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1
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Holail J, Sukkarieh HH, Aljada A. Expanding the Role of Heparin Derivatives in Oncology: From Anticoagulation to Antitumor Activity. Pharmaceuticals (Basel) 2025; 18:396. [PMID: 40143176 PMCID: PMC11944584 DOI: 10.3390/ph18030396] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/13/2025] [Revised: 03/07/2025] [Accepted: 03/09/2025] [Indexed: 03/28/2025] Open
Abstract
Current research demonstrates the expanding therapeutic potential of heparin derivatives in oncology, extending beyond traditional anticoagulation mechanisms. This systematic analysis examines the structural characteristics, molecular mechanisms, and therapeutic applications of heparin-based compounds in malignancy treatment. The essential antithrombin binding pentasaccharide sequence has enabled development of specialized molecular variants, particularly fractionated heparins and their non-anticoagulant counterparts. These agents exert antineoplastic effects via multiple pathways, particularly through modulation of heparanase enzymatic activity and specific protein-glycosaminoglycan interactions. Evidence from pivotal clinical trials (FRAGMATIC, MAGNOLIA, GASTRANOX) confirms efficacy in managing cancer-associated thrombosis while indicating potential enhancement of chemotherapeutic outcomes. The preparation methods utilize enzymatic cleavage reactions and selective chemical derivatization to generate structurally modified heparins exhibiting unique molecular characteristics and biological activities. Analysis of the glycosaminoglycan analog dociparstat sodium reveals significant activity in myeloid malignancies, mediated by specific interference with CXCL12/CXCR4 signaling cascades. Significant challenges remain in manufacturing scale-up, analytical validation, and long-term safety assessment. Future studies must address dose optimization, combination strategies, and controlled clinical trials to determine the full therapeutic potential of these compounds in clinical oncology.
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Affiliation(s)
- Jasmine Holail
- Department of Biochemistry and Molecular Medicine, College of Medicine, Alfaisal University, Riyadh 11533, Saudi Arabia;
| | - Hatouf Husni Sukkarieh
- Department of Pharmacology, College of Medicine, Alfaisal University, Riyadh 11533, Saudi Arabia;
| | - Ahmad Aljada
- Department of Biochemistry and Molecular Medicine, College of Medicine, Alfaisal University, Riyadh 11533, Saudi Arabia;
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2
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Le Pennec J, Picart C, Vivès RR, Migliorini E. Sweet but Challenging: Tackling the Complexity of GAGs with Engineered Tailor-Made Biomaterials. ADVANCED MATERIALS (DEERFIELD BEACH, FLA.) 2024; 36:e2312154. [PMID: 38011916 DOI: 10.1002/adma.202312154] [Citation(s) in RCA: 3] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Received: 11/14/2023] [Indexed: 11/29/2023]
Abstract
Glycosaminoglycans (GAGs) play a crucial role in tissue homeostasis by regulating the activity and diffusion of bioactive molecules. Incorporating GAGs into biomaterials has emerged as a widely adopted strategy in medical applications, owing to their biocompatibility and ability to control the release of bioactive molecules. Nevertheless, immobilized GAGs on biomaterials can elicit distinct cellular responses compared to their soluble forms, underscoring the need to understand the interactions between GAG and bioactive molecules within engineered functional biomaterials. By controlling critical parameters such as GAG type, density, and sulfation, it becomes possible to precisely delineate GAG functions within a biomaterial context and to better mimic specific tissue properties, enabling tailored design of GAG-based biomaterials for specific medical applications. However, this requires access to pure and well-characterized GAG compounds, which remains challenging. This review focuses on different strategies for producing well-defined GAGs and explores high-throughput approaches employed to investigate GAG-growth factor interactions and to quantify cellular responses on GAG-based biomaterials. These automated methods hold considerable promise for improving the understanding of the diverse functions of GAGs. In perspective, the scientific community is encouraged to adopt a rational approach in designing GAG-based biomaterials, taking into account the in vivo properties of the targeted tissue for medical applications.
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Affiliation(s)
- Jean Le Pennec
- U1292 Biosanté, INSERM, CEA, Univ. Grenoble Alpes, CNRS EMR 5000 Biomimetism and Regenerative Medicine, Grenoble, F-38054, France
| | - Catherine Picart
- U1292 Biosanté, INSERM, CEA, Univ. Grenoble Alpes, CNRS EMR 5000 Biomimetism and Regenerative Medicine, Grenoble, F-38054, France
| | | | - Elisa Migliorini
- U1292 Biosanté, INSERM, CEA, Univ. Grenoble Alpes, CNRS EMR 5000 Biomimetism and Regenerative Medicine, Grenoble, F-38054, France
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3
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Motta JM, Micheli KVA, Roberto-Fernandes C, Hermsdorff-Brandt M, Guedes AL, Frattani FS, Mourão PAS, Pereira MS. A low-anticoagulant heparin suppresses metastatic dissemination through the inhibition of tumor cell-platelets association. Biomed Pharmacother 2024; 171:116108. [PMID: 38218079 DOI: 10.1016/j.biopha.2023.116108] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/25/2023] [Revised: 12/23/2023] [Accepted: 12/28/2023] [Indexed: 01/15/2024] Open
Abstract
Metastasis is the leading cause of cancer-related deaths. Despite this relevance, there is no specific therapy targeting metastasis. The interaction of the tumor cell with platelets, forming microemboli is crucial for successful hematogenous dissemination. Heparin disrupts it by a P-selectin-mediated event. However, its clinical use for this purpose is hindered by the requirement of high doses, leading to anticoagulant-related side effects. In this study, we obtained a low-anticoagulant heparin through the fractionation of a pharmaceutical bovine heparin. This derivative was referred to as LA-hep and we investigated its efficacy in inhibiting metastases and explored its capacity of suppressing the interaction between tumor cells and platelets. Our data revealed that LA-hep is as efficient as porcine unfractionated heparin in attenuating lung metastases from melanoma and colon adenocarcinoma cells in an assay with a single intravenous administration. It also prevents platelet arrest shortly after cell injection in wild-type mice and suppresses melanoma-platelets interaction in vitro. Moreover, LA-hep blocks P-selectin's direct binding to tumor cells and platelet aggregation, providing further evidence for the role of P-selectin as a molecular target. Even in P-selectin-depleted mice which developed a reduced number of metastatic foci, both porcine heparin and LA-hep further inhibited metastasis burden. This suggests evidence of an additional mechanism of antimetastatic action. Therefore, our results indicate a dissociation between the heparin anticoagulant and antimetastatic effects. Considering the simple and highly reproducible methodology used to purify LA-hep along with the data presented here, LA-hep emerges as a promising drug for future use in preventing metastasis in cancer patients.
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Affiliation(s)
- Juliana M Motta
- Programa de Glicobiologia, Instituto de Bioquímica Médica Leopoldo de Meis and Hospital Universitário Clementino Fraga Filho, Universidade Federal do Rio de Janeiro, Rio de Janeiro 21941-913, Brazil
| | - Kayene V A Micheli
- Programa de Glicobiologia, Instituto de Bioquímica Médica Leopoldo de Meis and Hospital Universitário Clementino Fraga Filho, Universidade Federal do Rio de Janeiro, Rio de Janeiro 21941-913, Brazil
| | - Carlos Roberto-Fernandes
- Programa de Glicobiologia, Instituto de Bioquímica Médica Leopoldo de Meis and Hospital Universitário Clementino Fraga Filho, Universidade Federal do Rio de Janeiro, Rio de Janeiro 21941-913, Brazil
| | - Michelle Hermsdorff-Brandt
- Programa de Glicobiologia, Instituto de Bioquímica Médica Leopoldo de Meis and Hospital Universitário Clementino Fraga Filho, Universidade Federal do Rio de Janeiro, Rio de Janeiro 21941-913, Brazil
| | - Alessandra L Guedes
- Instituto de Bioquímica Médica Leopoldo de Meis, Universidade Federal do Rio de Janeiro, Rio de Janeiro, RJ, CEP 21941-902, Brazil
| | - Flávia S Frattani
- Departamento de Análises Clínicas e Toxicológicas, Faculdade de Farmácia, Centro de Ciências da Saúde, Universidade Federal do Rio de Janeiro, Rio de Janeiro 21941-599, Brazil
| | - Paulo A S Mourão
- Programa de Glicobiologia, Instituto de Bioquímica Médica Leopoldo de Meis and Hospital Universitário Clementino Fraga Filho, Universidade Federal do Rio de Janeiro, Rio de Janeiro 21941-913, Brazil
| | - Mariana S Pereira
- Programa de Glicobiologia, Instituto de Bioquímica Médica Leopoldo de Meis and Hospital Universitário Clementino Fraga Filho, Universidade Federal do Rio de Janeiro, Rio de Janeiro 21941-913, Brazil.
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4
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Monitoring the stability of heparin: NMR evidence for the rearrangement of sulfated iduronate in phosphate buffer. Carbohydr Polym 2023; 308:120649. [PMID: 36813341 DOI: 10.1016/j.carbpol.2023.120649] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/19/2022] [Revised: 01/11/2023] [Accepted: 01/30/2023] [Indexed: 02/05/2023]
Abstract
Heparin, a major anticoagulant drug, comprises a complex mixture of motifs. Heparin is isolated from natural sources while being subjected to a variety of conditions but the detailed effects of these on heparin structure have not been studied in depth. Therefore, the result of exposing heparin to a range of buffered environments, ranging pH values from 7 to 12, and temperatures of 40, 60 and 80 °C were examined. There was no evidence of significant N-desulfation or 6-O-desulfation in glucosamine residues, nor of chain scission, however, stereochemical re-arrangement of α-L-iduronate 2-O-sulfate to α-L-galacturonate residues occurred in 0.1 M phosphate buffer at pH 12/80 °C. The results confirm the relative stability of heparin in environments like those during extraction and purification processes; on the other hand, the sensitivity of heparin to pH 12 in buffered solution at high temperature is highlighted, providing an important insight for heparin manufacturers.
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5
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Wang Y, Li T, Li N, Huang C, Xiong X, Xie X, Wu M, Wang L, Jiang J. 6-O-desulfated heparin attenuates myocardial ischemia/reperfusion injury in mice through the regulation of miR-199a-5p/klotho axis. Glycoconj J 2022; 39:747-758. [PMID: 36107266 DOI: 10.1007/s10719-022-10081-9] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/21/2021] [Revised: 08/20/2022] [Accepted: 09/06/2022] [Indexed: 12/14/2022]
Abstract
Heparin has been documented to reduce myocardial injury caused by ischemia/reperfusion (I/R), but its clinical application is limited due to its strong intrinsic anticoagulant property. Some desulfated derivatives of heparin display low anticoagulant activity and may have potential value as therapeutic agents for myocardial I/R injury. In this study, we observed that 6-O-desulfated heparin, a desulfated derivative of heparin, shortened the activated partial thromboplastin time and exhibited lower anticoagulant activity compared with heparin or 2-O-desulfated heparin (another desulfated derivative of heparin). Then, we explored whether 6-O-desulfated heparin could protect against myocardial I/R injury, and elucidated its possible mechanisms. Administration of 6-O-desulfated heparin significantly reduced creatine kinase activity, myocardial infarct size and cell apoptosis in mice subjected to 30 min of myocardial ischemia following 2 h of reperfusion, accompanied by a reverse in miR-199a-5p elevation, klotho downregulation and reactive oxygen species (ROS) accumulation. In cultured H9c2 cells, the mechanism of 6-O-desulfated heparin against myocardial I/R injury was further explored. Consistent with the results in vivo, 6-O-desulfated heparin significantly ameliorated hypoxia/reoxygenation-induced injury, upregulated klotho and decreased miR-199a-5p levels and ROS accumulation, and these effects were reversed by miR-199a-5p mimics. In conclusion, these results suggested that 6-O-desulfated heparin with lower anticoagulant activity attenuated myocardial I/R injury through miR-199a-5p/klotho and ROS signaling. Our study may also indicate that 6-O-desulfated heparin, as an excellent heparin derivative, is a potential therapeutic agent for myocardial I/R injury.
