Lau Hoi Yan. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2003. / Includes bibliographical references (leaves 134-157). / Abstracts in English and Chinese. / PUBLICATIONS BASED ON WORK IN THIS THESIS --- p.I / ABSTRACT --- p.II / ACKNOWLEDGEMENTS --- p.VI / TABLE OF CONTENTS --- p.VIII / Chapter CHAPTER 1 --- INTRODUCTION --- p.1 / Chapter 1.1 --- General Introduction --- p.1 / Chapter 1.2 --- Emesis --- p.3 / Chapter 1.2.1 --- Introduction --- p.3 / Chapter 1.2.2 --- Retching & Vomiting --- p.3 / Chapter 1.2.3 --- Nausea --- p.4 / Chapter 1.2.4 --- Motor Components of Emetic Reflex --- p.5 / Chapter 1.2.4.1 --- Pre-ejection Phase --- p.5 / Chapter 1.2.4.2 --- Ejection Phase --- p.5 / Chapter 1.2.4.3 --- Post-ejection Phase --- p.6 / Chapter 1.2.5 --- Components of Emetic Reflex --- p.6 / Chapter 1.2.5.1 --- Area Postrema (AP) --- p.6 / Chapter 1.2.5.2 --- Nucleus Tractus Solitarius (NTS) --- p.7 / Chapter 1.2.5.3 --- Vomiting Centre --- p.8 / Chapter 1.2.5.4 --- Vestibular System --- p.10 / Chapter 1.2.5.5 --- Abdominal Visceral Afferents --- p.10 / Chapter 1.2.5.6 --- Forebrain --- p.11 / Chapter 1.2.6 --- Neurotransmitters & Receptors --- p.12 / Chapter 1.2.7 --- Anti-emetics --- p.13 / Chapter 1.3 --- Models of Nausea --- p.16 / Chapter 1.3.1 --- Introduction --- p.16 / Chapter 1.3.2 --- Conditioned Taste Aversion --- p.18 / Chapter 1.3.3 --- Pica Behaviour --- p.20 / Chapter 1.3.4 --- Studies of the Involvement of Vasopressin --- p.21 / Chapter 1.3.5 --- Tachygastria --- p.24 / Chapter 1.3.6 --- Locomotor Activity --- p.26 / Chapter 1.4 --- Markers of Neuronal Activity --- p.27 / Chapter 1.4.1 --- General Comments --- p.27 / Chapter 1.4.2 --- c-fos Expression as a Marker of Neuronal Activity --- p.28 / Chapter 1.4.2.1 --- What is c-fos? --- p.28 / Chapter 1.4.2.2 --- Regulation of c-fos Expression --- p.30 / Chapter 1.4.2.2.1 --- Calcium Response Element --- p.31 / Chapter 1.4.2.2.2 --- Serum Response Element --- p.32 / Chapter 1.4.2.3 --- Types of Receptors Involved in c-fos Expression --- p.32 / Chapter 1.4.2.4 --- Feasibility of Using c-fos Expression as Marker of Cellular Activity --- p.36 / Chapter 1.4.2.5 --- Identification of Emetic Pathway by c-fos Immunohistochemistry --- p.36 / Chapter 1.5 --- Aims & Objectives --- p.37 / Chapter CHAPTER 2 --- METHODS --- p.42 / Chapter 2.1 --- Animals --- p.42 / Chapter 2.1.1 --- Ferrets --- p.42 / Chapter 2.1.2 --- Suncus murinus --- p.42 / Chapter 2.2 --- Measurement of Animal Behaviour --- p.43 / Chapter 2.2.1 --- Experiment Design --- p.43 / Chapter 2.2.2 --- Recording of Animal Behaviour --- p.43 / Chapter 2.2.3 --- Calibration of Equipment Used to Record Spontaneous Locomotor Activity --- p.44 / Chapter 2.2.4 --- Behaviour Recorded by the Observer --- p.45 / Chapter 2.3 --- Administration of Drugs --- p.46 / Chapter 2.3.1 --- Ferrets --- p.46 / Chapter 2.3.1.1 --- General Comments --- p.46 / Chapter 2.3.1.2 --- Drug Antagonism Studies --- p.47 / Chapter 2.3.2 --- Suncus murinus --- p.47 / Chapter 2.3.2.1 --- General Comments --- p.47 / Chapter 2.3.2.2 --- Dose-Response Studies --- p.48 / Chapter 2.3.2.3 --- Drug Antagonism Studies --- p.48 / Chapter 2.4 --- c-fos Expression Studies in Ferret Brainstems --- p.50 / Chapter 2.4.1 --- Animals and Anaesthesia --- p.50 / Chapter 2.4.2 --- Perfusion and fixation --- p.50 / Chapter 2.4.3 --- Dehydration of brains --- p.51 / Chapter 2.4.4 --- Embedding of tissue --- p.52 / Chapter 2.4.5 --- Sectioning --- p.52 / Chapter 2.4.6 --- Staining --- p.52 / Chapter 2.4.7 --- Antibodies used --- p.55 / Chapter 2.4.8 --- Positive Control Slides --- p.55 / Chapter 2.5 --- Experimental Design and Statistics --- p.56 / Chapter 2.5.1 --- Randomization of Treatments --- p.56 / Chapter 2.5.2 --- Statistics --- p.57 / Chapter 2.5.2.1 --- Ferrets --- p.57 / Chapter 2.5.2.2 --- Suncus murinus --- p.59 / Chapter 2.6 --- Drugs and Chemicals Used --- p.60 / Chapter 2.6.1 --- Drugs Used --- p.60 / Chapter 2.6.2 --- Chemicals Used --- p.62 / Chapter CHAPTER 3 --- RESULTS --- p.63 / Chapter 3.1 --- Ferret --- p.63 / Chapter 3.1.1 --- "The Effect of Ondansetron and CP-99,994 on Emesis and Locomotor Activity Changes Induced by Cisplatin in the Ferret" --- p.63 / Chapter 3.1.2 --- The Effect of Domperidone on Emesis and Locomotor Activity Changes Induced by Apomorphine in the Ferret --- p.69 / Chapter 3.1.3 --- "The Effect of CP-99,994 on Emesis and Locomotor Activity Changes Induced by Apomorphine in the Ferret" --- p.74 / Chapter 3.1.4 --- c-fos Expression Studies in Ferret Brainstems --- p.79 / Chapter 3.1.4.1 --- Cisplatin-treated Ferrets --- p.79 / Chapter 3.1.4.2 --- Positive Control Slides --- p.84 / Chapter 3.2 --- Suncus murinus --- p.88 / Chapter 3.2.1 --- The Emetic Potential of Nicotine and its Effects on the Spontaneous Locomotor Activity of Suncus murinus --- p.88 / Chapter 3.2.2 --- "The Effect of CP-99,994 on Emesis and Locomotor Activity Changes Induced by Nicotine in Suncus murinus" --- p.92 / Chapter 3.2.3 --- The Emetic Potential of Copper Sulphate and its Effects on the Spontaneous Locomotor Activity of Suncus murinus --- p.95 / Chapter 3.2.4 --- "The Effect of CP-99,994 on Emesis and Locomotor Activity Changes Induced by Copper Sulphate in Suncus murinus" --- p.98 / Chapter 3.2.5 --- The Emetic Potential of Cisplatin and its Effects on the Spontaneous Locomotor Activity of Suncus murinus --- p.101 / Chapter 3.2.6 --- The Effect of Ondansetron on Emesis and Locomotor Activity Changes Induced by Cisplatin in Suncus murinus --- p.104 / Chapter 3.2.7 --- "The Effect of CP-99,994 on Emesis and Locomotor Activity Changes Induced by Cisplatin in Suncus murinus" --- p.107 / Chapter 3.2.8 --- "The Effects of Ondansetron and CP-99,994 on Locomotor Activity in Suncus murinus" --- p.110 / Chapter CHAPTER 4 --- DISCUSSION --- p.113 / Chapter CHAPTER 5 --- GENERAL SUMMARY --- p.130 / REFERENCES --- p.134

Choy Wai Yan. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2005. / Includes bibliographical references (leaves 98-122). / Abstracts in English and Chinese. / Thesis committee --- p.i / Statement --- p.ii / Abstract --- p.iii / Acknowledgements --- p.vi / General abbreviations --- p.viii / Abbreviations of chemicals --- p.x / Table of contents --- p.xi / List of figures --- p.xv / List of tables --- p.xviii / Chapter 1 --- Introduction --- p.1 / Chapter 1.1 --- Severe acute respiratory syndrome (SARS) - An overview --- p.1 / Chapter 1.1.1 --- Epidemiology of SARS --- p.1 / Chapter 1.1.2 --- Clinical presentation of SARS --- p.2 / Chapter 1.1.3 --- Diagnostic tests of SARS --- p.5 / Chapter 1.1.4 --- Treatment of SARS --- p.7 / Chapter 1.2 --- Severe acute respiratory syndrome coronavirus (SARS- CoV) --- p.8 / Chapter 1.2.1 --- The etiological