Tang Yin Nei. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2001. / Includes bibliographical references (leaves xv-xxv). / Abstracts in English and Chinese. / ACKNOWLEDGEMENT --- p.i / ABBREVIATIONS --- p.ii-iii / INDEX FOR FIGURES --- p.iv-v / INDEX FOR TABLES --- p.vi / ABSTRACT (Chinese and English) --- p.vii-xi / TABLE OF CONTENTS --- p.xii-xiv / Chapter CHAPTER I 226}0ؤ --- INTRODUCTION --- p.1 / Chapter 1.1 --- Background --- p.2 / Chapter 1.2 --- Tendon / Chapter 1.2.1 --- Structure and function --- p.3 / Chapter 1.2.2 --- Tendon fibroblast --- p.6 / Chapter 1.2.3 --- Components of the extracellular matrix --- p.7 / Chapter 1.2.3.1 --- Collagen --- p.8 / Chapter 1.2.3.2 --- Proteoglycan --- p.9 / Chapter 1.2.3.3 --- Non-collagenous structural glycoprotein --- p.10 / Chapter 1.3 --- Inflammation disorders of tendon / Chapter 1.3.1 --- Inflammation --- p.11 / Chapter 1.3.2 --- Treatment --- p.12 / Chapter 1.3.2.1 --- Glucocorticoid as an anti-inflammatory agent --- p.12 / Chapter 1.3.2.2 --- Dexamethasone --- p.14 / Chapter 1.3.3 --- Clinical occurrence of tendon rupture --- p.15 / Chapter 1.3.4 --- Animal research related to glucocorticoids and tendon rupture --- p.18 / Chapter 1.4 --- Platelet-derived growth factor isoform B (PDGFBB) / Chapter 1.4.1 --- Structure and function --- p.21 / Chapter 1.4.2 --- PDGFbb effects on connective tissue --- p.22 / Chapter CHAPTER II 226}0ؤ --- AIM OF THE STUDY --- p.23 / Chapter 2.1 --- Limitations of the past researches --- p.24 / Chapter 2.2 --- Hypothesis of this study --- p.25 / Chapter 2.3 --- Objectives --- p.26 / Chapter 2.4 --- Long term significance --- p.26 / Chapter CHAPTER III 226}0ؤ --- METHODOLOGY --- p.27 / Chapter 3.1 --- Chemicals and materials used / Chapter 3.1.1 --- Chemicals --- p.28 / Chapter 3.1.2 --- Materials --- p.28 / Chapter 3.2 --- Specimen collection and preparation / Chapter 3.2.1 --- Collection --- p.29 / Chapter 3.2.2 --- Preparation and isolation --- p.30 / Chapter 3.2.3 --- Cell culture --- p.31 / Chapter 3.3 --- Reagent preparation / Chapter 3.3.1 --- Charcoal-stripped serum --- p.32 / Chapter 3.3.2 --- Phenol-red free DMEM --- p.33 / Chapter 3.3.3 --- MTT --- p.33 / Chapter 3.3.4 --- Dexamethasone --- p.34 / Chapter 3.3.5 --- PDGFbb --- p.34 / Chapter 3.3.6 --- Trypan blue --- p.35 / Chapter 3.3.7 --- TCA/Tannic acid --- p.35 / Chapter 3.3.8 --- Collagenase buffer --- p.35 / Chapter 3.4 --- Morphology / Chapter 3.4.1 --- Inverted phase contrast light microscopy --- p.36 / Chapter 3.4.2 --- Scanning electron microscopy --- p.36 / Chapter 3.5 --- Biological assays / Chapter 3.5.1 --- "MTT (3-[4,5-Dimethylthiazol-2-yl]2,5-diphenyltetrazolium bromide) assay" --- p.38 / Chapter 3.5.1.1 --- Correlation between MTT assay and trypan blue dye method --- p.38 / Chapter 3.5.1.2 --- Growth kinetics for tendon fibroblasts --- p.41 / Chapter 3.5.1.3 --- Cell viability --- p.43 / Chapter 3.5.2 --- Brdu (5-bromo-2'-deoxyuridine) assay --- p.44 / Chapter 3.5.3 --- Flow cytometry --- p.45 / Chapter 3.5.4 --- Apoptosis --- p.47 / Chapter 3.5.5 --- 3H-Proline incorporation assay --- p.48 / Chapter 3.5.6 --- 35Sulfate incorporation assay --- p.51 / Chapter 3.5.7 --- Immunocytochemistry (PDGF-β receptor) --- p.54 / Chapter 3.6 --- Statistical analysis / Chapter 3.6.1 --- Dose-response curve of dexamethasone on cell viability and proliferation --- p.55 / Chapter 3.6.2 --- Comparison among various treatments of fibroblasts --- p.55 / Chapter CHAPTER I´Vؤ --- RESULTS --- p.56 / Chapter 4.1 --- In vitro effect of dexamethasone on rat tendon fibroblasts / Chapter 4.1.1 --- Viable cell number between two sexes --- p.57 / Chapter 4.2 --- In vitro effect of dexamethasone and PDGFBB on human tendon fibroblasts / Chapter 4.2.1 --- Gross morphology --- p.58 / Chapter 4.2.2 --- Cell cycle --- p.60 / Chapter 4.2.3 --- Apoptosis --- p.61 / Chapter 4.2.4 --- Viable cell number / Chapter 4.2.4.1 --- Effect of dexamethasone --- p.62 / Chapter 4.2.4.2 --- Effect of PDGFBB --- p.63 / Chapter 4.2.5 --- Cell proliferation / Chapter 4.2.5.1 --- Effect of dexamethasone --- p.65 / Chapter 4.2.5.2 --- Effect of PDGFbb --- p.67 / Chapter 4.2.6 --- Collagen synthesis --- p.68 / Chapter 4.2.7 --- Proteoglycan synthesis --- p.72 / Chapter 4.2.8 --- PDGF-rβ expression --- p.74 / Chapter CHAPTER V 226}0ؤ --- DISCUSSION --- p.75 / Chapter 5.1 --- Dexamethasone and PDGFBB induced change of cell morphology --- p.77 / Chapter 5.2 --- Dexamethasone retarded cell growth of human tendon fibroblast --- p.80 / Chapter 5.3 --- Dexamethasone inhibited collagen synthesis --- p.82 / Chapter 5.4 --- Dexamethasone inhibited proteoglycan synthesis --- p.86 / Chapter 5.5 --- PDGFbb could counteract the inhibitory effects of dexamethasone --- p.88 / Chapter 5.6 --- Expression of PDGF-(3 receptor is regulated by dexamethasone and PDGFBB --- p.90 / Chapter 5.7 --- Limitations of this study / Chapter 5.7.1 --- Not enough sample to differentiate different between two sexes --- p.92 / Chapter 5.7.2 --- Small sample size and few assays --- p.92 / Chapter 5.7.3 --- Limitations of the cell culture model --- p.93 / Chapter 5.7.4 --- Difficult to further in vivo study on human --- p.93 / Chapter 5.8 --- Contributions of this study / Chapter 5.8.1 --- Improve the limitation of the past research --- p.94 / Chapter 5.8.1.1 --- Human tendon specimen --- p.94 / Chapter 5.8.1.2 --- In vitro system --- p.94 / Chapter 5.8.2 --- Understand the effect of dexmaethasone on human tendon fibroblasts --- p.95 / Chapter 5.8.3 --- Counteract the deleterious effects of dexamethasone by PDGFBB --- p.95 / Chapter CHAPTER VÍؤ --- CONCLUSION & FUTURE STUDY --- p.96 / Chapter 6.1 --- Conclusion --- p.97 / Chapter 6.2 --- Future study --- p.98 / Chapter 6.2.1 --- Study the balance between matrix synthesis and degradation --- p.98 / Chapter 6.2.2 --- Determine collagen typing --- p.99 / Chapter 6.2.3 --- Further explore the effect of glucocorticoid in organ culture model --- p.100 / Chapter 6.2.4 --- Investigate molecular mechanism of dexamethasone and PDGFBB --- p.100 / REFERENCES --- p.xv-xxv / APPENDIX --- p.xxvi

Growth factor delivery may be useful to accelerate the rate of tendon healing. We studied Platelet Concentrate, which in effect can be regarded as a cocktail of growth factors relevant for tendon healing. In a rat Achilles tendon transection model, one postoperative injection of Platelet Concentrate resulted in increased strength even 3 weeks later. Mechanical stimulation improves the repair of ruptured tendons. We studied the effects of platelets upon Achilles tendon regenerates in rats 3, 5 and 14 days after transection, either unloaded or mechanically stimulated. At 14 days, physical activity and platelets increased repair independently. Unloading decreased the mechanical properties of the repair tissue to less than half of normal. Moreover, the platelets had no effect without loading. Thrombin, which we used for platelet activation, improved healing of the rat Achilles tendon by itself. Conversely, continuous inhibition of thrombin by low molecular weight heparin (LMWH) inhibited tendon repair. However, intermittent inhibition, similar to clinical thromboprophylaxis, had no effect on tendon healing. Cartilage Derived Morphogenetic Protein-2 (CDMP-2) can improve tendon healing in loaded defect models. We now studied unloaded repair in a rabbit patellar tendon model. Two hours postoperative, the rabbits received CDMP-2 injected into the haematoma. The healing tendon became 65 % stronger than controls. We then studied Achilles tendon healing with CDMP-2 injections in sheep, to get a bigger animal model. There was an unexpectedly high variation of repair in these animals, and the study turned out to be underpowered. Spontaneous ruptures in humans have a more variable geometry than in our sheep model, so humans can also be expected to vary a lot in mechanical characteristics of Achilles tendon repair. This accentuates the importance of individualized rehabilitation programs. In conclusion, both platelet concentrate and CDMP-2 injections might be of interest for clinical use as a complement to surgical or conservative treatment of tendon ruptures. Platelet treatment for tendon ruptures should probably be combined with early physiotherapy.

