Tendonopathy, resulting in the loss of mechanical strength of a tendon, is a serious health problem affecting many people. The common symptom of tendonopathy is pain – patients' daily activities, their participation in sport and exercise, and their ability to work are greatly compromised. Tendonopathy is considered to be a degenerative disorder caused by repetitive injury of the tendon. The most common tendon lesions are Achilles tendon rupture, lateral epicondylitis (tennis elbow) and rotator cuff tear. However, in spite of its clinical significance, our knowledge about tendonopathy is still very poor. This research was undertaken to investigate the pathology of tendonopathy. It is proposed that apoptosis, autophagic cell death and myofibroblasts play a role in the progression of tendonopathy in the rotator cuff; the aim of this study was therefore to determine if this was indeed the case. Tendon tissues were collected from 30 patients suffering from rotator cuff tears. A terminal deoxynucleotidyl transferase biotin-dUTP nick end labelling (TUNEL assay) was performed to detect apoptosis. Autophagic cell death of the tenocytes in the ruptured rotator cuff tendon was detected by immunohistochemical staining for ubiquitin. Myofibroblasts were identified immunohistochemically with anti-alpha-smooth muscle actin (anti--SMA) antibody. The distribution of apoptosis, autophagic cell death and myofibroblasts, as well as the total cell density, were assessed respectively and were correlated using a four-category (i.e. graded from 0-3) degeneration of collagen matrix. – 6 – The results showed that apoptosis, autophagic cell death and myofibroblasts were observed in all of the samples. The highest percentage of autophagic cell death was evidenced in the Grade 2 matrix, while the percentage of apoptosis increased significantly with the increase of matrix degeneration from Grade 0-3; a similar pattern was found for myofibroblasts. The total cell numbers varied among the matrix grades, with the maximum and minimum percentages occurring in Grades 1 and 3, respectively. It can be concluded that apoptosis, autophagic cell death and myofibroblasts might be closely related to the damage of the extracellular matrix (ECM) structure.

Rotator cuff injuries cause pain, disability, and loss of shoulder function in over 17 million individuals in the United States that result in over 500,000 surgeries performed annually, though with alarming failure rates of 20-94% (Colvin et al., 2012; Galatz et al., 2004; Harryman et al., 1991; Jain et al., 2014; Mather et al., 2013; Oh et al., 2007; Vitale et al., 2007; Yamaguchi et al., 2006). These surgeries involve repair or reconstruction of the damaged rotator cuff tendon(s) along with enlargement of the subacromial space by debriding the overlying bone (acromion) and removing the subacromial bursa (Beard et al., 2018; Burkhart et al., 2016; Dines et al., 2006; Lo & Burkhart, 2003; Rossi & Ranalletta, 2020). The subacromial bursa is a synovial-like tissue that is situated between the acromion and the tendons of the rotator cuff. This tissue has been long understood to serve a primarily mechanical role by providing cushioning and protecting from friction-wear from the acromion on the underlying tendons. More recently, the identification of a robust vascular network within the bursa, a resident population of mesenchymal stem cells, and inflammatory responsiveness to rotator cuff pathology have supported the existence of a biological role of this tissue in addition to its mechanical one (Blevins et al., 1997; Gotoh et al., 1998, 2001; Põldoja et al., 2017; Rathbun & Macnab, 1970; Steinert et al., 2015; Yepes et al., 2007). These observations make surgical excision of the bursa problematic, given our current lack of understanding of the implications of removing the bursa on the biological response to tendon injury. Therefore, the goals of this dissertation were three-fold: (1) to determine the role of the subacromial bursa in the rotator cuff tendon response to injury and healing, (2) to interrogate patterns of cellular crosstalk between the subacromial bursa and the rotator cuff following injury, and (3) to demonstrate therapeutic potential of targeting the subacromial bursa for modulating inflammation and improving tendon healing.Motivated by clinically observed phenotypic changes in the subacromial bursa with rotator cuff pathology, the profiles of human bursa and rotator cuff tendon tissues were assessed using histology, proteomics, and transcriptomics. This data set, analyzed in the context of patient demographics and diagnoses, revealed distinct bursa proteomes according to tissue phenotype (i.e., fibrous, vascular, or fatty), patient age, and presence of a tear in the underlying rotator cuff. These results suggested the presence of crosstalk between the rotator cuff and the bursa that had not been previously appreciated. Employing multiple methods of validation, including histology, microcomputed tomography, gene expression, and flow cytometry, the rat bursa was established as an appropriate animal model of the human bursa. Therefore, we used the rat model to investigate the role of the bursa in tendon injury response and healing; tendon injuries were created surgically with or without a subsequent repair to study healing and responses to injury, respectively. The role of the bursa in the response to injury was assessed using gene expression, transcriptomics, and histology. The bursa promoted inflammatory gene expression in the injured supraspinatus but resolved inflammatory gene expression in the intact infraspinatus. The role of the bursa in tendon healing was assessed using gene expression, histology, microcomputed tomography, and tensile mechanical testing of the cuff tendons. Consistent with responses during the inflammatory phase of healing, the bursa promoted expression of genes related to aberrant, scar-mediated healing in the supraspinatus, whereas it promoted tenogenic and tendon extracellular matrix gene expression in the intact infraspinatus. Mechanical testing demonstrated that the bursa protected the infraspinatus from the inflammatory environment caused by the supraspinatus injury but had a limited functional effect on the healing supraspinatus. Microcomputed tomography also indicated bursa-dependence in cortical and trabecular bone remodeling following tendon injury. Cross-talk between the bursa and the tendon was then studied in a novel tissue explant co-culture platform using gene expression and nitric oxide release as outcome measures. These experiments revealed that the activated bursa engaged in immunomodulation of tendon fibroblast responses to inflammatory stimulus. The in vitro platform also established the glucocorticoid dexamethasone as a viable therapeutic candidate for bursa-targeted treatment based on its capacity to regulate the bursa’s response to an inflammatory stimulus and enhance the bursa’s immunomodulatory potential. Therefore, in the final component of this thesis, dexamethasone was delivered via PLGA microspheres in vivo to the bursa to modulate the post-injury inflammatory response in the supraspinatus and the infraspinatus tendons. Results supported the therapeutic potential of this treatment approach to improve rotator cuff healing outcomes. This body of work demonstrated a robust involvement of the bursa in rotator cuff responses to injury, with distinct roles in the underlying injured and intact tendons. This work also established, for the first time, the immunomodulatory capacity of the activated bursa and provided strong evidence against the clinical practice of bursectomy. Finally, use of sustained-release dexamethasone to dampen the inflammatory responses to rotator cuff injury offers a new direction for harnessing the inherent properties of the subacromial bursa therapeutically for improved rotator cuff tendon healing.

