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Artikel ; Online: Methods for Hyaluronan Molecular Mass Determination by Agarose Gel Electrophoresis.

Cowman, Mary K

Methods in molecular biology (Clifton, N.J.)

2019 Band 1952, Seite(n) 91–102

Abstract: The average molecular mass of hyaluronan (HA) in most healthy biological fluids and tissues is usually about 6000-8000 kDa, but the biosynthetic mechanism results in a polydisperse mixture of sizes. Subsequent enzymatic degradation, or the action of ... ...

Abstract	The average molecular mass of hyaluronan (HA) in most healthy biological fluids and tissues is usually about 6000-8000 kDa, but the biosynthetic mechanism results in a polydisperse mixture of sizes. Subsequent enzymatic degradation, or the action of reactive oxygen and nitrogen species, can further increase polydispersity and decrease the average size. Fragmented HA can be a biomarker of inflammation. In addition, reductions in HA size are associated with tissue remodeling and repair processes. Some cell-surface receptor proteins have been reported to have HA-binding affinities that are size specific, and participate in activation of signaling cascades controlling multiple aspects of cell behavior. Here we describe simple agarose gel electrophoresis protocols for the determination of the molecular mass distribution of HA isolated from tissues and fluids.
Mesh-Begriff(e)	Acetates/chemistry ; Boric Acids/chemistry ; Densitometry/methods ; Edetic Acid/chemistry ; Electrophoresis, Agar Gel/methods ; Ethylenediamines/chemistry ; Humans ; Hyaluronic Acid/chemistry ; Hyaluronic Acid/isolation & purification ; Molecular Weight ; Staining and Labeling/methods ; Tromethamine/chemistry
Chemische Substanzen	Acetates ; Boric Acids ; Ethylenediamines ; Tris-borate-EDTA buffer ; tris-acetate-EDTA buffer ; Tromethamine (023C2WHX2V) ; Hyaluronic Acid (9004-61-9) ; Edetic Acid (9G34HU7RV0)
Sprache	Englisch
Erscheinungsdatum	2019-03-01
Erscheinungsland	United States
Dokumenttyp	Journal Article
ISSN	1940-6029
ISSN (online)	1940-6029
DOI	10.1007/978-1-4939-9133-4_8
Datenquelle	MEDical Literature Analysis and Retrieval System OnLINE

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Artikel ; Online: Hyaluronan and Hyaluronan Fragments.

Cowman, Mary K

Advances in carbohydrate chemistry and biochemistry

2017 Band 74, Seite(n) 1–59

Abstract: The glycosaminoglycan hyaluronan (HA) is a key component of the microenvironment surrounding cells. In healthy tissues, HA molecules have extremely high molecular mass and consequently large hydrodynamic volumes. Tethered to the cell surface by clustered ...

Abstract	The glycosaminoglycan hyaluronan (HA) is a key component of the microenvironment surrounding cells. In healthy tissues, HA molecules have extremely high molecular mass and consequently large hydrodynamic volumes. Tethered to the cell surface by clustered receptor proteins, HA molecules crowd each other, as well as other macromolecular species. This leads to severe nonideality in physical properties of the biomatrix, because steric exclusion leads to an increase in effective concentration of the macromolecules. The excluded volume depends on both polymer concentration and hydrodynamic volume/molecular mass. The biomechanical properties of the extracellular matrix, tissue hydration, receptor clustering, and receptor-ligand interactions are strongly affected by the presence of HA and by its molecular mass. In inflammation, reactive oxygen and nitrogen species fragment the HA chains. Depending on the rate of chain degradation relative to the rates of new synthesis and removal of damaged chains, short fragments of the HA molecules can be present at significant levels. Not only are the physical properties of the extracellular matrix affected, but the HA fragments decluster their primary receptors and act as endogenous danger signals. Bioanalytical methods to isolate and quantify HA fragments have been developed to determine profiles of HA content and size in healthy and diseased biological fluids and tissues. These methods have potential use in medical diagnostic tests. Therapeutic agents that modulate signaling by HA fragments show promise in wound healing and tissue repair without fibrosis.
Mesh-Begriff(e)	Animals ; Humans ; Hyaluronic Acid/chemistry ; Hyaluronic Acid/therapeutic use ; Particle Size ; Surface Properties
Chemische Substanzen	Hyaluronic Acid (9004-61-9)
Sprache	Englisch
Erscheinungsdatum	2017
Erscheinungsland	Netherlands
Dokumenttyp	Journal Article ; Review
ISSN	2162-5530
ISSN (online)	2162-5530
DOI	10.1016/bs.accb.2017.10.001
Datenquelle	MEDical Literature Analysis and Retrieval System OnLINE

