Glucocorticosteroids inhibit degradation in bovine cartilage explants stimulated with concomitant plasminogen and interleukin-1

1997 ◽  
Vol 46 (2) ◽  
pp. 60-64 ◽  
Author(s):  
A. J. Augustine ◽  
J. Oleksyszyn
Keyword(s):  
Interleukin 1 ◽  
Cartilage Explants ◽  
Bovine Cartilage

scholarly journals Chondrocyte-mediated catabolism of aggrecan: aggrecanase-dependent cleavage induced by interleukin-1 or retinoic acid can be inhibited by glucosamine

1998 ◽  
Vol 335 (1) ◽  
pp. 59-66 ◽  
Author(s):  
John D. SANDY ◽  
Dan GAMETT ◽  
Vivian THOMPSON ◽  
Christie VERSCHAREN
Keyword(s):  
Retinoic Acid ◽  
Core Protein ◽  
Interleukin 1 ◽  
Cartilage Explants ◽  
Biosynthetic Activity ◽  
Fragment Analysis ◽  
Analysis System ◽  
Chondrosarcoma Cell Line ◽  
Bovine Cartilage ◽  
Aggrecan Core Protein

A rat chondrosarcoma cell line and bovine cartilage explants have been used to study the control of aggrecan degradation by chondrocytes treated with interleukin-1 (IL-1) or retinoic acid (RA). Aggrecan fragment analysis with anti-neo-epitope antibodies suggests that aggrecanase (an as yet unidentified enzyme) is the only aggrecan-degrading proteinase active in these cultures. With rat cells, aggrecanase converts the aggrecan core protein into two major G1-domain-bearing products (60 kDa with a C-terminal Glu-373, and 220 kDa with a C-terminal Glu-1459). Both products were quantified on a standardized Western analysis system with a G1-specific antibody. Immunoblots were analysed by scanning densitometry and the sensitivity, linearity and reproducibility of the assay were established. With rat cells the aggrecanase response to IL-1 was optimal at about 2 mM glutamine, but was progressively inhibited at higher concentrations, with about 90% inhibition at 10 mM glutamine. Such inhibition by glutamine was not, however, observed with bovine explants. On the other hand, marked inhibition of aggrecanase-dependent cleavage was observed with both rat cells and bovine explants when d(+)-glucosamine was included at concentrations above 2 mM. Inhibition was apparently not due to cytotoxicity or interference with IL-1 signalling, since biosynthetic activity was not inhibited and inhibition of the aggrecanase response was also obtained when RA was used as the catabolic stimulator. Possible mechanisms for the inhibition of the aggrecanase response by glucosamine in chondrocytes treated with IL-1 or RA are discussed.

scholarly journals Monoclonal antibodies that specifically recognize neoepitope sequences generated by ‘aggrecanase’ and matrix metalloproteinase cleavage of aggrecan: application to catabolism in situ and in vitro

1995 ◽  
Vol 305 (3) ◽  
pp. 799-804 ◽  
Author(s):  
C E Hughes ◽  
B Caterson ◽  
A J Fosang ◽  
P J Roughley ◽  
J S Mort
Keyword(s):  
Monoclonal Antibodies ◽  
Retinoic Acid ◽  
Degradation Products ◽  
Interleukin 1 ◽  
Proteolytic Cleavage ◽  
Cartilage Explants ◽  
Reactive Material ◽  
Culture Systems ◽  
Bovine Cartilage

Monoclonal antibodies have been prepared that react specifically with the neoepitopes present on proteoglycan degradation products generated from the proteolytic cleavage of aggrecan in the interglobular domain. Antibody BC-3 recognizes the new N-terminus (ARGSV...) on aggrecan degradation products produced by the action of the as yet uncharacterized proteolytic activity, ‘aggrecanase’, and antibody BC-4 recognizes the new C-terminus (...DIPEN) generated by the proteolytic action of matrix metalloproteinases. Specificity for these neoepitope sequences was determined in competitive e.l.i.s.a. using synthetic peptide antigens as inhibitors. Antibody BC-3 was used in the detection of aggrecan degradation products in the culture medium obtained from two different in vitro culture systems: bovine cartilage explants treated with either retinoic acid or interleukin-1, and secondly, rat chondrosarcoma cells treated with retinoic acid. Both interleukin-1 and retinoic acid treatment caused an increase in aggrecan catabolism resulting in an increased release to the medium of specific aggrecan degradation products containing the BC-3 neoepitope generated by the action of ‘aggrecanase'. However, several additional aggrecan catabolites were present that were not immunoreactive with antibody BC-3. In addition, under control conditions, in the bovine cartilage cultures the BC-3 epitope was found on some of these aggrecan catabolites. In contrast, no immune-reactive material was found in the aggrecan degradation products present in control media of rat chondrosarcoma cells cultured in the absence of retinoic acid. Collectively, these results demonstrate that ‘aggrecanase’ activity is not a constitutive event in all cartilage culture systems and also suggest that proteolytic agents other than ‘aggrecanase’ are involved in aggrecan catabolism in normal turnover compared with pathological conditions. Antibody BC-4 was used to demonstrate the identity of the G1 domain of aggrecan following proteolytic cleavage of a purified G1-G2 preparation with collagenase, gelatinase A or stromelysin. The G2 product of this cleavage did not react with antibody BC-3, indicating that, under the experimental conditions used, none of these enzymes exhibited ‘aggrecanase’ activity. It is expected that both of these antibodies will play a pivotal role in detailed studies elucidating molecular mechanisms of aggrecan degradation and they will be particularly useful for the sensitive monitoring of aggrecan degradation products in tissue extracts and body fluids.

