The Connection of Composition, Structure, and Dynamic Processes to Tendon Mechanics: Structure-Function Relationships in Collagen V Deficient Tendons

Tendons are able to withstand the broad range of stresses and strains via their finely tuned composition and structure. In addition, tendons undergo a coordinated set of dynamic responses, specifically collagen uncrimping, re-alignment, sliding and deformation, within the matrix. To date, a complete understanding of the hierarchical structure-function relationships in tendon is lacking. Therefore, the overall goal of this thesis was to measure tendon structure and function in a mouse supraspinatus model of altered structure, and to analyze links between mechanical properties, dynamic processes and composition/structure using a series of statistical analyses. In the studies presented here, we used novel and established methods to measure the multi-scale composition, structure and mechanical function of mouse supraspinatus tendons from wild type, collagen V heterozygous and collagen V null mice. Overall, we found that the experimental groups were mechanically inferior to the wild type group, with larger changes in both macroscale function and the dynamic responses (re-alignment, crimp, deformation, sliding). In addition, while fibril morphology was altered at both locations, the insertion site also exhibited alterations in cell and fiber morphology as well as extracellular matrix composition. Finally, using a novel regression approach, we found that the contribution of composition and structure as well as the contribution of dynamic processes to determining macroscale mechanical function was highly dependent on location and that the dynamic processes were significant mediators of the relationship between composition/structure and mechanical properties. Overall, we conclude that although collagen V is a quantitatively minor component in mature tendon/ligament, it is a major regulator of composition and structure during development which ultimately leads to mechanical function. Furthermore, we conclude that the dynamic responses to load are crucial factors in ultimately determining regionally-dependent mechanical function. This information will help to guide clinicians in developing preventative techniques and appropriate rehabilitation strategies, as well as help to define the appropriate and important parameters on which to base tissue engineering efforts for tendon augmentation or replacement. Finally, this work presents a strong foundation on which to develop future experimental and modeling efforts in order to fully understand the complex structure-function relationships present in tendon

Senescence contributes to death resistance of aged tenocytes in a model of secondary joint damage

NIH/National Institutes of Health; The American Association of University Wome

Differential effect of acute compressive injury on young and aged tendons

5R00AG063896-04 REVISED - NIH/National Institute on Agin

Earlier proteoglycan turnover promotes higher efficiency matrix remodeling in MRL/MpJ tendons

While most mammalian tissue regeneration is limited, the Murphy Roths Large (MRL/MpJ) mouse has been identified to regenerate several tissues, including tendon. Recent studies have indicated that this regenerative response is innate to the tendon tissue and not reliant on a systemic inflammatory response. Therefore, we hypothesized that MRL/MpJ mice may also exhibit a more robust homeostatic regulation of tendon structure in response to mechanical loading. To assess this, MRL/MpJ and C57BL/6J Flexor digitorum longus tendon explants were subjected to stress-deprived conditions in vitro for up to 14 days. Explant tendon health (metabolism, biosynthesis, and composition), matrix metalloproteinase activity, gene expression, and tendon biomechanics were assessed periodically. We found a more robust response to a loss of mechanical stimulus in the MRL/MpJ tendon explants, exhibiting an increase in collagen production and MMP activity consistent with previous in vivo studies. This greater collagen turnover was preceded by an early expression of small leucine rich proteoglycans and proteoglycan-degrading MMP-3, promoting efficient regulation and organization of newly synthesized collagen and allowing for more efficient overall turnover in MRL/MpJ tendons. Therefore, mechanisms of MRL/MpJ matrix homeostasis may be fundamentally different from that of B6 tendons and may indicate better recovery from mechanical microdamage in MRL/MpJ tendons. We demonstrate here the utility of the MRL/MpJ model in elucidating mechanisms of efficient matrix turnover and its potential to shed light on new targets for more effective treatments for degenerative matrix changes brought about by injury, disease, or aging. This article is protected by copyright. All rights reserved.NIH/National Institutes of Healt

In situ AFM-based nanoscale rheology reveals regional non-uniformity in viscoporoelastic mechanical behavior of the murine periodontal ligament