O objetivo deste estudo foi avaliar o efeito dos tempos de aplicação 3, 4 e 5 minutos por ERA do ultrassom terapêutico (UST) na organização das fibras de colágeno em lesão do tendão do calcâneo de ratos. Foram utilizados quarenta ratos machos Wistar, dos quais 32 sofreram tenotomia total do tendão do calcâneo e foram divididos em 5 grupos: GC, sem tenotomia e tratamento; GT, com tenotomia e sem tratamento; UST3, UST4 e UST5 submetidos à tenotomia e tratados com UST nos tempos de 3, 4 e 5 minutos por área de radiação efetiva respectivamente. Os animais foram submetidos à primeira aplicação do UST foi 24 horas após a cirurgia de tenotomia. A irradiação ultrassônica foi aplicada com os seguintes parâmetros: 1 MHz, modo pulsado com 20% do ciclo de trabalho (2 ms de emissão / 8 ms de intervalo), frequência de 100 Hz, 0,5 W / cm² de intensidade e ERA de 0,5 cm². A aplicação foi realizada 1x/dia. Os animais foram sacrificados após a 10ª sessão de tratamento, no 12º dia pós-operatório. Os tendões foram retirados cirurgicamente para análise da organização das fibras colágenas através do método de birrefringência (retardo óptico - OR). As fibras colágenas mostraram melhor agregação e organização no grupo UST3, UST4 e UST5 quando comparado ao GT (p<0.05) e o UST5 apresentou melhor resposta na comparação intergrupos. Conclui-se que o UST, aplicado no tempo de 5 minutos por área de radiação efetiva, apresentou a melhor dose-resposta quanto à organização das fibras colágenas no reparo tecidual de tendões de ratos / The aim of this study was to evaluate the effect of application times 3, 4 and 5 minutes ERA of therapeutic ultrasound in the organization of the collagen fibers in rat calcaneal tendon injury. Forty male Wistar rats were used, of which 32 underwent complete tenotomy of the calcaneal tendon and were divided into 5 groups: GC without tenotomy and treatment; GT tenotomy with and without treatment; UST3, UST4 UST5 and submitted to tenotomy treated with therapeutic ultrasound at times 3, 4 and 5 minutes per effective radiating area respectively. The animals were submitted to the first application of therapy US tenotomy 24 hours after surgery. Ultrasonic irradiation was applied with the following parameters: 1 MHz, pulsed mode at 20% duty cycle (2ms transmission / 8 ms interval), frequency 100 Hz, 0.5 W / cm² intensity and ERA 0.5 cm². The application was performed 1x / day. The animals were sacrificed after the 10th treatment session, on the 12th postoperative day. The tendons were surgically removed for analysis of the organization of the collagen fibers through birefringence method (optical delay - OR). The collagen fibers showed better aggregation and organization in group UST3, UST4 and UST5 when compared to the GT (p <0.05) and UST5 showed better response in the intergroup comparison. We conclude that the UST, applied in time of 5 minutes for effective radiation area, presented the best dose-response as the organization of the collagen fibers in tissue repair of rat tendons

O objetivo deste estudo foi avaliar o efeito de diferentes tempos de tratamento do ultrassom terapêutico na cicatrização de lesão tendínea. Quarenta ratos machos Wistar (300 ± 45g), dos quais 32 sofreram tenotomia do tendão do calcâneo, foram divididos em 5 grupos: grupo C, sem tenotomia e tratamento, grupo T, com tenotomia e sem tratamento, US1,US2 e US3 submetidos à tenotomia e tratados com UST nos tempos de 1, 2 e 3 minutos por área de transdutor. Os animais foram mortos no 12º dia pósoperatório e os tendões retirados cirurgicamente para análise da organização das fibras colágenas utilizou-se o método de birrefringência (retardo óptico - OR). As fibras colágenas mostraram melhor agregação e organização no grupo US3 quando comparado ao grupo T (p<0.05). Concluise que o UST, aplicado no tempo de 3 minutos por área tratada, melhorou a organização das fibras colágenas no reparo tecidual de tendões de ratos / The aim of this study was to evaluate the effects of different treatment times of therapeutic ultrasound (US) on tendon injury healing. Forty male Wistar rats were selected (300 ± 45g) and 32, who underwent tenotomy of the Achilles tendon, were divided into five groups: Control, without tenotomy nor any treatment; tenotomy group, with tenotomy and without treatment; US groups (US1, US2, and US3), submitted to tenotomy and treated respectively with US for one, two, and three minutes per area of the transducer. The animals were sacrificed on the 12th post-operative day and the tendons were surgically removed for analyses of the collagen fiber organization by means of birefringence analyses, or optical retard. The collagen fibers showed better aggregation and organization in the US3 group, compared to the tenotomy group (p<0.05). The findings indicated that the US applied for three minutes per treated area improved the organization of the collagen fibers in the tendon repair of rats