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Artikel ; Online: The Role of Hyaluronan/Receptor for Hyaluronan-Mediated Motility Interactions in the Modulation of Macrophage Polarization and Cartilage Repair.

Bianchini, Emilia / Ashley Sin, Yun Jin / Lee, You Jin / Lin, Charles / Anil, Utkarsh / Hamill, Cassie / Cowman, Mary K / Kirsch, Thorsten

The American journal of pathology

2024 Band 194, Heft 6, Seite(n) 1047–1061

Abstract: Hyaluronan (HA), a negatively charged linear glycosaminoglycan, is a key macromolecular component of the articular cartilage extracellular matrix. The differential effects of HA are determined by a spatially/temporally regulated display of HA receptors, ... ...

Abstract	Hyaluronan (HA), a negatively charged linear glycosaminoglycan, is a key macromolecular component of the articular cartilage extracellular matrix. The differential effects of HA are determined by a spatially/temporally regulated display of HA receptors, such as CD44 and receptor for hyaluronan-mediated motility (RHAMM). HA signaling through CD44 with RHAMM has been shown to stimulate inflammation and fibrotic processes. This study shows an increased expression of RHAMM in proinflammatory macrophages. Interfering with HA/RHAMM interactions using a 15-mer RHAMM-mimetic, HA-binding peptide, together with high-molecular-weight (HMW) HA reduced the expression and release of inflammatory markers and increased the expression of anti-inflammatory markers in proinflammatory macrophages. HA/RHAMM interactions were interfered in vivo during the regeneration of a full-thickness cartilage defect after microfracture surgery in rabbits using three intra-articular injections of 15-mer RHAMM-mimetic. HA-binding peptide together with HMWHA reduced the number of proinflammatory macrophages and increased the number of anti-inflammatory macrophages in the injured knee joint and greatly improved the repair of the cartilage defect compared with intra-articular injections of HMWHA alone. These findings suggest that HA/RHAMM interactions play a key role in cartilage repair/regeneration via stimulating inflammatory and fibrotic events, including increasing the ratio of proinflammatory/anti-inflammatory macrophages. Interfering with these interactions reduced inflammation and greatly improved cartilage repair.
Mesh-Begriff(e)	Animals ; Hyaluronan Receptors/metabolism ; Macrophages/metabolism ; Macrophages/drug effects ; Rabbits ; Cartilage, Articular/metabolism ; Cartilage, Articular/pathology ; Hyaluronic Acid/metabolism ; Hyaluronic Acid/pharmacology ; Extracellular Matrix Proteins/metabolism ; Cell Polarity/drug effects ; Cell Polarity/physiology ; Regeneration/drug effects ; Regeneration/physiology ; Inflammation/metabolism ; Inflammation/pathology
Sprache	Englisch
Erscheinungsdatum	2024-02-24
Erscheinungsland	United States
Dokumenttyp	Journal Article
ZDB-ID	2943-9
ISSN	1525-2191 ; 0002-9440
ISSN (online)	1525-2191
ISSN	0002-9440
DOI	10.1016/j.ajpath.2024.01.020
Datenquelle	MEDical Literature Analysis and Retrieval System OnLINE

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Artikel ; Online: Protective Effects of a Hyaluronan-Binding Peptide (P15-1) on Mesenchymal Stem Cells in an Inflammatory Environment.

