Annals of Biomedical Engineering

, Volume 43, Issue 9, pp 2036–2046 | Cite as

Cathepsins in Rotator Cuff Tendinopathy: Identification in Human Chronic Tears and Temporal Induction in a Rat Model

  • Song P. Seto
  • Akia N. Parks
  • Yongzhi Qiu
  • Louis J. Soslowsky
  • Spero Karas
  • Manu O. Platt
  • Johnna S. TemenoffEmail author


While overuse of the supraspinatus tendon is a leading factor in rotator cuff injury, the underlying biochemical changes have not been fully elucidated. In this study, torn human rotator cuff (supraspinatus) tendon tissue was analyzed for the presence of active cathepsin proteases with multiplex cysteine cathepsin zymography. In addition, an overuse injury to supraspinatus tendons was induced through downhill running in an established rat model. Histological analysis demonstrated that structural damage occurred by 8 weeks of overuse compared to control rats in the region of tendon insertion into bone. In both 4- and 8-week overuse groups, via zymography, there was approximately a 180% increase in cathepsin L activity at the insertion region compared to the controls, while no difference was found in the midsubstance area. Additionally, an over 400% increase in cathepsin K activity was observed for the insertion region of the 4-week overused tendons. More cathepsin K and L immunostaining was observed at the insertion region of the overuse groups compared to controls. These results provide important information on a yet unexplored mechanism for tendon degeneration that may operate alone or in conjunction with other proteases to contribute to chronic tendinopathy.


Supraspinatus tendon Proteases Tendon tear Cathepsins 



Matrix metalloproteinase


Optimum cutting temperature


Hematoxylin and eosin


Sodium dodecyl sulfate



The authors thank Jennifer Lei for assistance in animal studies, Meredith Fay and Ang (Kevin) Li for help in tissue processing, and Bernard Kippelen’s laboratory at Georgia Tech for use of the circular polarized microscope. This study was supported by a National Football League Charities Medical Grant, a Regenerative Engineering and Medicine Seed Grant (REM) from Georgia Tech and Emory University through the Atlanta Clinical & Translational Science Institute (Advancing Translational Sciences of the National Institutes of Health, UL1TR000454) and the National Institute of Arthritis and Musculoskeletal and Skin Diseases of the National Institutes of Health under Award Number R01AR063692. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.


