Characterizing local collagen fiber re-alignment and crimp behavior throughout mechanical testing in a mature mouse supraspinatus tendon model

Abstract

Background: Collagen fiber re-alignment and uncrimping are two postulated mechanisms of tendon structural response to load. Recent studies have examined structural changes in response to mechanical testing in a postnatal development mouse supraspinatus tendon model (SST), however, those changes in the mature mouse have not been characterized. The objective of this study was to characterize collagen fiber re-alignment and crimp behavior throughout mechanical testing in a mature mouse SST.

Method of approach: A tensile mechanical testing set-up integrated with a polarized light system was utilized for alignment and mechanical analysis. Local collagen fiber crimp frequency was quantified immediately following the designated loading protocol using a traditional tensile set up and a flash-freezing method. The effect of number of preconditioning cycles on collagen fiber re-alignment, crimp frequency and mechanical properties in midsubstance and insertion site locations were examined.

Results: Decreases in collagen fiber crimp frequency were identified at the toe-region of the mechanical test at both locations. The insertion site re-aligned throughout the entire test, while the midsubstance re-aligned during preconditioning and the test's linear-region. The insertion site demonstrated a more disorganized collagen fiber distribution, lower mechanical properties and a higher cross-sectional area compared to the midsubstance location.

Conclusions: Local collagen fiber re-alignment, crimp behavior and mechanical properties were characterized in a mature mouse SST model. The insertion site and midsubstance respond differently to mechanical load and have different mechanisms of structural response. Additionally, results support that collagen fiber crimp is a physiologic phenomenon that may explain the mechanical test toe-region.

Fig. 1

Crimp throughout the mechanical test at insertion site and midsubstance locations demonstrates that collagen fiber crimp decreased at the toe-region with all preconditioning protocols. (*=p<0.016 , ** =p<0.001. ***=p<0.0001).

Fig. 2

Histology demonstrates that while collagen fiber uncrimping occurs primarily in the toe-region, a mild decrease in crimp frequency is also identified with increasing number of preconditioning cycles.

Fig. 3

Circular variance (VAR) for a representative sample at the insertion site and midsubstance locations shows significant collagen fiber re-alignment throughout the entire test at the insertion site and during preconditioning and the linear-region at the midsubstance. A larger value of circular variance indicates a more disorganized collagen fiber distribution.

Fig. 4

Circular variance values throughout the test demonstrate that the insertion site location is more disorganized throughout the entire mechanical test compared to the midsubstance location. (BP: Before preconditioning; AP: After preconditioning; Trans: Point at the transition strain; Linear: Point in the linear-region). (***=p<0.0001).

Fig. 5

A lower linear modulus was present at the insertion site compared to the midsubstance location. (***=p<0.0001).