | - Ying Luu
- Department of Medicine, VA San Diego Healthcare System and the University of California, San Diego, CA 92161, USA
| | - Kevin Ramirez
- Department of Cellular and Molecular Medicine, University of California-San Diego, La Jolla, CA 92093, USA
| | - Samuel B Ho
- Department of Medicine, VA San Diego Healthcare System and the University of California, San Diego, CA 92161, USA
| | - Minoru Fukuda
- Burnham Institute for Medical Research, La Jolla, CA 92037, USA
| | - Jamey D Marth
- Center for Nanomedicine, Sanford-Burnham Medical Research Institute, University of California-Santa Barbara, 2324 Life Sciences Building-9625, Santa Barbara, CA 93106-9625, USA
| | - Stuart M Haslam
- Division of Molecular Biosciences, Faculty of Natural Sciences, Imperial College London, London SW7 2AZ, UK
| | - Anne Dell
- Division of Molecular Biosciences, Faculty of Natural Sciences, Imperial College London, London SW7 2AZ, UK
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Fujita M, Tsuchida A, Hirata A, Kobayashi N, Goto K, Osumi K, Hirose Y, Nakayama J, Yamanoi T, Ashida H, Mizuno M. Glycoside hydrolase family 89 alpha-N-acetylglucosaminidase from Clostridium perfringens specifically acts on GlcNAc alpha1,4Gal beta1R at the non-reducing terminus of O-glycans in gastric mucin. J Biol Chem 2010; 286:6479-89. [PMID: 21177247 DOI: 10.1074/jbc.m110.206722] [Citation(s) in RCA: 26] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/30/2023] Open
Abstract
In mammals, α-linked GlcNAc is primarily found in heparan sulfate/heparin and gastric gland mucous cell type mucin. α-N-acetylglucosaminidases (αGNases) belonging to glycoside hydrolase family 89 are widely distributed from bacteria to higher eukaryotes. Human lysosomal αGNase is well known to degrade heparin and heparan sulfate. Here, we reveal the substrate specificity of αGNase (AgnC) from Clostridium perfringens strain 13, a bacterial homolog of human αGNase, by chemically synthesizing a series of disaccharide substrates containing α-linked GlcNAc. AgnC was found to release GlcNAc from GlcNAcα1,4Galβ1pMP and GlcNAcα1pNP substrates (where pMP and pNP represent p-methoxyphenyl and p-nitrophenyl, respectively). AgnC also released GlcNAc from porcine gastric mucin and cell surface mucin. Because AgnC showed no activity against any of the GlcNAcα1,2Galβ1pMP, GlcNAcα1,3Galβ1pMP, GlcNAcα1,6Galβ1pMP, and GlcNAcα1,4GlcAβ1pMP substrates, this enzyme may represent a specific glycosidase required for degrading α-GlcNAc-capped O-glycans of the class III mucin secreted from the stomach and duodenum. Deletion of the C-terminal region containing several carbohydrate-binding module 32 (CBM32) domains significantly reduced the activity for porcine gastric mucin; however, activity against GlcNAcα1,4Galβ1pMP was markedly enhanced. Dot blot and ELISA analyses revealed that the deletion construct containing the C-terminal CBM-C2 to CBM-C6 domains binds strongly to porcine gastric mucin. Consequently, tandem CBM32 domains located near the C terminus of AgnC should function by increasing the affinity for branched or clustered α-GlcNAc-containing glycans. The agnC gene-disrupted strain showed significantly reduced growth on the class III mucin-containing medium compared with the wild type strain, suggesting that AgnC might have an important role in dominant growth in intestines.
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Affiliation(s)
- Masaya Fujita
- Noguchi Institute, 1-8-1 Kaga, Itabashi, Tokyo 173-0003, Japan.
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Reis CA, Osorio H, Silva L, Gomes C, David L. Alterations in glycosylation as biomarkers for cancer detection. J Clin Pathol 2010; 63:322-9. [PMID: 20354203 DOI: 10.1136/jcp.2009.071035] [Citation(s) in RCA: 334] [Impact Index Per Article: 22.3] [Reference Citation Analysis] [Abstract] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/15/2023]
Abstract
Glycoconjugates constitute a major class of biomolecules which include glycoproteins, glycosphingolipids and proteoglycans. Glycans are involved in several physiological and pathological conditions, such as host-pathogen interactions, cell differentiation, migration, tumour invasion and metastisation, cell trafficking and signalling. Cancer is associated with glycosylation alterations in glycoproteins and glycolipids. This review describes various aspects of protein glycosylation with the focus on alterations associated with human cancer. The application of these glycosylation modifications as biomarkers for cancer detection in tumour tissues and serological assays is summarised.