Kirsch, Thorsten / Zhang, Fenglin / Braender-Carr, Olivia / Cowman, Mary K

International journal of molecular sciences

2021 Band 22, Heft 13

Abstract: Mesenchymal stem cells (MSCs) obtained from various sources, including bone marrow, have been proposed as a therapeutic strategy for the improvement of tissue repair/regeneration, including the repair of cartilage defects or lesions. Often the highly ... ...

Abstract	Mesenchymal stem cells (MSCs) obtained from various sources, including bone marrow, have been proposed as a therapeutic strategy for the improvement of tissue repair/regeneration, including the repair of cartilage defects or lesions. Often the highly inflammatory environment after injury or during diseases, however, greatly diminishes the therapeutic and reparative effectiveness of MSCs. Therefore, the identification of novel factors that can protect MSCs against an inflammatory environment may enhance the effectiveness of these cells in repairing tissues, such as articular cartilage. In this study, we investigated whether a peptide (P15-1) that binds to hyaluronan (HA), a major component of the extracellular matrix of cartilage, protects bone-marrow-derived MSCs (BMSCs) in an inflammatory environment. The results showed that P15-1 reduced the mRNA levels of catabolic and inflammatory markers in interleukin-1beta (IL-1β)-treated human BMSCs. In addition, P15-1 enhanced the attachment of BMSCs to HA-coated tissue culture dishes and stimulated the chondrogenic differentiation of the multipotential murine C3H/10T1/2 MSC line in a micromass culture. In conclusion, our findings suggest that P15-1 may increase the capacity of BMSCs to repair cartilage via the protection of these cells in an inflammatory environment and the stimulation of their attachment to an HA-containing matrix and chondrogenic differentiation.
Mesh-Begriff(e)	Animals ; Anti-Inflammatory Agents/chemistry ; Anti-Inflammatory Agents/pharmacology ; Cell Culture Techniques ; Cell Differentiation/drug effects ; Cell Line ; Chondrogenesis ; Cyclooxygenase 2/genetics ; Extracellular Matrix Proteins/chemistry ; Gene Expression Regulation ; Humans ; Hyaluronan Receptors/chemistry ; Hyaluronic Acid/metabolism ; Interleukin-1beta/adverse effects ; Interleukin-6/genetics ; Mesenchymal Stem Cells/cytology ; Mesenchymal Stem Cells/drug effects ; Mesenchymal Stem Cells/metabolism ; Metalloproteases/genetics ; Mice ; Peptides/chemistry ; Peptides/pharmacology
Chemische Substanzen	Anti-Inflammatory Agents ; Extracellular Matrix Proteins ; Hyaluronan Receptors ; Interleukin-1beta ; Interleukin-6 ; Peptides ; hyaluronan-mediated motility receptor ; Hyaluronic Acid (9004-61-9) ; Cyclooxygenase 2 (EC 1.14.99.1) ; Metalloproteases (EC 3.4.-)
Sprache	Englisch
Erscheinungsdatum	2021-06-30
Erscheinungsland	Switzerland
Dokumenttyp	Journal Article
ZDB-ID	2019364-6
ISSN	1422-0067 ; 1422-0067 ; 1661-6596
ISSN (online)	1422-0067
ISSN	1422-0067 ; 1661-6596
DOI	10.3390/ijms22137058
Datenquelle	MEDical Literature Analysis and Retrieval System OnLINE

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Artikel ; Online: Protective Effects of a Hyaluronan-Binding Peptide (P15-1) on Mesenchymal Stem Cells in an Inflammatory Environment

