Effect of Preconditioning and Stress Relaxation on Local Collagen Fiber Re-Alignment: Inhomogeneous Properties of Rat Supraspinatus Tendon

McKay Orthopaedic Research Laboratory, University of Pennsylvania, Philadelphia, PA 19104-6081, USA.
Journal of Biomechanical Engineering (Impact Factor: 1.78). 03/2012; 134(3):031007. DOI: 10.1115/1.4006340
Source: PubMed


Repeatedly and consistently measuring the mechanical properties of tendon is important but presents a challenge. Preconditioning can provide tendons with a consistent loading history to make comparisons between groups from mechanical testing experiments. However, the specific mechanisms occurring during preconditioning are unknown. Previous studies have suggested that microstructural changes, such as collagen fiber re-alignment, may be a result of preconditioning. Local collagen fiber re-alignment is quantified throughout tensile mechanical testing using a testing system integrated with a polarized light setup, consisting of a backlight, 90 deg-offset rotating polarizer sheets on each side of the test sample, and a digital camera, in a rat supraspinatus tendon model, and corresponding mechanical properties are measured. Local circular variance values are compared throughout the mechanical test to determine if and where collagen fiber re-alignment occurred. The inhomogeneity of the tendon is examined by comparing local circular variance values, optical moduli and optical transition strain values. Although the largest amount of collagen fiber re-alignment was found during preconditioning, significant re-alignment was also demonstrated in the toe and linear regions of the mechanical test. No significant changes in re-alignment were seen during stress relaxation. The insertion site of the supraspinatus tendon demonstrated a lower linear modulus and a more disorganized collagen fiber distribution throughout all mechanical testing points compared to the tendon midsubstance. This study identified a correlation between collagen fiber re-alignment and preconditioning and suggests that collagen fiber re-alignment may be a potential mechanism of preconditioning and merits further investigation. In particular, the conditions necessary for collagen fibers to re-orient away from the direction of loading and the dependency of collagen reorganization on its initial distribution must be examined.

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Available from: Kristin Suzanne Miller
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    • "From a structural perspective, stress relaxation at low strain levels occurs predominantly through sliding between collagen fibres, while at high strain levels fibril reorganization and relaxation might play a major role, with interfibrillar linkages eventually failing and leading to subsequent excessive fibril sliding [25]. However, no change in collagen fibre realignment was found during a 600 s long static stress-relaxation test with rat supraspinatus tendon [28]. In terms of fluid interactions within the material, several studies suggest the involvement of water movement in the stress response of tendon tissue. "
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    ABSTRACT: Cyclic and static loading regimes are commonly used to study tenocyte metabolism in vitro and to improve our understanding of exercise associated tendon pathologies. The aims of our study were to investigate if cyclic and static stress relaxation affected the mechanical properties of tendon fascicles differently, if this effect was reversible after a recovery period, and if the removal of glycosaminoglycans (GAGs) affected sample recovery. Tendon fascicles were dissected from bovine-foot extensors and subjected to 14% cyclic (1Hz) or static tensile strain for 30 minutes. Additional fascicles were incubated overnight in buffer with 0.5U Chondroitinase ABC or in buffer alone prior to the static stress relaxation regime. To assess the effect of different stress relaxation regimes, a quasi-static test to failure was carried out, either directly post loading or after a 2 hour recovery period, and compared with unloaded control fascicles. Both stress relaxation regimes led to a significant reduction in fascicle failure stress and strain, but this was more pronounced in the cyclically loaded specimens. Removal of GAGs led to more stress relaxation and greater reductions in failure stress after static loading compared to controls. The reduction in mechanical properties was partially reversible in all samples, given a recovery period of 2 hours. This has implications for mechanical testing protocols, as a time delay between fatiguing specimens and characterization of mechanical properties will affect the results. GAGs appear to protect tendon fascicles from fatigue effects, possibly by enabling sample hydration.
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    • "Interestingly, unlike the mature rat SST (Miller et al., 2012c), the midsubstance and insertion site in the mouse SST demonstrated different re-alignment behaviors (Fig. 3). This may suggest that the locations experience different loading conditions in vivo, which may initiate local remodeling, resulting in different fiber network configurations including alterations in collagen fiber alignment as well as cross-links, fiber–fiber and fiber–matrix interactions. "
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    ABSTRACT: 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. 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. 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. 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.
    Full-text · Article · Jul 2012 · Journal of Biomechanics
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    ABSTRACT: Collagen fiber re-alignment is one postulated mechanism of tendon structural response to load. While collagen fiber distribution has been shown to vary by tendon location in the supraspinatus tendon (SST), changes in local re-alignment behavior have not been examined throughout postnatal development. Postnatal tendons, with immature collagen fibrils, may respond to load in a much different manner than collagen fibers with mature fiber-fiber and fiber-matrix connections. Local collagen fiber re-alignment is quantified throughout tensile mechanical testing in a developmental mouse SST model and corresponding mechanical properties measured. Collagen fiber re-alignment occurred during preconditioning for 28 day old tendons, at the toe-region for 10 day tendons and at the linear-region for 4 day tendon midsubstance. Mechanical properties increased with developmental age. Linear modulus was lower at the insertion site compared to the midsubstance location at all time points. Local differences in collagen fiber distributions were found at 10 and 28 days for all mechanical testing points (except the 10 day transition point). This study found that collagen fiber re-alignment depends on developmental age and suggests that collagen fibrillogenesis may influence the tendon's ability to structurally respond to load. Additionally, results indicate that the insertion site and tendon midsubstance locations develop differently.
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