Suture or bone anchors are used to reattach a tendon or ligament after it has been torn away from the bone. Anchors provide secure attachments to bone during trauma or reconstructive surgery, holding the ligament or tendon in place and potentially allowing greater mobility during recovery. Computer modelling techniques are used to investigate both established bone anchor technology, such as threaded implants, and emerging technologies such as cement augmentation or sonic-fusion. Sonic fusion is an ultrasound-assisted anchoring method which has recently been introduced in low load maxillofacial applications, and is expected to be used in other low load applications such as hallux valgus alignment procedures and suture attachment. Threaded anchors were examined using two Finite Element (FE) models of human cancellous bone, representing both “normal” and “weaker” bone. Simulation and analysis revealed the critical nature of modelling the microstructure of bone. Changing the direction of loading in the model leads to significant changes in the response of the construct, and this cannot be represented in continuum models, or in physical models using artificial cancellous bone. Rapid prototyping (RP) using 3d printing was used for validation of the FE models. While this method has previously been implemented to create physical bone models, testing an assembly model and comparing it to FE results for inclined loading had not been attempted. RP models were created of the threaded anchor in both “normal” and “weaker” bone, and a sonic fusion model in the normal bone was also created. These models were then subjected to mechanical testing. Results produced from the simulation correlated with the physical results. The importance of a cortical layer was re-confirmed. At the apparent densities simulated, engagement with the cortical layer increases pull-out force dramatically. Engaging the anchor even with a thin cortical layer can produce a significant improvement to pull-out strength. Novel sonic fusion FE models were created from a CT scan of animal bone, and the geometry for both the sonic-fusion pin and bone were taken from the CT scan. Computer generated geometry was used to build pin concepts of varying shapes. It was shown that if good engagement is made with bone, as in the case of all of the concepts created, then sonic fusion can produce a good holding power - comparable with that of a threaded anchor. The results showed that sonic-fusion requires less drill penetration into the bone, meaning less of the inherent bone structure is removed – vital for patients with poor bone quality. Bone cement models were investigated. Bone augmentation models were created, and the addition of cement demonstrated an improvement in anchor holding power. The research showed that there are benefits to using FEA as a tool to evaluate the mechanical aspects of cement distribution. The results proved the hypothesis that augmentation will likely increase the holding power of anchor, and its distribution will affect pull-out significantly. This work has created a method for modelling and evaluating both established and novel bone anchor technology in CT bone geometry, a procedure which could be expanded to other bone implants. It has been validated using the innovative approach of rapid prototyping.

O alcoolismo é considerado uma doença crônica, trazendo prejuízos para saúde do indivíduo. O tecido ósseo também sofre sérios danos com o consumo crônico do álcool, pois induz defeitos na mineralização e redução da espessura do osso medular e cortical. Devido à grande procura por tratamentos reconstrutivos para lesões e perdas ósseas, os enxertos xenógenos vêm sendo uma boa opção para o tratamento regenerativo. Em vista da procura de pacientes que fazem consumo crônico do álcool e apresentam perdas ósseas, necessitando de enxertos xenógenos, este trabalho teve como objetivo comparar, por meio de análise histomorfológica e histomorfométrica, a ação de dois biomateriais, o Bio-Oss® e o OrthoGen®, no processo de reparo de defeitos ósseos em tíbia de ratos submetidos ou não ao alcoolismo experimental. Foram utilizados 80 ratos machos (Rattus norvegicus) da linhagem Wistar, com 60 dias de idade, separados aleatoriamente em quatro grupos experimentais contendo 20 animais cada: Grupo Água Bio-Oss® (GAB), Grupo Etanol Bio-Oss® (GEB), Grupo Água OrthoGen® (GAO) e Grupo Etanol OrthoGen® (GEO). Os animais dos grupos GEB e GEO foram submetidos à adaptação gradativa ao álcool e depois permaneceram a 25% por 90 dias. Após esse período, todos os animais foram submetidos à cirurgia experimental. Foi realizada uma cavidade cirúrgica de aproximadamente 3 mm de diâmetro na epífise proximal da tíbia e nos animais dos grupos experimentais GAB e GEB se fez o preenchimento com o biomaterial Bio-Oss®. Os animais dos grupos experimentais GAO e GEO receberam o preenchimento da cavidade com o biomaterial OrthoGen®. Decorridos os períodos de 10, 20, 40 e 60 dias pós-cirúrgico, cinco animais de cada grupo, por período, foram eutanasiados. Os resultados histomorfológicos