Thorsten Kirsch / Fenglin Zhang / Olivia Braender-Carr / Mary K. Cowman

International Journal of Molecular Sciences, Vol 22, Iss 7058, p

2021 Band 7058

Abstract	Mesenchymal stem cells (MSCs) obtained from various sources, including bone marrow, have been proposed as a therapeutic strategy for the improvement of tissue repair/regeneration, including the repair of cartilage defects or lesions. Often the highly inflammatory environment after injury or during diseases, however, greatly diminishes the therapeutic and reparative effectiveness of MSCs. Therefore, the identification of novel factors that can protect MSCs against an inflammatory environment may enhance the effectiveness of these cells in repairing tissues, such as articular cartilage. In this study, we investigated whether a peptide (P15-1) that binds to hyaluronan (HA), a major component of the extracellular matrix of cartilage, protects bone-marrow-derived MSCs (BMSCs) in an inflammatory environment. The results showed that P15-1 reduced the mRNA levels of catabolic and inflammatory markers in interleukin-1beta (IL-1β)-treated human BMSCs. In addition, P15-1 enhanced the attachment of BMSCs to HA-coated tissue culture dishes and stimulated the chondrogenic differentiation of the multipotential murine C3H/10T1/2 MSC line in a micromass culture. In conclusion, our findings suggest that P15-1 may increase the capacity of BMSCs to repair cartilage via the protection of these cells in an inflammatory environment and the stimulation of their attachment to an HA-containing matrix and chondrogenic differentiation.
Schlagwörter	mesenchymal stem cells ; hyaluronan ; peptide ; inflammation ; articular cartilage ; Biology (General) ; QH301-705.5 ; Chemistry ; QD1-999
Thema/Rubrik (Code)	616
Sprache	Englisch
Erscheinungsdatum	2021-06-01T00:00:00Z
Verlag	MDPI AG
Dokumenttyp	Artikel ; Online
Datenquelle	BASE - Bielefeld Academic Search Engine (Lebenswissenschaftliche Auswahl)

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Artikel ; Online: Methods for isolating and analyzing physiological hyaluronan: a review.

Rivas, Felipe / Erxleben, Dorothea / Smith, Ian / Rahbar, Elaheh / DeAngelis, Paul L / Cowman, Mary K / Hall, Adam R

American journal of physiology. Cell physiology

2022 Band 322, Heft 4, Seite(n) C674–C687

Abstract: The carbohydrate hyaluronan (or hyaluronic acid, HA) is found in all human tissues and biofluids where it has wide-ranging functions in health and disease that are dictated by both its abundance and size. Consequently, hyaluronan evaluation in ... ...

Abstract	The carbohydrate hyaluronan (or hyaluronic acid, HA) is found in all human tissues and biofluids where it has wide-ranging functions in health and disease that are dictated by both its abundance and size. Consequently, hyaluronan evaluation in physiological samples has significant translational potential. Although the analytical tools and techniques for probing other biomolecules such as proteins and nucleic acids have become standard approaches in biochemistry, those available for investigating hyaluronan are less well established. In this review, we survey methods related to the assessment of native hyaluronan in biological specimens, including protocols for separating it from biological matrices and technologies for determining its concentration and molecular weight.
Mesh-Begriff(e)	Humans ; Hyaluronan Receptors/metabolism ; Hyaluronic Acid ; Molecular Weight
Chemische Substanzen	Hyaluronan Receptors ; Hyaluronic Acid (9004-61-9)
Sprache	Englisch
Erscheinungsdatum	2022-02-23
Erscheinungsland	United States
Dokumenttyp	Journal Article ; Review ; Research Support, N.I.H., Extramural
ZDB-ID	392098-7
ISSN	1522-1563 ; 0363-6143
ISSN (online)	1522-1563
ISSN	0363-6143
DOI	10.1152/ajpcell.00019.2022
Datenquelle	MEDical Literature Analysis and Retrieval System OnLINE

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Artikel ; Online: A Hyaluronan-binding Peptide (P15-1) Reduces Inflammatory and Catabolic Events in IL-1β-treated Human Articular Chondrocytes.

