Cheuk, Yau Chuk. / Thesis submitted in: Dec 2008. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2009. / Includes bibliographical references (leaves 132-144). / Abstracts in English and Chinese. / ABSTRACT --- p.i / 論文摘要 --- p.iii / PUBLICATIONS --- p.v / ACKNOWLEDGEMENT --- p.vi / LIST OF ABBREBIVIATIONS --- p.vii / INDEX FOR FIGURES --- p.x / INDEX FOR TABLES --- p.xiv / TABLE OF CONTENTS --- p.xv / Chapter CHAPTER ONE - --- INTRODUCTION / Chapter 1.1 --- "Joint function, structure and biochemistry" / Chapter 1.1.1 --- Function of joint --- p.1 / Chapter 1.1.2 --- Types of cartilage --- p.1 / Chapter 1.1.3 --- Composition and structure of articular cartilage --- p.2 / Chapter 1.1.4 --- The subchondral bone --- p.3 / Chapter 1.1.5 --- Maturation of articular cartilage and subchondral bone --- p.3 / Chapter 1.2 --- Osteochondral defect / Chapter 1.2.1 --- Clinical problem --- p.6 / Chapter 1.2.2 --- Spontaneous repair --- p.7 / Chapter 1.2.3 --- Current treatment strategies --- p.7 / Chapter 1.2.4 --- Limitations of current treatment strategies --- p.8 / Chapter 1.2.5 --- Treatments under development --- p.11 / Chapter 1.2.6 --- Potential and limitations in cell therapies --- p.14 / Chapter 1.3 --- The 3-D scaffold-free cartilage / Chapter 1.3.1 --- Fabrication of scaffold-free cartilage --- p.16 / Chapter 1.3.2 --- Scaffold-free cartilage for chondral / osteochondral defect repair --- p.18 / Chapter 1.3.3 --- Scaffold-free bioengineered chondrocyte pellet from our group --- p.20 / Chapter 1.3.4 --- BCP as a possible treatment for OCD --- p.21 / Chapter 1.4 --- The objectives of the study --- p.22 / Chapter 1.5 --- The study plan / Chapter 1.5.1 --- Design of the study --- p.23 / Chapter 1.5.2 --- Choice of animal model --- p.23 / Chapter 1.5.3 --- Selection of evaluation time points --- p.24 / Chapter 1.5.4 --- Choice and modification of histological scoring system --- p.24 / Chapter CHAPTER TWO - --- METHODOLOGY / Chapter 2.1 --- Preparation of reagents and materials for tissue culture and histology --- p.26 / Chapter 2.2 --- Creation of osteochondral defect model --- p.28 / Chapter 2.3 --- Synthesis of scaffold-free cartilage using 3-D chondrocyte pellet culture / Chapter 2.3.1 --- Isolation of rabbit costal chondrocytes --- p.31 / Chapter 2.3.2 --- Three-dimensional chondrocyte pellet culture --- p.31 / Chapter 2.3.3 --- BrdU labeling for cell fate tracing --- p.32 / Chapter 2.4 --- Further characterization of the 3-D scaffold-free chondrocyte pellet / Chapter 2.4.1 --- Gross appearance --- p.35 / Chapter 2.4.2 --- Cell viability / Chapter 2.4.2.1 --- Alamar blue reduction assay --- p.35 / Chapter 2.4.3 --- Preparation of samples for histology --- p.36 / Chapter 2.4.4 --- General morphology and histomorphology / Chapter 2.4.4.1 --- H&E staining --- p.36 / Chapter 2.4.5 --- Cartilage properties / Chapter 2.4.5.1 --- Safranin O /Fast Green staining --- p.37 / Chapter 2.4.5.2 --- Immunohistochemistry of type II collagen --- p.37 / Chapter 2.4.5.3 --- Immunohistochemistry of type I collagen --- p.38 / Chapter 2.4.6 --- Angiogenic properties / Chapter 2.4.6.1 --- Immunohistochemistry of VEGF --- p.40 / Chapter 2.4.7 --- Osteogenic properties / Chapter 2.4.7.1 --- ALP staining --- p.40 / Chapter 2.5 --- Implantation of scaffold-free cartilage into osteochondral defect model / Chapter 2.5.1 --- Surgical procedures --- p.41 / Chapter 2.5.2 --- Experimental groups --- p.42 / Chapter 2.6 --- Assessment of osteochondral defect healing / Chapter 2.6.1 --- Macroscopic evaluation --- p.43 / Chapter 2.6.2 --- Preparation of samples for histology --- p.43 / Chapter 2.6.3 --- Histology for general morphology / Chapter 2.6.3.1 --- H&E staining --- p.45 / Chapter 2.6.4 --- Histological scoring / Chapter 2.6.4.1 --- Modification of the scoring system --- p.45 / Chapter 2.6.4.2 --- Procedures of scoring and validation --- p.45 / Chapter 2.6.5 --- Cell proliferation / Chapter 2.6.5.1 --- Immunohistochemistry of PCNA --- p.49 / Chapter 2.6.6 --- Cartilage regeneration / Chapter 2.6.6.1 --- Safranin O /Fast Green staining --- p.49 / Chapter 2.6.6.2 --- Immunohistochemistry of type II collagen --- p.49 / Chapter 2.6.6.3 --- Immunohistochemistry of type I collagen --- p.50 / Chapter 2.6.6.4 --- Polarized light