Submitted in partial compliance with the requirements for the Master’s Degree in Technology: Chiropractic, Department of Chiropractic, Durban University of Technology, Durban, South Africa, 2015. / Background: The most common shoulder complaint seen by physicians is shoulder impingement syndrome. There has been limited success with the current conservative treatment that has been provided for this condition. Thoracic spine and rib manipulation is purported to aid in the treatment of this condition; however there is a paucity of controlled investigations. The purpose of this study was to determine the short term efficacy of thoracic and prone rib manipulation on shoulder impingement syndrome. Methods: A randomised, placebo controlled pre-test post-test experimental design was used. Informed consent was obtained and 30 participants were recruited according to inclusion criteria and allocated to either a placebo or intervention group. Intervention consisted of thoracic spinal and rib manipulation. Data was collected, pre and post the first treatment and at a 48 hours follow up. SPSS was used to analyse the data with a p value of 0.05. Results: No statistically significant differences were seen between the groups for pain rating, range of motion of the glenohumeral joint, lateral scapula slide test or scapula isometric pinch test. The shoulder pain and disability index (SPADI) showed significant (p = 0.04) differences between the groups in terms of disability scores, with the intervention group having a great improvement in disability. No clinically significant differences were observed between the groups. Conclusion: Thoracic spine and rib manipulation appears to improve the disability associated with shoulder impingement syndrome, however further research is required with a larger sample size.

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Surgical repair of torn rotator cuff tendons have a high rate of failure and does not address the underlying pathophysiology. Tissue engineering strategies, employing the use of multipotent progenitor cells or growth factors, represent potential therapies to improve the outcome of rotator cuff surgery. The use of glycosaminoglycan-based biomaterials in these therapies may enhance the effectiveness of cell and growth factor delivery techniques. Furthermore, understanding the cellular and molecular mediators in tendon overuse can help elucidate the causes of tendon degeneration. Thus the overall goals of this dissertation were to 1) develop heparin-based biomaterials to enhance cell pre-culture and maintain growth factor bioactivity and 2) characterize the histological and enzymatic changes in a supraspinatus tendon overuse model. To investigate the use of heparin in enhancing dynamic signaling, mesenchymal stem cells (MSCs) were encapsulated in heparin-containing hydrogels and evaluated for differentiation markers when cocultured with a small population of differentiated cells. To probe the effect of sulfation of heparin on the interactions with protein, selectively desulfated heparin species were synthesized and evaluated for their ability to bind and protect proteins. Finally, to develop a tendon overuse model that can become a test bed for testing future targeted therapeutics, an animal model was evaluated for tissue damage and protease activity. Together these studies represent a multi-pronged approach to understanding how tendon tissues become degenerative and for developing technologies to improve the biological fixation of tendon to bone in order to reduce the need for revision surgeries.