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Affiliation(s)
- Yujie Wang
- Department of Pharmacology, Xiangya School of Pharmaceutical Sciences, Central South University, Changsha, 410078, Hunan, China
- Department of Pharmacy, People's Hospital of Rizhao, Rizhao, 276826, Shandong, China
| | - Ting Li
- Department of Pharmacology, Xiangya School of Pharmaceutical Sciences, Central South University, Changsha, 410078, Hunan, China
| | - Niansheng Li
- Department of Pharmacology, Xiangya School of Pharmaceutical Sciences, Central South University, Changsha, 410078, Hunan, China
| | - Chuyi Huang
- Department of Pharmacology, Xiangya School of Pharmaceutical Sciences, Central South University, Changsha, 410078, Hunan, China
| | - Xiaoming Xiong
- Department of Pharmacology, Xiangya School of Pharmaceutical Sciences, Central South University, Changsha, 410078, Hunan, China
| | - Xu Xie
- Department of Pharmacology, Xiangya School of Pharmaceutical Sciences, Central South University, Changsha, 410078, Hunan, China
| | - Meiting Wu
- Department of Pharmacology, Xiangya School of Pharmaceutical Sciences, Central South University, Changsha, 410078, Hunan, China
| | - Lianchun Wang
- Department of Molecular Pharmacology and Physiology, Morsani School of Medicine, Byrd Alzheimer's Research Institute, University of South Florida, FL, 33613, Tampa, USA
| | - Junlin Jiang
- Department of Pharmacology, Xiangya School of Pharmaceutical Sciences, Central South University, Changsha, 410078, Hunan, China.
- Hunan Provincial Key Laboratory of Cardiovascular Research, Central South University, Changsha, 410078, Hunan, China.
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6
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Chitosan sulfate-lysozyme hybrid hydrogels as platforms with fine-tuned degradability and sustained inherent antibiotic and antioxidant activities. Carbohydr Polym 2022; 291:119611. [DOI: 10.1016/j.carbpol.2022.119611] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/23/2022] [Revised: 05/06/2022] [Accepted: 05/09/2022] [Indexed: 12/14/2022]
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7
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Metabolism of a hybrid algal galactan by members of the human gut microbiome. Nat Chem Biol 2022; 18:501-510. [PMID: 35289327 DOI: 10.1038/s41589-022-00983-y] [Citation(s) in RCA: 18] [Impact Index Per Article: 6.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/28/2021] [Accepted: 01/27/2022] [Indexed: 12/12/2022]
Abstract
Native porphyran is a hybrid of porphryan and agarose. As a common element of edible seaweed, this algal galactan is a frequent component of the human diet. Bacterial members of the human gut microbiota have acquired polysaccharide utilization loci (PULs) that enable the metabolism of porphyran or agarose. However, the molecular mechanisms that underlie the deconstruction and use of native porphyran remains incompletely defined. Here, we have studied two human gut bacteria, porphyranolytic Bacteroides plebeius and agarolytic Bacteroides uniformis, that target native porphyran. This reveals an exo-based cycle of porphyran depolymerization that incorporates a keystone sulfatase. In both PULs this cycle also works together with a PUL-encoded agarose depolymerizing machinery to synergistically reduce native porphyran to monosaccharides. This provides a framework for understanding the deconstruction of a hybrid algal galactan, and insight into the competitive and/or syntrophic relationship of gut microbiota members that target rare nutrients.
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8
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Role of HSPGs in Systemic Bacterial Infections. Methods Mol Biol 2021. [PMID: 34626410 DOI: 10.1007/978-1-0716-1398-6_46] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 08/11/2023]
Abstract
Heparan sulfate proteoglycans (HSPGs) are at the forefront of host-microbe interactions. Cell surface HSPGs are thought to promote infection as attachment and internalization receptors for many bacterial pathogens and as soluble inhibitors of host immunity when released from the cell surface by ectodomain shedding. However, the importance of HSPG-pathogen interactions in vivo has yet to be clearly established. Here we describe several representative methods to study the role of HSPGs in systemic bacterial infections, such as bacteremia and sepsis. The overall experimental strategy is to use mouse models to establish the physiological significance of HSPGs, to determine the identity of HSPGs that specifically promote infection, and to define key structural features of HSPGs that enhance bacterial virulence in systemic infections.
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9
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Chemical Modification of Glycosaminoglycan Polysaccharides. Molecules 2021; 26:molecules26175211. [PMID: 34500644 PMCID: PMC8434129 DOI: 10.3390/molecules26175211] [Citation(s) in RCA: 12] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/20/2021] [Revised: 08/20/2021] [Accepted: 08/21/2021] [Indexed: 12/16/2022] Open
Abstract
The linear anionic class of polysaccharides, glycosaminoglycans (GAGs), are critical throughout the animal kingdom for developmental processes and the maintenance of healthy tissues. They are also of interest as a means of influencing biochemical processes. One member of the GAG family, heparin, is exploited globally as a major anticoagulant pharmaceutical and there is a growing interest in the potential of other GAGs for diverse applications ranging from skin care to the treatment of neurodegenerative conditions, and from the treatment and prevention of microbial infection to biotechnology. To realize the potential of GAGs, however, it is necessary to develop effective tools that are able to exploit the chemical manipulations to which GAGs are susceptible. Here, the current knowledge concerning the chemical modification of GAGs, one of the principal approaches for the study of the structure-function relationships in these molecules, is reviewed. Some additional methods that were applied successfully to the analysis and/or processing of other carbohydrates, but which could be suitable in GAG chemistry, are also discussed.
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10
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Mitra D, Hasan MH, Bates JT, Bierdeman MA, Ederer DR, Parmar RC, Fassero LA, Liang Q, Qiu H, Tiwari V, Zhang F, Linhardt RJ, Sharp JS, Wang L, Tandon R. The degree of polymerization and sulfation patterns in heparan sulfate are critical determinants of cytomegalovirus entry into host cells. PLoS Pathog 2021; 17:e1009803. [PMID: 34352038 PMCID: PMC8384199 DOI: 10.1371/journal.ppat.1009803] [Citation(s) in RCA: 21] [Impact Index Per Article: 5.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/24/2021] [Revised: 08/24/2021] [Accepted: 07/15/2021] [Indexed: 01/10/2023] Open
Abstract
Several enveloped viruses, including herpesviruses attach to host cells by initially interacting with cell surface heparan sulfate (HS) proteoglycans followed by specific coreceptor engagement which culminates in virus-host membrane fusion and virus entry. Interfering with HS-herpesvirus interactions has long been known to result in significant reduction in virus infectivity indicating that HS play important roles in initiating virus entry. In this study, we provide a series of evidence to prove that specific sulfations as well as the degree of polymerization (dp) of HS govern human cytomegalovirus (CMV) binding and infection. First, purified CMV extracellular virions preferentially bind to sulfated longer chain HS on a glycoarray compared to a variety of unsulfated glycosaminoglycans including unsulfated shorter chain HS. Second, the fraction of glycosaminoglycans (GAG) displaying higher dp and sulfation has a larger impact on CMV titers compared to other fractions. Third, cell lines deficient in specific glucosaminyl sulfotransferases produce significantly reduced CMV titers compared to wild-type cells and virus entry is compromised in these mutant cells. Finally, purified glycoprotein B shows strong binding to heparin, and desulfated heparin analogs compete poorly with heparin for gB binding. Taken together, these results highlight the significance of HS chain length and sulfation patterns in CMV attachment and infectivity.
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Affiliation(s)
- Dipanwita Mitra
- Department of Microbiology and Immunology, University of Mississippi Medical Center, Jackson, Mississippi, United States of America
| | - Mohammad H. Hasan
- Department of Microbiology and Immunology, University of Mississippi Medical Center, Jackson, Mississippi, United States of America
| | - John T. Bates
- Department of Microbiology and Immunology, University of Mississippi Medical Center, Jackson, Mississippi, United States of America
- Department of Medicine, University of Mississippi Medical Center, Jackson, Mississippi, United States of America
| | - Michael A. Bierdeman
- Department of Medicine, University of Mississippi Medical Center, Jackson, Mississippi, United States of America
| | - Dallas R. Ederer
- Department of Microbiology and Immunology, University of Mississippi Medical Center, Jackson, Mississippi, United States of America
| | - Rinkuben C. Parmar
- Department of Microbiology and Immunology, University of Mississippi Medical Center, Jackson, Mississippi, United States of America
| | - Lauren A. Fassero
- Department of Microbiology and Immunology, University of Mississippi Medical Center, Jackson, Mississippi, United States of America
| | - Quntao Liang
- Biomolecular Sciences, School of Pharmacy, University of Mississippi, Oxford, Mississippi, United States of America
- College of Biological Science and Engineering, University of Fuzhou, Fujian, China
| | - Hong Qiu
- Complex Carbohydrate Research Center, University of Georgia, Athens, Georgia, United States of America
| | - Vaibhav Tiwari
- Department of Microbiology and Immunology, Midwestern University, Downers Grove, Illinois, United States of America
| | - Fuming Zhang
- Center for Biotechnology and Interdisciplinary Studies, Rensselaer Polytechnic Institute, Troy, New York, United States of America
| | - Robert J. Linhardt
- Center for Biotechnology and Interdisciplinary Studies, Rensselaer Polytechnic Institute, Troy, New York, United States of America
| | - Joshua S. Sharp
- Biomolecular Sciences, School of Pharmacy, University of Mississippi, Oxford, Mississippi, United States of America
| | - Lianchun Wang
- Department of Molecular Pharmacology and Physiology, University of South Florida, Tampa, Florida, United States of America
| | - Ritesh Tandon
- Department of Microbiology and Immunology, University of Mississippi Medical Center, Jackson, Mississippi, United States of America
- Department of Medicine, University of Mississippi Medical Center, Jackson, Mississippi, United States of America
- Biomolecular Sciences, School of Pharmacy, University of Mississippi, Oxford, Mississippi, United States of America
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11
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Heparan sulfate analogues regulate tumor-derived exosome formation that attenuates exosome functions in tumor processes. Int J Biol Macromol 2021; 187:481-491. [PMID: 34298051 DOI: 10.1016/j.ijbiomac.2021.07.110] [Citation(s) in RCA: 11] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [Key Words] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/25/2021] [Revised: 07/03/2021] [Accepted: 07/16/2021] [Indexed: 12/13/2022]
Abstract
Heparan sulfate (HS) is involved in many biological activities, including the biogenesis and uptake of exosomes, which are related to the occurrence and development of tumors. This study investigated the role of HS analogues (heparin, low molecular weight heparin, and 6-O-desulfated heparin) in modulating exosome secretion, composition and functions. Exosomes derived from B16F10 cells exposed to different HS analogues were isolated and characterized by TEM, western blotting and Nanosight analyses. The number, size and protein cargo of exosomes secreted by HS analogues-induced B16F10 cells were detected. The findings indicated the reduced tumor-derived exosome secretion and protein cargo as reflected by lower levels of CD63, TSG101, heparinase and IL-6 in exosomes derived from heparin-induced B16F10 cells as compared with 6-O-desulfated heparin-induced tumor cells. Further functional assays demonstrated that exosomes from tumor cells exposed to heparin weakened tumor proliferation, migration and invasion most significantly among various exosomes derived from B16F10 cells treated with different HS analogues. Moreover, the sulfate group at 6-O position of heparan sulfate has been proved to play an important role in tumor-derived exosome formation and functions. This study suggested a vital view to develop more specific and efficient HS-based strategies in cancer treatment for targeting tumor-derived exosomes.
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12
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De Rossi G, Vähätupa M, Cristante E, Arokiasamy S, Liyanage SE, May U, Pellinen L, Uusitalo-Järvinen H, Bainbridge JW, Järvinen TA, Whiteford JR. Pathological Angiogenesis Requires Syndecan-4 for Efficient VEGFA-Induced VE-Cadherin Internalization. Arterioscler Thromb Vasc Biol 2021; 41:1374-1389. [PMID: 33596666 PMCID: PMC7613699 DOI: 10.1161/atvbaha.121.315941] [Citation(s) in RCA: 14] [Impact Index Per Article: 3.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/18/2022]
Abstract
[Figure: see text].