In summary, the biomechanical stimulations can augment osteotendinous healing processes by facilitating better fibrocartilagious transitional zone regeneration as well as the restoration of proprioceptions, and the early application showed the more beneficial effects. However, further experimental and clinical studies are still needed to explore the optimal timing, intensity, frequency, and duration of the proposed postoperative biomechanical stimulation protocols. / LIPUS is a "non-contact" biomechanical stimulation, which can provide a direct mechanical stimulation through cavitation and acoustic microstreaming effects to improve tissue healing in a less-than-rigid biomechanical environment. So the mechanical stimulation induced from LIPUS could be applied immediately after surgery without worrying about the mechanical strain exceed the structural property at the osteotendinous healing interface in the early phase of repair. In this part of study, we also examined the effects of the regime of biomechanical stimulations applying immediately after repair on the osteotendinous healing interface. By using the same healing junction model, forty-two female New Zealand white rabbits were randomly divided into two groups; daily mechanical stimulation was applied immediately after surgery lasting up to post-operative 12 weeks on the healing interface in the treatment group. The regime of mechanical stimulations included by LIPUS was 20 minutes, 5 days per week for 4 weeks, followed by cyclic mechanical stimulation generated from quadriceps muscles induced by FES for 8 weeks. Results showed that early application of biomechanical stimulations on the osteotendinous healing interface were significantly better radiologically, histologically and biomechanically than that of not any or later application of the biomechanical stimulations during the osteotendinous healing processes when assessing at the same healing time point. In addition, the early application of biomechanical stimulations showed the better functional recovery in terms of the restoration of the proprioceptions, which an increased numbers of sensory nerve endings labeled by calcitonin gene-relate peptide (CGRP) was detected in the whole osteotendinous healing complex. / Sports or trauma injuries around osteotendinous junctions are common; treatments usually require surgical reattachment of the involved tendon to bone. Restoration of osteotendinous junction after repair is slow and difficult due to regenerating the intermitted fibrocartilage zone to connect two different characteristic tissues, tendon to bone. Although the factors influencing fibrocartilage zone regeneration and remodeling during osteotendinous repair are poorly understood, however, is believed that the mechanical environment plays an important role in such healing process. In present study, the effects of mechanical stimulation on osteotendinous healing process were examined, in the way of mechanical stimulations induced by biophysical stimulations, surface functional electric stimulation (FES) and low intensity pulsed ultrasound (LIPUS), applying on the patellar tendon to patellar bone healing interface in an established partial patellectomy model in rabbits. / The mechanotransductive stimulation linked to the transmission of forces across osteotendinous junction can be generated from its muscle contraction induced by FES. In the partial patellectomy model, thirty-five female New Zealand white rabbits were randomly divided into two groups with initial immobilization for 6 weeks, daily FES was applied to quadriceps muscles for 30 minutes, 5 days per week for 6 weeks in treatment group and compared with non-treatment control group at postoperative week 6, 12 and 18, radiologically, histologically and biomechanically. Results showed that FES-induced cyclic mechanical stimulation significantly increased new bone formation and its bone mineral density. An elevated expression of tenascin C and TGFbeta1; an increased proteoglycant stainability; mature fibrocartilage zone formation with better resumptions of biomechanical properties also observed on the osteotendinous healing interface, indicating that the post-operative programmed cyclic mechanical stimulation generated from its muscle contraction has beneficial effects on osteotendinous healing processes by facilitating the fibrocartilagious transitional zone regeneration. / by Wang Wen. / Advisers: Kai Ming Chan; Ling Qin. / Source: Dissertation Abstracts International, Volume: 68-03, Section: B, page: 1550. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2006. / Includes bibliographical references (p. 159-175). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Electronic reproduction. [Ann Arbor, MI] : ProQuest Information and Learning, [200-] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstracts in English and Chinese. / School code: 1307.