demostraram que os animais dos grupos GAB e GEB apresentaram células sanguíneas e tecido conjuntivo nos períodos de 10 e 20 dias, e nos períodos de 40 e 60 dias ocorreu a formação de novo osso, sendo que o grupo GEB apresentou um maior retardo na formação quando comparado ao GAB. Nos grupos GAO e GEO o início da formação de tecido ósseo ocorreu nos períodos de 40 e 60 dias, com um retardo na formação quando comparado ao biomaterial Bio-Oss®. Na análise histomorfométrica, comparando os grupos que receberam a dieta líquida não alcóolica com biomaterias diferentes (GAB e GAO) ocorreu diferença significante em formação óssea em todos os períodos sendo que o grupo Água Bio-Oss® apresentou uma maior formação óssea. Nos grupos que receberam a dieta alcóolica, com biomateriais diferentes (GEB e GEO), ocorreu diferença em formação óssea nos períodos de 20, 40 e 60 dias, onde o biomaterial Bio-Oss® apresentou maiores médias. Comparando os grupos que receberam o mesmo biomaterial, mas com dieta líquida diferente (GABXGEB e GAOXGEO), evidenciou-se diferença significante em formação óssea em todos os períodos (exceto GAOXGEO; 10 dias), sendo que os grupos que receberam dieta líquida apenas com água apresentaram melhores resultados. Conclui-se que o biomaterial Bio-Oss® mostrou-se superior na neoformação óssea quando comparado ao Ortoghen®, e que a dieta alcoólica interferiu de forma negativa no processo de reparação. / Alcoholism is considered a chronic disease, which brings harm to an individuals health. Bones also suffer serious damage resulting from the chronic consumption of alcohol, since it induces defects in the mineralization and reduction of the thickness of the cortical and the cancellous bone. Xenografts have been a good option for regenerative treatments in response to the high search for reconstructive treatments for bone losses and lesions. Given the demand of patients who make chronic use of alcohol and present bone loss, needing xenografts, the aim of this study was to compare, by means of a histomorphological and histomorphometric analysis, the action of the biomaterials Bio-Oss® and OrthoGen® in the repair process of bone defects in the tibia of rats submitted or not to experimental alcoholism. Eighty male Wistar rats (Rattus norvegicus) aged 60 days were randomly separated into four experimental groups with 20 animals each: Bio-Oss® Water Group (GAB), Bio-Oss® Ethanol Group (GEB), OrthoGen® Water Group (GAO) and OrthoGen® Ethanol Group (GEO). The animals in the GEB and GEO were submitted to gradual adaptation to alcohol and later kept at 25% for 90 days. After this period, all animals were submitted to experimental surgery. A surgical cavity with approximately 3 mm of diameter was made in the proximal tibial epiphysis and in animals from the experimental groups GAB and GEB it was filled with the Bio-Oss® biomaterial. Animals from the experimental groups GAO and GEO had their cavity filled with the OrthoGen® biomaterial. After 10, 20, 40 and 60 days post-surgery, five animals in each group, per period, were euthanized. The histomorphological results showed that the animals from the GAB and GEB groups presented blood cells and connective tissue in the 10 and 20 days periods, and in the periods of 40 and 60 days there was formation of new bone, with the GEB group presenting greater delay in formation when compared to the GAB. In the GAO and GEO groups, bone formation started in the periods of 40 and 60 days, with a delay in formation when compared to the Bio-Oss® biomaterial. In the histomorphometric analysis, comparing the groups that received nonalcoholic liquid diet with different biomaterials (GAB and GAO), there was a significant difference in bone formation in all periods, with the Bio-Oss® Water Group presenting greater bone formation. In the groups that received an alcoholic diet, with different biomaterials (GEB and GEO), there was a difference in bone formation in the periods of 20, 40 and 60 days, in which the Bio-Oss® biomaterial presented greater means. Comparing the groups that received the same biomaterial, but with a different liquid diet (GABXGEB and GAOXGEO), there was a significant difference in bone formation in all periods (except for GAOXGEO; 10 days), in which the groups that received a liquid diet with only water presented better results. In conclusion, the Bio-Oss® biomaterial presented better performance in bone neoformation when compared to the Ortoghen®, and the alcoholic diet affected negatively the repair process.

No description available.

