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International journal of molecular sciences2020; 21(6); 2155; doi: 10.3390/ijms21062155

Development of a Cartilage Oligomeric Matrix Protein Neo-Epitope Assay for the Detection of Intra-Thecal Tendon Disease.

Abstract: The diagnosis of tendon injury relies on clinical signs and diagnostic imaging but imaging is subjective and does not always correlate with clinical signs. A molecular marker would potentially offer a sensitive and specific diagnostic tool that could also provide objective assessment of healing for the comparison of different treatments. Cartilage Oligomeric Matrix Protein (COMP) has been used as a molecular marker for osteoarthritis in humans and horses but assays for the protein in tendon sheath synovial fluids have shown overlap between horses affected by tendinopathy and controls. We hypothesized that quantifying a COMP neoepitope would be more discriminatory of injury. COMP fragments were purified from synovial fluids of horses with intra-thecal tendon injuries and media from equine tendon explants, and mass spectrometry of a consistent and abundant fragment revealed a ~100 kDa COMP fragment with a new N-terminus at the 78th amino-acid (NH-TPRVSVRP) located just outside the junctional region of the protein. A competitive inhibition ELISA based on a polyclonal antibody raised to this sequence yielded more than a 10-fold rise in the mean neoepitope levels for tendinopathy cases compared to controls (5.3 ± 1.3 µg/mL ( = 7) versus 58.8 ± 64.3 µg/mL ( = 13); = 0.002). However, there was some cross-reactivity of the neoepitope polyclonal antiserum with intact COMP, which could be blocked by a peptide spanning the neoepitope. The modified assay demonstrated a lower concentration but a significant > 500-fold average rise with tendon injury (2.5 ± 2.2 ng/mL ( = 6) versus 1029.8 ± 2188.8 ng/ml ( = 14); = 0.013). This neo-epitope assay therefore offers a potentially useful marker for clinical use.
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Publication Date: 2020-03-20 PubMed ID: 32245107PubMed Central: PMC7139564DOI: 10.3390/ijms21062155Google Scholar: Lookup
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  • Journal Article

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.

The research focuses on developing a method that uses Cartilage Oligomeric Matrix Protein (COMP) as a molecular marker to detect tendon injury in horses. The study suggests that this new technique could be far more specific and sensitive in diagnosing such injuries, comparing favourably to current imaging-based methods.

Objective of the Study

  • The objective of the study was to develop a sensitive and specific diagnostic tool for the detection of tendon injury in horses that could possibly replace subjective imaging methods.
  • The researchers chose to test COMP as a potential molecular marker, as previous studies have shown it to be effective in diagnosing osteoarthritis in humans and horses.

Research Methods

  • The researchers hypothesized that quantifying a COMP neoepitope (a new antigen that is generated after a protein undergoes a certain process) would be more indicative of tendon injury.
  • COMP fragments were extracted from the synovial fluid (lubricating fluid between joints) of horses that had tendon injuries (intra-thecal), and from tendon explants in an equine laboratory.
  • A competitive inhibition ELISA (a type of analysis used for measuring the concentration of antibodies in samples) was used to discover the concentration of the new COMP neoepitope.

Results and Findings

  • Through mass spectrometry (a powerful technique used for identifying and quantifying substances within a sample), they found a COMP fragment with a new end at the 78th amino acid.
  • An ELISA based on an antibody raised to this sequence resulted in a more than ten-fold rise in the levels of the neoepitope in the cases of tendinopathy (a tendon disorder) when compared to the control group.
  • There occurred some cross-reactivity between the neoepitope and the intact COMP, this was counteracted with a peptide overlapping the neoepitope.
  • After refinement in the assay, it still showed a significant average rise in concentration (of over 500-fold) during a tendon injury.

Conclusions and Outlook

  • Overall, the study demonstrated that this modified neo-epitope assay could act as a powerful indicator of tendon injury, presenting a potential new diagnostic tool.
  • This research could have considerable impact on clinical methods for treating and managing tendon diseases in horses.