scholarly journals A potential key role for alpha-haemolysin ofStaphylococcus aureusin mediating chondrocyte death in septic arthritis

2018 ◽  
Vol 7 (7) ◽  
pp. 457-467 ◽  
Author(s):  
I. D. M. Smith ◽  
K. M. Milto ◽  
C. J. Doherty ◽  
S. G. B. Amyes ◽  
A. H. R. W. Simpson ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Septic Arthritis ◽  
Cartilage Damage ◽  
Laser Scanning Microscopy ◽  
Cartilage Explants ◽  
Confocal Laser ◽  
Dead Cell ◽  
Chondrocyte Viability ◽  
Chondrocyte Death ◽  
Bovine Cartilage

ObjectivesStaphylococcus aureus (S. aureus) is the most commonly implicated organism in septic arthritis, a condition that may be highly destructive to articular cartilage. Previous studies investigating laboratory and clinical strains of S. aureus have demonstrated that potent toxins induced significant chondrocyte death, although the precise toxin or toxins that were involved was unknown. In this study, we used isogenic S. aureus mutants to assess the influence of alpha (Hla)-, beta (Hlb)-, and gamma (Hlg)-haemolysins, toxins considered important for the destruction of host tissue, on in situ bovine chondrocyte viability.MethodsBovine cartilage explants were cultured with isogenic S. aureus mutants and/or their culture supernatants. Chondrocyte viability was then assessed within defined regions of interest in the axial and coronal plane following live- and dead-cell imaging using the fluorescent probes 5-chloromethylfluorescein diacetate and propidium iodide, respectively, and confocal laser-scanning microscopy.ResultsHla-producing mutants caused substantial chondrocyte death compared with the toxin-deficient control (Hla-Hlb-Hlg-), whilst mutants producing Hlb and Hlg in the absence of Hla induced minimal chondrocyte death. Coronal studies established that Hla-induced chondrocyte death started in the superficial zone of cartilage and spread to deeper layers, whereas Hlb and Hlg toxins were without significant effect.ConclusionThis study identified Hla as a highly potent S. aureus toxin that caused rapid chondrocyte death in bovine cartilage, with other toxins or metabolic products produced by the bacteria playing a minor role. The identification of Hla in mediating chondrocyte death may assist in the development of therapeutic strategies aimed at reducing the extent of cartilage damage during and after an episode of septic arthritis. Cite this article: I. D. M. Smith, K. M. Milto, C. J. Doherty, S. G. B. Amyes, A. H. R. W. Simpson, A. C. Hall. A potential key role for alpha-haemolysin of Staphylococcus aureus in mediating chondrocyte death in septic arthritis. Bone Joint Res 2018;7:457–467. DOI: 10.1302/2046-3758.77.BJR-2017-0165.R1.

scholarly journals Lysosomal cysteine endopeptidases mediate interleukin 1-stimulated cartilage proteoglycan degradation

1992 ◽  
Vol 287 (2) ◽  
pp. 657-661 ◽  
Author(s):  
D J Buttle ◽  
J Saklatvala
Keyword(s):  
Interleukin 1 ◽  
Active Form ◽  
Growth Factor Receptor ◽  
Test System ◽  
Significant Inhibition ◽  
Cartilage Explants ◽  
Cartilage Proteoglycan ◽  
Proteoglycan Degradation ◽  
Cysteine Endopeptidases ◽  
Proteoglycan Release