by Cheng Sze Lok, Alfred. / Thesis (M.Phil.)--Chinese University of Hong Kong, 1998. / Includes bibliographical references (leaves 104-109). / Abstract also in Chinese. / DECLARATION --- p.i / ABSTRACT --- p.ii / ACKNOWLEDGEMENT --- p.vii / ABBREVIATIONS --- p.ix / LIST OF FIGURES --- p.x / LIST OF TABLES --- p.xii / TABLE OF CONTENTS --- p.xiii / Chapter CHAPTER ONE 226}0ؤ --- INTRODUCTION / Chapter 1.1 --- The Growth Plate / Chapter 1.1.1 --- "Function, Structure and Biochemistry of the Growth Plate" --- p.1 / Chapter 1.1.2 --- Extracellular Matrix of the Growth Plate Cartilage --- p.4 / Chapter 1.1.3 --- Vascular Supply to the Growth Plate --- p.9 / Chapter 1.1.4 --- Endochondral Ossification --- p.10 / Chapter 1.2 --- Growth Plate Damage and the Contemporary Reconstruction Models --- p.13 / Chapter 1.3 --- The 3-D Chondrocyte Pellet Culture --- p.15 / Chapter 1.4 --- The Study Plan --- p.16 / Chapter 1.5 --- The Objectives of the Study --- p.18 / Chapter CHAPTER TWO 一 --- METHODOLOGY / Chapter 2.1 --- Biosynthesis of Artificial Growth Plate using 3-D Chondrocyte Pellet Culture / Chapter 2.1.1 --- Isolation of Rabbit Costal Resting Chondrocytes --- p.19 / Chapter 2.1.2 --- Chondrocyte Monolayer Culture --- p.20 / Chapter 2.1.3 --- Three-dimensional Chondrocyte Pellet Culture --- p.20 / Chapter 2.1.4 --- Optimization of 3-D Chondrocyte Pellet Culture System --- p.20 / Chapter 2.2 --- Characterization of the 3-D Chondrocyte Pellet Culture and Monolayer Culture / Chapter 2.2.1 --- Histomorphology --- p.22 / Chapter 2.2.2 --- Alkaline Phosphatase Histochemistry --- p.22 / Chapter 2.2.3 --- Collagen Typing --- p.23 / Chapter 2.2.3.1 --- Labeling and extraction of newly synthesized collagen / Chapter 2.2.3.2 --- SDS-PAGE and autoradiography / Chapter 2.2.4 --- Growth Rate --- p.25 / Chapter 2.2.4.1 --- Total DNA content determination / Chapter 2.2.4.2 --- Thymidine incorporation assay / Chapter 2.3 --- Implantation of Artificial Growth Plate and Assessment / Chapter 2.3.1 --- Implantation of Artificial Growth Plate into Partial Growth Plate Defect Model --- p.27 / Chapter 2.3.1.1 --- Animals / Chapter 2.3.1.2 --- Surgical procedure / Chapter 2.3.1.3 --- Experimental groups / Chapter 2.3.2 --- Histology --- p.30 / Chapter 2.3.3 --- Metabolism of Artificial Growth Plate In Vivo --- p.31 / Chapter 2.3.3.1 --- Radio sulfate labeling / Chapter 2.3.3.2 --- Liquid emulsion and autoradiography / Chapter CHAPTER THREE 一 --- RESULTS / Chapter 3.1 --- Biosynthesis of Artificial Growth Plate using 3-D Chondrocyte Pellet Culture / Chapter 3.1.1 --- Morphology of the Isolated Rabbit Chondrocyte --- p.32 / Chapter 3.1.2 --- Three-dimensional Chondrocyte Pellet Culture --- p.32 / Chapter 3.1.3 --- Optimization of 3-D Chondrocyte Pellet Culture System --- p.35 / Chapter 3.2 --- Characterization of the 3-D Chondrocyte Pellet Culture and Monolayer Culture / Chapter 3.2.1 --- Histomorphology --- p.38 / Chapter 3.2.2 --- Alkaline Phosphatase Histochemistry --- p.43 / Chapter 3.2.3 --- Collagen Typing --- p.47 / Chapter 3.2.4 --- Growth Rate --- p.50 / Chapter 3.2.4.1 --- Total DNA content determination / Chapter 3.2.4.2 --- Thymidine incorporation assay / Chapter 3.3 --- Implantation of Artificial Growth Plate and Assessment / Chapter 3.3.1 --- Histology --- p.54 / Chapter 3.3.2 --- Metabolism of Artificial Growth Plate In Vivo --- p.65 / Chapter CHAPTER FOUR 一 --- DISCUSSION / Chapter 4.1 --- Optimal Condition for 3-D Chondrocyte Pellet Culture System --- p.67 / Chapter 4.1.1 --- Some Critical Characteristics of the Growth Plate --- p.68 / Chapter 4.1.2 --- Selection of Animal Model --- p.69 / Chapter 4.1.3 --- Optimization of Culturing Conditions 226}0ؤ Screening Based on Morphological Studies --- p.69 / Chapter 4.2 --- Characterization of the 3-D Chondrocyte Pellet Culture and Monolayer Culture --- p.73 / Chapter 4.2.1 --- Development of the 3-D Chondrocyte Pellet Culture --- p.73 / Chapter 4.2.2 --- Development of the Chondrocyte Monolayer Culture --- p.78 / Chapter 4.2.3 --- Comparing the 3-D Chondrocyte Pellet Culture and Monolayer Culture --- p.79 / Chapter 4.2.3.1 --- Cellular organization / Chapter 4.2.3.2 --- Terminal differentiation of chondrocytes / Chapter 4.2.3.3 --- Cell division potential / Chapter 4.2.3.4 --- Production of cartilaginous matrix / Chapter 4.3 --- Resumption of Physeal Characteristics by Artificial Growth Plate In Vivo --- p.86 / Chapter 4.3.1 --- Three Stages of In Vivo Development of the Artificial Growth Plate --- p.86 / Chapter 4.3.1.1 --- Incorporation of artificial growth plate with host tissues / Chapter 4.3.1.2 --- Growth of the artificial growth plate invivo / Chapter 4.3.1.3 --- Resumption of endochondral ossification in the artificial growth plate / Chapter 4.3.2 --- Significance of Development of the 3-D Pellet Culture on its In Vivo Development --- p.89 / Chapter 4.3.2.1 --- 3-D pellet culture processes similar extracellular matrix with host / Chapter 4.3.2.2 --- 3-D pellet culture acquires growth plate-like cellular organization and differentiation pattern / Chapter 4.3.3 --- Effect of Host Microenvironment on Artificial Growth Plate Development --- p.90 / Chapter 4.3.3.1 --- Orientation of artificial growth plate implants / Chapter 4.3.3.2 --- Evidence from development of 3-D pellet culture in longer period of culture / Chapter 4.4 --- Comparison with other Growth Plate Reconstruction Models --- p.93 / Chapter 4.4.1 --- Implantation of Biologic or Inert Fillers --- p.93 / Chapter 4.4.2 --- Physeal Transplantation --- p.94 / Chapter 4.4.3 --- Transplantation of Cartilage Allografts --- p.95 / Chapter 4.4.4 --- Transplantation of High-density Chondrocyte Culture --- p.96 / Chapter CHAPTER FIVE 一 --- SUMMARY AND CONCLUSION --- p.98 / Chapter CHAPTER SIX 一 --- FURTHER STUDIES --- p.102 / REFERENCES --- p.104