The periodontal ligament (PDL) is a critical player in the maintenance of tooth health, acting as the primary stabilizer of tooth position. Recent studies have identified two unique regions within the PDL, the ‘dense collar’ region and the ‘furcation’ region, which exhibit distinct structural and compositional differences. However, specific functional differences between these regions have yet to be investigated. We adapted an AFM-based nanoscale rheology method to regionally assess mechanical properties and poroelasticity in the mouse PDL while minimizing the disruption of the 3-dimensional native boundary conditions, and then explored tissue mechanical function in four different regions within the dense collar as well as in the furcation region. We found significant differences between the collar and furcation regions, with the collar acting as a stabilizing ligamentous structure and the furcation acting as both a compressive cushion for vertical forces and a conduit for nutrient transport. While this finding supports our hypothesis, based on previous studies investigating structural and compositional differences, we also found surprising inhomogeneity within the collar region itself. This inhomogeneity supports previous findings of a tilting movement in the buccal direction of mandibular molar teeth and the structural adaptation to prevent lingual movement. Future work will aim to understand how different regions of the PDL change functionally during biological or mechanical perturbations, such as orthodontic tooth movement, development, or aging, with the ultimate goal of better understanding the mechanobiology of the PDL function in health and disease.Accepted manuscrip

The effect of myostatin deficiency on achilles tendon structural and material behavior in male mice

Myostatin (MSTN) is a secreted protein that acts as a negative regulator of skeletal muscle growth. Deficiencies in this protein have been shown to increase muscle mass in a variety of animal models. Consequently, clinical suppression of myostatin is now being pursued as a therapeutic strategy to counteract the muscle wasting that occurs in patients with degenerative neuromuscular diseases such as Duchenne Muscular Dystrophy (DMD). Although research supports the use of myostatin suppression therapy to increase muscle mass, investigation into the effects of myostatin on tendon is limited. The aim of this study was to investigate the effects of myostatin deficiency by characterizing the structural, material and compositional properties of the Achilles tendons from mature (16 week old) male myostatin deficient (MSTN-/-), wild type (MSTN+/+), and heterozygous (MSTN+/-) mice. Specifically, we tested the hypothesis that myostatin deficiency is associated with stiffer and stronger tendons, that these effects are dose dependent, and that structural and material differences can be explained by differences in tendon composition. The experimental model consisted of sixty male mice, thirty of which were used for Achilles tendon tensile mechanical testing and thirty for tendon biochemical compositional analysis. Results demonstrated that there were no significant differences in tendon geometric, structural or material properties. There was a statistically significant difference in total body mass, with a larger mass in the myostatin null group, but there was no difference in Achilles tendon wet weight itself. DNA, glycosaminoglycan (GAG), and hydroxyproline (indicative of total collagen) content were also assessed. Myostatin null animals were found to have less DNA per wet weight, but more GAG per cell than their wild type counterparts. There were no significant differences in collagen between any of the genotypes. These data do not support the conclusion that myostatin deficiency causes stiffer and stronger tendons, but suggest that myostatin levels have no effect on mature mouse Achilles tendon mechanical properties. Further study is necessary to better understand the role of myostatin in tendon composition and mechanical function

Lose‐Dose Administration of Dexamethasone is Beneficial in Preventing Secondary Tendon Damage in a Stress‐Deprived Joint Injury Explant Model

© 2019 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. Secondary joint damage is the process by which a single injury can lead to detrimental changes in adjacent tissue structures, typically through the spread of inflammatory responses. We recently developed an in vitro model of secondary joint damage using a murine rotator cuff explant system, in which injuries to muscle and bone cause massive cell death in otherwise uninjured tendon. The purpose of the present study was to test the ability cytokine-targeted and broad-spectrum therapeutics to prevent cell death and tissue degeneration associated with secondary joint damage. We treated injured bone-tendon-muscle explants with either interleukin-1 receptor antagonist, etanercept, or dexamethasone (DEX) for up to 7 days in culture. Only the low-dose DEX treatment was able to prevent cell death and tissue degeneration. We then identified a critical window between 24 and 72 h following injury for maximal benefit of DEX treatment through timed administration experiments. Finally, we performed two tendon-only explant studies to identify mechanistic effects on tendon health. Interestingly, DEX did not prevent cell death and degeneration in a model of cytokine-induced damage, suggesting other targets of DEX activity. Future studies will aim to identify factors in joint inflammation that may be targeted by DEX treatment, as well as to investigate novel delivery strategies. Statement of clinical significance: Overall, this work demonstrates beneficial effects of DEX administration on preventing tenocyte death and extracellular matrix degeneration in an explant model of secondary joint damage, supporting the clinical use of low-dose glucocorticoids for short-term treatment of joint inflammation. © 2019 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 38:139–149, 2020