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Investigation of the composition, turnover, and thermal properties of ruptured cranial cruciate ligaments of dogs

Eithne J. Comerford MVB, PhD1, John F. Innes BVSc, PhD2, John F. Tarlton PhD3, and Allen J. Bailey PhD, ScD4
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  • 1 Department of Clinical Veterinary Science, University of Bristol, Langford, Bristol, BS40 5DU, UK.
  • | 2 Department of Veterinary Clinical Science, University of Liverpool, Liverpool, L7 7EX, UK.
  • | 3 Department of Clinical Veterinary Science, University of Bristol, Langford, Bristol, BS40 5DU, UK.
  • | 4 Department of Clinical Veterinary Science, University of Bristol, Langford, Bristol, BS40 5DU, UK.

Abstract

Objective—To assess different components of the extracellular matrix with regard to their thermal properties, composition, and turnover in ruptured cranial cruciate ligaments (CCLs) of dogs, compared with components of intact CCLs from a breed predisposed to CCL failure.

Sample Population—Ruptured CCLs obtained from 8 dogs of breeds predisposed to ruptured CCLs and intact CCLs from 12 cadaveric Labrador Retrievers.

Procedure—Ruptured and intact CCLs were analyzed for water content; collagen content and collagen cross-links were evaluated via hydroxyproline and amino-acid analyses, respectively. Glycosaminoglycan (GAG) content was analyzed via dimethylmethylene blue and uronic acid assays. Matrix metalloproteinases (MMPs)-2 and -9 and the tissue inhibitors of metalloproteinases (TIMPs)-1 and -2 were detected via gelatin SDS-PAGE zymography and reverse gelatin zymography. Thermal analysis of ligaments was performed by use of differential scanning calorimetry.

Results—Ruptured CCLs had significantly higher lamounts of immature cross-links, total and sulfated GAGs, and water content, compared with that of the intact ligaments. Compared with intact CCLs, concentration of pro–MMP-2 was significantly higher in ruptured CCLs; the maximum temperature of collagen denaturation was significantly lower in the ruptured CCLs.

Conclusions and Clinical Relevance—The extracellular matrix of ruptured CCLs had an increased matrix turnover indicated by increased collagen and GAG synthesis, compared with that of intact CCLs. Although the extracellular matrix changes may have occurred before ligament rupture, it is possible that these observed changes may be part of a reparative process after rupture. (Am J Vet Res 2004;65:1136–1141)

Abstract

Objective—To assess different components of the extracellular matrix with regard to their thermal properties, composition, and turnover in ruptured cranial cruciate ligaments (CCLs) of dogs, compared with components of intact CCLs from a breed predisposed to CCL failure.

Sample Population—Ruptured CCLs obtained from 8 dogs of breeds predisposed to ruptured CCLs and intact CCLs from 12 cadaveric Labrador Retrievers.

Procedure—Ruptured and intact CCLs were analyzed for water content; collagen content and collagen cross-links were evaluated via hydroxyproline and amino-acid analyses, respectively. Glycosaminoglycan (GAG) content was analyzed via dimethylmethylene blue and uronic acid assays. Matrix metalloproteinases (MMPs)-2 and -9 and the tissue inhibitors of metalloproteinases (TIMPs)-1 and -2 were detected via gelatin SDS-PAGE zymography and reverse gelatin zymography. Thermal analysis of ligaments was performed by use of differential scanning calorimetry.

Results—Ruptured CCLs had significantly higher lamounts of immature cross-links, total and sulfated GAGs, and water content, compared with that of the intact ligaments. Compared with intact CCLs, concentration of pro–MMP-2 was significantly higher in ruptured CCLs; the maximum temperature of collagen denaturation was significantly lower in the ruptured CCLs.

Conclusions and Clinical Relevance—The extracellular matrix of ruptured CCLs had an increased matrix turnover indicated by increased collagen and GAG synthesis, compared with that of intact CCLs. Although the extracellular matrix changes may have occurred before ligament rupture, it is possible that these observed changes may be part of a reparative process after rupture. (Am J Vet Res 2004;65:1136–1141)