microscopy --- p.50 / Chapter 2.6.7 --- Expression of angiogenic factor / Chapter 2.6.7.1 --- Immunohistochemistry of VEGF --- p.50 / Chapter 2.6.8 --- Bone regeneration / Chapter 2.6.8.1 --- μCT analysis --- p.50 / Chapter 2.6.9 --- Histomorphometric analysis of cartilage and bone regeneration --- p.53 / Chapter 2.6.10 --- BrdU detection for cell fate tracing --- p.55 / Chapter 2.6.11 --- Statistical analysis --- p.55 / Chapter CHAPTER THREE - --- RESULTS / Chapter 3.1 --- Further characterization of the 3-D chondrocyte pellet culture / Chapter 3.1.1 --- Gross examination --- p.57 / Chapter 3.1.2 --- Cell viability --- p.57 / Chapter 3.1.3 --- Cartilage properties --- p.61 / Chapter 3.1.4 --- Angiogenic properties --- p.63 / Chapter 3.1.5 --- Osteogenic properties --- p.64 / Chapter 3.2 --- Implantation of scaffold-free cartilage and assessment / Chapter 3.2.1 --- Gross examination --- p.65 / Chapter 3.2.2 --- General morphology --- p.67 / Chapter 3.2.3 --- Histological scores --- p.71 / Chapter 3.2.4 --- Cell proliferation --- p.75 / Chapter 3.2.5 --- Cartilage regeneration --- p.78 / Chapter 3.2.6 --- Expression of angiogenic factor --- p.90 / Chapter 3.2.7 --- Bone regeneration --- p.93 / Chapter 3.2.8 --- Histomorphometric analysis on cartilage and bone regeneration --- p.96 / Chapter 3.2.9 --- Cell fate tracing --- p.100 / Chapter CHAPTER FOUR - --- DISCUSSION / Chapter 4.1 --- Summary of key findings / Chapter 4.1.1 --- Further characterization of BCP and determination of implantation time --- p.102 / Chapter 4.1.2 --- Implantation of BCP in OCD --- p.102 / Chapter 4.2 --- Spontaneous healing in osteochondral defect / Chapter 4.2.1 --- Findings from the current study --- p.104 / Chapter 4.2.2 --- Comparison with other studies --- p.104 / Chapter 4.2.3 --- Factors affecting spontaneous healing --- p.105 / Chapter 4.3 --- Fabrication and further characterization of the 3-D chondrocyte pellet / Chapter 4.3.1 --- Comparison of different methods of producing scaffold-free cartilage construct --- p.106 / Chapter 4.3.2 --- Cartilage phenotype of the BCP --- p.107 / Chapter 4.3.3 --- Angiogenic and osteogenic potential of the BCP --- p.108 / Chapter 4.3.4 --- Role of mechanical stimulation on tissue-engineered cartilage --- p.109 / Chapter 4.4 --- Repair of osteochondral defect with allogeneic scaffold-free cartilage / Chapter 4.4.1 --- Advantages of the current scaffold-free chondrocyte pellet --- p.111 / Chapter 4.4.2 --- Remodeling of BCP after implantation --- p.111 / Chapter 4.4.3 --- Effect of BCP on cartilage repair --- p.112 / Chapter 4.4.4 --- Effect of BCP on bone regeneration / Chapter 4.4.4.1 --- Findings in the present study --- p.113 / Chapter 4.4.4.2 --- Possible reasons of slow bone repair --- p.114 / Chapter 4.4.4.3 --- Effect of BCP on bone region peripheral to defect --- p.115 / Chapter 4.4.5 --- Immunorejection-free properties of the BCP --- p.116 / Chapter 4.4.6 --- Comparison with other animal studies using scaffold-free cartilage --- p.117 / Chapter 4.4.7 --- Possibility of implanting a BCP cultured for shorter or longer period --- p.118 / Chapter 4.4.8 --- Scaffold-free cartilage construct and construct with scaffold for OCD repair --- p.119 / Chapter 4.4.9 --- Chondrocytes and stem cells for OCD repair --- p.120 / Chapter 4.5 --- Limitations of the study / Chapter 4.5.1 --- Animal model --- p.122 / Chapter 4.5.2 --- Histomorphometric analysis --- p.122 / Chapter 4.5.3 --- Lack of quantitative data analysis --- p.122 / Chapter 4.5.4 --- BrdU labeling of cells --- p.123 / Chapter 4.5.5 --- Lack of biomechanical test --- p.123 / Chapter 4.5.6 --- Small sample size --- p.123 / Chapter CHAPTER FIVE - --- CONCLUSION --- p.124 / Chapter CHAPTER SIX - --- FUTURE STUDIES / Chapter 6.1 --- Identification of factors affecting bone repair after OCD treatment --- p.125 / Chapter 6.2 --- Modifications of BCP treatment --- p.125 / Chapter 6.3 --- Alternative cell source --- p.126 / Chapter 6.4 --- Alternative cell tracking methods --- p.126 / Chapter 6.5 --- Inclusion of biomechanical test --- p.126 / APPENDICES / Appendix 1. Conference paper 1 --- p.129 / Appendix 2: Conference paper 2 --- p.130 / Appendix 3: Animal experimentation ethics approval --- p.131 / BIBLIOGRAPHY --- p.132

