Characterisation of tendons at different length scales using atomic force microscopy and polarised light microscopy may provide insight into tendon disease

© 2014 by Nova Science Publishers, Inc. Chronic, non- inflammatory tendon disorders are widespread and debilitating. The aetiology of these disorders is unclear but is believed to be related to tissue degeneration associated with altered mechanical loading. For example, studies have reported biological and cellular adaptations in torn tendons indicative of increased amounts of compressive and shear loading. However, due to a lack of suitable characterisation techniques, the role of mechanical loading in tendon degeneration and tearing is largely unknown. Given the strong correlation between age and tendinopathy, and unprecedented population aging, these disorders will become increasingly prevalent. Improved understanding of noninflammatory tendon disorders is therefore urgently needed to aid clinician decisionmaking.As the fibrous tissues that connect muscles to bones, tendons must be strong and flexible whilst facilitating efficient energy storage and return. This is achieved through the use of a fibre-reinforced composite material with a hierarchical, rope-like structure consisting of collagen fibrils, fibres and fascicles embedded in a mucopolysaccharide matrix. Like all biological materials, tendons exhibit a dynamic relationship between loading environment and material properties; cells tailor the proportions and dimensions of the extra-fibrillar matrix and fibrous elements of tendons to better suit the mechanical environment they experience. Given this strong, dynamic relationship between loading and material properties, knowledge of the material adaptations of healthy and torn tendons could provide significant insight into the role of the mechanical environment in the aetiology and pathology of tendon disorders. To date, the material properties of healthy and torn tendons, and in particular the structural adaptations, have been largely unreported. This is largely caused by a lack of techniques suitable for characterising the material adaptations of small, surgically obtained, formalin-fixed tissue samples. Historically, structural investigations at small length scales (ultrastructure) have been restricted to transmission electron microscopy, a technique which requires glutaraldehyde fixed samples and complex sample preparation, and provides a limited amount of structural information and no direct mechanical information. Recently, however, it has been demonstrated that atomic force microscopy is a highly suitable technique for biological characterisation, requiring little, if any, sample preparation and providing a wealth of structural and mechanical information, even for formalin-fixed samples. In this chapter, atomic force microscopy, polarised light microscopy and instrumented indentation are used to characterise the adaptations of small, formalin-fixed animal tendon samples that have been exposed to different loading environments in vivo, to gain insight into the effect of mechanical loading on the structural properties of tendons. The results demonstrate that atomic force microscopy and polarised light microscopy are highly suitable techniques for characterising the structural properties of surgically obtained, formalin-fixed tendon samples; loading environment is demonstrated to significantly affect the structural adaptations of healthy tendons with regions exposed to compressive stresses exhibit thinner fibres, shorter crimp lengths and thinner, less aligned fibrils compared with regions exposed to tensile and shear stresses. However, comparison of mechanical data obtained using AFM and instrumented indentation reveal that a great deal of additional work is required in order to establish appropriate methodologies and data analysis algorithms before these techniques can be reliably used to characterise the micro-mechanical properties of tendon. The application of these techniques to human samples could provide information regarding the material adaptations of healthy, degenerative and torn tendons that could provide valuable information regarding the role of mechanical environment in the aetiology and pathology of tendon disorders.