Shortt, Claire / Luyt, Leonard G / Turley, Eva A / Cowman, Mary K / Kirsch, Thorsten

Scientific reports

2020 Band 10, Heft 1, Seite(n) 1441

Abstract: Inflammation plays a critical role in osteoarthritis (OA). It stimulates catabolic events in articular chondrocytes and prevents chondrogenic precursor cells from repairing cartilage lesions, leading to accelerated cartilage degradation. Therefore, the ... ...

Abstract	Inflammation plays a critical role in osteoarthritis (OA). It stimulates catabolic events in articular chondrocytes and prevents chondrogenic precursor cells from repairing cartilage lesions, leading to accelerated cartilage degradation. Therefore, the identification of novel factors that reduce catabolic events in chondrocytes and enhances chondrogenic differentiation of precursor cells in an inflammatory environment may provide novel therapeutic strategies for the treatment of OA. The goal of this study was to determine whether a hyaluronan (HA)-binding peptide (P15-1), via interacting with high molecular weight (HMW)HA can enhance the anti-inflammatory properties of HMWHA and decrease catabolic events in interleukin-1beta (IL-1β)-treated human articular chondrocytes. Treatment with P15-1 decreased catabolic events and stimulated anabolic events in articular chondrocytes cultured in an inflammatory environment. P15-1 pre-mixed with HMWHA was more effective in inhibiting catabolic events and stimulating anabolic events than P15-1 or HMWHA alone. Our findings suggest that P15-1 together with HMWHA inhibits catabolic events in articular chondrocytes via the inhibition of p38 mitogen-activated protein kinases (MAPK) and increasing the thickness of the pericellular matrix (PCM) around chondrocytes thereby decreasing catabolic signaling. Finally, conditioned medium from IL-1β and P15-1-treated human articular chondrocytes was less inhibitory for chondrogenic differentiation of precursor cells than conditioned medium from chondrocytes treated with IL-1β alone. In conclusion, P15-1 is proposed to function synergistically with HMWHA to enhance the protective microenvironment for chondrocytes and mesenchymal stem cells during inflammation and regeneration.
Mesh-Begriff(e)	Adult ; Cartilage/pathology ; Cell Differentiation ; Cells, Cultured ; Chondrocytes/metabolism ; Chondrocytes/pathology ; Chondrogenesis ; Culture Media, Conditioned/pharmacology ; Extracellular Matrix/metabolism ; Humans ; Hyaluronan Receptors/metabolism ; Hyaluronic Acid/metabolism ; Inflammation/metabolism ; Osteoarthritis/metabolism ; Signal Transduction ; p38 Mitogen-Activated Protein Kinases/metabolism
Chemische Substanzen	Culture Media, Conditioned ; Hyaluronan Receptors ; Hyaluronic Acid (9004-61-9) ; p38 Mitogen-Activated Protein Kinases (EC 2.7.11.24)
Sprache	Englisch
Erscheinungsdatum	2020-01-29
Erscheinungsland	England
Dokumenttyp	Journal Article
ZDB-ID	2615211-3
ISSN	2045-2322 ; 2045-2322
ISSN (online)	2045-2322
ISSN	2045-2322
DOI	10.1038/s41598-020-57586-7
Datenquelle	MEDical Literature Analysis and Retrieval System OnLINE

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Artikel ; Online: A Hyaluronan-binding Peptide (P15-1) Reduces Inflammatory and Catabolic Events in IL-1β-treated Human Articular Chondrocytes

Claire Shortt / Leonard G. Luyt / Eva A. Turley / Mary K. Cowman / Thorsten Kirsch

Scientific Reports, Vol 10, Iss 1, Pp 1-

2020 Band 11

Abstract: Abstract Inflammation plays a critical role in osteoarthritis (OA). It stimulates catabolic events in articular chondrocytes and prevents chondrogenic precursor cells from repairing cartilage lesions, leading to accelerated cartilage degradation. ... ...

