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American journal of veterinary research2012; 73(7); 1047-1056; doi: 10.2460/ajvr.73.7.1047

Effects of cleavage by a disintegrin and metalloproteinase with thrombospondin motifs-4 on gene expression and protein content of versican and aggrecan in the digital laminae of horses with starch gruel-induced laminitis.

Abstract: To determine whether increased gene expression of a disintegrin and metalloproteinase with thrombospondin motifs-4 (ADAMTS-4) in laminae of horses with starch gruel-induced laminitis was accompanied by increased enzyme activity and substrate degradation. Methods: Laminae from the forelimb hooves of 8 healthy horses and 17 horses with starch gruel-induced laminitis (6 at onset of fever, 6 at onset of Obel grade 1 lameness, and 5 at onset of Obel grade 3 lameness). Methods: Gene expression was determined by use of cDNA and real-time quantitative PCR assay. Protein expression and processing were determined via SDS-PAGE and quantitative western blotting. Protein distribution and abundance were determined via quantitative immunofluorescent staining. Results: ADAMTS-4 gene expression was increased and that of versican decreased in laminitic laminae, compared with expression in healthy laminae. Catalytically active ADAMTS-4 also was increased in the tissue, as were ADAMTS-4-cleavage fragments of versican. Immunofluorescent analyses indicated that versican was depleted from the basal epithelia of laminae of horses at onset of Obel grade 3 lameness, compared with results for healthy laminae, and this was accompanied by regional separation of basal epithelial cells from the basement membrane. Aggrecan gene and protein expression were not significantly affected. Conclusions: Changes in gene and protein expression of ADAMTS-4 and versican in the basal epithelium of laminitic laminae indicated a fundamental change in the physiology of basal epithelial cells. This was accompanied by and may have caused detachment of these cells from the basement membrane.
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Publication Date: 2012-06-29 PubMed ID: 22738057PubMed Central: PMC3535458DOI: 10.2460/ajvr.73.7.1047Google Scholar: Lookup
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  • Journal Article
  • Research Support
  • N.I.H.
  • Extramural
  • 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 investigates the role of a specific enzyme, ADAMTS-4, in the onset of laminitis in horses, a disease causing inflammation and degradation in hoof tissue. The study found that the increase in ADAMTS-4 activity correlates with greater tissue damage and altered gene expression which may lead to detachment of cells within the hoof structure.

Research Methodology

  • The study was conducted on a total of 25 horses. Eight of these were healthy while 17 suffered from starch gruel-induced laminitis. The affected horses were subdivided based on the stages of the disease – onset of fever (6 horses), onset of Obel grade 1 lameness (6 horses), and onset of Obel grade 3 lameness (5 horses).
  • The gene expression was determined using cDNA and real-time quantitative PCR assay, a technique that quantifies RNA in a sample via conversion to DNA.
  • Protein analysis was conducted using SDS-PAGE and quantitative western blotting. These techniques help to separate, identify, and quantify proteins within the tissue samples.
  • Immunofluorescent staining was used to determine the distribution and abundance of proteins.

Research Findings

  • There was an increased gene expression of ADAMTS-4 and decrease of versican gene expression in the laminitic laminae, when compared with healthy tissue.
  • Active ADAMTS-4 and ADAMTS-4-cleavage fragments of versican were observed more frequently in the tissue of the affected horses, indicating heightened enzyme activity.
  • Immunofluorescent analysis showed depleted versican protein in the basal epithelium of laminae of horses at the onset of Obel grade 3 lameness. This depletion was accompanied by the separation of these cells from the basement membrane.
  • There was a region-specific change in the gene and protein expression of ADAMTS-4 and versican, highlighting a fundamental alteration in the physiology of basal epithelial cells during the disease progression.
  • Interestingly, the gene and protein expression of Aggrecan were not significantly affected during the process.

Conclusions

  • The study proposes that the increase in the activity of enzyme ADAMTS-4 may cause the degradation of hoof tissue. This alteration in gene and protein expression affects cellular behavior which possibly leads to their detachment from the hoof structure.
  • These findings suggest that ADAMTS-4 might play a key role in the development of laminitis and may provide a targeted approach for the treatment of this condition in horses.

Cite This Article

APA
Wang L, Pawlak E, Johnson PJ, Belknap JK, Alfandari D, Black SJ. (2012). Effects of cleavage by a disintegrin and metalloproteinase with thrombospondin motifs-4 on gene expression and protein content of versican and aggrecan in the digital laminae of horses with starch gruel-induced laminitis. Am J Vet Res, 73(7), 1047-1056. https://doi.org/10.2460/ajvr.73.7.1047

Publication

American journal of veterinary research
ISSN: 1943-5681
NlmUniqueID: 0375011
Country: United States
Language: English
Volume: 73
Issue: 7
Pages: 1047-1056

Researcher Affiliations

Wang, Le
  • Department of Veterinary and Animal Sciences, College of Natural Sciences, University of Massachusetts, Amherst, MA 01003, USA.
Pawlak, Erica
    Johnson, Philip J
      Belknap, James K
        Alfandari, Dominique
          Black, Samuel J

            MeSH Terms

            • ADAM Proteins / genetics
            • ADAM Proteins / metabolism
            • ADAMTS4 Protein
            • Aggrecans / genetics
            • Aggrecans / metabolism
            • Animals
            • Blotting, Western / veterinary
            • Enzyme Activation
            • Foot Diseases / genetics
            • Foot Diseases / metabolism
            • Foot Diseases / veterinary
            • Gene Expression Profiling / veterinary
            • Hoof and Claw / metabolism
            • Horse Diseases / genetics
            • Horse Diseases / metabolism
            • Horses
            • Procollagen N-Endopeptidase / genetics
            • Procollagen N-Endopeptidase / metabolism
            • RNA / chemistry
            • RNA / genetics
            • Real-Time Polymerase Chain Reaction / veterinary
            • Statistics, Nonparametric
            • Versicans / genetics
            • Versicans / metabolism

            Grant Funding

            • R01 DE016289 / NIDCR NIH HHS
            • DE016289 / NIDCR NIH HHS

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            Citations

            This article has been cited 6 times.
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