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Affiliation(s)
- Giulia De Rossi
- William Harvey Research Institute, Barts and The London School of Medicine and Dentistry, Queen Mary University of London, Charterhouse Square, London EC1M 6BQ, United Kingdom
- UCL Institute of Ophthalmology, Department of Cell Biology, 11-43 Bath Street, London EC1V 9EL, UK
| | - Maria Vähätupa
- Faculty of Medicine & Health Technology, Tampere University, 33014 Tampere, Finland & Departments of Orthopedics & Traumatology and Tampere Eye Centre, Tampere University Hospital, 33521 Tampere, Finland
| | - Enrico Cristante
- UCL Institute of Ophthalmology, Genetics department, 11-43 Bath Street, London EC1V 9EL, UK
| | - Samantha Arokiasamy
- William Harvey Research Institute, Barts and The London School of Medicine and Dentistry, Queen Mary University of London, Charterhouse Square, London EC1M 6BQ, United Kingdom
| | - Sidath E. Liyanage
- UCL Institute of Ophthalmology, Genetics department, 11-43 Bath Street, London EC1V 9EL, UK
| | - Ulrike May
- Faculty of Medicine & Health Technology, Tampere University, 33014 Tampere, Finland & Departments of Orthopedics & Traumatology and Tampere Eye Centre, Tampere University Hospital, 33521 Tampere, Finland
| | - Laura Pellinen
- Faculty of Medicine & Health Technology, Tampere University, 33014 Tampere, Finland & Departments of Orthopedics & Traumatology and Tampere Eye Centre, Tampere University Hospital, 33521 Tampere, Finland
| | - Hannele Uusitalo-Järvinen
- Faculty of Medicine & Health Technology, Tampere University, 33014 Tampere, Finland & Departments of Orthopedics & Traumatology and Tampere Eye Centre, Tampere University Hospital, 33521 Tampere, Finland
| | - James W. Bainbridge
- UCL Institute of Ophthalmology, Genetics department, 11-43 Bath Street, London EC1V 9EL, UK
- NIHR Biomedical Research Centre at Moorfields Eye Hospital NHS Foundation Trust, City Road, London EC1V 2PD, UK
| | - Tero A.H. Järvinen
- Faculty of Medicine & Health Technology, Tampere University, 33014 Tampere, Finland & Departments of Orthopedics & Traumatology and Tampere Eye Centre, Tampere University Hospital, 33521 Tampere, Finland
| | - James R. Whiteford
- William Harvey Research Institute, Barts and The London School of Medicine and Dentistry, Queen Mary University of London, Charterhouse Square, London EC1M 6BQ, United Kingdom
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13
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Kozlowski AM, Yates EA, Roubroeks JP, Tømmeraas K, Smith AM, Morris GA. Hydrolytic Degradation of Heparin in Acidic Environments: Nuclear Magnetic Resonance Reveals Details of Selective Desulfation. ACS APPLIED MATERIALS & INTERFACES 2021; 13:5551-5563. [PMID: 33471995 DOI: 10.1021/acsami.0c20198] [Citation(s) in RCA: 4] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 06/12/2023]
Abstract
Heparin is a complex glycosaminoglycan, derived mainly from pig mucosa, used therapeutically for its anticoagulant activity. Yet, owing largely to the chain complexity, the progressive effects of environmental conditions on heparin structure have not been fully described. A systematic study of the influence of acidic hydrolysis on heparin chain length and substitution has therefore been conducted. Changes in the sulfation pattern, monitored via 2D NMR, revealed initial de-N-sulfation of the molecule (pH 1/ 40 °C) and unexpectedly identified the secondary sulfate of iduronate as more labile than the 6-O-sulfate of glucosamine residues under these conditions (pH 1/ 60 °C). Additionally, the loss of sulfate groups, rather than depolymerization, accounted for most of the reduction in molecular weight. This provides an alternative route to producing partially 2-O-de-sulfated heparin derivatives that avoids using conventional basic conditions and may be of value in the optimization of processes associated with the production of heparin pharmaceuticals.
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Affiliation(s)
- Aleksandra M Kozlowski
- Biopolymer Research Centre, School of Applied Sciences, University of Huddersfield, Huddersfield HD1 3DH, United Kingdom
| | - Edwin A Yates
- Department of Biochemistry and Systems Biology, Institute of Systems, Molecular and Integrative Biology, University of Liverpool, Liverpool, L69 7ZB, United Kingdom
| | | | | | - Alan M Smith
- Biopolymer Research Centre, School of Applied Sciences, University of Huddersfield, Huddersfield HD1 3DH, United Kingdom
| | - Gordon A Morris
- Biopolymer Research Centre, School of Applied Sciences, University of Huddersfield, Huddersfield HD1 3DH, United Kingdom
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14
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Effective Inhibition of SARS-CoV-2 Entry by Heparin and Enoxaparin Derivatives. J Virol 2021; 95:JVI.01987-20. [PMID: 33173010 DOI: 10.1128/jvi.01987-20] [Citation(s) in RCA: 159] [Impact Index Per Article: 39.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/07/2020] [Accepted: 11/05/2020] [Indexed: 12/12/2022] Open
Abstract
Severe acute respiratory syndrome-related coronavirus 2 (SARS-CoV-2) has caused a pandemic of historic proportions and continues to spread globally, with enormous consequences to human health. Currently there is no vaccine, effective therapeutic, or prophylactic. As with other betacoronaviruses, attachment and entry of SARS-CoV-2 are mediated by the spike glycoprotein (SGP). In addition to its well-documented interaction with its receptor, human angiotensin-converting enzyme 2 (hACE2), SGP has been found to bind to glycosaminoglycans like heparan sulfate, which is found on the surface of virtually all mammalian cells. Here, we pseudotyped SARS-CoV-2 SGP on a third-generation lentiviral (pLV) vector and tested the impact of various sulfated polysaccharides on transduction efficiency in mammalian cells. The pLV vector pseudotyped SGP efficiently and produced high titers on HEK293T cells. Various sulfated polysaccharides potently neutralized pLV-S pseudotyped virus with clear structure-based differences in antiviral activity and affinity to SGP. Concentration-response curves showed that pLV-S particles were efficiently neutralized by a range of concentrations of unfractionated heparin (UFH), enoxaparin, 6-O-desulfated UFH, and 6-O-desulfated enoxaparin with 50% inhibitory concentrations (IC50s) of 5.99 μg/liter, 1.08 mg/liter, 1.77 μg/liter, and 5.86 mg/liter, respectively. In summary, several sulfated polysaccharides show potent anti-SARS-CoV-2 activity and can be developed for prophylactic as well as therapeutic purposes.IMPORTANCE The emergence of severe acute respiratory syndrome coronavirus (SARS-CoV-2) in Wuhan, China, in late 2019 and its subsequent spread to the rest of the world has created a pandemic situation unprecedented in modern history. While ACE2 has been identified as the viral receptor, cellular polysaccharides have also been implicated in virus entry. The SARS-CoV-2 spike glycoprotein (SGP) binds to glycosaminoglycans like heparan sulfate, which is found on the surface of virtually all mammalian cells. Here, we report structure-based differences in antiviral activity and affinity to SGP for several sulfated polysaccharides, including both well-characterized FDA-approved drugs and novel marine sulfated polysaccharides, which can be developed for prophylactic as well as therapeutic purposes.
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15
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Lin J, Zheng L, Liang Q, Jiang L, Wei Z. Preparation and characterization of partial de-O-sulfation of heparin oligosaccharide library. Carbohydr Res 2021; 499:108226. [PMID: 33401230 DOI: 10.1016/j.carres.2020.108226] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/12/2020] [Revised: 12/07/2020] [Accepted: 12/18/2020] [Indexed: 10/22/2022]
Abstract
The O-sulfation, including 2-O- and 6-O-sulfation, in heparan sulfate (HS) have important biological and pathophysiological roles. Therefore, the ability to chemically generate a series of oligosaccharides, which have a similar structure to the naturally-occurring, 2-O- and 6-O-sulfating oligosaccharides from HS, would greatly contribute to investigating their natural role in HS. In this study, a heparin oligosaccharide library, including dp2, dp4 and dp6, were prepared from the chemical modification of the fully sulfated dp2, dp4 and dp6. Chemical reaction conditions were optimized to generate different patterns of 2-O- and 6-O-sulfated oligosaccharides, then the disaccharide composition and structure of the library was detected by high-performance liquid chromatography-ion trap/time-of-flight mass spectrometry (LC-IT-TOF/MS) analysis. This provides a foundation for further structural and functional studies of O-sulfated groups in HS.
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Affiliation(s)
- Jianghui Lin
- College of Chemistry, Fu Zhou University, FuZhou, 350002, PR China; Institute of Glycobiochemistry, National Engineering Research Centre of Chemical Fertilizer Catalyst, Fu Zhou University, FuZhou, 350002, PR China
| | - Liyang Zheng
- College of Chemistry, Fu Zhou University, FuZhou, 350002, PR China; Institute of Glycobiochemistry, National Engineering Research Centre of Chemical Fertilizer Catalyst, Fu Zhou University, FuZhou, 350002, PR China
| | - Quntao Liang
- College of Biological Science and Engineering, Fu Zhou University, FuZhou, 350002, PR China.
| | - Lilong Jiang
- Institute of Glycobiochemistry, National Engineering Research Centre of Chemical Fertilizer Catalyst, Fu Zhou University, FuZhou, 350002, PR China
| | - Zheng Wei
- Institute of Glycobiochemistry, National Engineering Research Centre of Chemical Fertilizer Catalyst, Fu Zhou University, FuZhou, 350002, PR China.
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16
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A heparin derivatives library constructed by chemical modification and enzymatic depolymerization for exploitation of non-anticoagulant functions. Carbohydr Polym 2020; 249:116824. [PMID: 32933671 DOI: 10.1016/j.carbpol.2020.116824] [Citation(s) in RCA: 14] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/22/2020] [Revised: 07/21/2020] [Accepted: 07/22/2020] [Indexed: 12/11/2022]
Abstract
Non-anticoagulant biological functions of heparin-based drugs have drawn increasing attention. However, the exploration into the non-anticoagulant activities of various low molecular weight heparins was associated with bleeding risks in clinical practice and often led to controversial conclusions due to the structural differences. In this study, we aimed to establish a process to produce a library of heparin derivatives with structural diversity and reduced/abolished anticoagulant activity through the combination of chemical modifications and enzymatic cleavage of heparins. The depolymerization characteristics of various selectively modified heparin derivatives by three heparinases were comprehensively analyzed. The order of periodate treatment and heparinase-I depolymerization was proved to significantly change the structural characteristics of the oligosaccharide products. Finally, among several heparin derivatives that screened in the bleomycin-induced cell apoptosis model, the low molecular weight partially 6-O-/N-desulfated heparins showed the strongest anti-apoptotic activities. This study provided a useful approach for future development of novel heparin-derivative medications.
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17
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Tandon R, Sharp JS, Zhang F, Pomin VH, Ashpole NM, Mitra D, Jin W, Liu H, Sharma P, Linhardt RJ. Effective Inhibition of SARS-CoV-2 Entry by Heparin and Enoxaparin Derivatives. BIORXIV : THE PREPRINT SERVER FOR BIOLOGY 2020:2020.06.08.140236. [PMID: 32577638 PMCID: PMC7302190 DOI: 10.1101/2020.06.08.140236] [Citation(s) in RCA: 21] [Impact Index Per Article: 4.2] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Indexed: 02/07/2023]
Abstract
Severe acute respiratory syndrome-related coronavirus 2 (SARS-CoV-2) has caused a pandemic of historic proportions and continues to spread globally, with enormous consequences to human health. Currently there is no vaccine, effective therapeutic or prophylactic. Like other betacoronaviruses, attachment and entry of SARS-CoV-2 is mediated by the spike glycoprotein (SGP). In addition to its well-documented interaction with its receptor, human angiotensin converting enzyme 2 (hACE2), SGP has been found to bind to glycosaminoglycans like heparan sulfate, which is found on the surface of virtually all mammalian cells. Here, we pseudotyped SARS-CoV-2 SGP on a third generation lentiviral (pLV) vector and tested the impact of various sulfated polysaccharides on transduction efficiency in mammalian cells. The pLV vector pseudotyped SGP efficiently and produced high titers on HEK293T cells. Various sulfated polysaccharides potently neutralized pLV-S pseudotyped virus with clear structure-based differences in anti-viral activity and affinity to SGP. Concentration-response curves showed that pLV-S particles were efficiently neutralized by a range of concentrations of unfractionated heparin (UFH), enoxaparin, 6-O-desulfated UFH and 6-O-desulfated enoxaparin with an IC50 of 5.99 μg/L, 1.08 mg/L, 1.77 μg/L, and 5.86 mg/L respectively. The low serum bioavailability of intranasally administered UFH, along with data suggesting that the nasal epithelium is a portal for initial infection and transmission, suggest that intranasal administration of UFH may be an effective and safe prophylactic treatment.