Establishment of animal model for studying treatment efficacy of low-intensity pulsed ultrasound stimulations for accelerating bone-tendon repair. Standard partial patellectomy was conducted in the 18-week old rabbits that were then divided into the LIPUS treatment and control groups. The animals were followed for 2, 4, 8, and 16 weeks for various tissue analyses. LIPUS was applied to the experimental animals from postoperative day 3 to 16 weeks. We demonstrated that the healing process of PPT junction was initiated through endochondral ossification. The results showed that the size and length of newly formed bone, and its bone mineral content (BMC), but not its bone mineral density (BMD) were correlated with the failure load, ultimate strength and energy at failure. Using radiographic, biomechanical, histomorphologic and biomechanical methods, it was found that LIPUS had significant accelerating effect on PPT junction repair. We validated our study hypothesis in that LIPUS enhances bone-tendon junction healing by stimulating angiogenesis, chondrogenesis and osteogenesis. / Establishment of in vitro model for mechanism study on effects of low-intensity pulsed ultrasound stimulations. An in vitro model of osteoblast-like cell line (SaOS-2 cells) was studied using cDNA microarray to explore the molecular mechanism mediated by LIPUS. This microarray analysis revealed a total of 165 genes that were regulated at 4 and 24 hours by LIPUS treatment in osteoblastic-like cells. These genes belonged to more than ten protein families based on their function and were involved in some signal transduction pathways. This study has validated the hypothesis that LIPUS can regulate a number of critical genes transient expressions in osteoblast cell line Saos-2. / Keywords. partial patellectomy model; bone-tendon junction repair; low intensity pulsed ultrasound stimulations (LIPUS); gene expression; complementary DNA microarray; rabbit. / This study explored the intact morphology, regular healing and the augmented healing under the effects of low intensity pulsed ultrasound stimulations (LIPUS) on the patella-patella tendon (PPT) junction in a rabbit partial patellectomy model. To probe its possible mechanism, the key genes involved in regulating osteogenesis mediated by LIPUS were identified using the state-of-the-art methods---complementary DNA microarray. / Lu Hongbin. / "June 2006." / Advisers: Ling Qin; Kwok Sui Leung. / Source: Dissertation Abstracts International, Volume: 68-03, Section: B, page: 1548. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2006. / Includes bibliographical references (p. 259-288). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Electronic reproduction. [Ann Arbor, MI] : ProQuest Information and Learning, [200-] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstracts in English and Chinese. / School code: 1307.

O objetivo deste estudo foi avaliar o efeito de diferentes tempos de tratamento do ultrassom terapêutico na cicatrização de lesão tendínea. Quarenta ratos machos Wistar (300 ± 45g), dos quais 32 sofreram tenotomia do tendão do calcâneo, foram divididos em 5 grupos: grupo C, sem tenotomia e tratamento, grupo T, com tenotomia e sem tratamento, US1,US2 e US3 submetidos à tenotomia e tratados com UST nos tempos de 1, 2 e 3 minutos por área de transdutor. Os animais foram mortos no 12º dia pósoperatório e os tendões retirados cirurgicamente para análise da organização das fibras colágenas utilizou-se o método de birrefringência (retardo óptico - OR). As fibras colágenas mostraram melhor agregação e organização no grupo US3 quando comparado ao grupo T (p<0.05). Concluise que o UST, aplicado no tempo de 3 minutos por área tratada, melhorou a organização das fibras colágenas no reparo tecidual de tendões de ratos / The aim of this study was to evaluate the effects of different treatment times of therapeutic ultrasound (US) on tendon injury healing. Forty male Wistar rats were selected (300 ± 45g) and 32, who underwent tenotomy of the Achilles tendon, were divided into five groups: Control, without tenotomy nor any treatment; tenotomy group, with tenotomy and without treatment; US groups (US1, US2, and US3), submitted to tenotomy and treated respectively with US for one, two, and three minutes per area of the transducer. The animals were sacrificed on the 12th post-operative day and the tendons were surgically removed for analyses of the collagen fiber organization by means of birefringence analyses, or optical retard. The collagen fibers showed better aggregation and organization in the US3 group, compared to the tenotomy group (p<0.05). The findings indicated that the US applied for three minutes per treated area improved the organization of the collagen fibers in the tendon repair of rats