Hung Wing Yin Vivian. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2004. / Includes bibliographical references (leaves 128-142). / Abstracts in English and Chinese ; appendix in Chinese. / ABSTRACT --- p.i / ABSTRACT (in Chinese) --- p.iv / ACKNOWLEDGMENT --- p.vii / TABLE OF CONTENTS --- p.viii / LIST OF TABLES --- p.xiv / LIST OF FIGURES --- p.xvi / LIST OF ABBREVIATIONS --- p.xix / Chapter I. --- INTRODUCTION --- p.1 / Chapter 1.1. --- Scoliosis --- p.1 / Chapter 1.1.1. --- Classification of scoliosis --- p.1 / Chapter 1.1.2. --- Idiopathic scoliosis --- p.1 / Chapter 1.1.3. --- Clinical examination --- p.2 / Chapter 1.1.4. --- Curve pattern --- p.2 / Chapter 1.2. --- Etiology of AIS --- p.3 / Chapter 1.2.1. --- Prevalence of AIS --- p.5 / Chapter 1.2.2. --- Anthropometric Measurement in AIS --- p.5 / Chapter 1.2.3. --- Bone mass --- p.6 / Chapter 1.2.4. --- Bone mineral density measurements --- p.6 / Chapter 1.2.5. --- Osteopenia in AIS --- p.7 / Chapter 1.3. --- Natural history ofAIS --- p.8 / Chapter 1.3.1. --- Curve progression --- p.9 / Chapter 1.3.2. --- Treatment of scoliosis --- p.11 / Chapter 1.4. --- Research questions --- p.12 / Chapter 1.5. --- Objectives --- p.13 / Chapter II. --- METHODOLOGY --- p.20 / Chapter 2.1 --- Study Design --- p.20 / Chapter 2.2 --- Subject recruitment --- p.20 / Chapter 2.2.1 --- AIS patients --- p.20 / Chapter 2.2.2 --- Inclusion criteria --- p.20 / Chapter 2.2.3 --- Exclusion criteria --- p.20 / Chapter 2.2.4 --- Informed consent --- p.21 / Chapter 2.3 --- Grouping for chronological age --- p.21 / Chapter 2.4 --- Radiography assessments --- p.21 / Chapter 2.4.1 --- Cobb angle measurement --- p.21 / Chapter 2.4.2 --- Curve pattern --- p.22 / Chapter 2.4.3 --- Risser grade --- p.22 / Chapter 2.5 --- Definition of curve progression --- p.22 / Chapter 2.6 --- Bone mineral density (BMD) measurements --- p.23 / Chapter 2.6.1 --- Dual energy X-ray Absorptiometry (DXA) --- p.23 / Chapter 2.6.2 --- Peripheral quantitative computed tomography (pQCT) --- p.24 / Chapter 2.6.3 --- Definition of osteopenia or low bone mass --- p.24 / Chapter 2.7 --- Anthropometric measurements --- p.25 / Chapter 2.7.1 --- Body height --- p.25 / Chapter 2.7.2 --- Body weight --- p.26 / Chapter 2.7.3 --- Arm span --- p.26 / Chapter 2.7.4 --- Sitting height --- p.27 / Chapter 2.8 --- Family history --- p.27 / Chapter 2.9 --- Menstrual status --- p.27 / Chapter 2.10 --- Medication and fracture history --- p.27 / Chapter 2.11 --- Statistical analysis --- p.27 / Chapter 2.11.1 --- Sample size power calculation --- p.28 / Chapter 2.11.2 --- Student t test --- p.28 / Chapter 2.11.3 --- Paired t-test --- p.28 / Chapter 2.11.4 --- Predicting the incidence of curve progression --- p.28 / Chapter 2.11.4.1 --- Predictive outcome --- p.28 / Chapter 2.11.4.2 --- Potential risk factors --- p.28 / Chapter 2.11.4.3 --- Coding system for categorical variables --- p.29 / Chapter 2.11.4.4 --- Univariate analysis --- p.30 / Chapter 2.11.4.5 --- Logistic regression --- p.30 / Chapter 2.11.4.6 --- Receiver operating characteristics (ROC) curves --- p.32 / Chapter III. --- RESULTS --- p.54 / Chapter 3.1 --- Patients Characteristics --- p.54 / Chapter 3.1.1 --- Sample size --- p.54 / Chapter 3.1.2 --- Distribution of patient characteristics --- p.54 / Chapter 3.1.3 --- Drop out --- p.54 / Chapter 3.1.4 --- Prevalence of osteopenia (BMDage-adjusted ≤ -1) and low bone mass (BMCage-adjusted ≤ -1) --- p.55 / Chapter 3.1.5 --- Comparison between the BMD of the bilateral hip and tibia --- p.55 / Chapter 3.2 --- Comparison of AIS patients with osteopenia and with normal bone status --- p.55 / Chapter 3.3 --- Univariate analysis --- p.56 / Chapter 3.3.1 --- Growth related factors --- p.56 / Chapter 3.3.2 --- "Skeletal related parameters (areal BMD, volumetric BMD and BMC)" --- p.56 / Chapter 3.3.2.1 --- DXA lumbar spine --- p.56 / Chapter 3.3.2.2 --- DXA proximal femur at the convex-side hip --- p.56 / Chapter 3.3.2.3 --- DXA proximal femur at the concave-side hip --- p.57 / Chapter 3.3.2.4 --- pQCT at non-dominant distal radius --- p.57 / Chapter 3.3.2.5 --- pQCT - vBMD at convex-side distal tibia --- p.57 / Chapter 3.3.2.6 --- pQCT - vBMD at concave-side distal tibia --- p.58 / Chapter 3.3.3 --- Curve related factors --- p.58 / Chapter 3.3.4 --- Anthropometrics parameters --- p.58 / Chapter 3.3.5 --- Family history --- p.58 / Chapter 3.3.6 --- Summary of univariate analysis --- p.59 / Chapter 3.4 --- Logistic regression model (single factor) --- p.59 / Chapter 3.5 --- Logistic regression model (multiple factors) --- p.60 / Chapter 3.5.1 --- BMD inclusive model --- p.60 / Chapter 3.5.2 --- BMC inclusive model --- p.61 / Chapter 3.5.3 --- Conventional model --- p.63 / Chapter 3.6 --- ROC curve --- p.63 / Chapter 3.6.1 --- BMD inclusive model --- p.64 / Chapter 3.6.2 --- Conventional model --- p.64 / Chapter 3.7 --- Predictive equation obtained from different logistic regression models --- p.64 / Chapter 3.7.1 --- BMD inclusive model --- p.65 / Chapter 3.7.2 --- Conventional model --- p.65 / Chapter IV. --- DISCUSSION --- p.105 / Chapter 4.1 --- Prognostic factors for curve progression --- p.105 / Chapter 4.1.1 --- Well-known prognostic factors --- p.105 / Chapter 4.1.1.1 --- Growth-related factors --- p.106 / Chapter 4.1.1.2 --- Initial curve magnitude --- p.107 / Chapter 4.1.2 --- A new predictor 一 Osteopenia --- p.107 / Chapter 4.2 --- Non-significant prognostic factors for curve progression --- p.109 / Chapter 4.2.1 --- Anthropometric parameters --- p.109 / Chapter 4.2.2 --- Family History --- p.110 / Chapter 4.2.3 --- Curve pattern --- p.110 / Chapter 4.3 --- Predictive model --- p.111 / Chapter 4.4 --- Comparison of predictive models between BMD inclusive model and conventional model derived from our population --- p.115 / Chapter 4.5 --- Possible relationship between osteopenia and etiopathogensis of AIS --- p.116 / Chapter 4.6 --- Axial measurement has a better predictive power in curve progression than peripheral measurement --- p.117 / Chapter 4.7 --- Discordance of BMD in bilateral hips --- p.118 / Chapter 4.8 --- Method justifications --- p.119 / Chapter 4.8.1 --- Definition of curve progression --- p.119 / Chapter 4.8.2 --- Incidence of progression as the outcome of prediction --- p.119 / Chapter 4.8.3 --- Selection on bone densitometers --- p.119 / Chapter 4.9 --- Clinical significance --- p.121 / Chapter 4.10 --- Limitations and Future Studies --- p.122 / Chapter 4.10.1 --- Limited follow-up time --- p.122 / Chapter 4.10.2 --- No defined cutoff value for 226}0´ببosteopenia 226}0ح or low BMC in paediatric area --- p.122 / Chapter 4.10.3 --- Predictive model could only applied in local population --- p.122 / Chapter 4.10.4 --- Intrinsic error in Risser grade measurement --- p.123 / Chapter 4.10.5 --- Further studies --- p.123 / Chapter 4.10.5.1 --- Validation of the newly developed predictive model --- p.123 / Chapter 4.10.5.2 --- Possible intervention of osteopenia --- p.124 / Chapter 4.10.5.3 --- Long term follow-up BMD measurements and fracture risk in AIS patients --- p.124 / Chapter 4.10.5.4 --- Discordance of bilateral hips BMD contributed by the shift of center of gravity --- p.125 / Chapter 4.10.5.5 --- Axial QCT can be an alternative method in assessing BMDin scoliotic patients --- p.125 / Chapter V. --- CONCLUSION --- p.126 / Chapter VI. --- APPENDIX --- p.127 / Chapter VII. --- BIBLIOGRAPHY --- p.128 / Chapter VIII. --- CONFERENCE PUBLICATIONS --- p.142

MR allows non-invasive means of evaluating the non-mineralized components of bone, particularly the bone marrow. This thesis focuses on potential changes occurring in bone marrow perfusion and marrow fat in osteoporosis, - changes which may improve our understanding of osteoporosis pathophysiology. We know from histology studies that as osteoporosis develops and bone tissue is lost, it is replaced by fat filling the vacated marrow space. MR allows non-invasive quantification of this fat component. Although fat content may be increasing, it is not known whether any change in fat composition occurs with osteoporosis i.e. does the type of fat within bone change. Epidemiological studies have indicated a link between arterial disease and osteoporosis. It is not known, however, whether any changes occur in bone perfusion in osteoporosis and how these may be related to increasing fat within the marrow. / The hypothesis to be tested is that: Advanced magnetic resonance (MR) techniques can be applied to investigate the non-mineralised components of bone tissue in osteoporosis thereby providing more information on bone physiology both in health and disease / This thesis is based on a series of eight studies designed to study the relationship between bone marrow perfusion, bone marrow fat content, bone marrow fat composition and bone mineral density. These studies showed that as bone mineral density decreased, bone marrow perfusion decreased. A strong reciprocal relationship was found between decreasing bone marrow perfusion and increasing marrow fat. The reduction in perfusion occurred only with bone and did not affect the extra-osseous tissues alongside bone with the same arterial supply. This indicates that the reduction in bone perfusion is not simply a reflection of a more generalized circulatory impairment in subjects with osteoporosis. This same effect is seen in both males and females and in the proximal femora as well as the spine. / In animal-based studies, we found that reduction in bone perfusion was apparent as little as two weeks after orchidectomy or oorphorectomy and closely paralleled features of impaired endothelial function as well as decreased bone mineral density and a hitherto unrecognized reduction in red marrow fraction within the medullary canal. Nitric oxide synthase, produced by the endothelium, is a potent stimulator of angiogenesis and osteoblastic activity. Mesenchymal stem cell differentiation may switch from osteoblastogenesis