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Home / Equine Research Bank / Can J Vet Res / Recombinant equine interleukin-1beta induces putative mediat...
Canadian journal of veterinary research = Revue canadienne de recherche veterinaire2002; 66(1); 19-25;

Recombinant equine interleukin-1beta induces putative mediators of articular cartilage degradation in equine chondrocytes.

Abstract: Interleukin-1 is considered a central mediator of cartilage loss in osteoarthritis in several species, however an equine recombinant form of this cytokine is not readily available for in vitro use in equine osteoarthritis research. Equine recombinant interleukin-1beta was cloned and expressed and its effects on the expression and activity of selected chondrocytic proteins implicated in cartilage matrix degradation were characterized. Reverse transcriptase polymerase chain reaction methods were used to amplify the entire coding region of the equine IL-1beta mRNA, which was cloned into an expression vector, expressed in E. coli, and purified using a Ni2+ chromatographic method. The effects of the recombinant peptide on chondrocyte gene expression were determined by Northern blotting using RNA from equine chondrocyte cultures hybridized to probes for matrix metalloproteinases (MMP 1, MMP 3, MMP 13), tissue inhibitor of matrix metalloproteinases 1 (TIMP 1) and cyclooxygenase 2 (COX 2). Effects on selected mediators of cartilage degradation (nitrite concentrations and MMP activity) were determined using conditioned medium from reIL-1beta-treated equine cartilage explant cultures. A recombinant peptide of approximately 21 kd was obtained. Northern blotting analyses revealed a marked up-regulation of expression of all MMPs, TIMP 1, and COX 2 in mRNA from treated chondrocytes. Furthermore, cartilage explants exposed to reIL-1beta had augmented collagenase/gelatinase and stromelysin activities as well as increased concentration of nitrite in conditioned media. The development of a biologically active, species-specific IL-1beta provides a valuable tool in the study of osteoarthritis pathophysiology and its treatment in horses.
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Publication Date: 2002-02-23 PubMed ID: 11858644PubMed Central: PMC226977
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  • Journal Article
  • Research Support
  • Non-U.S. Gov't
  • Research Support
  • U.S. Gov't
  • Non-P.H.S.

Summary

This research summary has been generated with artificial intelligence and may contain errors and omissions. Refer to the original study to confirm details provided. Submit correction.

This research is about the development and testing of a species-specific version of a protein called interleukin-1beta, which is believed to play a critical role in cartilage loss in osteoarthritis, specifically in horses.

Objective of the Research

The goal of this study was to produce and evaluate the effects of a horse-specific form of interleukin-1beta, a protein thought to be a key player in the degradation of cartilage in osteoarthritis. Since an equine version of this protein wasn’t readily available for lab work, the researchers sought to clone and express it themselves.

Methodology

  • The researchers used reverse transcriptase polymerase chain reaction methods to amplify the genetic coding for equine interleukin-1beta, which they then cloned into an expression vector.
  • They expressed the gene in E.coli and then purified the produced protein using a Ni2+ chromatographic method.
  • Once they had the protein, they evaluated its effects on the gene expression of horse chondrocytes (cartilage cells) by testing it on equine chondrocyte cultures and conducting Northern blotting to analyze the expression of matrix metalloproteinases (MMP 1, MMP 3, MMP 13) and other related genes.

Findings

  • The team successfully produced a recombinant peptide of about 21 kilodaltons in size.
  • Their tests revealed that treating the chondrocytes with the peptide resulted in a sharp increase in the expression of all tested matrix metalloproteinases and other associated genes. Matrix metalloproteinases are enzymes that break down the extracellular matrix in normal physiological processes.
  • Exposure to the peptide also boosted collagenase/gelatinase activities and elevated the concentration of nitrite in the conditioned media, indicating enhanced cartilage degradation.

Conclusion

The research team successfully developed an equine-specific version of interleukin-1beta and showed that it promotes the activity of factors implicated in cartilage loss. This form of the protein could serve as a vital tool in studying the pathophysiology of osteoarthritis in horses and in seeking potential treatments.

Cite This Article

APA
Tung JT, Fenton JI, Arnold C, Alexander L, Yuzbasiyan-Gurkan V, Venta PJ, Peters TL, Orth MW, Richardson DW, Caron JP. (2002). Recombinant equine interleukin-1beta induces putative mediators of articular cartilage degradation in equine chondrocytes. Can J Vet Res, 66(1), 19-25.

Publication

Canadian journal of veterinary research = Revue canadienne de recherche veterinaire
ISSN: 0830-9000
NlmUniqueID: 8607793
Country: Canada
Language: English
Volume: 66
Issue: 1
Pages: 19-25

Researcher Affiliations

Tung, J T
  • Department of Animal Science, Michigan State University, East Lansing 48824, USA.
Fenton, J I
    Arnold, C
      Alexander, L
        Yuzbasiyan-Gurkan, V
          Venta, P J
            Peters, T L
              Orth, M W
                Richardson, D W
                  Caron, J P

                    MeSH Terms

                    • Animals
                    • Blotting, Northern / veterinary
                    • Cartilage, Articular / drug effects
                    • Cartilage, Articular / enzymology
                    • Cartilage, Articular / pathology
                    • Cells, Cultured
                    • Chondrocytes / drug effects
                    • Chondrocytes / enzymology
                    • Cyclooxygenase 2
                    • Gene Expression Regulation, Enzymologic
                    • Horse Diseases / physiopathology
                    • Horses
                    • Interleukin-1 / pharmacology
                    • Interleukin-1 / physiology
                    • Isoenzymes / genetics
                    • Isoenzymes / metabolism
                    • Matrix Metalloproteinases / genetics
                    • Matrix Metalloproteinases / metabolism
                    • Osteoarthritis / physiopathology
                    • Osteoarthritis / veterinary
                    • Prostaglandin-Endoperoxide Synthases / genetics
                    • Prostaglandin-Endoperoxide Synthases / metabolism
                    • RNA, Messenger / metabolism
                    • Recombinant Proteins / metabolism
                    • Recombinant Proteins / pharmacology
                    • Reverse Transcriptase Polymerase Chain Reaction / veterinary
                    • Tissue Inhibitor of Metalloproteinase-1 / genetics
                    • Tissue Inhibitor of Metalloproteinase-1 / metabolism

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