O objetivo deste estudo foi avaliar o efeito dos tempos de aplicação 3, 4 e 5 minutos por ERA do ultrassom terapêutico (UST) na organização das fibras de colágeno em lesão do tendão do calcâneo de ratos. Foram utilizados quarenta ratos machos Wistar, dos quais 32 sofreram tenotomia total do tendão do calcâneo e foram divididos em 5 grupos: GC, sem tenotomia e tratamento; GT, com tenotomia e sem tratamento; UST3, UST4 e UST5 submetidos à tenotomia e tratados com UST nos tempos de 3, 4 e 5 minutos por área de radiação efetiva respectivamente. Os animais foram submetidos à primeira aplicação do UST foi 24 horas após a cirurgia de tenotomia. A irradiação ultrassônica foi aplicada com os seguintes parâmetros: 1 MHz, modo pulsado com 20% do ciclo de trabalho (2 ms de emissão / 8 ms de intervalo), frequência de 100 Hz, 0,5 W / cm² de intensidade e ERA de 0,5 cm². A aplicação foi realizada 1x/dia. Os animais foram sacrificados após a 10ª sessão de tratamento, no 12º dia pós-operatório. Os tendões foram retirados cirurgicamente para análise da organização das fibras colágenas através do método de birrefringência (retardo óptico - OR). As fibras colágenas mostraram melhor agregação e organização no grupo UST3, UST4 e UST5 quando comparado ao GT (p<0.05) e o UST5 apresentou melhor resposta na comparação intergrupos. Conclui-se que o UST, aplicado no tempo de 5 minutos por área de radiação efetiva, apresentou a melhor dose-resposta quanto à organização das fibras colágenas no reparo tecidual de tendões de ratos / The aim of this study was to evaluate the effect of application times 3, 4 and 5 minutes ERA of therapeutic ultrasound in the organization of the collagen fibers in rat calcaneal tendon injury. Forty male Wistar rats were used, of which 32 underwent complete tenotomy of the calcaneal tendon and were divided into 5 groups: GC without tenotomy and treatment; GT tenotomy with and without treatment; UST3, UST4 UST5 and submitted to tenotomy treated with therapeutic ultrasound at times 3, 4 and 5 minutes per effective radiating area respectively. The animals were submitted to the first application of therapy US tenotomy 24 hours after surgery. Ultrasonic irradiation was applied with the following parameters: 1 MHz, pulsed mode at 20% duty cycle (2ms transmission / 8 ms interval), frequency 100 Hz, 0.5 W / cm² intensity and ERA 0.5 cm². The application was performed 1x / day. The animals were sacrificed after the 10th treatment session, on the 12th postoperative day. The tendons were surgically removed for analysis of the organization of the collagen fibers through birefringence method (optical delay - OR). The collagen fibers showed better aggregation and organization in group UST3, UST4 and UST5 when compared to the GT (p <0.05) and UST5 showed better response in the intergroup comparison. We conclude that the UST, applied in time of 5 minutes for effective radiation area, presented the best dose-response as the organization of the collagen fibers in tissue repair of rat tendons

Until today equine flexor tendon injuries represent a major entity within the scope of equine medicine. Since their response to treatment is often poor, they remain an important therapeutic challenge for researchers as well as clinicians. It is of great importance to emphasize the tedious nature of the natural healing response within the tendon. This inevitably ends in a repair including scar tissue formation which is inferior to the original tendon tissue especially regarding its biomechanical properties and therefore, predisposing to reinjury. The wide range of treatment strategies for tendinopathies or suspensory ligament pathologies reflects the tenacious nature of the disease as well as lack of universal therapeutic effectiveness. The use of regenerative treatment strategies opened up new prospects for the treatment of tendon injuries and showed promising results. However, research efforts are still warranted to objectively asses their efficacy. At the same time basic science concerning aetiopathogenesis and pathophysiology of equine tendon injuries is still a factor of major importance regarding the development of effective strategies for therapy and prevention. Recently, the use of regenerative medicine has rapidly increased, progressively upstaging traditional surgical treatment options for