to adipocytogenesis under hypoxic conditions, while haematopoetic stem cells also supply endothelial stem cells. Potentially endothelial dysfunction, mesenchymal stem cell differentiation and haematopoetic stem cell maturation may be implemented in the development of osteoporosis. / In normal subjects, blood perfusion was markedly reduced in the femoral head compared to the femoral neck. In osteoporotic subjects, a further reduction in blood perfusion occurred in both areas. Overall perfusion indices reduced relatively more in the femoral neck than the femoral head region. These changes in bone perfusion help explain why subjects with osteoporosis have impaired healing of femoral neck fractures though do not seem to be at increased risk of avascular necrosis. At a micro-architectural level, reduced bone perfusion may also help explain the impaired healing of microfractures seen in subjects with osteoporosis, a feature likely to contribute to reduce bone strength, microfracture accumulation and eventually clinical insufficiency fracture. / Marrow fat was increased in subjects with osteoporosis. Our studies showed that percentage marrow fat content increased even allowing for the quantitative effect increased marrow fat has on the bone densitometry measurements. This effect was shown to be of negligible clinical significance. We found a strong reciprocal relationship between increasing fat and decreasing bone perfusion in both the proximal femur and vertebral body. Although fat content increased, very little difference in marrow fat composition was apparent between normal subjects and those with osteoporosis. We found no difference was apparent in either the N3/N6 marrow fat ratio or the spectrum of individual fatty acids in the marrow of subjects with either normal bone mineral density or osteoporosis. This suggests that alternation of marrow fat composition is not likely to be a direct contributory factor in osteoporosis. Also marrow fat increase was shown to be due to an increase in number rather than size of marrow fat cells. This suggests that marrow fat increases as a result of a switch in mesenchymal cell maturation to adipocytes rather than osteoblasts. / Below average perfusion indices in the acetabulum and adductor muscle is predictive of more pronounced bone loss at the femoral neck over the ensuing four years. Perfusion indices can also predict between fast and slow losers with a high sensitivity / To summarise, in the eight studies presented, it was shown that osteoporosis is associated with a significant reduction in bone perfusion and a reciprocal increase in marrow fat content though no change in marrow fat composition. Reduction in bone perfusion is most likely due to an accompanying reduction in functioning marrow fraction. Marrow fat increase is most likely the result of a switch in mesenchymal cell maturation to adipocytes rather than osteoblasts. The studies present in this thesis confirmed the initial hypothesis that “Advanced magnetic resonance techniques can be applied to investigate the non-mineralised components of bone tissue in osteoporosis thereby providing more information on bone physiology both in health and disease. / Griffith, James Frances. / "June 2009." / Thesis (M.D.)--Chinese University of Hong Kong, 2012. / Includes bibliographical references (leaves 227-250). / Appendix includes Chinese. / PREFACE AND DECLARATION --- p.1 / DEDICATION --- p.2 / ACKNOWLEDGEMENT --- p.3 / PRECIS --- p.4 / PUBLICATIONS AND PRESENTATIONS OF STUDIES RELATED TO THIS THESIS --- p.8 / INTRODUCTION --- p.16 / Chapter STUDY 1 --- What is the relationship between bone perfusion, marrow fat and bone mineral density? --- p.76 / Chapter STUDY 2 --- Vertebral marrow fat content, molecular diffusion, and perfusion indices in women with varying bone density, including osteoporosis: MR evaluation --- p.94 / Chapter STUDY 3 --- Could the results of Study 1 and Study 2 be spurious due to the effect of increasing marrow fat lowering BMD estimation by DEXA? --- p.111 / Chapter STUDY 4 --- Are the same changes in perfusion and marrow fat seen in the proximal femur as were seen in the lumbar spine (Study 1 & Study 2)? --- p.128 / Chapter STUDY 5 --- What is the reproducibility of MR perfusion studies and 1H spectroscopy of bone marrow? --- p.150 / Chapter STUDY 6 --- Marrow fat content increases but does the composition of marrow fat change in osteoporosis? --- p.159 / Chapter STUDY 7 --- Likely causes of reduced bone perfusion in osteoporosis: novel findings in an ovariectomy rat model --- p.180 / Chapter STUDY 8 --- Do perfusion indices or marrow fat content predict rate of bone loss? --- p.204 / SUMMARY --- p.222 / REFERENCES --- p.227 / ABBREVIATION LIST --- p.251 / APPENDIX --- p.253

Chan Chun Wai. / "August 2004." / Thesis (Ph.D.)--Chinese University of Hong Kong, 2004. / Includes bibliographical references. / 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 Company, [200-] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Mode of access: World Wide Web. / Abstracts in English and Chinese.

