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Increased Proteolysis of Collagen in an In Vitro Tensile Overload Tendon Model

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Abstract

Presently, there is a lack of fundamental understanding regarding changes in collagen’s molecular state due to mechanical damage. The bovine tail tendon (BTT; steers approximately 30 months) was characterized and used as an in vitro model for investigating the effect of tensile mechanical overload on collagen susceptibility to proteolysis by acetyltrypsin and α-chymotrypsin. Two strain rates with a 1000-fold difference (0.01 and 10 s−1) were used, since molecular mechanisms that determine mechanical behavior were presumed to be strain rate dependent. First, it was determined that the BTTs were normal but immature tendons. Water content and collagen content (approx. 60% of wet weight and 80% of dry weight, respectively) and mechanical properties were all within the expected range. The collagen crosslinking was dominated by the intermediate crosslink hydroxylysinonorleucine. Second, tensile overload damage significantly enhanced proteolysis by acetyltrypsin and, to a lesser degree, by α-chymotrypsin. Interestingly, proteolysis by acetyltrypsin was greatest for specimens ruptured at 0.01 s−1 and seemed to occur throughout the specimen. Understanding damage is important for insight into injuries (as in sports and trauma) and for better understanding of collagen fiber stability, durability, and damage mechanisms, aiding in the development of durable tissue-based products for mechanically demanding surgical applications.

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Abbreviations

BTT:

Bovine tail tendon

DSC:

Differential scanning calorimetry

HHL:

Histidinohydroxylysinonorleucine

HIT:

Hydrothermal isometric tension

HLKNL:

Hydroxylysinoketonorleucine

HLNL:

Hydroxylysinonorleucine

NaBH4 :

Sodium borohydride

OH-Pyr:

Hydroxylysyl-pyridinoline

PMSF:

Phenylmethylsulfonyl fluoride

RGD:

Arginine–glycine–aspartic acid

TLD:

Thermally labile domain

TLCK:

1-chloro-3-tosylamido-7-amino-2-heptanone

UTS:

Ultimate tensile stress

A :

Specimen cross-sectional area

ε :

Strain

F :

Force

L :

Length

σ :

True stress

T d :

Denaturation temperature

T o :

Onset temperature

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Acknowledgments

The authors would like to thank Ms. Maxine Langman, Dr. Paul F. Gratzer, Mr. Hong Tang, and Dr. Mary Anne White for valuable technical assistance, advice, and access to laboratory equipment. We would also like to thank our funding sources: The Natural Science and Engineering Research Council of Canada (J.M. Lee and T.L. Willett) and the Canadian Institutes for Heath Research Strategic Training Program in Cell Signaling in Mucosal Inflammation and Pain (STP-53877; T.L. Willett).

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Authors and Affiliations

  1. School of Biomedical Engineering, Dalhousie University, Halifax, NS, Canada, B3M 3J5

    Thomas L. Willett, Rosalind S. Labow & J. Michael Lee

  2. Department of Surgery, University of Ottawa Heart Institute, Ottawa, ON, Canada

    Rosalind S. Labow

  3. Department of Biochemistry, Microbiology and Immunology, University of Ottawa, Ottawa, ON, Canada

    Rosalind S. Labow

  4. Matrix Biology Research Group, University of Bristol, Bristol, United Kingdom

    Nicholas C. Avery

  5. Department of Applied Oral Sciences, Dalhousie University, Halifax, NS, Canada

    J. Michael Lee

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  1. Thomas L. Willett
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Correspondence to Thomas L. Willett.

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Willett, T.L., Labow, R.S., Avery, N.C. et al. Increased Proteolysis of Collagen in an In Vitro Tensile Overload Tendon Model. Ann Biomed Eng 35, 1961–1972 (2007). https://doi.org/10.1007/s10439-007-9375-x

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  • DOI: https://doi.org/10.1007/s10439-007-9375-x

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