Cite This Article

APA
Smith R, Önnerfjord P, Holmgren K, di Grado S, Dudhia J. (2020). Development of a Cartilage Oligomeric Matrix Protein Neo-Epitope Assay for the Detection of Intra-Thecal Tendon Disease. Int J Mol Sci, 21(6), 2155. https://doi.org/10.3390/ijms21062155

Publication

International journal of molecular sciences
ISSN: 1422-0067
NlmUniqueID: 101092791
Country: Switzerland
Language: English
Volume: 21
Issue: 6
PII: 2155

Researcher Affiliations

Smith, Roger
  • Dept of Clinical Sciences and Services, The Royal Veterinary College, Hawkshead Lane, North Mymms, Hatfield, Herts AL9 7TA, UK.
Önnerfjord, Patrik
  • Department of Clinical Sciences, Rheumatology and Molecular Skeletal Biology, University of Lund, 22184 Lund, Sweden.
Holmgren, Kristin
  • Department of Clinical Sciences, Rheumatology and Molecular Skeletal Biology, University of Lund, 22184 Lund, Sweden.
di Grado, Shacko
  • Dept of Clinical Sciences and Services, The Royal Veterinary College, Hawkshead Lane, North Mymms, Hatfield, Herts AL9 7TA, UK.
  • Anicura Regiondjursjukhuset Bagarmossen, Ljusnevägen 17, 128 48 Bagarmossen, Sweden.
Dudhia, Jayesh
  • Dept of Clinical Sciences and Services, The Royal Veterinary College, Hawkshead Lane, North Mymms, Hatfield, Herts AL9 7TA, UK.

MeSH Terms

  • Amino Acid Sequence
  • Animals
  • Antibodies / immunology
  • Biological Assay / methods
  • Cartilage Oligomeric Matrix Protein / chemistry
  • Cartilage Oligomeric Matrix Protein / immunology
  • Cartilage Oligomeric Matrix Protein / metabolism
  • Cross Reactions / immunology
  • Epitopes / metabolism
  • Horses
  • Spinal Cord / pathology
  • Synovial Fluid / metabolism
  • Tendon Injuries / diagnosis
  • Tendon Injuries / metabolism
  • Tendons / pathology

Grant Funding

  • VET/PRJ/674 and VET/PRJ/703 / Horserace Betting Levy Board

Conflict of Interest Statement

There are no conflicts of interest.