Bone metastases are a major cause of morbidity in patients of advanced breast cancer. Development of osteolytic bone metastasis depends on the interaction between malignant tumour cells and bone microenvironment. Receptor Activator of Nuclear Factor Kappa B (RANK) is a member of tumour necrosis factor (TNF) superfamily that is expressed by osteoclasts (the bone resorbing cells) and primary breast tumour cells. Previous studies demonstrated that RANK receptor and its ligand (RANKL) play an important role in bone remodelling, mammary gland development and immune system. RANKL was also found to serve as a chemotactic factor that facilitates breast tumour metastasis to bone. However, the role of the RANK receptor in breast cancer cell metastatic behaviour in bone is not fully understood. Therefore, the aim of this thesis was to explore the role of the RANK receptor in parental and bone-tropic breast cancer cell growth, motility and invasion, and assess these cells influence on breast cancer cell induced osteoclastogenesis. Functional studies in breast cancer cells showed that RANKL (100 - 300 ng/ ml) significantly enhanced parental human MDA-231 (MDA-231P) and mouse 4T1 breast cancer cell spreading within minutes. RANKL induced chemotactic cell migration of MDA-231P cells in vitro. I also found that RANKL significantly stimulated random and directional 2D and 3D cell migration of parental MDA-231P and bone-tropic (MDA-231BT) breast cancer cells in vitro. These effects were observed at concentrations (100 – 300 ng/ml) that were sufficient to induce osteoclast formation in the presence and absence of breast cancer cells in vitro. In contrast, high concentrations of RANKL (1000 ng/ ml) dramatically suppressed human MDA-231P breast cancer cell invasion in vitro. These data indicate that the RANK receptor in the breast cancer cell lines tested influences cancer cell spreading, migration and invasion in vitro. Thus, targeting RANK in tumour cells may be of value in the prevention of tumour burden associated with breast cancer bone metastasis. Mechanistic studies revealed that RANKL stimulated the phosphorylation of p38 kinase in human and mouse breast cancer cells. Interestingly, RANKL had no effect on NFᴋB, JNK and AKT pathways in parental human MDA-231 and mouse 4T1 breast cancer cells at concentrations up to 300 ng/ ml. These data implies that the RANK receptor modulates human and mouse breast cancer cell metastatic behaviour via p38 activation and independently of the NFᴋB and PI3K/AKT pathways. Silencing of the RANK receptor in the bone-tropic human breast cancer cells MDA- 231BT2 reduced directional cell migration without affecting cell viability and growth. Functional studies in osteoclast and breast cancer cell revealed that knockdown of RANK expression in both parental and bone-tropic human breast cancer cells significantly inhibited the ability of these cells to stimulate osteoclast formation. Although, I cannot exclude the possibility of the involvement of other signalling pathways downstream of the RANK receptor, these studies suggest that the RANK/P38 signalling in osteoclast and breast cancer cells contributes significantly to breast cancer cell behaviour in bone. Genetic analysis of the RANK gene in human parental and bone-tropic MDA-231 breast cancer cells showed a number of polymorphisms. One variant detected was found to be deleterious for the RANK protein. This variant changes the amino acid sequence from alanine to threonine (Ala ˃ Thr) and only appeared in the RANK gene in the parental human MDA-231P breast cancer cells. Moreover, of the four known RANK isoforms that were detected in the parental and bone-tropic breast cancer cells tested, two lacked the TRAF6 binding motifs associated with NFκB activation. All RANK isoforms detected on the bone-tropic MDA-231BT breast cancer cells expressed the P38 binding motifs. Altogether, these findings support the role of the RANK/P38 signalling pathway in breast cancer cell behaviour in bone. Epigenetic analysis in parental human MDA-231P breast cancer cells showed that continuous and long-term exposure to RANKL (10 and 100 ng/ ml) for up to 50 passages (approximately 120 days) did not induce epigenetic changes, particularly DNA methylation, in the RANK gene. However, I found DNA methylation changes in a set of genes that are known to be involved in cell development and regulation. The methylation status of the altered CpG loci either hypermethylated or hypomethylated are located at different parts in the CpG islands. Whole genome DNA methylation pattern of the bone-tropic breast cancer cells showed a number of genes that appeared in both bone-tropic variants are correlated with different biological function of the cells. I also found that long-term exposure of human MDA-231P to RANKL (100 ng/ ml) enhanced the ability of these cells to stimulate osteoclastogenesis in vitro. These data together indicate that long-term exposure to RANKL induces “boney” epigenetic changes in a set of genes that enhances breast cancer cell behaviour in bone. Overall, this thesis illustrated that the RANK receptor on human parental and bone-tropic breast cancer cells plays an important role in cell motility and ability of these cells to influence osteoclastogenesis and ultimately osteolysis. Therefore, agents that selectively target the RANK receptor may be of value in the treatment of both tumour burden and osteolytic bone disease associated with breast cancer. However, the role of the RANK receptor in bone metastasis will require further in vivo investigation.