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Affiliation(s)
- Celso A Reis
- Institute of Molecular Pathology and Immunology of the University of Porto - IPATIMUP, Porto 4200-465, Portugal.
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Zheng Z, Jia Y, Hou L, Persson C, Yeager M, Lissowska J, Chanock SJ, Blaser M, Chow WH, Ye W. Genetic variation in a4GnT in relation to Helicobacter pylori serology and gastric cancer risk. Helicobacter 2009; 14:120-5. [PMID: 19751437 PMCID: PMC3008782 DOI: 10.1111/j.1523-5378.2009.00708.x] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/09/2022]
Abstract
BACKGROUND Helicobacter pylori, a known risk factor of gastric cancer, rarely colonize the deeper portion of normal gastric glands, where the mucus is rich in alpha-1,4-linked N-acetylglucosamine capped O-glycans, that strongly inhibit H. pylori growth in vitro. MATERIALS AND METHODS We investigated the association between genetic variation in the O-glycan transferase encoding gene (a4GnT) and H. pylori infection and gastric cancer risk using a Polish population-based case-control study (273 gastric cancer patients and 377 controls). RESULTS A haplotype at the rs2622694-rs397266 locus was associated with H. pylori infection, with the A-A haplotype associated with a higher risk compared with the most frequent G-G haplotype (odds ratio 2.30; 95% confidence interval 1.35-3.92). The association remained significant after correction for multiple tests (global p value: nominal 0.002, empirical 0.045). Neither this haplotype nor the tagSNPs were associated with overall gastric cancer risk. CONCLUSION a4GnT genetic variation may be relevant to H. pylori infection, but not to gastric cancer risk.
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Affiliation(s)
- Zongli Zheng
- Department of Medical Epidemiology and Biostatistics, Karolinska Institutet, Stockholm, Sweden
| | - Yanbin Jia
- Department of Medical Epidemiology and Biostatistics, Karolinska Institutet, Stockholm, Sweden, Department of Basic Medicine, Baotou Medical College, Baotou, China
| | - Lifang Hou
- Division of Cancer Epidemiology and Genetics, National Cancer Institute, Bethesda, MD
| | - Christina Persson
- Department of Medical Epidemiology and Biostatistics, Karolinska Institutet, Stockholm, Sweden
| | - Meredith Yeager
- Core Genotyping Facility, Advanced Technology Center, National Cancer Institute, Gaithersburg, MD
| | - Jolanta Lissowska
- Department of Cancer Epidemiology and Prevention, The M. Sklodowska-Curie Memorial Cancer Center and Institute of Oncology, Warsaw, Poland
| | - Stephen J. Chanock
- Core Genotyping Facility, Advanced Technology Center, National Cancer Institute, Gaithersburg, MD
| | - Martin Blaser
- Department of Medicine, New York University School of Medicine, New York, NY
| | - Wong-Ho Chow
- Division of Cancer Epidemiology and Genetics, National Cancer Institute, Bethesda, MD
| | - Weimin Ye
- Department of Medical Epidemiology and Biostatistics, Karolinska Institutet, Stockholm, Sweden,Correspondence to: Weimin Ye Department of Medical Epidemiology and Biostatistics Karolinska Institutet Box 281 SE-17177 Stockholm Sweden
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Nakajima M, Kato T, Kanamasa S, Park EY. Molecular chaperone-assisted production of human alpha-1,4-N-acetylglucosaminyltransferase in silkworm larvae using recombinant BmNPV bacmids. Mol Biotechnol 2009; 43:67-75. [PMID: 19418270 DOI: 10.1007/s12033-009-9174-8] [Citation(s) in RCA: 10] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/04/2009] [Accepted: 04/15/2009] [Indexed: 02/05/2023]
Abstract
In this study, human alpha-1,4-N-acetylglucosaminyltransferase (alpha4GnT) fused with GFP(uv) (GFP(uv)-alpha4GnT) was expressed using both a transformed cell system and silkworm larvae. A Tn-pXgp-GFP(uv)-alpha4GnT cell line, isolated after expression vector transfection, produced 106 mU/ml of alpha4GnT activity in suspension culture. When Bombyx mori nucleopolyhedrovirus containing a GFP(uv)-alpha4GnT fusion gene (BmNPV-CP (-)/GFP(uv)-alpha4GnT) bacmid was injected into silkworm larvae, alpha4GnT activity in larval hemolymph was 352 mU/ml, which was 3.3-fold higher than that of the Tn-pXgp-GFP(uv)-alpha4GnT cell line. With human calnexin (CNX) or human immunoglobulin heavy chain-binding protein (BiP, GRP78) coexpressed under the control of the ie-2 promoter, alpha4GnT activity in larval hemolymph increased by 1.4-2.0-fold. Moreover, when BmNPV-CP (-)/GFP(uv)-alpha4GnT bacmid injection was delayed for 3 h after BmNPV-CP (-)/CNX injection, the alpha4GnT activity increased significantly to 922 mU/ml, which was 8.7-fold higher than that of the Tn-pXgp-GFP(uv)-alpha4GnT cell line. Molecular chaperone assisted-expression in silkworm larvae using the BmNPV bacmid is a promising tool for recombinant protein production. This system could lead to large-scale production of more complex recombinant proteins.