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Affiliation(s)
- Ritesh Tandon
- Department of Microbiology and Immunology, University of Mississippi Medical Center, Jackson, MS 39216
| | - Joshua S. Sharp
- Department of BioMolecular Sciences, University of Mississippi, Oxford, MS 38677
- Department of Chemistry and Biochemistry, University of Mississippi, Oxford, MS 38677
| | - Fuming Zhang
- Center for Biotechnology and Interdisciplinary Studies, Rensselaer Polytechnic Institute, Troy, NY, 12180
| | - Vitor H. Pomin
- Department of BioMolecular Sciences, University of Mississippi, Oxford, MS 38677
| | - Nicole M. Ashpole
- Department of BioMolecular Sciences, University of Mississippi, Oxford, MS 38677
| | - Dipanwita Mitra
- Department of Microbiology and Immunology, University of Mississippi Medical Center, Jackson, MS 39216
| | - Weihua Jin
- Center for Biotechnology and Interdisciplinary Studies, Rensselaer Polytechnic Institute, Troy, NY, 12180
| | - Hao Liu
- Department of BioMolecular Sciences, University of Mississippi, Oxford, MS 38677
| | - Poonam Sharma
- Department of Microbiology and Immunology, University of Mississippi Medical Center, Jackson, MS 39216
| | - Robert J. Linhardt
- Center for Biotechnology and Interdisciplinary Studies, Rensselaer Polytechnic Institute, Troy, NY, 12180
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18
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Du Z, Jia X, Chen J, Zhou S, Chen J, Liu X, Cao X, Zhong S, Hong P. Isolation and Characterization of a Heparin-Like Compound with Potent Anticoagulant and Fibrinolytic Activity from the Clam Coelomactra antiquata. Mar Drugs 2019; 18:E6. [PMID: 31861572 PMCID: PMC7024239 DOI: 10.3390/md18010006] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/19/2019] [Revised: 11/23/2019] [Accepted: 12/03/2019] [Indexed: 12/18/2022] Open
Abstract
Heparin from mollusks with unique sulfated glycosaminoglycan exhibits strong anti-thrombotic activities. This study reports on a purified heparinoid from Coelomactra antiquata, which shows potent anticoagulant and fibrinolytic abilities. Its structure was characterized by infrared spectroscopy, high-performance liquid chromatography, and one-dimensional and two-dimensional nuclear magnetic resonance spectroscopy. Its fibrinolytic activity was determined in vitro and in vivo. Its anticoagulant activity was determined by activated partial thromboplastin time (APTT), prothrombin time (PT), and thrombin time (TT). The results indicated that clam heparinoid was a homogeneous glycosaminoglycan with a molecular weight of 30.99 kDa, mainly composed of →4)-α-IdoA2S-(1→4)-α-GlcNS3S6S (or GlcNS6S)-(1→4)-β-GlcA-(1→4)-α-GlcNS6S (or GlcNAC)-(1→. Furthermore, this heparinoid showed a highly anticoagulant titer and fibrinolytic value of 149.63 IU/mg and 1.96 IU/mg, respectively. In summary, clam heparinoid shows great potential for application in the clinic and antithrombotic drugs industry.
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Affiliation(s)
- ZhenXing Du
- School of Food Science and Technology, Guangdong Ocean University, Zhanjiang 524088, China; (Z.D.); (X.J.); (J.C.); (S.Z.); (J.C.); (X.L.); (X.C.); (P.H.)
- Shenzhen institute, Guangdong Ocean University, Shenzhen 518108, China
- Guangdong Provincial Key Laboratory of Aquatic Products Processing and Safety, Guangdong Province Engineering Laboratory for Marine Biological Products, Zhanjiang 524088, China
| | - XueJing Jia
- School of Food Science and Technology, Guangdong Ocean University, Zhanjiang 524088, China; (Z.D.); (X.J.); (J.C.); (S.Z.); (J.C.); (X.L.); (X.C.); (P.H.)
- Guangdong Provincial Key Laboratory of Aquatic Products Processing and Safety, Guangdong Province Engineering Laboratory for Marine Biological Products, Zhanjiang 524088, China
| | - Jing Chen
- School of Food Science and Technology, Guangdong Ocean University, Zhanjiang 524088, China; (Z.D.); (X.J.); (J.C.); (S.Z.); (J.C.); (X.L.); (X.C.); (P.H.)
- Shenzhen institute, Guangdong Ocean University, Shenzhen 518108, China
- Guangdong Provincial Key Laboratory of Aquatic Products Processing and Safety, Guangdong Province Engineering Laboratory for Marine Biological Products, Zhanjiang 524088, China
| | - SiYi Zhou
- School of Food Science and Technology, Guangdong Ocean University, Zhanjiang 524088, China; (Z.D.); (X.J.); (J.C.); (S.Z.); (J.C.); (X.L.); (X.C.); (P.H.)
- Shenzhen institute, Guangdong Ocean University, Shenzhen 518108, China
- Guangdong Provincial Key Laboratory of Aquatic Products Processing and Safety, Guangdong Province Engineering Laboratory for Marine Biological Products, Zhanjiang 524088, China
| | - JianPing Chen
- School of Food Science and Technology, Guangdong Ocean University, Zhanjiang 524088, China; (Z.D.); (X.J.); (J.C.); (S.Z.); (J.C.); (X.L.); (X.C.); (P.H.)
| | - XiaoFei Liu
- School of Food Science and Technology, Guangdong Ocean University, Zhanjiang 524088, China; (Z.D.); (X.J.); (J.C.); (S.Z.); (J.C.); (X.L.); (X.C.); (P.H.)
| | - XiaoHuang Cao
- School of Food Science and Technology, Guangdong Ocean University, Zhanjiang 524088, China; (Z.D.); (X.J.); (J.C.); (S.Z.); (J.C.); (X.L.); (X.C.); (P.H.)
| | - SaiYi Zhong
- School of Food Science and Technology, Guangdong Ocean University, Zhanjiang 524088, China; (Z.D.); (X.J.); (J.C.); (S.Z.); (J.C.); (X.L.); (X.C.); (P.H.)
- Shenzhen institute, Guangdong Ocean University, Shenzhen 518108, China
- Guangdong Provincial Key Laboratory of Aquatic Products Processing and Safety, Guangdong Province Engineering Laboratory for Marine Biological Products, Zhanjiang 524088, China
- Collaborative Innovation Center of Seafood Deep Processing, Dalian Polytechnic University, Dalian 116034, China
| | - PengZhi Hong
- School of Food Science and Technology, Guangdong Ocean University, Zhanjiang 524088, China; (Z.D.); (X.J.); (J.C.); (S.Z.); (J.C.); (X.L.); (X.C.); (P.H.)
- Shenzhen institute, Guangdong Ocean University, Shenzhen 518108, China
- Guangdong Provincial Key Laboratory of Aquatic Products Processing and Safety, Guangdong Province Engineering Laboratory for Marine Biological Products, Zhanjiang 524088, China
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19
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Ishihara M, Nakamura S, Sato Y, Takayama T, Fukuda K, Fujita M, Murakami K, Yokoe H. Heparinoid Complex-Based Heparin-Binding Cytokines and Cell Delivery Carriers. Molecules 2019; 24:molecules24244630. [PMID: 31861225 PMCID: PMC6943580 DOI: 10.3390/molecules24244630] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/18/2019] [Revised: 12/10/2019] [Accepted: 12/11/2019] [Indexed: 12/20/2022] Open
Abstract
Heparinoid is the generic term that is used for heparin, heparan sulfate (HS), and heparin-like molecules of animal or plant origin and synthetic derivatives of sulfated polysaccharides. Various biological activities of heparin/HS are attributed to their specific interaction and regulation with various heparin-binding cytokines, antithrombin (AT), and extracellular matrix (ECM) biomolecules. Specific domains with distinct saccharide sequences in heparin/HS mediate these interactions are mediated and require different highly sulfated saccharide sequences with different combinations of sulfated groups. Multivalent and cluster effects of the specific sulfated sequences in heparinoids are also important factors that control their interactions and biological activities. This review provides an overview of heparinoid-based biomaterials that offer novel means of engineering of various heparin-binding cytokine-delivery systems for biomedical applications and it focuses on our original studies on non-anticoagulant heparin-carrying polystyrene (NAC-HCPS) and polyelectrolyte complex-nano/microparticles (N/MPs), in addition to heparin-coating devices.
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Affiliation(s)
- Masayuki Ishihara
- Division of Biomedical Engineering, Research Institute, National Defense Medical College, 3-2 Namiki, Tokorazawa, Saitama 359-8513, Japan; (S.N.); (Y.S.); (K.F.)
- Correspondence: ; Tel.: +81-429-95-1211 (ext. 2610)
| | - Shingo Nakamura
- Division of Biomedical Engineering, Research Institute, National Defense Medical College, 3-2 Namiki, Tokorazawa, Saitama 359-8513, Japan; (S.N.); (Y.S.); (K.F.)
| | - Yoko Sato
- Division of Biomedical Engineering, Research Institute, National Defense Medical College, 3-2 Namiki, Tokorazawa, Saitama 359-8513, Japan; (S.N.); (Y.S.); (K.F.)
| | - Tomohiro Takayama
- Department of Oral and Maxillofacial Surgery, National Defense Medical College, 3-2 Namiki, Tokorozawa, Saitama 359-8513, Japan; (T.T.); (K.M.); (H.Y.)
| | - Koichi Fukuda
- Division of Biomedical Engineering, Research Institute, National Defense Medical College, 3-2 Namiki, Tokorazawa, Saitama 359-8513, Japan; (S.N.); (Y.S.); (K.F.)
| | - Masanori Fujita
- Division of Environmental Medicine, Research Institute, National Defense Medical College, 3-2 Namiki, Tokorozawa, Saitama 359-1324, Japan;
| | - Kaoru Murakami
- Department of Oral and Maxillofacial Surgery, National Defense Medical College, 3-2 Namiki, Tokorozawa, Saitama 359-8513, Japan; (T.T.); (K.M.); (H.Y.)
| | - Hidetaka Yokoe
- Department of Oral and Maxillofacial Surgery, National Defense Medical College, 3-2 Namiki, Tokorozawa, Saitama 359-8513, Japan; (T.T.); (K.M.); (H.Y.)
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20
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Pan Q, Zhang C, Wu X, Chen Y. Identification of a heparosan heptasaccharide as an effective anti-inflammatory agent by partial desulfation of low molecular weight heparin. Carbohydr Polym 2019; 227:115312. [PMID: 31590876 DOI: 10.1016/j.carbpol.2019.115312] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/27/2019] [Revised: 09/01/2019] [Accepted: 09/09/2019] [Indexed: 10/26/2022]
Abstract
Low molecular weight heparin (LMWH) possesses a dual function of anticoagulation and anti-inflammation. While the structures and mechanisms on its anticoagulation have been widely studied, the structural features responsible for the anti-inflammatory activity of LMWH remain to be explored. In the present study, guided by an anti-inflammation assay, a non-anticoagulant species was generated from partial desulfation of LMWH to fully retain the anti-inflammatory activity, from which five fractions were further separated and three of them were characterized by enzymatic degradation, hydrophobic labeling, C18-based HPLC and LC-MS/MS analyses. The structure-activity relationship revealed that the sulfate groups in LMWH are critical to distinguish and separate the activities of anticoagulation and anti-inflammation, leading to the identification of a synthetic heparosan-type heptasaccharide as a potent anti-inflammatory agent. The present strategy enables the simplification of complex polysaccharides to bioactive synthetic oligosaccharides for therapeutic utility.