Abstract	Abstract Inflammation plays a critical role in osteoarthritis (OA). It stimulates catabolic events in articular chondrocytes and prevents chondrogenic precursor cells from repairing cartilage lesions, leading to accelerated cartilage degradation. Therefore, the identification of novel factors that reduce catabolic events in chondrocytes and enhances chondrogenic differentiation of precursor cells in an inflammatory environment may provide novel therapeutic strategies for the treatment of OA. The goal of this study was to determine whether a hyaluronan (HA)-binding peptide (P15-1), via interacting with high molecular weight (HMW)HA can enhance the anti-inflammatory properties of HMWHA and decrease catabolic events in interleukin-1beta (IL-1β)-treated human articular chondrocytes. Treatment with P15-1 decreased catabolic events and stimulated anabolic events in articular chondrocytes cultured in an inflammatory environment. P15-1 pre-mixed with HMWHA was more effective in inhibiting catabolic events and stimulating anabolic events than P15-1 or HMWHA alone. Our findings suggest that P15-1 together with HMWHA inhibits catabolic events in articular chondrocytes via the inhibition of p38 mitogen-activated protein kinases (MAPK) and increasing the thickness of the pericellular matrix (PCM) around chondrocytes thereby decreasing catabolic signaling. Finally, conditioned medium from IL-1β and P15-1-treated human articular chondrocytes was less inhibitory for chondrogenic differentiation of precursor cells than conditioned medium from chondrocytes treated with IL-1β alone. In conclusion, P15-1 is proposed to function synergistically with HMWHA to enhance the protective microenvironment for chondrocytes and mesenchymal stem cells during inflammation and regeneration.
Schlagwörter	Medicine ; R ; Science ; Q
Thema/Rubrik (Code)	616
Sprache	Englisch
Erscheinungsdatum	2020-01-01T00:00:00Z
Verlag	Nature Publishing Group
Dokumenttyp	Artikel ; Online
Datenquelle	BASE - Bielefeld Academic Search Engine (Lebenswissenschaftliche Auswahl)

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Artikel ; Online: Extracellular Vesicles Released From Articular Chondrocytes Play a Major Role in Cell-Cell Communication.

Liu, Xiaoming / Shortt, Claire / Zhang, Fenglin / Bater, Mariah Q / Cowman, Mary K / Kirsch, Thorsten

Journal of orthopaedic research : official publication of the Orthopaedic Research Society

2019 Band 38, Heft 4, Seite(n) 731–739

Abstract: The purpose of this investigation was to determine the role of extracellular vesicles (EVs), released from articular chondrocytes in a physiological or pathological state, in cell-cell communication with other articular chondrocytes or chondrocyte ... ...

Abstract	The purpose of this investigation was to determine the role of extracellular vesicles (EVs), released from articular chondrocytes in a physiological or pathological state, in cell-cell communication with other articular chondrocytes or chondrocyte precursor cells. The conditioned medium from interleukin-1β (IL-1β)-treated human articular chondrocytes stimulated catabolic events and inhibited type II collagen expression in articular chondrocytes to a much greater degree than medium from IL-1β-treated chondrocytes after complete removal of EVs. The vehicle-treated and IL-1β-treated human articular chondrocytes released EVs of similar size; however, the number of EVs released by IL-1β-treated chondrocytes was markedly higher than the number of EVs released from the vehicle-treated cells. Furthermore, our findings demonstrate that similar to medium from IL-1β-treated chondrocytes containing EVs, EVs isolated from medium of IL-1β-treated chondrocytes stimulated catabolic events in articular chondrocytes, whereas EVs isolated from the medium of vehicle-treated chondrocytes inhibited catabolic events and increased messenger RNA levels of aggrecan and type II collagen in IL-1β-treated chondrocytes. Furthermore, the medium containing EVs from vehicle-treated articular chondrocytes or EVs isolated from this medium stimulated chondrogenesis of C3H10T1/2 cells, whereas medium containing EVs from IL-1β-treated chondrocytes or EVs isolated from this medium inhibited chondrogenesis. Our findings suggest that EVs released by articular chondrocytes play a key role in the communication between joint cells and ultimately in joint homeostasis, maintenance, pathology, and repair. © 2019 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 38:731-739, 2020.
Mesh-Begriff(e)	Aged ; Animals ; Cartilage, Articular/cytology ; Cell Communication ; Cell Differentiation ; Cell Line ; Chondrocytes/physiology ; Extracellular Vesicles/physiology ; Humans ; Mice ; Middle Aged ; Primary Cell Culture
Sprache	Englisch
Erscheinungsdatum	2019-11-27
Erscheinungsland	United States
Dokumenttyp	Journal Article ; Research Support, N.I.H., Extramural ; Research Support, Non-U.S. Gov't
ZDB-ID	605542-4
ISSN	1554-527X ; 0736-0266
ISSN (online)	1554-527X
ISSN	0736-0266
DOI	10.1002/jor.24525
Datenquelle	MEDical Literature Analysis and Retrieval System OnLINE