References

This article includes 42 references
  1. Avella CS, Ely ER, Verheyen KL, Price JS, Wood JL, Smith RK. Ultrasonographic assessment of the superficial digital flexor tendons of National Hunt racehorses in training over two racing seasons.. Equine Vet J 2009 May;41(5):449-54.
    doi: 10.2746/042516409X391042pubmed: 19642404google scholar: lookup
  2. Suchak AA, Bostick G, Reid D, Blitz S, Jomha N. The incidence of Achilles tendon ruptures in Edmonton, Canada.. Foot Ankle Int 2005 Nov;26(11):932-6.
    doi: 10.1177/107110070502601106pubmed: 16309606google scholar: lookup
  3. Kujala UM, Sarna S, Kaprio J. Cumulative incidence of achilles tendon rupture and tendinopathy in male former elite athletes.. Clin J Sport Med 2005 May;15(3):133-5.
    doi: 10.1097/01.jsm.0000165347.55638.23pubmed: 15867554google scholar: lookup
  4. Järvinen TA, Kannus P, Maffulli N, Khan KM. Achilles tendon disorders: etiology and epidemiology.. Foot Ankle Clin 2005 Jun;10(2):255-66.
    doi: 10.1016/j.fcl.2005.01.013pubmed: 15922917google scholar: lookup
  5. Patterson-Kane JC, Rich T. Achilles tendon injuries in elite athletes: lessons in pathophysiology from their equine counterparts.. ILAR J 2014;55(1):86-99.
    doi: 10.1093/ilar/ilu004pubmed: 24936032google scholar: lookup
  6. Lui PP, Maffulli N, Rolf C, Smith RK. What are the validated animal models for tendinopathy?. Scand J Med Sci Sports 2011 Feb;21(1):3-17.
    doi: 10.1111/j.1600-0838.2010.01164.xpubmed: 20673247google scholar: lookup
  7. Happonen KE, Heinegård D, Saxne T, Blom AM. Interactions of the complement system with molecules of extracellular matrix: relevance for joint diseases.. Immunobiology 2012 Nov;217(11):1088-96.
    doi: 10.1016/j.imbio.2012.07.013pubmed: 22964234google scholar: lookup
  8. Garvican ER, Salavati M, Smith RKW, Dudhia J. Exposure of a tendon extracellular matrix to synovial fluid triggers endogenous and engrafted cell death: A mechanism for failed healing of intrathecal tendon injuries.. Connect Tissue Res 2017 Sep;58(5):438-446.
    doi: 10.1080/03008207.2016.1245726pubmed: 27726447google scholar: lookup
  9. Smith MR, Wright IM, Smith RK. Endoscopic assessment and treatment of lesions of the deep digital flexor tendon in the navicular bursae of 20 lame horses.. Equine Vet J 2007 Jan;39(1):18-24.
    doi: 10.2746/042516407X151095pubmed: 17228590google scholar: lookup
  10. Smith MR, Wright IM. Noninfected tenosynovitis of the digital flexor tendon sheath: a retrospective analysis of 76 cases.. Equine Vet J 2006 Mar;38(2):134-41.
    doi: 10.2746/042516406776563350pubmed: 16536382google scholar: lookup
  11. Smith MRW, Wright IM, Smith RKW, Bladon B, Schramme MC. Treatment options for DDFT lesions: Evidence based results. Proceedings of the 2009 ECVS Annual Conference Nantes, France. 2–4 July 2009.
  12. Young-Min S, Cawston T, Marshall N, Coady D, Christgau S, Saxne T, Robins S, Griffiths I. Biomarkers predict radiographic progression in early rheumatoid arthritis and perform well compared with traditional markers.. Arthritis Rheum 2007 Oct;56(10):3236-47.
    doi: 10.1002/art.22923pubmed: 17907159google scholar: lookup
  13. Lindqvist E, Eberhardt K, Bendtzen K, Heinegård D, Saxne T. Prognostic laboratory markers of joint damage in rheumatoid arthritis.. Ann Rheum Dis 2005 Feb;64(2):196-201.
    doi: 10.1136/ard.2003.019992pmc: PMC1755350pubmed: 15458956google scholar: lookup
  14. Månsson B, Geborek P, Saxne T. Cartilage and bone macromolecules in knee joint synovial fluid in rheumatoid arthritis: relation to development of knee or hip joint destruction.. Ann Rheum Dis 1997 Feb;56(2):91-6.
    doi: 10.1136/ard.56.2.91pmc: PMC1752317pubmed: 9068280google scholar: lookup
  15. Månsson B, Carey D, Alini M, Ionescu M, Rosenberg LC, Poole AR, Heinegård D, Saxne T. Cartilage and bone metabolism in rheumatoid arthritis. Differences between rapid and slow progression of disease identified by serum markers of cartilage metabolism.. J Clin Invest 1995 Mar;95(3):1071-7.
    doi: 10.1172/JCI117753pmc: PMC441442pubmed: 7533784google scholar: lookup
  16. Saxne T, Heinegård D. Cartilage oligomeric matrix protein: a novel marker of cartilage turnover detectable in synovial fluid and blood.. Br J Rheumatol 1992 Sep;31(9):583-91.