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Affiliation(s)
- Qi Pan
- State Key Laboratory of Natural Medicines and Laboratory of Chemical Biology, China Pharmaceutical University, Nanjing, Jiangsu 210009, China
| | - Chengchang Zhang
- State Key Laboratory of Natural Medicines and Laboratory of Chemical Biology, China Pharmaceutical University, Nanjing, Jiangsu 210009, China
| | - Xuri Wu
- State Key Laboratory of Natural Medicines and Laboratory of Chemical Biology, China Pharmaceutical University, Nanjing, Jiangsu 210009, China
| | - Yijun Chen
- State Key Laboratory of Natural Medicines and Laboratory of Chemical Biology, China Pharmaceutical University, Nanjing, Jiangsu 210009, China.
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21
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Benito-Arenas R, Doncel-Pérez E, Fernández-Gutiérrez M, Garrido L, García-Junceda E, Revuelta J, Bastida A, Fernández-Mayoralas A. A holistic approach to unravelling chondroitin sulfation: Correlations between surface charge, structure and binding to growth factors. Carbohydr Polym 2018; 202:211-218. [DOI: 10.1016/j.carbpol.2018.08.120] [Citation(s) in RCA: 16] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/27/2018] [Revised: 08/12/2018] [Accepted: 08/27/2018] [Indexed: 01/08/2023]
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22
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Atallah P, Schirmer L, Tsurkan M, Putra Limasale YD, Zimmermann R, Werner C, Freudenberg U. In situ-forming, cell-instructive hydrogels based on glycosaminoglycans with varied sulfation patterns. Biomaterials 2018; 181:227-239. [DOI: 10.1016/j.biomaterials.2018.07.056] [Citation(s) in RCA: 27] [Impact Index Per Article: 3.9] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/18/2018] [Revised: 07/20/2018] [Accepted: 07/28/2018] [Indexed: 01/11/2023]
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23
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Tadai K, Shioiri T, Tsuchimoto J, Nagai N, Watanabe H, Sugiura N. Interaction of receptor type of protein tyrosine phosphatase sigma (RPTPσ) with a glycosaminoglycan library. J Biochem 2018; 164:41-51. [PMID: 29420785 DOI: 10.1093/jb/mvy027] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/06/2017] [Accepted: 01/31/2018] [Indexed: 12/14/2022] Open
Abstract
Receptor type of protein tyrosine phosphatase sigma (RPTPσ) functions as a glycosaminoglycan (GAG) receptor of neuronal cells in both the central and peripheral nervous systems. Both chondroitin sulphate (CS) and heparan sulphate (HS) are important constituents of GAG ligands for RPTPσ, although they have opposite effects on neuronal cells. CS inhibits neurite outgrowth and neural regeneration through RPTPσ, whereas HS enhances them. We prepared recombinant RPTPσ N-terminal fragment containing the GAG binding site and various types of biotin-conjugated GAG (CS and HS) with chemical modification and chemo-enzymatic synthesis. Then interaction of the RPTPσ N-terminal fragment was analysed using GAG-biotin immobilized on streptavidin sensor chips by surface plasmon resonance. Interaction of RPTPσ with the CS library was highly correlated to the degree of disulphated disaccharide E unit, which had two sulphate groups at C-4 and C-6 positions of the N-acetylgalactosamine residue (CSE). The optimum molecular mass of CSE was suggested to be approximately 10 kDa. Heparin showed higher affinity to RPTPσ than the CS library. Our GAG library will not only contribute to the fields of carbohydrate science and cell biology, but also provide medical application to regulate neural regeneration.
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Affiliation(s)
- Kouki Tadai
- Institute for Molecular Science of Medicine, Aichi Medical University, 1-1 Yazakokarimata, Nagakute, Aichi 480-1195, Japan.,Faculty of Health and Nutrition, Shubun University, 6 Nikko-cho, Ichinomiya, Aichi 491-0938, Japan
| | - Tatsumasa Shioiri
- Institute for Molecular Science of Medicine, Aichi Medical University, 1-1 Yazakokarimata, Nagakute, Aichi 480-1195, Japan
| | - Jun Tsuchimoto
- Institute for Molecular Science of Medicine, Aichi Medical University, 1-1 Yazakokarimata, Nagakute, Aichi 480-1195, Japan
| | - Naoko Nagai
- Institute for Molecular Science of Medicine, Aichi Medical University, 1-1 Yazakokarimata, Nagakute, Aichi 480-1195, Japan
| | - Hideto Watanabe
- Institute for Molecular Science of Medicine, Aichi Medical University, 1-1 Yazakokarimata, Nagakute, Aichi 480-1195, Japan
| | - Nobuo Sugiura
- Institute for Molecular Science of Medicine, Aichi Medical University, 1-1 Yazakokarimata, Nagakute, Aichi 480-1195, Japan
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24
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A review of chemical methods for the selective sulfation and desulfation of polysaccharides. Carbohydr Polym 2017; 174:1224-1239. [DOI: 10.1016/j.carbpol.2017.07.017] [Citation(s) in RCA: 60] [Impact Index Per Article: 7.5] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/01/2016] [Revised: 05/22/2017] [Accepted: 07/06/2017] [Indexed: 11/24/2022]
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25
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Baumann H. Biological Effects of Heparan Sulfates and Regioselectively Modified Heparin-Heparan Mimetics. J BIOACT COMPAT POL 2016. [DOI: 10.1177/0883911503018001006] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/16/2022]
Abstract
The heparan sulfates (HS) are structurally the most complex but information rich biopolymers known. They are composed of polysaccharides containing regioselectively distributed carboxyl, sulfate ester, acetyl, amino, and N-sulfonyl groups with sequence- and domain-like arrangements. HS are found ubiquitously on cell surfaces and in extracellular matrices where they are covalently anchored via restricted protein cores. They modulate numerous development cell processes and the pathology of living organisms. HS concentration is extremely low on endothelial cell surfaces (1 pmol/cm2), therefore, they are difficult to isolate and evaluate. Furthermore, their sequence variability is extremely high and the sequence analysis is in its infancy. HS acts as a low affinity receptor which plays a central role in the reception and modulation of a wide range of effector proteins such as growth factors, morphogens, chemokines, enzymes, protease inhibitors. Water soluble fragments of HS and heparin (HE) enzymatically released or synthetic sequences, analogs of heparinoids and heparanoids (HH) mimetics regioselectively modified oligo- and polysaccharides with HE/HS like functional groups, and nonsaccharide containing structures can modulate effector proteins and influence some of the development and pathological processes. Modulation effects are described for anticoagulant antiproliferative properties, for reducing platelet and plasma protein adhesion as well as inhibition or activating growth factors by the influence of HH mimetics. The advantage of defined high molecular weight substrates are discussed and compared to the low molecular weight mimetics. The potential of HH mimetics opens new approaches and strategies for therapeutic treatment.
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Affiliation(s)
- H. Baumann
- Institute for Technical and Macromolecular Chemistry Hemocompatible and Biocompatible Biomaterials RWTH Aachen, Worringer Weg 1, D-52074 Aachen, Germany
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26
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Bedini E, Laezza A, Iadonisi A. Chemical Derivatization of Sulfated Glycosaminoglycans. European J Org Chem 2016. [DOI: 10.1002/ejoc.201600108] [Citation(s) in RCA: 23] [Impact Index Per Article: 2.6] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/27/2022]
Affiliation(s)
- Emiliano Bedini
- Department of Chemical Sciences; University of Naples Federico II; Complesso Universitario Monte S. Angelo; via Cintia 4 80126 Napoli Italy
| | - Antonio Laezza
- Department of Chemical Sciences; University of Naples Federico II; Complesso Universitario Monte S. Angelo; via Cintia 4 80126 Napoli Italy
| | - Alfonso Iadonisi
- Department of Chemical Sciences; University of Naples Federico II; Complesso Universitario Monte S. Angelo; via Cintia 4 80126 Napoli Italy
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27
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Abstract
Glycosaminoglycans (GAGs) have been shown to bind to a wide variety of microbial pathogens, including viruses, bacteria, parasites, and fungi in vitro. GAGs are thought to promote pathogenesis by facilitating pathogen attachment, invasion, or evasion of host defense mechanisms. However, the role of GAGs in infectious disease has not been extensively studied in vivo and therefore their pathophysiological significance and functions are largely unknown. Here we describe methods to directly investigate the role of GAGs in infections in vivo using mouse models of bacterial lung and corneal infection. The overall experimental strategy is to establish the importance and specificity of GAGs, define the essential structural features of GAGs, and identify a biological activity of GAGs that promotes pathogenesis.
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Affiliation(s)
- Akiko Jinno
- Division of Respiratory Diseases, Children's Hospital, Harvard Medical School, 320 Longwood Avenue, Enders-461, Boston, MA, 02115, USA,
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28
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Zieris A, Dockhorn R, Röhrich A, Zimmermann R, Müller M, Welzel PB, Tsurkan MV, Sommer JU, Freudenberg U, Werner C. Biohybrid Networks of Selectively Desulfated Glycosaminoglycans for Tunable Growth Factor Delivery. Biomacromolecules 2014; 15:4439-46. [DOI: 10.1021/bm5012294] [Citation(s) in RCA: 38] [Impact Index Per Article: 3.5] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/29/2022]
Affiliation(s)
- Andrea Zieris
- Leibniz Institute of Polymer Research Dresden, Hohe Strasse 6, 01069 Dresden, Germany
| | - Ron Dockhorn
- Leibniz Institute of Polymer Research Dresden, Hohe Strasse 6, 01069 Dresden, Germany
- Institute
for Theoretical Physics, Technische Universität Dresden, Zellescher Weg
17, 01069 Dresden, Germany
| | - Anika Röhrich
- B CUBE
Center for Molecular Bioengineering, Technische Universität Dresden, Arnoldstrasse 18, 01307 Dresden, Germany
| | - Ralf Zimmermann
- Leibniz Institute of Polymer Research Dresden, Hohe Strasse 6, 01069 Dresden, Germany
| | - Martin Müller
- Leibniz Institute of Polymer Research Dresden, Hohe Strasse 6, 01069 Dresden, Germany
| | - Petra B. Welzel
- Leibniz Institute of Polymer Research Dresden, Hohe Strasse 6, 01069 Dresden, Germany
| | - Mikhail V. Tsurkan
- Leibniz Institute of Polymer Research Dresden, Hohe Strasse 6, 01069 Dresden, Germany
| | - Jens-Uwe Sommer
- Leibniz Institute of Polymer Research Dresden, Hohe Strasse 6, 01069 Dresden, Germany
- Institute
for Theoretical Physics, Technische Universität Dresden, Zellescher Weg
17, 01069 Dresden, Germany
| | - Uwe Freudenberg
- Leibniz Institute of Polymer Research Dresden, Hohe Strasse 6, 01069 Dresden, Germany
- Center
for Regenerative Therapies Dresden, Technische Universität Dresden, Fetscherstrasse 105, 01307 Dresden, Germany
| | - Carsten Werner
- Leibniz Institute of Polymer Research Dresden, Hohe Strasse 6, 01069 Dresden, Germany
- Center
for Regenerative Therapies Dresden, Technische Universität Dresden, Fetscherstrasse 105, 01307 Dresden, Germany
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Babazada H, Yamashita F, Yanamoto S, Hashida M. Self-assembling lipid modified glycol-split heparin nanoparticles suppress lipopolysaccharide-induced inflammation through TLR4-NF-κB signaling. J Control Release 2014; 194:332-40. [PMID: 25234820 DOI: 10.1016/j.jconrel.2014.09.011] [Citation(s) in RCA: 25] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/02/2014] [Revised: 08/15/2014] [Accepted: 09/08/2014] [Indexed: 01/23/2023]
Abstract
Self-assembling heparin nanoparticles have attracted much attention as promising drug carriers for various drugs, genes and imaging agents. In the present investigation, we found that heparin nanoparticles are selective Toll-like receptor 4 (TLR-4) antagonists and have a much greater anti-inflammatory effect than native heparin. More specifically, we developed self-assembling nanoparticles composed of glycol-split heparin/D-erythro-sphingosine conjugates (NAHNP), characterized their physicochemical properties and anti-inflammatory effect in vitro. Unlike native heparin, NAHNP significantly inhibited lipopolysaccharide-induced activation of MyD88-dependent NF-κB signaling pathway and production of pro-inflammatory cytokines such as TNF-alpha from mouse macrophages with IC50 = 0.019 mg/mL. Furthermore, we investigated the structure-activity relationship of the conjugates and identified the length of attached alkyl chains of d-erythro-sphingosine to be critical for anti-inflammatory effect. Decrease in alkyl chain length of NAHNP resulted in loss of inhibitory activity. In line with these findings, 6-O-sulfate groups of D-glucosamine residue were essential for effective inhibition, while removal of 2-O-sulfo and 3-O-sulfo groups as well as replacement of N-sulfo groups with N-acetyl did not alter anti-inflammatory activity. Therefore, NAHNP would be a promising candidate in acute and chronic inflammatory disorders, in addition to the nature of a drug carrier.