agent of SARS --- p.8 / Chapter 1.2.2 --- The coronaviruses --- p.9 / Chapter 1.2.3 --- Genome of SARS-CoV --- p.11 / Chapter 1.3 --- Spike (S) glycoprotein of SARS-CoV --- p.14 / Chapter 1.3.1 --- Functions of SARS-CoV S glycoprotein --- p.15 / Chapter 1.3.2 --- Receptors for S glycoprotein of SARS-CoV --- p.17 / Chapter 1.4 --- 3C-like protease (3CLPro) of SARS-CoV --- p.20 / Chapter 1.4.1 --- Extensive proteolytic processing of SARS-CoV replicase polyproteins --- p.20 / Chapter 1.4.2 --- SARS-CoV 3CLPro --- p.21 / Chapter 1.4.3 --- Substrate specificity of SARS-CoV 3CLPro --- p.22 / Chapter 1.5 --- Combating SARS - Vaccine and drug development --- p.24 / Chapter 1.5.1 --- Vaccine development against SARS --- p.24 / Chapter 1.5.2 --- Drug development against SARS --- p.25 / Chapter 1.6 --- Project objectives of this thesis --- p.27 / Chapter 1.6.1 --- Synthetic Peptide Studies on SARS-CoV S glycoprotein --- p.27 / Chapter 1.6.2 --- Synthetic Peptide Studies on SARS-CoV 3CLPro --- p.28 / Chapter 2 --- Materials and Methods --- p.30 / Chapter 2.1 --- Synthetic peptide studies on SARS-CoV S glycoprotein --- p.30 / Chapter 2.1.1 --- Bioinformatics analyses of SARS-CoV S gly- coprotein --- p.30 / Chapter 2.1.2 --- Peptide design and molecular modeling --- p.32 / Chapter 2.1.3 --- Solid phase peptide synthesis (SPPS) --- p.33 / Chapter 2.1.4 --- Peptide conjugation --- p.35 / Chapter 2.1.5 --- Immunization in rabbits and monkeys --- p.36 / Chapter 2.1.6 --- ELISA analysis --- p.37 / Chapter 2.1.7 --- Immunofluorescent confocal microscopy --- p.39 / Chapter 2.2 --- Synthetic peptide studies on SARS-CoV 3CLpro --- p.40 / Chapter 2.2.1 --- Protein expression and purification --- p.40 / Chapter 2.2.2 --- Solid phase peptide synthesis (SPPS) --- p.41 / Chapter 2.2.3 --- Peptide cleavage assay --- p.44 / Chapter 2.2.4 --- Molecular docking --- p.46 / Chapter 3 --- Results --- p.48 / Chapter 3.1 --- Synthetic peptide studies on SARS-CoV S glycoprotein --- p.48 / Chapter 3.1.1 --- General features and structural analyses of the S glycoprotein --- p.48 / Chapter 3.1.2 --- Peptides design and synthesis --- p.53 / Chapter 3.1.3 --- ELISA analysis and immunofluorescent con- focal microscopy --- p.55 / Chapter 3.2 --- Synthetic peptide studies on SARS-CoV 3CLpro --- p.62 / Chapter 3.2.1 --- Substrate specificity of SARS-CoV 3CLPro . . --- p.62 / Chapter 3.2.2 --- Molecular docking of SARS-CoV 3CLPro and peptide substrates --- p.74 / Chapter 4 --- Discussion --- p.78 / Chapter 4.1 --- Synthetic peptide studies on SARS-CoV S glycoprotein --- p.78 / Chapter 4.1.1 --- Synthetic peptides elicited SARS-CoV specific antibodies --- p.78 / Chapter 4.1.2 --- Factors affecting the specificity and antigenic- ity of synthetic peptides --- p.80 / Chapter 4.1.3 --- Next step towards vaccine development --- p.83 / Chapter 4.1.4 --- A synthetic peptide-based approach --- p.84 / Chapter 4.2 --- Synthetic peptide studies on SARS-CoV 3CLpro --- p.86 / Chapter 4.2.1 --- A comprehensive overview of the substrate specificity of SARS-CoV 3CLpro --- p.87 / Chapter 4.2.2 --- Sequence comparison between SARS-CoV 3CLpro cleavage sites --- p.90 / Chapter 4.2.3 --- A rapid and high throughput approach to screen protease substrate specificity --- p.94 / Bibliography --- p.98