    doi: 10.1093/rheumatology/31.9.583pubmed: 1381980google scholar: lookup
  17. Åhrman E, Lorenzo P, Holmgren K, Grodzinsky AJ, Dahlberg LE, Saxne T, Heinegård D, Önnerfjord P. Novel cartilage oligomeric matrix protein (COMP) neoepitopes identified in synovial fluids from patients with joint diseases using affinity chromatography and mass spectrometry.. J Biol Chem 2014 Jul 25;289(30):20908-16.
    doi: 10.1074/jbc.M114.554683pmc: PMC4110297pubmed: 24917676google scholar: lookup
  18. Lorenzo P, Aspberg A, Saxne T, Önnerfjord P. Quantification of cartilage oligomeric matrix protein (COMP) and a COMP neoepitope in synovial fluid of patients with different joint disorders by novel automated assays.. Osteoarthritis Cartilage 2017 Sep;25(9):1436-1442.
    doi: 10.1016/j.joca.2017.04.004pubmed: 28473207google scholar: lookup
  19. Arai K, Misumi K, Carter SD, Shinbara S, Fujiki M, Sakamoto H. Analysis of cartilage oligomeric matrix protein (COMP) degradation and synthesis in equine joint disease.. Equine Vet J 2005 Jan;37(1):31-6.
    doi: 10.2746/0425164054406784pubmed: 15651731google scholar: lookup
  20. Arai K, Tagami M, Hatazoe T, Nishimatsu E, Shimizu Y, Fujiki M, Misumi K. Analysis of cartilage oligomeric matrix protein (COMP) in synovial fluid, serum and urine from 51 racehorses with carpal bone fracture.. J Vet Med Sci 2008 Sep;70(9):915-21.
    doi: 10.1292/jvms.70.915pubmed: 18840965google scholar: lookup
  21. Misumi K, Tagami M, Kamimura T, Miyakoshi D, Helal IE, Arai K, Fujiki M. Urine cartilage oligomeric matrix protein (COMP) measurement is useful in discriminating the osteoarthritic Thoroughbreds.. Osteoarthritis Cartilage 2006 Nov;14(11):1174-80.
    doi: 10.1016/j.joca.2006.04.017pubmed: 16895759google scholar: lookup
  22. Taylor SE, Weaver MP, Pitsillides AA, Wheeler BT, Wheeler-Jones CP, Shaw DJ, Smith RK. Cartilage oligomeric matrix protein and hyaluronan levels in synovial fluid from horses with osteoarthritis of the tarsometatarsal joint compared to a control population.. Equine Vet J 2006 Nov;38(6):502-7.
    doi: 10.2746/042516406X156073pubmed: 17124839google scholar: lookup
  23. Misumi K, Vilim V, Hatazoe T, Murata T, Fujiki M, Oka T, Sakamoto H, Carter SD. Serum level of cartilage oligomeric matrix protein (COMP) in equine osteoarthritis.. Equine Vet J 2002 Sep;34(6):602-8.
    doi: 10.2746/042516402776180205pubmed: 12358001google scholar: lookup
  24. Misumi K, Vilim V, Clegg PD, Thompson CC, Carter SD. Measurement of cartilage oligomeric matrix protein (COMP) in normal and diseased equine synovial fluids.. Osteoarthritis Cartilage 2001 Feb;9(2):119-27.
    doi: 10.1053/joca.2000.0367pubmed: 11237659google scholar: lookup
  25. Yamanokuchi K, Tagami M, Nishimatsu E, Shimizu Y, Hirose Y, Komatsu K, Misumi K. Sandwich ELISA system for cartilage oligomeric matrix protein in equine synovial fluid and serum.. Equine Vet J 2009 Jan;41(1):41-6.
    doi: 10.2746/042516408X330356pubmed: 19301580google scholar: lookup
  26. Skiöldebrand E, Lorenzo P, Zunino L, Rucklidge GJ, Sandgren B, Carlsten J, Ekman S. Concentration of collagen, aggrecan and cartilage oligomeric matrix protein (COMP) in synovial fluid from equine middle carpal joints.. Equine Vet J 2001 Jul;33(4):394-402.
    doi: 10.2746/042516401776249480pubmed: 11469774google scholar: lookup
  27. Skiöldebrand E, Heinegård D, Eloranta ML, Nilsson G, Dudhia J, Sandgren B, Ekman S. Enhanced concentration of COMP (cartilage oligomeric matrix protein) in osteochondral fractures from racing Thoroughbreds.. J Orthop Res 2005 Jan;23(1):156-63.
    doi: 10.1016/j.orthres.2004.05.013pubmed: 15607888google scholar: lookup
  28. Skiöldebrand E, Ekman S, Mattsson Hultén L, Svala E, Björkman K, Lindahl A, Lundqvist A, Önnerfjord P, Sihlbom C, Rüetschi U. Cartilage oligomeric matrix protein neoepitope in the synovial fluid of horses with acute lameness: A new biomarker for the early stages of osteoarthritis.. Equine Vet J 2017 Sep;49(5):662-667.
    doi: 10.1111/evj.12666pmc: PMC5573946pubmed: 28097685google scholar: lookup
  29. Smith RK, Zunino L, Webbon PM, Heinegård D. The distribution of cartilage oligomeric matrix protein (COMP) in tendon and its variation with tendon site, age and load.. Matrix Biol 1997 Nov;16(5):255-71.