Orientador: Jorge Kennety Silva Formiga / Coorientadora: Vivian Silveira dos Santos Silva Bardini / Banca : João Maurício Ferraz da Silva / Banca: Denilson Paulo Souza dos Santos / Resumo: Os modelos matemáticos utilizados atualmente para análise variação da densidade óssea quando submetido a estímulos mecânicos, consideram apenas carregamentos estáticos ou a variação dentro de um curto espaço de tempo e permanecendo estático novamente. Esse artigo tem o objetivo de desenvolver dois novos modelos que permitam simular o comportamento do tecido ósseo de remodelagem e determinar as faixas de subcarga e sobrecarga onde ocorrem a reabsorção óssea, quando submetido a estímulos mecânicos variados oscilando no tempo. Tradicionalmente os estudos realizados para determinação da variação da densidade óssea, vem utilizando um modelo adotando uma equação diferencial ordinária (EDO) para analisar essa variação. A partir do modelo dessa EDO, foram desenvolvidas duas novas equações matemáticas para simular o comportamento do tecido ósseo quando submetido à estímulos mecânicos variados no tempo. A primeira equação utiliza uma variação da tensão seguindo o padrão de oscilação de uma onda senoidal e a segunda equação utiliza um padrão de onda resultante da combinação linear entre seno e cosseno. A resolução dessa EDO foi feita utilizando o método de Runge-Kutta de 5ª ordem, um integrador de maior precisão e permitindo uma melhor análise do comportamento do tecido ósseo através dos resultados obtidos com maior precisão para diversos níveis de tensão. A análise dos resultados obtidos através das simulações matemáticas empregando três tipos de carregamentos, estático, variável de ac... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract : The mathematical models currently used for bone density variation analysis when subjected to mechanical stimuli, consider only static loading or variation within a short time and remaining static again. This article aims to develop two new models that allow simulating the behavior of bone remodeling tissue and determine the underload and overload ranges where bone resorption occurs when subjected to varying mechanical stimuli oscillating over time. Traditionally, studies performed to determine bone density variation have been using a model using an ordinary differential equation (ODE) to analyze this variation. From the model of this ODE, two new mathematical equations were developed to simulate the behavior of bone tissue when subjected to varying mechanical stimuli over time. The first equation uses a voltage variation following the sine wave oscillation pattern and the second equation uses a wave pattern resulting from the linear combination of sine and cosine. The resolution of this ODE was performed using the 5th order Runge-Kutta method, a more accurate integrator and allowing a better analysis of bone tissue behavior through the results obtained with greater precision for various stress levels. The analysis of the results obtained through mathematical simulations employing three types of loads, static, variable according to the oscillation of a sine wave and variable through the oscillation of a sine-cosine wave, demonstrated that the behavior of bone tissue in relation to the stimuli Mechanics vary by type of loading. The stress levels applied for the three simulations were the same, but the bone remodeling behavior response was different for each type of loading. The resorption tension range, remodeling tension range and overload range change according to the type of loading applied, demonstrating that bone tissue behavior may ....(Complete abstract click electronic access below) / Mestre

The foundation for osteoporosis risk is established during the time periods of childhood, adolescence, and young adulthood, periods of development when bone mass is being accrued rapidly. The relative quantity of bone mass accrued is influenced by both lifestyle and genetic factors. The purpose of this dissertation project was to discover single nucleotide polymorphisms (SNPs) associated with: (1) The rate of hip bone accrual (measured as bone mineral content or BMC) during the adolescent growth spurt, and (2) Total hip bone mass measured as BMC around the age of 19 when the amount of bone accrued is approximately at its peak. Additionally, SNP × longitudinal lifestyle factor (calcium intake per day, vitamin D intake per day, and minutes of moderate to vigorous physical activity (MVPA) per day) multiplicative interaction effects were assessed. Each cohort member’s vector of longitudinal physical activity measurements was summarized as belonging to one of a set of specific trajectory groups using finite mixture modeling. The same was then done for calcium intake and vitamin D intake. The source of the data utilized was the Iowa Bone Development Study (IBDS), which includes genetic and longitudinal bone measurement information. To discover SNPs, a genome-wide association study (GWAS) design was utilized. Females and males were analyzed separately and together. The association between SNPs and the rate of hip bone accrual during the adolescent growth spurt was assessed using linear mixed models controlling for body size, and the association between SNPs and peak hip bone mass was assessed using an ordinary linear regression model, also controlling for body size. Approximately 500,000 SNPs were tested in each GWA analysis; significance was assessed at a familywise error rate of 0.05, the individual test cutoff of which was determined by using SimpleM, a modified Šídák correction. No statistically significant SNPs were detected at the 0.05 familywise error rate threshold established by SimpleM (p < 1.76×10-7); however genes near suggestive SNPs (24 total) were assessed for biological relevance. Of most biological relevance were two suggestive SNPs (rs2051756 and rs2866908, p-values of 1.25×10-6 and 4.28×10-6, respectively) that were detected in an intron of the DKK2 gene through the GWA analysis exploring peak bone mass in females. The DKK2 gene is part of the Wnt signaling pathway and is associated with embryonic development; additionally, it is expressed more highly in osteoarthritic osteoblasts than in normal osteoblasts. No statistically significant results were found from the SNP × lifestyle factor multiplicative interaction effect tests. The potential importance of the DKK2 gene to peak hip bone mass accrual in females should be studied further in order to understand the pathophysiology of this suggested novel association identified during a discovery GWA analysis.