  1. 1.
    Archambault, J. M., S. A. Jelinsky, S. P. Lake, et al. Rat supraspinatus tendon expresses cartilage markers with overuse. J. Orthop. Res. 25(5):617–624, 2007.PubMedCrossRefGoogle Scholar
  2. 2.
    Attia, M., A. Scott, A. Duchesnay, et al. Alterations of overused supraspinatus tendon: a possible role of glycosaminoglycans and HARP/pleiotrophin in early tendon pathology. J. Orthop. Res. 30(1):61–71, 2011.PubMedCrossRefGoogle Scholar
  3. 3.
    Attia, M., E. Huet, C. Gossard, et al. Early events of overused supraspinatus tendons involve matrix metalloproteinases and EMMPRIN/CD147 in the absence of inflammation. Am. J. Sports Med. 41(4):908–917, 2013.PubMedCrossRefGoogle Scholar
  4. 4.
    Barbato, J. C., L. G. Koch, A. Darvish, et al. Spectrum of aerobic endurance running performance in eleven inbred strains of rats. J. Appl. Physiol. 85(2):530–536, 1998.PubMedGoogle Scholar
  5. 5.
    Benjamin, M., H. Toumi, J. R. Ralphs, et al. Where tendons and ligaments meet bone: attachment sites (‘entheses’) in relation to exercise and/or mechanical load. J. Anat. 208(4):471–490, 2006.PubMedCentralPubMedCrossRefGoogle Scholar
  6. 6.
    Berglund, M. E., D. A. Hart, C. Reno, and M. Wiig. Growth factor and protease expression during different phases of healing after rabbit deep flexor tendon repair. J. Orthop. Res. 29(6):886–892, 2011.PubMedCrossRefGoogle Scholar
  7. 7.
    Blevins, F. T. Rotator cuff pathology in athletes. Sports Med. 24(3):205–220, 1997.PubMedCrossRefGoogle Scholar
  8. 8.
    Boileau, P., N. Brassart, D. J. Watkinson, et al. Arthroscopic repair of full-thickness tears of the supraspinatus: does the tendon really heal? J. Bone Joint Surg. Am. 87(6):1229–1240, 2005.PubMedCrossRefGoogle Scholar
  9. 9.
    Bromme, D., and F. Lecaille. Cathepsin K inhibitors for osteoporosis and potential off-target effects. Expert Opin. Investig. Drugs 18(5):585–600, 2009.PubMedCentralPubMedCrossRefGoogle Scholar
  10. 10.
    Bromme, D., Z. Li, M. Barnes, and E. Mehler. Human cathepsin V functional expression, tissue distribution, electrostatic surface potential, enzymatic characterization, and chromosomal localization. Biochemistry 38(8):2377–2385, 1999.PubMedCrossRefGoogle Scholar
  11. 11.
    Cook, J. L., J. A. Feller, S. F. Bonar, and K. M. Khan. Abnormal tenocyte morphology is more prevalent than collagen disruption in asymptomatic athletes’ patellar tendons. J. Orthop. Res. 22(2):334–338, 2004.PubMedCrossRefGoogle Scholar
  12. 12.
    Cunnane, G., O. FitzGerald, K. M. Hummel, et al. Collagenase, cathepsin B and cathepsin L gene expression in the synovial membrane of patients with early inflammatory arthritis. Rheumatology 38(1):34–42, 1999.PubMedCrossRefGoogle Scholar
  13. 13.
    Dejica, V. M., J. S. Mort, S. Laverty, et al. Cleavage of type II collagen by cathepsin K in human osteoarthritic cartilage. Am. J. Pathol. 173(1):161–169, 2008.PubMedCentralPubMedCrossRefGoogle Scholar
  14. 14.
    Deval, C., S. Mordier, C. Obled, et al. Identification of cathepsin L as a differentially expressed message associated with skeletal muscle wasting. Biochem. J. 360(Pt 1):143, 2001.PubMedCentralPubMedCrossRefGoogle Scholar
  15. 15.
    Eeckhout, Y., and G. Vaes. Further studies on the activation of procollagenase, the latent precursor of bone collagenase. Effects of lysosomal cathepsin B, plasmin and kallikrein, and spontaneous activation. Biochem. J. 166(1):21–31, 1977.PubMedCentralPubMedGoogle Scholar
  16. 16.
    Fu, S. C., B. P. Chan, W. Wang, et al. Increased expression of matrix metalloproteinase 1 (MMP1) in 11 patients with patellar tendinosis. Acta Orthop. Scand. 73(6):658–662, 2002.PubMedGoogle Scholar
  17. 17.