    doi: 10.1016/S0945-053X(97)90014-7pubmed: 9501326google scholar: lookup
  30. Smith RK, Birch HL, Goodman S, Heinegård D, Goodship AE. The influence of ageing and exercise on tendon growth and degeneration--hypotheses for the initiation and prevention of strain-induced tendinopathies.. Comp Biochem Physiol A Mol Integr Physiol 2002 Dec;133(4):1039-50.
    doi: 10.1016/S1095-6433(02)00148-4pubmed: 12485691google scholar: lookup
  31. Smith RK, Birch H, Patterson-Kane J, Firth EC, Williams L, Cherdchutham W, van Weeren WR, Goodship AE. Should equine athletes commence training during skeletal development?: changes in tendon matrix associated with development, ageing, function and exercise.. Equine Vet J Suppl 1999 Jul;(30):201-9.
    doi: 10.1111/j.2042-3306.1999.tb05218.xpubmed: 10659252google scholar: lookup
  32. Smith RK, Heinegård D. Cartilage oligomeric matrix protein (COMP) levels in digital sheath synovial fluid and serum with tendon injury.. Equine Vet J 2000 Jan;32(1):52-8.
    doi: 10.2746/042516400777612053pubmed: 10661386google scholar: lookup
  33. Smith MR, Wright IM, Minshall GJ, Dudhia J, Verheyen K, Heinegård D, Smith RK. Increased cartilage oligomeric matrix protein concentrations in equine digital flexor tendon sheath synovial fluid predicts intrathecal tendon damage.. Vet Surg 2011 Jan;40(1):54-8.
    doi: 10.1111/j.1532-950X.2010.00751.xpubmed: 21077920google scholar: lookup
  34. Johnson A, Smith R, Saxne T, Hickery M, Heinegård D. Fibronectin fragments cause release and degradation of collagen-binding molecules from equine explant cultures.. Osteoarthritis Cartilage 2004 Feb;12(2):149-59.
    doi: 10.1016/j.joca.2003.10.008pubmed: 14723874google scholar: lookup
  35. Dakin SG, Smith RK, Heinegård D, Önnerfjord P, Khabut A, Dudhia J. Proteomic analysis of tendon extracellular matrix reveals disease stage-specific fragmentation and differential cleavage of COMP (cartilage oligomeric matrix protein).. J Biol Chem 2014 Feb 21;289(8):4919-27.
    doi: 10.1074/jbc.M113.511972pmc: PMC3931053pubmed: 24398684google scholar: lookup
  36. Peffers MJ, Thornton DJ, Clegg PD. Characterization of neopeptides in equine articular cartilage degradation.. J Orthop Res 2016 Jan;34(1):106-20.
    doi: 10.1002/jor.22963pmc: PMC4737130pubmed: 26124002google scholar: lookup
  37. Thur J, Rosenberg K, Nitsche DP, Pihlajamaa T, Ala-Kokko L, Heinegård D, Paulsson M, Maurer P. Mutations in cartilage oligomeric matrix protein causing pseudoachondroplasia and multiple epiphyseal dysplasia affect binding of calcium and collagen I, II, and IX.. J Biol Chem 2001 Mar 2;276(9):6083-92.
    doi: 10.1074/jbc.M009512200pubmed: 11084047google scholar: lookup
  38. Danfelter M, Onnerfjord P, Heinegård D. Fragmentation of proteins in cartilage treated with interleukin-1: specific cleavage of type IX collagen by matrix metalloproteinase 13 releases the NC4 domain.. J Biol Chem 2007 Dec 21;282(51):36933-41.
    doi: 10.1074/jbc.M702491200pubmed: 17951262google scholar: lookup
  39. Gough MR, Munroe GA, Mayhew G. Urea as a measure of dilution of equine synovial fluid.. Equine Vet J 2002 Jan;34(1):76-9.
    doi: 10.2746/042516402776181213pubmed: 11817555google scholar: lookup
  40. Brink P, Smith RK, Tverdal A, Dolvik NI. Changes in synovial fluid biomarker concentrations following arthroscopic surgery in horses with osteochondritis dissecans of the distal intermediate ridge of the tibia.. Am J Vet Res 2015 Jul;76(7):599-607.
    doi: 10.2460/ajvr.76.7.599pubmed: 26111089google scholar: lookup
  41. Wright IM, McMahon PJ. Tenosynovitis associated with longitudinal tears of the digital flexor tendons in horses: a report of 20 cases.. Equine Vet J 1999 Jan;31(1):12-8.
    doi: 10.1111/j.2042-3306.1999.tb03785.xpubmed: 9952324google scholar: lookup
  42. Jackson BE, Smith RK, Price JS. A molecular marker of type I collagen metabolism reflects changes in connective tissue remodelling associated with injury to the equine superficial digital flexor tendon.. Equine Vet J 2003 Mar;35(2):211-3.
    doi: 10.2746/042516403776114135pubmed: 12638801google scholar: lookup