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Artikel: Noncovalent hyaluronan crosslinking by TSG-6: Modulation by heparin, heparan sulfate, and PRG4.

Sin, Yun Jin Ashley / MacLeod, Rebecca / Tanguay, Adam P / Wang, Andrew / Braender-Carr, Olivia / Vitelli, Teraesa M / Jay, Gregory D / Schmidt, Tannin A / Cowman, Mary K

Frontiers in molecular biosciences

2022 Band 9, Seite(n) 990861

Abstract: The size, conformation, and organization of the glycosaminoglycan hyaluronan (HA) affect its interactions with soluble and cell surface-bound proteins. HA that is induced to form stable networks has unique biological properties relative to unmodified ... ...

Abstract	The size, conformation, and organization of the glycosaminoglycan hyaluronan (HA) affect its interactions with soluble and cell surface-bound proteins. HA that is induced to form stable networks has unique biological properties relative to unmodified soluble HA. AlphaLISA assay technology offers a facile and general experimental approach to assay protein-mediated networking of HA in solution. Connections formed between two end-biotinylated 50 kDa HA (bHA) chains can be detected by signal arising from streptavidin-coated donor and acceptor beads being brought into close proximity when the bHA chains are bridged by proteins. We observed that incubation of bHA with the protein TSG-6 (tumor necrosis factor alpha stimulated gene/protein 6, TNFAIP/TSG-6) leads to dimerization or higher order multimerization of HA chains in solution. We compared two different heparin (HP) samples and two heparan sulfate (HS) samples for the ability to disrupt HA crosslinking by TSG-6. Both HP samples had approximately three sulfates per disaccharide, and both were effective in inhibiting HA crosslinking by TSG-6. HS with a relatively high degree of sulfation (1.75 per disaccharide) also inhibited TSG-6 mediated HA networking, while HS with a lower degree of sulfation (0.75 per disaccharide) was less effective. We further identified Proteoglycan 4 (PRG4, lubricin) as a TSG-6 ligand, and found it to inhibit TSG-6-mediated HA crosslinking. The effects of HP, HS, and PRG4 on HA crosslinking by TSG-6 were shown to be due to HP/HS/PRG4 inhibition of HA binding to the Link domain of TSG-6. Using the AlphaLISA platform, we also tested other HA-binding proteins for ability to create HA networks. The G1 domain of versican (VG1) effectively networked bHA in solution but required a higher concentration than TSG-6. Cartilage link protein (HAPLN1) and the HA binding protein segment of aggrecan (HABP, G1-IGD-G2) showed only low and variable magnitude HA networking effects. This study unambiguously demonstrates HA crosslinking in solution by TSG-6 and VG1 proteins, and establishes PRG4, HP and highly sulfated HS as modulators of TSG-6 mediated HA crosslinking.
Sprache	Englisch
Erscheinungsdatum	2022-10-05
Erscheinungsland	Switzerland
Dokumenttyp	Journal Article
ZDB-ID	2814330-9
ISSN	2296-889X
ISSN	2296-889X
DOI	10.3389/fmolb.2022.990861
Datenquelle	MEDical Literature Analysis and Retrieval System OnLINE

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