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Affiliation(s)
- Makoto Nakajima
- Department of Applied Biological Chemistry, Faculty of Agriculture, Shizuoka University, 836 Ohya, Suruga-ku, Shizuoka 422-8529, Japan
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Kobayashi M, Lee H, Nakayama J, Fukuda M. Roles of gastric mucin-type O-glycans in the pathogenesis of Helicobacter pylori infection. Glycobiology 2009; 19:453-61. [PMID: 19150806 DOI: 10.1093/glycob/cwp004] [Citation(s) in RCA: 47] [Impact Index Per Article: 2.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/16/2022] Open
Abstract
Helicobacter pylori is a Gram-negative bacterium that infects over 50% of the world's population. This organism causes various gastric diseases such as chronic gastritis, peptic ulcer, and gastric cancer. H. pylori possesses lipopolysaccharides that share structural similarity to Lewis blood group antigens in gastric mucosa. Such antigenic mimicry could result in immune tolerance against antigens of this pathogen. On the other hand, H. pylori colonizes gastric mucosa by utilizing adhesins that bind Lewis blood group antigen-related carbohydrates expressed on gastric epithelial cells. After colonization, H. pylori induces acute inflammatory responses mainly by neutrophils. This acute phase is gradually replaced by a chronic inflammatory response. In chronic gastritis, lymphocytes infiltrate the lamina propria, and such infiltration is facilitated by the interaction between L-selectin on lymphocytes and peripheral lymph node addressin (PNAd), which contains 6-sulfo sialyl Lewis X-capped O-glycans, on high endothelial venule (HEV)-like vessels. H. pylori barely colonizes gland mucous cell-derived mucin where alpha1,4-GlcNAc-capped O-glycans exist. In vitro experiments show that alpha1,4-GlcNAc-capped O-glycans function as a natural antibiotic to inhibit H. pylori growth. These findings show that distinct sets of carbohydrates expressed in the stomach are closely associated with pathogenesis and prevention of H. pylori-related diseases, providing therapeutic potentialities based on specific carbohydrate modulation.
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Affiliation(s)
- Motohiro Kobayashi
- Department of Molecular Pathology, Shinshu University Graduate School of Medicine, Matsumoto 390-8621, Japan
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Kobayashi M, Lee H, Nakayama J, Fukuda M. Carbohydrate-dependent defense mechanisms against Helicobacter pylori infection. Curr Drug Metab 2009; 10:29-40. [PMID: 19149511 PMCID: PMC2666621 DOI: 10.2174/138920009787048428] [Citation(s) in RCA: 16] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/07/2023]
Abstract
Helicobacter pylori is a Gram-negative bacterium that infects over 50% of the world's population. This organism causes various gastric diseases such as chronic gastritis, peptic ulcer, and gastric cancer. H. pylori possesses lipopolysaccharide, which shares structural similarity to Lewis blood group antigens in gastric mucosa. Such antigenic mimicry could result in immune tolerance against antigens of this pathogen. On the other hand, H. pylori colonize gastric mucosa by utilizing adhesins, which bind Lewis blood group antigen-related carbohydrates expressed on gastric epithelial cells. In chronic gastritis, lymphocytes infiltrate the lamina propria, and such infiltration is facilitated by 6-sulfo sialyl Lewis X-capped O-glycans, peripheral lymph node addressin (PNAd), on high endothelial venule (HEV)-like vessels. The number of HEV-like vessels increases as chronic inflammation progresses. Furthermore, PNAd formed on HEV-like vessels disappear once H. pylori is eradicated. These results indicate that PNAd plays an important role in H. pylori-associated inflammation. H. pylori barely colonizes gland mucous cell-derived mucin where alpha1,4-GlcNAc-capped O-glycans exist. In vitro experiments show that alpha1,4-GlcNAc-capped O-glycans function as a natural antibiotic to inhibit H. pylori growth. We recently identified cholesterol alpha-glucosyltransferase (CHLalphaGcT) using an expression cloning strategy and showed that this enzyme is specifically inhibited by mucin-type O-glycans like those present in deeper portions of the gastric mucosa. These findings show that a battery of carbohydrates expressed in the stomach is closely associated with pathogenesis and also prevention of H. pylori-related diseases.