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Affiliation(s)
- Hasan Babazada
- Department of Drug Delivery Research, Graduate School of Pharmaceutical Sciences, Kyoto University, Yoshidashimoadachi-cho, Sakyo-ku, Kyoto 606-8501, Japan
| | - Fumiyoshi Yamashita
- Department of Drug Delivery Research, Graduate School of Pharmaceutical Sciences, Kyoto University, Yoshidashimoadachi-cho, Sakyo-ku, Kyoto 606-8501, Japan
| | - Shinya Yanamoto
- Department of Drug Delivery Research, Graduate School of Pharmaceutical Sciences, Kyoto University, Yoshidashimoadachi-cho, Sakyo-ku, Kyoto 606-8501, Japan
| | - Mitsuru Hashida
- Department of Drug Delivery Research, Graduate School of Pharmaceutical Sciences, Kyoto University, Yoshidashimoadachi-cho, Sakyo-ku, Kyoto 606-8501, Japan; Institute for Integrated Cell-Material Sciences, Kyoto University, Yoshidaushinomiya-cho, Sakyo-ku, Kyoto 606-8501, Japan.
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30
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Structural analysis of isomeric chondroitin sulfate oligosaccharides using regioselective 6-O-desulfation method and tandem mass spectrometry. Anal Chim Acta 2014; 843:27-37. [DOI: 10.1016/j.aca.2014.07.027] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/14/2014] [Revised: 07/17/2014] [Accepted: 07/20/2014] [Indexed: 01/24/2023]
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31
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Troeberg L, Lazenbatt C, Anower-E-Khuda MF, Freeman C, Federov O, Habuchi H, Habuchi O, Kimata K, Nagase H. Sulfated glycosaminoglycans control the extracellular trafficking and the activity of the metalloprotease inhibitor TIMP-3. ACTA ACUST UNITED AC 2014; 21:1300-1309. [PMID: 25176127 PMCID: PMC4210636 DOI: 10.1016/j.chembiol.2014.07.014] [Citation(s) in RCA: 62] [Impact Index Per Article: 5.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/25/2014] [Revised: 07/15/2014] [Accepted: 07/16/2014] [Indexed: 12/15/2022]
Abstract
Tissue inhibitor of metalloproteinase 3 (TIMP-3) is an important regulator of extracellular matrix (ECM) turnover. TIMP-3 binds to sulfated ECM glycosaminoglycans or is endocytosed by cells via low-density lipoprotein receptor-related protein 1 (LRP-1). Here, we report that heparan sulfate (HS) and chondroitin sulfate E (CSE) selectively regulate postsecretory trafficking of TIMP-3 by inhibiting its binding to LRP-1. HS and CSE also increased TIMP-3 affinity for glycan-binding metalloproteinases, such as adamalysin-like metalloproteinase with thrombospondin motifs 5 (ADAMTS-5), by reducing the dissociation rate constants. The sulfation pattern was crucial for these activities because monosulfated or truncated heparin had a reduced ability to bind to TIMP-3 and increase its affinity for ADAMTS-5. Therefore, sulfation of ECM glycans regulates the levels and inhibitory activity of TIMP-3 and modulates ECM turnover, and small mimicries of sulfated glycans may protect the tissue from the excess destruction seen in diseases such as osteoarthritis, cancer, and atherosclerosis.
The metalloprotease inhibitor TIMP-3 binds to sulfated extracellular glycans This inhibits cellular uptake of TIMP-3 by the endocytic receptor LRP-1 Glycans also increase TIMP-3 affinity for selected target proteases The sulfation of matrix glycans therefore modulates TIMP-3 activity and ECM turnover
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Affiliation(s)
- Linda Troeberg
- Arthritis Research UK Centre for Osteoarthritis Pathogenesis, Kennedy Institute of Rheumatology, University of Oxford, Roosevelt Drive, Oxford OX3 7FY, UK.
| | - Christopher Lazenbatt
- Arthritis Research UK Centre for Osteoarthritis Pathogenesis, Kennedy Institute of Rheumatology, University of Oxford, Roosevelt Drive, Oxford OX3 7FY, UK
| | - Md Ferdous Anower-E-Khuda
- Aichi Medical University Research Complex for Medicine Frontiers, Aichi Medical University, Nagakute, Aichi 480-1195, Japan
| | - Craig Freeman
- Division of Immunology and Genetics, John Curtin School of Medical Research, Australian National University, Canberra ACT 2601, Australia
| | - Oleg Federov
- Structural Genomics Consortium, Nuffield Department of Clinical Medicine, University of Oxford, Oxford OX3 7FZ, UK
| | - Hiroko Habuchi
- Advanced Medical Research Centre, Aichi Medical University, Nagakute, Aichi 480-1195, Japan
| | - Osami Habuchi
- Advanced Medical Research Centre, Aichi Medical University, Nagakute, Aichi 480-1195, Japan
| | - Koji Kimata
- Aichi Medical University Research Complex for Medicine Frontiers, Aichi Medical University, Nagakute, Aichi 480-1195, Japan
| | - Hideaki Nagase
- Arthritis Research UK Centre for Osteoarthritis Pathogenesis, Kennedy Institute of Rheumatology, University of Oxford, Roosevelt Drive, Oxford OX3 7FY, UK
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32
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Hwang SR, Seo DH, Al-Hilal TA, Jeon OC, Kang JH, Kim SH, Kim HS, Chang YT, Kang YM, Yang VC, Byun Y. Orally active desulfated low molecular weight heparin and deoxycholic acid conjugate, 6ODS-LHbD, suppresses neovascularization and bone destruction in arthritis. J Control Release 2012; 163:374-84. [DOI: 10.1016/j.jconrel.2012.09.013] [Citation(s) in RCA: 20] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/19/2012] [Revised: 07/31/2012] [Accepted: 09/21/2012] [Indexed: 12/21/2022]
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33
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Gill VL, Wang Q, Shi X, Zaia J. Mass spectrometric method for determining the uronic acid epimerization in heparan sulfate disaccharides generated using nitrous acid. Anal Chem 2012; 84:7539-46. [PMID: 22873817 DOI: 10.1021/ac3016054] [Citation(s) in RCA: 26] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/19/2023]
Abstract
Heparan sulfate (HS) glycosaminoglycans (GAGs) regulate a host of biological functions. To better understand their biological roles, it is necessary to gain understanding about the structure of HS, which requires identification of the sulfation pattern as well as the uronic acid epimerization. In order to model HS structure, it is necessary to quantitatively profile depolymerization products. To date, liquid chromatography-mass spectrometry (LC-MS) methods for profiling heparin lyase decomposition products have been shown. These enzymes, however, destroy information about uronic acid epimerization. Deaminative cleavage using nitrous acid (HONO) is a classic method for GAG depolymerization that retains uronic acid epimerization. Several chromatographic methods have been used for analysis of deaminative cleavage products. The chromatographic methods have the disadvantage that there is no direct readout on the structures producing the observed peaks. This report demonstrates a porous graphitized carbon (PGC)-MS method for the quantification of HONO generated disaccharides to obtain information about the sulfation pattern and uronic acid epimerization. Here, we demonstrate the separation and identification of uronic acid epimers as well as geometric sulfation isomers. The results are comparable to those expected for benchmark HS and heparin samples. The data demonstrate the utility of PGC-MS for quantification of HS nitrous acid depolymerization products for structural analysis of HS and heparin.
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Affiliation(s)
- Vanessa Leah Gill
- Department of Biochemistry, Boston University School of Medicine, Boston, Massachusetts, United States
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34
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35
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He Y, Li J, Chen Y. Catalytic Effects of Different Heparin Analogs on the Hydrolysis of Auramine O. JOURNAL OF BIOMATERIALS SCIENCE. POLYMER EDITION 2011; 22:253-261. [PMID: 20557699 DOI: 10.1163/092050609x12601848092242] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 05/29/2023]
Abstract
Glycosaminoglycans, such as heparin, are a class of biomaterials with a variety of important biological functions. They were found to catalyze the hydrolysis of 4,4'-(imidocarbonyl)-bis(N,N-dimethylaniline) monohydrochloride to 4,4'-bis(dimethy-lamino) benzophenone, but the catalytic mechanism of this interesting reaction is poorly understood, mainly due to the structural complexity of polysaccharides. In order to study such an unusual reaction, a pentasaccharide, fondaparinux, with defined chemical structure was chosen as a probe to investigate the hydrolytic characteristics, along with other heparin analogs. The hydrolytic reaction was catalyzed by fondaparinux, low-molecular-weight heparins, heparin and different desulfated heparins at various rates. The present results demonstrated that the length of polysaccharide chain, the N- or O-sulfo groups are critical to the hydrolysis of 4,4'-(imidocarbonyl)-bis(N,N-dimethy-laniline) monohydrochloride in addition to pH and ionic strength, which provides new insights into the catalytic phenomenon of polysaccharides.
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Affiliation(s)
- Yunmian He
- a Laboratory of Chemical Biology, China Pharmaceutical University, 24 Tongjia Street, Nanjing, Jiangsu 210009, P. R. China; Key Lab of Analytical Chemistry for Life Science, School of Chemistry and Chemical Engineering, Nanjing University, Nanjing, Jiangsu, 210093, P. R. China
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36
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Becher J, Möller S, Weiss D, Schiller J, Schnabelrauch M. Synthesis of New Regioselectively Sulfated Hyaluronans for Biomedical Application. ACTA ACUST UNITED AC 2010. [DOI: 10.1002/masy.201051060] [Citation(s) in RCA: 16] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/11/2022]
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37
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Jastrebova N, Vanwildemeersch M, Lindahl U, Spillmann D. Heparan sulfate domain organization and sulfation modulate FGF-induced cell signaling. J Biol Chem 2010; 285:26842-26851. [PMID: 20576609 DOI: 10.1074/jbc.m109.093542] [Citation(s) in RCA: 61] [Impact Index Per Article: 4.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/24/2023] Open
Abstract
Heparan sulfates (HSs) modulate various developmental and homeostatic processes by binding to protein ligands. We have evaluated the structural characteristics of porcine HS in cellular signaling induced by basic fibroblast growth factor (FGF2), using CHO745 cells devoid of endogenous glycosaminoglycans as target. Markedly enhanced stimulation of cell signaling, measured as phosphorylation of ERK1/2 and protein kinase B, was only observed with the shortest HS chains isolated from liver, whereas the longer chains from either liver or intestine essentially prolonged duration of signals induced by FGF2 in the absence of polysaccharide. Structural analysis showed that contiguous sulfated domains were most abundant in the shortest HS chains and were more heavily sulfated in HS from liver than in HS from intestine. Moreover, the shortest chains from either source entered into ternary complexes with FGF2 and FGF receptor-1c more efficiently than the corresponding longer chains. In addition to authentic HSs, decasaccharide libraries generated by chemo-enzymatic modification of heparin were probed for effect on FGF2 signaling. Only the most highly sulfated decamers, previously found most efficient in ternary complex formation (Jastrebova, N., Vanwildemeersch, M., Rapraeger, A. C., Giménez-Gallego, G., Lindahl, U., and Spillmann, D. (2006) J. Biol. Chem. 281, 26884-26892), promoted FGF2 cellular signaling as efficiently as short HS chains from liver. Together these results suggest that the effects of HS on FGF2 signaling are determined by both the structure of the highly sulfated domains and by the organization/availability of such domains within the HS chain. These findings underpin the need for regulation of HS biosynthesis in relation to control of growth factor-induced signaling pathways.