equine tendon or suspensory ligament pathologies. The aim of the present study on the one hand was the analysis of the results and evaluation of the surgical treatment options percutaneous desmoplasty and osteostixis at the origin of the suspensory ligament by means of retrospective clinical case studies. On the other hand, success of these techniques was evaluated depending on different success parameters. These included the age of the horse, duration of lameness prior to surgery, observance of a given controlled exercise program by the owners and ground conditions. In case of percutaneous osteostixis also presence of a hindlimb conformation predisposing for proximal suspensory desmitis was investigated. Part of the first retrospective study examining percutaneous desmoplasty (tendon splitting) as a single surgical treatment were 71 horses (n=85). Desmoplasty was performed at the origin of the suspensory ligament (41 cases), inferior check ligament (21 cases), superficial flexor tendon (13 cases), branch of the suspensory ligament (6 cases) or at the body of the suspensory ligament (4 cases). The second retrospective study investigated the implementation of percutaneous osteostixis at the origin of the suspensory ligament in 14 horses (n=16). Percutaneous osteostixis was carried out alone (1 case), in conjunction with desmoplasty at the origin of the suspensory ligament (7 cases) and combined with desmoplasty, neurectomy of the deep branch of the lateral plantar nerve and plantar fasciotomy (8 cases). Concerning percutaneous osteostixis overall successful outcome was achieved in 11 horses (78.6 %) respectively 12 cases (75.0 %). The various evaluated parameters showed no statistically significant influence on the success of treatment. For percutaneous desmoplasty overall successful outcome was achieved in 55 cases (67.9 %, n=81). Observance of the controlled exercise program showed a statistically significant influence on the success of treatment (n=79, p<0.05). In summary the present paper illustrates the important role of percutaneous desmoplasty in nowadays` management of tendon and suspensory ligament injuries in the horse. Furthermore, percutaneous osteostixis proved to be a valuable alternative to conservative treatment for osseous as well as soft tissue injuries at the origin of the suspensory ligament, specifically in case of recurring lesions not responding to desmoplasty alone. Therefore, despite the rapidly increasing significance of regenerative treatment strategies these traditional surgical approaches should not fall into oblivion for they still have unchanged effectiveness. Especially postoperative implementation of a controlled and gradually increasing exercise program represents an essential part of rehabilitation, highlighted by the significant connection with successfull outcome of desmoplasty shown in the present paper, as a key factor greatly contributing to the success of each form of therapy:1 EINLEITUNG 1 2 LITERATURÜBERSICHT 3 2.1 Einführung in die Thematik der equinen Sehnenerkrankungen 3 2.1.1 Anatomische Grundlagen 3 2.1.2 Histologie der Sehne 4 2.1.3 Biomechanik der Sehne 7 2.1.4 Ätiologie und Pathogenese von Sehnenerkrankungen 11 2.1.5 Mechanismus der Sehnenheilung 20 2.1.6 Grundsätzliches zur Therapie von Sehnenerkrankungen 29 2.1.7 Definitionen 32 2.2 Das perkutane Sehnensplitting beim Pferd 33 2.2.1 Grundprinzip und Indikationen 33 2.2.2 Bedeutung des postoperativen Managements 35 2.3 Einsatzgebiete der perkutanen Osteostixis im Bereich des Fesselträgerursprungs 38 2.3.1 Allgemeines zur Methodik der Osteostixis 38 2.3.2 Anatomische Verhältnisse im Bereich des FTRU 40 2.3.3 Klinisches Erscheinungsbild der PSD 42 2.3.4 Sonographische Untersuchung 43 2.3.5 Röntgenologische Untersuchung 46 2.3.6 Szintigraphische Untersuchung 47 2.3.7 Magnetresonanztomographische Untersuchung 48 2.3.8 Innervation des FTRU 49 2.3.9 Diagnostische Anästhesien 49 2.3.10 Begleitoperationen zur Osteostixis: Faziotomie, Neurektomie und Splitting 50 2.3.11 Knochenschmerz 55 2.3.12 Prädispositionen für die Entwicklung einer PSD 55 3 TIERE, MATERIAL UND METHODEN 57 3.1 Sehnensplitting 57 3.1.1 Material 57 3.1.1.1 Einschlusskriterien 57 3.1.1.2 Alters-, Geschlechts-, Nutzungs- und Rassenverteilung 57 3.1.1.3 Erkrankte Gliedmaßen und Strukturen 57 3.1.1.4 Begleitende Eingriffe 58 3.1.1.5 Vorbehandlung 58 3.1.1.6 Begleitende Behandlung 58 3.1.1.7 Folgeinformationen 58 3.1.2 Methoden 58 3.1.2.1 Diagnostik 58 3.1.2.1.1 Klinische Lahmheitsuntersuchung 58 3.1.2.1.2 Befunderhebung mittels bildgebender