    Garnero, P., O. Borel, I. Byrjalsen, et al. The collagenolytic activity of cathepsin K is unique among mammalian proteinases. J. Biol. Chem. 273(48):32347–32352, 1998.PubMedCrossRefGoogle Scholar
  18. 18.
    Gimbel, J. A., J. P. Van Kleunen, S. Mehta, et al. Supraspinatus tendon organizational and mechanical properties in a chronic rotator cuff tear animal model. J. Biomech. 37(5):739–749, 2004.PubMedCrossRefGoogle Scholar
  19. 19.
    Goretzki, L., M. Schmitt, K. Mann, et al. Effective activation of the proenzyme form of the urokinase-type plasminogen activator (pro-uPA) by the cysteine protease cathepsin L. FEBS Lett. 297(1–2):112–118, 1992.PubMedCrossRefGoogle Scholar
  20. 20.
    Gotoh, M., K. Hamada, H. Yamakawa, et al. Significance of granulation tissue in torn supraspinatus insertions: an immunohistochemical study with antibodies against interleukin-1β, cathepsin D, and matrix metalloprotease-1. J. Orthop. Res. 15(1):33–39, 1997.PubMedCrossRefGoogle Scholar
  21. 21.
    Joshi, S. K., H. T. Kim, B. T. Feeley, and X. Liu. Differential ubiquitin-proteasome and autophagy signaling following rotator cuff tears and suprascapular nerve injury. J. Orthop. Res. 32(1):138–144, 2014.PubMedCrossRefGoogle Scholar
  22. 22.
    Kannus, P., and L. Jozsa. Histopathological changes preceding spontaneous rupture of a tendon. A controlled study of 891 patients. J. Bone Joint Surg. Am. 73(10):1507–1525, 1991.PubMedGoogle Scholar
  23. 23.
    Kirschke, H., B. Wiederanders, D. Bromme, and A. Rinne. Cathepsin S from bovine spleen. Purification, distribution, intracellular localization and action on proteins. Biochem. J. 264(2):467–473, 1989.PubMedCentralPubMedGoogle Scholar
  24. 24.
    Kozawa, E., Y. Nishida, X. W. Cheng, et al. Osteoarthritic change is delayed in a Ctsk-knockout mouse model of osteoarthritis. Arthritis Rheum. 64(2):454–464, 2012.PubMedCrossRefGoogle Scholar
  25. 25.
    Li, W. A., Z. T. Barry, J. D. Cohen, et al. Detection of femtomole quantities of mature cathepsin K with zymography. Anal. Biochem. 401(1):91–98, 2010.PubMedCrossRefGoogle Scholar
  26. 26.
    Lo, I. K. Y., L. L. Marchuk, R. Hollinshead, et al. Matrix metalloproteinase and tissue inhibitor of matrix metalloproteinase mRNA levels are specifically altered in torn rotator cuff tendons. Am. J. Sport Med. 32(5):1223–1229, 2004.CrossRefGoogle Scholar
  27. 27.
    Maffulli, N., U. G. Longo, F. Franceschi, et al. Movin and Bonar scores assess the same characteristics of tendon histology. Clin. Orthop. Relat. Res. 466(7):1605–1611, 2008.PubMedCentralPubMedCrossRefGoogle Scholar
  28. 28.
    Maganaris, C. N., M. V. Narici, L. C. Almekinders, and N. Maffulli. Biomechanics and pathophysiology of overuse tendon injuries: ideas on insertional tendinopathy. Sports Med. 34(14):1005–1017, 2004.PubMedCrossRefGoogle Scholar
  29. 29.
    Masarachia, P. J., B. L. Pennypacker, M. Pickarski, et al. Odanacatib reduces bone turnover and increases bone mass in the lumbar spine of skeletally mature ovariectomized rhesus monkeys. J. Bone Miner. Res. 27(3):509–523, 2012.PubMedCrossRefGoogle Scholar
  30. 30.
    Millar, N. L., A. Q. Wei, T. J. Molloy, et al. Heat shock protein and apoptosis in supraspinatus tendinopathy. Clin. Orthop. Relat. Res. 466(7):1569–1576, 2008.PubMedCentralPubMedCrossRefGoogle Scholar
  31. 31.
    Morko, J. P., M. Soderstrom, A. M. Saamanen, et al. Up regulation of cathepsin K expression in articular chondrocytes in a transgenic mouse model for osteoarthritis. Ann. Rheum. Dis. 63(6):649–655, 2004.PubMedCentralPubMedCrossRefGoogle Scholar
  32. 32.