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Affiliation(s)
- Nadja Jastrebova
- Department of Medical Biochemistry and Microbiology, Uppsala University, SE-751 23 Uppsala, Sweden
| | - Maarten Vanwildemeersch
- Department of Medical Biochemistry and Microbiology, Uppsala University, SE-751 23 Uppsala, Sweden
| | - Ulf Lindahl
- Department of Medical Biochemistry and Microbiology, Uppsala University, SE-751 23 Uppsala, Sweden
| | - Dorothe Spillmann
- Department of Medical Biochemistry and Microbiology, Uppsala University, SE-751 23 Uppsala, Sweden.
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38
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Kulkarni AA, Weiss AA, Iyer SS. Glycan-based high-affinity ligands for toxins and pathogen receptors. Med Res Rev 2010; 30:327-93. [DOI: 10.1002/med.20196] [Citation(s) in RCA: 55] [Impact Index Per Article: 3.7] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/11/2022]
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39
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Zhang F, Zhang Z, Lin X, Beenken A, Eliseenkova AV, Mohammadi M, Linhardt RJ. Compositional analysis of heparin/heparan sulfate interacting with fibroblast growth factor.fibroblast growth factor receptor complexes. Biochemistry 2009; 48:8379-86. [PMID: 19591432 PMCID: PMC3348549 DOI: 10.1021/bi9006379] [Citation(s) in RCA: 60] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/28/2022]
Abstract
Heparan sulfate (HS) proteoglycans (PGs) interact with a number of extracellular signaling proteins, thereby playing an essential role in the regulation of many physiological processes. One major function of HS is to interact with fibroblast growth factors (FGFs) and their receptors (FGFRs) and form FGF.HS.FGFR signaling complexes. Past studies primarily examined the selectivity of HS for FGF or FGFR. In this report, we used a new strategy to study the structural specificity of HS binding to 10 different FGF.FGFR complexes. Oligosaccharide libraries prepared from heparin, 6-desulfated heparin, and HS were used for the interaction studies by solution competition surface plasmon resonance (SPR) and filter trapping assays. Specific oligosaccharides binding to FGF.FGFR complexes were subjected to polyacrylamide gel electrophoresis (PAGE) analysis and disaccharide compositional analysis using liquid chromatography and mass spectrometry. The competition SPR studies using sized oligosaccharide mixtures showed that binding of each of the tested FGFs or FGF.FGFR complexes to heparin immobilized to an SPR chip was size-dependent. The 6-desulfated heparin oligosaccharides exhibited a reduced level of inhibition of FGF and FGF.FGFR complex binding to heparin in the competition experiments. Heparin and the 6-desulfated heparin exhibited higher levels of inhibition of the FGF.FGFR complex binding to heparin than of FGF binding to heparin. In the filter trapping experiments, PAGE analysis showed different affinities between the FGF.FGFR complexes and oligosaccharides. Disaccharide analysis showed that HS disaccharides with a degree of polymerization of 10 (dp10) had high binding selectivity, while dp10 heparin and dp10 6-desulfated heparin showed reduced or no selectivity for the different FGF.FGFR complexes tested.
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Affiliation(s)
- Fuming Zhang
- Department of Chemistry and Chemical Biology, Center for Biotechnology and Interdisciplinary Studies, Rensselaer Polytechnic Institute, Troy, New York 12180, USA.
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40
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Modulation of the proteolytic activity of the complement protease C1s by polyanions: implications for polyanion-mediated acceleration of interaction between C1s and SERPING1. Biochem J 2009; 422:295-303. [PMID: 19522701 DOI: 10.1042/bj20090198] [Citation(s) in RCA: 11] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/17/2022]
Abstract
The complement system plays crucial roles in the immune system, but incorrect regulation causes inflammation and targeting of self-tissue, leading to diseases such as systemic lupus erythematosus, rheumatoid arthritis and age-related macular degeneration. In vivo, the initiating complexes of the classical complement and lectin pathways are controlled by SERPING1 [(C1 inhibitor) serpin peptidase inhibitor, clade G, member 1], which inactivates the components C1s and MASP-2 (mannan-binding lectin serine peptidase 2). GAGs (glycosaminoglycan) and DXS (dextran sulfate) are able to significantly accelerate SERPING1-mediated inactivation of C1s, the key effector enzyme of the classical C1 complex, although the mechanism is poorly understood. In the present study we have shown that C1s can bind to DXS and heparin and that these polyanions enhanced C1s proteolytic activity at low concentrations and inhibited it at higher concentrations. The recent determination of the crystal structure of SERPING1 has given rise to the hypothesis that both the serpin (serine protease inhibitor)-polyanion and protease-polyanion interactions might be required to accelerate the association rate of SERPING1 and C1s. To determine what proportion of the acceleration was due to protease-polyanion interactions, a chimaeric mutant of alpha1-antitrypsin containing the P4-P1 residues from the SERPING1 RCL (reactive-centre loop) was produced. Like SERPING1, this molecule is able to effectively inhibit C1s, but is unable to bind polyanions. DXS exerted a biphasic effect on the association rate of C1s which correlated strongly with the effect of DXS on C1s proteolytic activity. Thus, whereas polyanions are able to bind C1s and modulate its activity, polyanion interactions with SERPING1 must also play a vital role in the mechanism by which these cofactors accelerate the C1s-SERPING1 reaction.
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Suzuki K, Yamamoto K, Kariya Y, Maeda H, Ishimaru T, Miyaura S, Fujii M, Yusa A, Joo EJ, Kimata K, Kannagi R, Kim YS, Kyogashima M. Generation and characterization of a series of monoclonal antibodies that specifically recognize [HexA(+/-2S)-GlcNAc]n epitopes in heparan sulfate. Glycoconj J 2008; 25:703-12. [PMID: 18461440 DOI: 10.1007/s10719-008-9130-z] [Citation(s) in RCA: 17] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/21/2008] [Revised: 03/25/2008] [Accepted: 03/26/2008] [Indexed: 11/29/2022]
Abstract
Five monoclonal antibodies AS17, 22, 25, 38 and 48, a single monoclonal antibody ACH55, and three monoclonal antibodies NAH33, 43, 46, that recognize acharan sulfate (IdoA2S-GlcNAc)n, acharan (IdoA-GlcNAc)n and N-acetyl-heparosan (GlcA-GlcNAc)n, respectively, were generated by immunization of mice with keyhole limpet hemocyanin-conjugated polysaccharides. Specificity tests were performed using a panel of biotinylated GAGs that included chemically modified heparins. Each antibody bound avidly to the immunized polysaccharide, but did not bind to chondroitin sulfates, keratan sulfate, chondroitin nor hyaluronic acid. AS antibodies did not bind to heparan sulfate or heparin, but bound to 6-O-desulfated, N-desulfated and re-N-acetylated heparin to varying degrees. ACH55 bound to tri-desulfated and re-N-acetylated heparin but hardly bound to other modified heparins. NAH antibodies did not bind to heparin and modified heparins but bound to heparan sulfate to varying degrees. NAH43 and NAH46 also bound to partially N-de-acetylated N-acetyl-heparosan. Immunohistochemical analysis in rat cerebella was performed with the antibodies. While NAH46 stained endothelia, where heparan sulfate is typically present, neither ACH55 nor AS25 stained endothelia. On the contrary ACH55 and AS25 stained the molecular layer of the rat cerebella. Furthermore, ACH55 specifically stained Purkinje cells. These results suggest that there is unordinary expression of IdoA2S-GlcNAc and IdoA-GlcNAc in specific parts of the nervous system.
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Affiliation(s)
- Kiyoshi Suzuki
- Central Research Laboratories, Seikagaku Corporation, 3-1253 Tateno, Higashiyamato, Tokyo, 207-0021, Japan
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42
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Knappe M, Bodevin S, Selinka HC, Spillmann D, Streeck RE, Chen XS, Lindahl U, Sapp M. Surface-exposed amino acid residues of HPV16 L1 protein mediating interaction with cell surface heparan sulfate. J Biol Chem 2007; 282:27913-22. [PMID: 17640876 DOI: 10.1074/jbc.m705127200] [Citation(s) in RCA: 108] [Impact Index Per Article: 6.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/06/2022] Open
Abstract
Efficient infection of cells by human papillomaviruses (HPVs) and pseudovirions requires primary interaction with cell surface proteoglycans with apparent preference for species carrying heparan sulfate (HS) side chains. To identify residues contributing to virus/cell interaction, we performed point mutational analysis of the HPV16 major capsid protein, L1, targeting surface-exposed amino acid residues. Replacement of lysine residues 278, 356, or 361 for alanine reduced cell binding and infectivity of pseudovirions. Various combinations of these amino acid exchanges further decreased cell attachment and infectivity with residual infectivity of less than 5% for the triple mutant, suggesting that these lysine residues cooperate in HS binding. Single, double, or triple exchanges for arginine did not impair infectivity, demonstrating that interaction is dependent on charge distribution rather than sequence-specific. The lysine residues are located within a pocket on the capsomere surface, which was previously proposed as the putative receptor binding site. Fab fragments of binding-neutralizing antibody H16.56E that recognize an epitope directly adjacent to lysine residues strongly reduced HS-mediated cell binding, further corroborating our findings. In contrast, mutation of basic surface residues located in the cleft between capsomeres outside this pocket did not significantly reduce interaction with HS or resulted in assembly-deficient proteins. Computer-simulated heparin docking suggested that all three lysine residues can form hydrogen bonds with 2-O-, 6-O-, and N-sulfate groups of a single HS molecule with a minimal saccharide domain length of eight monomer units. This prediction was experimentally confirmed in binding experiments using capsid protein, heparin molecules of defined length, and sulfate group modifications.
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Affiliation(s)
- Maren Knappe
- Institute for Medical Microbiology, University of Mainz, D-55101 Mainz, Germany
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43
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Shipp EL, Hsieh-Wilson LC. Profiling the Sulfation Specificities of Glycosaminoglycan Interactions with Growth Factors and Chemotactic Proteins Using Microarrays. ACTA ACUST UNITED AC 2007; 14:195-208. [PMID: 17317573 DOI: 10.1016/j.chembiol.2006.12.009] [Citation(s) in RCA: 148] [Impact Index Per Article: 8.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/30/2006] [Revised: 12/13/2006] [Accepted: 12/28/2006] [Indexed: 12/24/2022]
Abstract
We report a carbohydrate microarray-based approach for the rapid, facile analysis of glycosaminoglycan-protein interactions. The key structural determinants responsible for protein binding, such as sulfate groups that participate in the interactions, were elucidated. Specificities were also readily compared across protein families or functional classes, and comparisons among glycosaminoglycan subclasses provided a more comprehensive understanding of protein specificity. To validate the approach, we showed that fibroblast growth factor family members have distinct sulfation preferences. We also demonstrated that heparan sulfate and chondroitin sulfate interact in a sulfation-dependent manner with various axon guidance proteins, including slit2, netrin1, ephrinA1, ephrinA5, and semaphorin5B. We anticipate that these microarrays will accelerate the discovery of glycosaminoglycan-binding proteins and provide a deeper understanding of their roles in regulating diverse biological processes.