Verfahren 59 3.1.2.2 Durchführung 60 3.1.2.2.1 Methodik des Sehnensplittings 60 3.1.2.2.2 Postoperatives Management 61 3.1.2.2.3 Kontrolliertes Bewegungsprogramm 61 3.1.2.3 Statistik 62 3.2. Osteostixis 63 3.2.1 Material 63 3.2.1.1 Einschlusskriterien 63 3.2.1.2 Alters-, Geschlechts-, Nutzungs- und Rassenverteilung 64 3.2.1.3 Erkrankte Gliedmaßen und begleitende Eingriffe 64 3.2.1.4 Vorbehandlung 64 3.2.1.5 Begleitende Behandlung 64 3.2.1.6 Folgeinformationen 64 3.2.2 Methoden 65 3.2.2.1 Diagnostik 65 3.2.2.1.1 Klinische Lahmheitsuntersuchung 65 3.2.2.1.2 Befunderhebung mittels bildgebender Verfahren 66 3.2.2.2 Durchführung 66 3.2.2.2.1 Methodik der perkutanen Osteostixis sowie der Begleitoperationen 66 3.2.2.2.2 Postoperatives Management 68 3.2.2.2.3 Kontrolliertes Bewegungsprogramm 68 3.2.2.3 Statistik 69 4 ERGEBNISSE 70 4.1 Ergebnisse Sehnensplitting 70 4.1.1 Sehnenübergreifende Ergebnisse 70 4.1.2 Sehnenspezifische Ergebnisse 72 4.1.2.1 Splitting des Fesselträgerkörpers (FTRK, 4 Fälle) 72 4.1.2.2 Splitting des Fesselträgerschenkels (FTRS, 6 Fälle) 72 4.1.2.3 Splitting der oberflächlichen Beugesehne (OBS, 13 Fälle) 73 4.1.2.4 Splitting des Unterstützungsbandes der tiefen Beugesehne (UB-TBS, 21 Fälle) 74 4.1.2.5 Splitting des Fesselträgerursprungs (FTRU, 41 Fälle) 75 4.2 Ergebnisse Osteostixis 78 4.2.1 Fallbetrachtungen Osteostixis 79 4.2.1.1 Osteostixis ohne Begleitoperationen (1 Fall) 79 4.2.1.2 Osteostixis mit Splitting des FTRU (7 Fälle) 79 4.2.1.3 Osteostixis mit Splitting sowie Fasziotomie und Neurektomie (8 Fälle) 81 5 DISKUSSION 84 5.1. Anmerkungen zum Studienaufbau 84 5.2 Beurteilung der Ergebnisse des perkutanen Sehnensplittings 84 5.3 Beurteilung der Ergebnisse der perkutanen Osteostixis 88 5.4 Schlussfolgerungen 91 6 ZUSAMMENFASSUNG 93 7 SUMMARY 95 8 LITERATURVERZEICHNIS 97 9 DANKSAGUNG 110

A direct bone tendon junction consists of four zones: tendon, uncalcified fibrocartilage, calcified fibrocartilage, and bone. The uncalcified and calcified fibrocartilage together forms the transition zone. This organization ensures a gradual transition in stiffness and material properties, and protects the junction from failure. Transition zone regeneration during bone tendon junction healing is important to restore this unique protective mechanism. / Bone tendon junction repair is involved in many orthopaedic reconstructive procedures. Healing is observed to be slow. The junction often heals by fibrous tissue formation. Previous attempts to enhance bone tendon junction healing have resulted in increased bone formation. However, fibrocartilage transition zone is not restored. / This thesis describes a series of studies on transition zone regeneration in bone tendon junction healing using two partial patellectomy animal models. The healing process inside a bone trough was first studied and characterized. Little transition zone regeneration was observed except near the articular cartilage cut surface. The possibility of using articular cartilage to stimulate transition zone regeneration was explored. Both articular cartilage autograft and allogeneic cultured chondrocyte pellet implantations resulted in significantly increased fibrocartilage transition zone regeneration. Cell tracking indicated that the regenerated tissue likely originated from host cells. To elucidate the mechanism of stimulation by allogeneic cultured chondrocyte pellet, the role of cellular and matrix component needed to be differentiated. Freezing and rapid freeze thaw cycles permanently devitalized the allogeneic cultured chondrocyte pellet, but retained its structural integrity and matrix contents. Preliminary results indicated that implantation of the devitalized allogeneic cultured chondrocyte pellet could still increase fibrocartilage transition zone regeneration. Cellular activity seemed not to be essential for the stimulatory effect. / With further research and development, it is envisioned that a cartilage-based stimulation method for fibrocartilage transition zone regeneration in bone tendon junction healing will be developed for clinical application. / Wong Wan Nar, Margaret. / Source: Dissertation Abstracts International, Volume: 70-06, Section: B, page: 3423. / Thesis (M.D.)--Chinese University of Hong Kong, 2008. / Includes bibliographical references (leaves 216-231). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Electronic reproduction. [Ann Arbor, MI] : ProQuest Information and Learning, [200-] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstracts in English and Chinese. / School code: 1307.