    Morko, J., R. Kiviranta, K. Joronen, et al. Spontaneous development of synovitis and cartilage degeneration in transgenic mice overexpressing cathepsin K. Arthritis Rheum. 52(12):3713–3717, 2005.PubMedCrossRefGoogle Scholar
  33. 33.
    Oliva, F., D. Barisani, A. Grasso, and N. Maffulli. Gene expression analysis in calcific tendinopathy of the rotator cuff. Eur. Cells Mater. 21:548–557, 2011.Google Scholar
  34. 34.
    Park, K. Y., W. A. Li, and M. O. Platt. Patient specific proteolytic activity of monocyte-derived macrophages and osteoclasts predicted with temporal kinase activation states during differentiation. Integr. Biol. (Camb.) 4(12):1459–1469, 2012.CrossRefGoogle Scholar
  35. 35.
    Patterson-Kane, J. C., A. M. Wilson, E. C. Firth, et al. Exercise-related alterations in crimp morphology in the central regions of superficial digital flexor tendons from young thoroughbreds: a controlled study. Equine Vet. J. 30(1):61–64, 1998.PubMedCrossRefGoogle Scholar
  36. 36.
    Riley, G. P., R. L. Harrall, C. R. Constant, et al. Tendon degeneration and chronic shoulder pain: changes in the collagen composition of the human rotator cuff tendons in rotator cuff tendinitis. Ann. Rheum. Dis. 53(6):359–366, 1994.PubMedCentralPubMedCrossRefGoogle Scholar
  37. 37.
    Scott, A., J. L. Cook, D. A. Hart, et al. Tenocyte responses to mechanical loading in vivo: a role for local insulin-like growth factor 1 signaling in early tendinosis in rats. Arthritis Rheum. 56(3):871–881, 2007.PubMedCrossRefGoogle Scholar
  38. 38.
    Sharma, P., and N. Maffulli. Tendon injury and tendinopathy: healing and repair. J. Bone Joint Surg. Am. 87(1):187–202, 2005.PubMedCrossRefGoogle Scholar
  39. 39.
    Sher, J. S. Anatomy, biomechanics, and pathophysiology of rotator cuff disease. In: Disorders of the Shoulder: Diagnosis and Management, edited by J. P. Iannotti, and G. R. Williams. Philadelphia: Lippincott, Williams, and Wilkins, 1999, pp. 3–29.Google Scholar
  40. 40.
    Silverstein, B., E. Welp, N. Nelson, and J. Kalat. Claims incidence of work-related disorders of the upper extremities: washington state, 1987 through 1995. Am. J. Public Health 88(12):1827–1833, 1998.PubMedCentralPubMedCrossRefGoogle Scholar
  41. 41.
    Soslowsky, L. J., J. E. Carpenter, C. M. DeBano, et al. Development and use of an animal model for investigations on rotator cuff disease. J. Shoulder Elb. Surg. 5(5):383–392, 1996.CrossRefGoogle Scholar
  42. 42.
    Soslowsky, L. J., S. Thomopoulos, S. Tun, et al. Neer award 1999 Overuse activity injures the supraspinatus tendon in an animal model: a histologic and biomechanical study. J. Shoulder Elb. Surg. 9(2):79–84, 2000.CrossRefGoogle Scholar
  43. 43.
    Wilder, C. L., K. Y. Park, P. M. Keegan, and M. O. Platt. Manipulating substrate and pH in zymography protocols selectively distinguishes cathepsins K, L, S, and V activity in cells and tissues. Arch. Biochem. Biophys. 516(1):52–57, 2011.PubMedCentralPubMedCrossRefGoogle Scholar

Copyright information

© Biomedical Engineering Society 2015

Authors and Affiliations

  • Song P. Seto
    • 1
  • Akia N. Parks
    • 1
  • Yongzhi Qiu
    • 1
  • Louis J. Soslowsky
    • 2
  • Spero Karas
    • 3
  • Manu O. Platt
    • 1
    • 4
  • Johnna S. Temenoff
    • 1
    • 4
    • 5
    Email author
  1. 1.W.H. Coulter Department of Biomedical EngineeringGeorgia Institute of Technology and Emory UniversityAtlantaUSA
  2. 2.McKay Orthopaedic Research LaboratoryUniversity of PennsylvaniaPhiladelphiaUSA
  3. 3.Department of OrthopaedicsEmory UniversityAtlantaUSA
  4. 4.Petit Institute for Bioengineering and BioscienceGeorgia Institute of TechnologyAtlantaUSA
  5. 5.W.H. Coulter Department of Biomedical EngineeringGeorgia Tech/Emory UniversityAtlantaUSA

Personalised recommendations