Citations

This article has been cited 6 times.
  1. Docheva D. Editorial for Special Issue: Achilles Curse and Remedy: Tendon Diseases from Pathophysiology to Novel Therapeutic Approaches. Int J Mol Sci 2020 Oct 9;21(20).
    doi: 10.3390/ijms21207454pubmed: 33050349google scholar: lookup
  2. Guadalupi M, Girelli CR, Della Tommasa S, Corte FD, Crovace AM, Fanizzi FP, Brehm W, Lacitignola L. Metabolomic analysis of synovial fluid from healthy and pathological equine joints and tendon sheaths using high-resolution (1)H Nuclear Magnetic Resonance. Front Vet Sci 2025;12:1671176.
    doi: 10.3389/fvets.2025.1671176pubmed: 41477162google scholar: lookup
  3. Iwasaki N, Llewellyn J, Brown J, Zamboulis DE, Finding EJT, Wheeler-Jones CPD, Thorpe CT. Immunolabelling and Micro-Computed Tomography Revealed Age-Related Alterations in 3D Microvasculature of Tendons. Aging Cell 2026 Jan;25(1):e70293.
    doi: 10.1111/acel.70293pubmed: 41250917google scholar: lookup
  4. Smith MM, Melrose J. COMP Is a Biomarker of Cartilage Destruction, Extracellular Matrix and Vascular Remodeling and Tissue Repair. Int J Mol Sci 2025 Sep 19;26(18).
    doi: 10.3390/ijms26189182pubmed: 41009743google scholar: lookup
  5. Li J, Wen M, Zhang S, Du L, Fan X, Liang H, Wang H, Sun J, Ding Y, Ge L, Ma J, Zhang J. Research Progress on the Preparation and Application of Decellularized Tendons. Curr Issues Mol Biol 2025 Apr 6;47(4).
    doi: 10.3390/cimb47040251pubmed: 40699650google scholar: lookup
  6. Li J, Sun H, Guan J, Li B, Jin C, Xie S, Liu Y. Immunogenicity of chondrocyte sheets: a review. Front Immunol 2025;16:1529384.
    doi: 10.3389/fimmu.2025.1529384pubmed: 40124370google scholar: lookup
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