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Affiliation(s)
- Eric L Shipp
- Division of Chemistry and Chemical Engineering and Howard Hughes Medical Institute, California Institute of Technology, Pasadena, CA 91125, USA
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44
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Jastrebova N, Vanwildemeersch M, Rapraeger AC, Giménez-Gallego G, Lindahl U, Spillmann D. Heparan sulfate-related oligosaccharides in ternary complex formation with fibroblast growth factors 1 and 2 and their receptors. J Biol Chem 2006; 281:26884-92. [PMID: 16807244 DOI: 10.1074/jbc.m600806200] [Citation(s) in RCA: 66] [Impact Index Per Article: 3.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/16/2023] Open
Abstract
Biosynthesis of heparan sulfate (HS) is strictly regulated to yield products with cell/tissue-specific composition. Interactions between HS and a variety of proteins, including growth factors and morphogens, are essential for embryonic development and for homeostasis in the adult. Fibroblast growth factors (FGFs) and their various receptors (FRs) form ternary complexes with HS, as required for receptor signaling. Libraries of HS-related, radiolabeled oligosaccharides were generated by chemo-enzymatic modification of heparin and tested for affinity to immobilized FR ectodomains in the presence of FGF1 or FGF2. Experiments were designed to enable assessment of N-sulfated 8- and 10-mers with defined numbers of iduronic acid 2-O-sulfate and glucosamine 6-O-sulfate groups. FGF1 and FGF2 were found to require similar oligosaccharides in complex formation with FR1c-3c, FGF2 affording somewhat more efficient oligosaccharide recruitment than FGF1. FR4, contrary to FR1c-3c, bound oligosaccharides at physiological ionic conditions even in the absence of FGFs, and this interaction was further promoted by FGF1 but not by FGF2. In all systems studied, the stability of FGF-oligosaccharide-FR complexes correlated with the overall level of saccharide O-sulfation rather than on the precise distribution of sulfate groups.
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Affiliation(s)
- Nadja Jastrebova
- Department of Medical Biochemistry and Microbiology, Uppsala University, SE-751 23 Uppsala, Sweden
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McDowell LM, Frazier BA, Studelska DR, Giljum K, Chen J, Liu J, Yu K, Ornitz DM, Zhang L. Inhibition or Activation of Apert Syndrome FGFR2 (S252W) Signaling by Specific Glycosaminoglycans. J Biol Chem 2006; 281:6924-30. [PMID: 16373332 DOI: 10.1074/jbc.m512932200] [Citation(s) in RCA: 42] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/06/2022] Open
Abstract
Most Apert syndrome patients harbor a single amino acid mutation (S252W) in fibroblast growth factor (FGF) receptor 2 (FGFR2), which leads to abnormal FGF/FGFR2 signaling. Here we show that specific combinations of FGFs and glycosaminoglycans activate both alternative splice forms of the mutant but not of the wild-type FGF receptors. More importantly, 2-O- and N-sulfated heparan sulfate, prepared by a combined chemical and enzymatic synthesis, antagonized the over-activated FGFR2b (S252W) to basal levels at nanomolar concentrations. These studies demonstrated that specific glycosaminoglycans could be useful in treating ligand-dependent FGFR signaling-related diseases, such as Apert syndrome and cancer.
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Affiliation(s)
- Lynda M McDowell
- Department of Pathology and Immunology, Washington University School of Medicine, St. Louis, MO 63110, USA
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Nakamura S, Ishihara M, Obara K, Masuoka K, Ishizuka T, Kanatani Y, Takase B, Matsui T, Hattori H, Sato T, Kariya Y, Maehara T. Controlled release of fibroblast growth factor-2 from an injectable 6-O-desulfated heparin hydrogel and subsequent effect onin vivo vascularization. J Biomed Mater Res A 2006; 78:364-71. [PMID: 16673389 DOI: 10.1002/jbm.a.30688] [Citation(s) in RCA: 33] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/13/2023]
Abstract
We prepared a 6-O-desulfated (DS-) heparin (Hep) hydrogel as an excellent carrier for the controlled release of Hep-binding growth factors, such as fibroblast growth factor (FGF)-2. This material, which is partially derived from photoreactive groups, such as cinnamate, is easily crosslinked upon ultraviolet light (UV)-irradiation, resulting in a water-insoluble, viscous, and injectable hydrogel. In the present study, we examined the capacity of 6-O-DS-Hep hydrogel to immobilize FGF-2, as well as the controlled release of FGF-2 molecules from this hydrogel in vitro and in vivo. Only 10% of FGF-2 was gradually released from the FGF-2-containing 6-O-DS-Hep hydrogel (photocrosslinked 6-O-DS-Hep (4%; w/w) hydrogel containing 50 microg/mL FGF-2) into PBS (phosphate-buffered saline) within first 7 days. The 6-O-DS-Hep hydrogel in vitro maintained the original form through 1 weeks incubation in PBS, but it was gradually fragmented and could not maintain the original form by 2-3 week-washing. When the FGF-2-containing 6-O-DS-Hep hydrogel was subcutaneously injected into the back of rats, significant neovascularization and fibrous tissue formation were induced near the injected site from day 3 after the injection. And, the hydrogel had been biodegraded and completely disappeared from the injected sites in vivo within about 15-20 days after the injection. These findings indicate a controlled release of biologically active FGF-2 molecules together with fragmentation and biodegradation of 6-O-DS-Hep hydrogel and the subsequent induction of neovascularization in vivo.
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Affiliation(s)
- Shingo Nakamura
- Department of Surgery II, National Defense Medical College, 3-2 Namiki, Tokorozawa, Saitama 359-8513, Japan.
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Mori H, Kanemura Y, Onaya J, Hara M, Miyake J, Yamasaki M, Kariya Y. Effects of heparin and its 6-O-and 2-O-desulfated derivatives with low anticoagulant activity on proliferation of human neural stem/progenitor cells. J Biosci Bioeng 2005; 100:54-61. [PMID: 16233851 DOI: 10.1263/jbb.100.54] [Citation(s) in RCA: 10] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/25/2005] [Accepted: 03/17/2005] [Indexed: 11/17/2022]
Abstract
Heparin binds various growth factors and activates them to interact with high-affinity cell surface receptors; a specific array of sulfate groups in the heparin backbone structure is very important for this interaction. In the present study, we evaluated the effects of two novel heparin derivatives, 6-O-desulfated heparin (6-DSH) and 2-O-desulfated heparin (2-DSH), on blood coagulation and the proliferation of human neural stem/progenitor cells (NSPCs). 6-DSH showed lower anticoagulant activity than intact heparin or 2-DSH, as measured by the activated partial thromboplastin time and thrombin time. In the presence of FGF-2, 6-DSH and 2-DSH promoted approximately the same rate of proliferation of human NSPCs, without noticeably changing the expression of nestin. The mitotic effects of 6-DSH and 2-DSH on human NSPCs were different from their effects on mouse hematopoietic stem cells and fibroblasts. These findings indicate that 6-DSH and 2-DSH have the same ability to promote the growth of human NSPCs as intact heparin. Our results suggest that these two novel heparin derivates, especially 6-DSH, could be used in clinical applications for ex vivo human NSPC culture, as a lower-risk growth co-adjuvant than intact heparin.
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Affiliation(s)
- Hideki Mori
- Research Institute for Cell Engineering, National Institute of Advanced Industrial Science and Technology, 3-11-46 Nakoji, Amagasaki, Hyogo 661-0974, Japan
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Ji SL, Cui HF, Shi F, Chi YQ, Cao JC, Geng MY, Guan HS. Inhibitory effect of heparin-derived oligosaccharides on secretion of interleukin-4 and interleukin-5 from human peripheral blood T lymphocytes. World J Gastroenterol 2004; 10:3490-4. [PMID: 15526371 PMCID: PMC4576233 DOI: 10.3748/wjg.v10.i23.3490] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/15/2022] Open
Abstract
AIM: To investigate the inhibitory effect of heparin-derived oligosaccharides (Oligs) on secretion of interleukin-4 (IL-4) and interleukin-5 (IL-5) from human peripheral blood T lymphocytes (PBTLs).
METHODS: Oligs were prepared by three different heparin depolymerization methods and separated by gel filtration chromatography. PBTLs from ten adult patients with allergic eosinophilic gastroenteritis were treated with phytahematoagglutinin (PHA) and Oligs. The supernatants from the cell culture of PBTLs were harvested and subjected to the determination of IL-4 and IL-5 contents by ELISA method.
RESULTS: At the concentration of 5 μg/mL, Oligs with different Mr had different effects on the secretion of IL-4 and IL-5. The tetrasaccharide with Mr of 1142, produced by depolymerizing heparin with hydrogen peroxide, had the strongest inhibitory effect on the secretion of IL-4. It decreased the IL-4 content from 375.6 ± 39.2 ng/L (PHA group) to 12.5 ± 5.7 ng/L (P < 0.01). The hexasaccharide with Mr of 1806, produced by depolymerizing heparin with β -elimination method, had the strongest inhibitory effect on the secretion of IL-5. It decreased the IL-5 content from 289.2 ± 33.4 ng/L (PHA group) to 22.0 ± 5.2 ng/L (P < 0.01).
CONCLUSION: The inhibitory activity of Oligs on the secretion of IL-4 and IL-5 from human PBTLs closely depends on their molecular structure, and there may be an essential structure to act as an inhibitor. The most effective inhibitors of IL-4 and IL-5 secretion are tetrasaccharides and hexasaccharides, respectively.
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Affiliation(s)
- Sheng-Li Ji
- Key Laboratory of Marine Drugs, Marine Drug and Food Institute, Ocean University of China, Qingdao 266003, Shandong Province, China.
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Ledin J, Staatz W, Li JP, Götte M, Selleck S, Kjellén L, Spillmann D. Heparan sulfate structure in mice with genetically modified heparan sulfate production. J Biol Chem 2004; 279:42732-41. [PMID: 15292174 DOI: 10.1074/jbc.m405382200] [Citation(s) in RCA: 205] [Impact Index Per Article: 9.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/06/2022] Open
Abstract
Using a high throughput heparan sulfate (HS) isolation and characterization protocol, we have analyzed HS structure in several tissues from mice/mouse embryos deficient in HS biosynthesis enzymes (N-deacetylase/N-sulfotransferase (NDST)-1, NDST-2, and C5-epimerase, respectively) and in mice lacking syndecan-1. The results have given us new information regarding HS biosynthesis with implications on the role of HS in embryonic development. Our main conclusions are as follows. 1) The HS content, disaccharide composition, and the overall degree of N- and O-sulfation as well as domain organization are characteristic for each individual mouse tissue. 2) Removal of a key biosynthesis enzyme (NDST-1 or C5-epimerase) results in similar structural alterations in all of the tissues analyzed. 3) Essentially no variation in HS tissue structure is detected when individuals of the same genotype are compared. 4) NDST-2, although generally expressed, does not contribute significantly to tissue-specific HS structures. 5) No change in HS structure could be detected in syndecan-1-deficient mice.
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Affiliation(s)
- Johan Ledin
- Department of Medical Biochemistry and Microbiology, University of Uppsala, SE-75123 Uppsala, Sweden
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Wei M, Tai G, Gao Y, Li N, Huang B, Zhou Y, Hao S, Zeng X. Modified Heparin Inhibits P-selectin-mediated Cell Adhesion of Human Colon Carcinoma Cells to Immobilized Platelets under Dynamic Flow Conditions. J Biol Chem 2004; 279:29202-10. [PMID: 15133030 DOI: 10.1074/jbc.m312951200] [Citation(s) in RCA: 47] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/15/2023] Open
Abstract
Accumulating evidence indicates that the formation of tumor cell platelet emboli complexes in the blood stream is a very important step during metastases and that the anti-metastasis effects of heparin are partially due to a blockade of P-selectin on platelets. In this study, heparin and chemically modified heparins were tested as inhibitors of three human colon carcinoma cell lines (COLO320, LS174T, and CW-2) binding to P-selectin, adhering to CHO cells expressing a transfected human P-selectin cDNA, and adhering to surface-anchored platelets expressing P-selectin under static and flow conditions. The aim was to screen for heparin derivatives with high anti-adhesion activity but negligible anticoagulant activity. In this study, four modified heparins with high anti-adhesion activity were identified including RO-heparin, CR-heparin, 2/3ODS-heparin, and N/2/3DS-heparin. NMR analysis proved the reliability of structure of the four modified heparins. Our findings suggested that the 6-O-sulfate group of glucosamine units in heparin is critical for the inhibition of P-selectin-mediated tumor cell adhesion. Heparan sulfate-like proteoglycans on these tumor cell surfaces are implicated in adhesion of the tumor cells to P-selectin. Some chemically modified heparins with low anticoagulant activities, such as 2/3ODS-heparin, may have potential value as therapeutic agents that block P-selectin-mediated cell adhesion and prevent tumor metastasis.
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Affiliation(s)
- Min Wei
- Institute of Genetics and Cytology, School of Life Science, Northeast Normal University, Changchun, 130024, Jilin, People's Republic of China
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