Chan, Lai Shan. / Thesis submitted in: Dec. 2008. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2009. / Includes bibliographical references (leaves 148-152). / Abstracts in English and Chinese. / ACKNOWLEDGEMENT --- p.I / PUBLICATIONS --- p.II / ABBREVIATION --- p.III-IV / INDEX OF FIGURES --- p.V-IX / INDEX OF TABLES --- p.X / ABSTRACT (English) --- p.XI-XIII / ABSTRACT (Chinese) --- p.XIV / Chapter CHAPTER 1 --- INTRODUCTION --- p.1-22 / Chapter CHAPTER 2 --- METHODOLOGY --- p.23-49 / Chapter CHAPTER 3 --- RESULTS --- p.50-97 / Chapter CHAPTER 4 --- DISCUSSION --- p.98-117 / BIBLIOGRAPHY / APPENDIX

Lee, Yuen Shan. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2007. / Includes bibliographical references (leaves 81-102). / Abstracts in English and Chinese. / Abstract --- p.i / Acknowledgements --- p.iv / Table of contents --- p.vi / List of Figures --- p.ix / List of Abbreviations --- p.xi / Chapter Chapter 1. --- General introduction --- p.1 / Chapter Chapter 2. --- Literature review / Chapter 2.1 --- History of food allergy research --- p.3 / Chapter 2.2 --- Prevalence of food allergy --- p.4 / Chapter 2.3 --- Clinical symptoms of food allergy --- p.6 / Chapter 2.4 --- Mechanism of food allergy --- p.6 / Chapter 2.4.1 --- Properties of food allergens --- p.7 / Chapter 2.4.2 --- Exposures to food allergens in the gastrointestinal tract --- p.8 / Chapter 2.4.3 --- Oral tolerance and its relationship to food allergy --- p.9 / Chapter 2.4.4 --- Cellular mechanism of food allergy --- p.13 / Chapter 2.5 --- Studies on seafood allergies and allergens --- p.17 / Chapter 2.6 --- Use of animal models in the study of food allergy --- p.22 / Chapter 2.6.1 --- Selection of species and strain for developing animal models --- p.22 / Chapter 2.6.2 --- Parameters of sensitization protocol --- p.25 / Chapter 2.6.3 --- Lessons from animal models --- p.27 / Chapter 2.6.3.1 --- Investigations on pathogenesis of food allergy --- p.27 / Chapter 2.6.3.2 --- Studies on development of therapeutic strategies --- p.28 / Chapter Chapter 3. --- Characterization of hypersensitive responses to recombinant shrimp tropomyosin in mice / Chapter 3.1 --- Introduction --- p.30 / Chapter 3.2 --- Materials and Methods / Chapter 3.2.1 --- Preparation of the recombinant shrimp tropomyosin / Chapter 3.2.1.1 --- Expression of the recombinant shrimp tropomyosin --- p.32 / Chapter 3.2.1.2 --- Extraction and purification of the recombinant protein under native condition --- p.32 / Chapter 3.2.1.3 --- Sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) --- p.33 / Chapter 3.2.1.4 --- Quantification of the recombinant protein and detection of level of endotoxin in the protein --- p.34 / Chapter 3.2.2 --- Characterization of hypersensitive responsesin mice / Chapter 3.2.2.1 --- Mice --- p.37 / Chapter 3.2.2.2 --- Sensitization and challenge of mice --- p.37 / Chapter 3.2.2.3 --- Assessment of systemic anaphylaxis responses --- p.38 / Chapter 3.2.2.4 --- Detection of shrimp tropomyosin specific IgE level --- p.39 / Chapter 3.2.2.5 --- Passive cutaneous anaphylaxis (PCA) test --- p.40 / Chapter 3.2.2.6 --- In vitro proliferation assay under stimulation of shrimp tropomyosin --- p.40 / Chapter 3.2.2.7 --- Cytokine profile of splenocytes --- p.42 / Chapter 3.2.2.8 --- Histological examination of small intestine --- p.44 / Chapter 3.2.2.9 --- Statistical analysis --- p.45 / Chapter 3.3 --- Results / Chapter 3.3.1 --- Preparation of the recombinant shrimp tropomyosin --- p.47 / Chapter 3.3.2 --- Induction of systemic anaphylaxis responses after challenge --- p.48 / Chapter 3.3.3 --- Elevated level of shrimp tropomyosin specific IgE --- p.49 / Chapter 3.3.4 --- Passive cutaneous anaphylaxis (PCA) reactions --- p.50 / Chapter 3.3.5 --- Proliferation response of splenocytes under in vitro stimulation --- p.54 / Chapter 3.3.6 --- Cytokine profiles of restimulated splenocytes --- p.58 / Chapter 3.3.7 --- Histology of small intestine --- p.65 / Chapter 3.4 --- Discussion --- p.68 / Chapter Chapter 4. --- General conclusion --- p.78 / References --- p.81