The peptidyl diazomethane inactivator of cysteine endopeptidases, benzyloxycarbonyl-Tyr-Ala-CHN2, was tested as an inhibitor of interleukin 1 alpha-stimulated release of proteoglycan from bovine nasal septum cartilage explants. Like the previously tested epoxidyl peptide proinhibitor trans-epoxysuccinyl-leucylamido-(3-methyl)butane ethyl ester, it proved to be an effective inhibitor of proteoglycan release from cartilage, with significant inhibition at a concentration of 1 microM. The inhibition did not seem to be due to a general toxic effect. The rates of inactivation of the bovine cysteine endopeptidases by the peptidyl diazomethane, the epoxidyl peptide proinhibitor and its active form were determined. Benzyloxycarbonyl-Tyr-Ala-CHN2 proved to be a rapid inactivator of cathepsins L, S and B, but reacted much more slowly with cathepsin H and calpain. Thus it would appear that the latter two enzymes are not implicated in proteoglycan release in our test system. The peptidyl diazomethane and epoxidyl peptide proinhibitor (above) were also tested for their effects on three other interleukin 1-mediated cellular events, namely epidermal growth factor receptor transmodulation, and interleukin 6 and prostaglandin E2 production. In all cases the inactivators did not interfere with the response to interleukin 1 in human gingival fibroblasts. We conclude that one or more of the lysosomal cysteine endopeptidases cathepsins B, L and S mediate interleukin 1-stimulated cartilage proteoglycan degradation without affecting signal transduction.

Biological Mechanisms for Cartilage Repair Using a BioCartilage Scaffold: Cellular Adhesion/Migration and Bioactive Proteins

Cartilage ◽  
2020 ◽  
pp. 194760351990080 ◽  
Author(s):  
Jacqueline Commins ◽  
Rebecca Irwin ◽  
Andrea Matuska ◽  
Margaret Goodale ◽  
Michelle Delco ◽  
...  
Keyword(s):  
Fibrin Glue ◽  
Cartilage Repair ◽  
Matrix Protein ◽  
Cell Attachment ◽  
Interleukin 1 ◽  
Cellular Adhesion ◽  
Cellular Migration ◽  
Cartilage Explants ◽  
Cartilage Defects ◽  
Bioactive Proteins

Objective. BioCartilage is a desiccated, particulated cartilage allograft used for repair of focal cartilage defects. It is mixed with a biologic such as bone marrow concentrate (BMC), pressed into a contained defect, and sealed with fibrin glue. The objective of this study was to assess if BioCartilage could serve as a bioactive scaffold by affecting cellular adhesion, cellular migration, or the release interleukin-1 receptor antagonist protein (IL-1RA), and to identify its full proteomic makeup. Design. Cartilage explants were used to model confined defects. BioCartilage was mixed with BMC, grafted into defects, and sealed with 1 of 5 fibrin glues. Constructs were cultured for 24 or 48 hours and then processed for live/dead microscopy. Chondrocyte and mesenchymal stem cell (MSC) adhesion on BioCartilage was assessed using scanning electron microscopy. Conditioned medium from cultures and the biologics used in the study were assayed for IL-1RA. The protein footprint of BioCartilage was determined using bottom-up proteomics. Results. BioCartilage supported chondrocyte and MSC attachment within 24 hours, and cell viability was retained in all constructs at 24 and 48 hours. Fibrin glue did not inhibit cell attachment. BMC had the highest concentration of IL-1RA. Proteomics yielded 254 proteins, including collagens, proteoglycans, and several bioactive proteins with known anabolic roles including cartilage oligomeric matrix protein. Conclusions. This study suggests that BioCartilage has the chemical composition and architecture to support cell adherence and migration and to provide bioactive proteins, which together should have biologics advantages in cartilage repair beyond its role as a scaffold.

Tissue Shear Deformation Stimulates Proteoglycan and Protein Biosynthesis in Bovine Cartilage Explants

2001 ◽  
Vol 395 (1) ◽  
pp. 41-48 ◽  
Author(s):  
Moonsoo Jin ◽  
Eliot H. Frank ◽  
Thomas M. Quinn ◽  
Ernst B. Hunziker ◽  
Alan J. Grodzinsky
Keyword(s):  
Shear Deformation ◽  
Protein Biosynthesis ◽  
Cartilage Explants ◽  
Bovine Cartilage

Biological Properties of Allogenic Articular Chondrocytes on the Surface of Bovine Cartilage Explants in vitro

2003 ◽  
Vol 50 (8) ◽  
pp. 418-423 ◽  
Author(s):  
G. Kim ◽  
M. Okumura ◽  
D. Bosnakovski ◽  
T. Ishiguro ◽  
T. Kadosawa ◽  
...  
Keyword(s):  
Articular Chondrocytes ◽  
Biological Properties ◽  
Cartilage Explants ◽  
Bovine Cartilage
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