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Affiliation(s)
- Motohiro Kobayashi
- Department of Molecular Pathology, Shinshu University Graduate School of Medicine, Matsumoto, Japan.
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Lee H, Wang P, Hoshino H, Ito Y, Kobayashi M, Nakayama J, Seeberger PH, Fukuda M. Alpha1,4GlcNAc-capped mucin-type O-glycan inhibits cholesterol alpha-glucosyltransferase from Helicobacter pylori and suppresses H. pylori growth. Glycobiology 2008; 18:549-58. [PMID: 18458030 PMCID: PMC2758100 DOI: 10.1093/glycob/cwn037] [Citation(s) in RCA: 51] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/29/2008] [Revised: 04/23/2008] [Accepted: 04/29/2008] [Indexed: 02/07/2023] Open
Abstract
Helicobacter pylori infects over half of the world's population and is thought to be a leading cause of gastric ulcer, gastric carcinoma, and gastric malignant lymphoma of mucosa-associated lymphoid tissue type. Previously, we reported that a gland mucin (MUC6) present in the lower portion of the gastric mucosa containing alpha1,4-N-acetylglucosamine (alpha1,4GlcNAc)-capped core 2-branched O-glycans suppresses H. pylori growth by inhibiting the synthesis of alpha-glucosyl cholesterol, a major constituent of the H. pylori cell wall (Kawakubo et al. 2004. Science. 305:1003-1006). Therefore, we cloned the genomic DNA encoding cholesterol alpha-glucosyltransferase (HP0421) and expressed its soluble form in Escherichia coli. Using this soluble HP0421, we show herein that HP0421 sequentially acts on uridine diphosphoglucose and cholesterol in an ordered Bi-Bi manner. We found that competitive inhibition of HP0421 by alpha1,4GlcNAc-capped core 2-branched O-glycan is much more efficient than noncompetitive inhibition by newly synthesized alpha-glucosyl cholesterol. Utilizing synthetic oligosaccharides, alpha-glucosyl cholesterol, and monosaccharides, we found that alpha1,4GlcNAc-capped core 2-branched O-glycan most efficiently inhibits H. pylori growth. These findings together indicate that alpha1,4GlcNAc-capped O-glycans suppress H. pylori growth by inhibiting HP0421, and that alpha1,4GlcNAc-capped core 2 O-glycans may be useful to treat patients infected with H. pylori.
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Affiliation(s)
- Heeseob Lee
- Tumor Microenvironment Program, Glycobiology Unit, Cancer Center, Burnham Institute for Medical Research, La Jolla, CA 92037, USA
| | - Ping Wang
- Tumor Microenvironment Program, Glycobiology Unit, Cancer Center, Burnham Institute for Medical Research, La Jolla, CA 92037, USA
| | - Hitomi Hoshino
- Department of Pathology, Shinshu University School of Medicine, Matsumoto 390-8621, Japan
| | - Yuki Ito
- Tumor Microenvironment Program, Glycobiology Unit, Cancer Center, Burnham Institute for Medical Research, La Jolla, CA 92037, USA
| | - Motohiro Kobayashi
- Department of Pathology, Shinshu University School of Medicine, Matsumoto 390-8621, Japan
| | - Jun Nakayama
- Department of Pathology, Shinshu University School of Medicine, Matsumoto 390-8621, Japan
| | - Peter H Seeberger
- Tumor Microenvironment Program, Glycobiology Unit, Cancer Center, Burnham Institute for Medical Research, La Jolla, CA 92037, USA
| | - Minoru Fukuda
- Tumor Microenvironment Program, Glycobiology Unit, Cancer Center, Burnham Institute for Medical Research, La Jolla, CA 92037, USA
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