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Thesis (PH.D. (Medical Sciences)) --University of Limpopo, 2010 / Refere to document

Abstract not available

Abstract not available

Pancreatic cancer kills over 1700 people each year in Australia. In 2000, there were 1908 new cases diagnosed and it remains one of the least treatable malignancies. In the USA, it was the fourth leading cause of cancer death in 2004, with 31,860 new cases and 31,270 recorded deaths. Photodynamic therapy (PDT) is a novel, potentially useful treatment for locally advanced pancreatic cancer with only limited research and clinical work addressing this until now. PDT induces non-thermal, cytotoxic and ischaemic injury to a targeted volume of tissue. During PDT, a photosensitizer is activated by non-thermal light in the presence of oxygen, generating cytotoxic oxygen species and inducing cellular injury and microvascular occlusion. The aim of this thesis was to conduct an animal safety study using two second generation photosensitizers, talaporfin sodium and verteporfin, to assess the risks of pancreatic PDT by looking at injury to organs adjacent to the pancreas and assessing recovery from PDT treatment of the pancreas. The Syrian Golden hamster animal model was used to compare the results of this research to previous work by other authors. The study design incorporated a number of additional experiments, including quantitative tissue fluorescence techniques, plasma level analysis and histopathology techniques. The methods for the animal safety study were similar to the approach used in the clinical setting and provided vital data on the likely risks and side effects of phototherapy in humans. The first study, looking at talaporfin sodium, found likely risks of duodenal injury, gastric injury and death with a limited effect on normal pancreas at photosensitizer doses likely to be employed for pancreatic cancer PDT. The second study, using verteporfin, found similar results with a more potent effect on the normal pancreas at studied drug doses. Both agents had short drug-light intervals, ranging from 15 minutes to 2 hours, reducing the need for pre-treatment hospitalization and short photosensitivity periods of about one to two weeks. Some animals suffered minor cutaneous photosensitivity injuries. A human pancreatic cancer PDT pilot study is feasible and the risks and complications should be acceptable.

Includes bibliographical references (leaves 142-168) Aim was the purification and identification of the early primitive ectoderm-like (EPL) cell induction signals within the medium conditioned by the human hepatocellular carcinoma cell line HepG2 and the localisation of the signals that induce EPL cell and primitive ectoderm formation.

Addenda slip inserted in back. Includes bibliographical references (leaves 178-231). To further the understanding of the molecular mechanisms that contribute to the skeletal pathology of mucopolysaccharidosis type VI and to investigate the production of organic matrix by mucopolysaccharidosis VI osteoblasts