Wong Chun Lung. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2005. / Includes bibliographical references (leaves 202-225). / Abstracts in English and Chinese. / Thesis Committee --- p.ii / Abstract --- p.iii / 摘要 --- p.vi / Acknowledgements --- p.ix / Table of Contents --- p.x / List of Abbreviations --- p.xv / List of Figures --- p.xix / List of Tables --- p.xx iii / Chapter Chapter 1 --- Introduction / Chapter 1.1 --- "Neurodegeneration, Inflammation and Gliosis" --- p.1 / Chapter 1.2 --- Anatomy of the CNS --- p.5 / Chapter 1.3 --- Astrocytes --- p.6 / Chapter 1.3.1 --- Morphology and Identification of Astrocytes --- p.6 / Chapter 1.3.2 --- Physiological Functions of Astrocytes in the CNS --- p.7 / Chapter 1.3.2.1 --- Induction of Blood-brain Barrier (BBB) --- p.7 / Chapter 1.3.2.2 --- Metabolism of Neurotransmitters --- p.9 / Chapter 1.3.2.3 --- Nursing Role of Astrocytes --- p.9 / Chapter 1.3.2.4 --- Immunological Functions of Astrocytes --- p.10 / Chapter 1.3.3 --- Neonatal Rat Cortical Astrocytes as In Vitro Model --- p.12 / Chapter 1.4 --- Cytokines in Brain Damage --- p.14 / Chapter 1.4.1 --- Lipopolysaccharides (LPS) --- p.16 / Chapter 1.4.2 --- Tumor Necrosis Factor-α (TNF-α) --- p.17 / Chapter 1.4.3 --- Interleukin-1 (IL-1) --- p.19 / Chapter 1.4.4 --- Interleukin-6 (IL-6) --- p.20 / Chapter 1.4.5 --- Interferon-γ (IFN-γ) --- p.21 / Chapter 1.5 --- Cytokines-induced Signaling Cascade --- p.22 / Chapter 1.5.1 --- TNF Receptors --- p.23 / Chapter 1.5.2 --- Ca2+ --- p.25 / Chapter 1.5.3 --- MAPK --- p.26 / Chapter 1.5.4 --- PICA --- p.27 / Chapter 1.5.5 --- NFkB --- p.29 / Chapter 1.6 --- Glucose Metabolism in the Brain and Glucose Transporters --- p.31 / Chapter 1.6.1 --- Glucose Transporters in the Brain --- p.32 / Chapter 1.6.2 --- Glucose Transporters in Brain Damage --- p.34 / Chapter 1.7 --- Ascorbic Acid Metabolism in the Brain --- p.36 / Chapter 1.8 --- Aim and Scope of this Project --- p.39 / Chapter Chapter 2 --- Materials and Methods / Chapter 2.1 --- Materials / Chapter 2.1.1 --- Neonatal Sprawley 一Dawley Rats --- p.43 / Chapter 2.1.2 --- Plain Dulbecco Modified Eagle Medium ´ؤ Formula 12 (pDF12) --- p.43 / Chapter 2.1.3 --- Complete DF-12(cDF12) --- p.43 / Chapter 2.1.4 --- Phosphate Buffered Saline (PBS) --- p.44 / Chapter 2.1.5 --- Hank's Buffer (HSB) --- p.44 / Chapter 2.1.6 --- D/L-Homocysteine Buffer --- p.44 / Chapter 2.1.7 --- "LPS, Cytokines and Pentoxifylline" --- p.45 / Chapter 2.1.8 --- Specific TNF Receptor Agonist: TNF antibodies --- p.45 / Chapter 2.1.9 --- Calcium Modulators --- p.45 / Chapter 2.1.10 --- PKA Modulators --- p.46 / Chapter 2.1.11 --- NFkB Inhibitors --- p.47 / Chapter 2.1.12 --- MAPK Inhibitors --- p.47 / Chapter 2.1.13 --- β-Adrenergic Receptor Modulators --- p.47 / Chapter 2.1.14 --- Reagents for RNA and Protein Isolation --- p.48 / Chapter 2.1.15 --- Reagents for Reverse Transcription-Polymerase Chain Reaction (RT-PCR) --- p.48 / Chapter 2.1.16 --- Reagents for DNA Electrophoresis --- p.49 / Chapter 2.1.17 --- Reagents for Real-time PCR --- p.51 / Chapter 2.1.18 --- Reagents for Western Blotting --- p.51 / Chapter 2.1.19 --- Reagents for MTT Assay --- p.51 / Chapter 2.1.20 --- Reagents for 3H-Thymidine Incorporation Assay --- p.52 / Chapter 2.1.21 --- Reagents for Glucose Uptake Assay --- p.52 / Chapter 2.1.22 --- Reagents for Ascorbic Acid Accumulation Assay --- p.53 / Chapter 2.1.23 --- Reagents for Immunostammg --- p.53 / Chapter 2.1.24 --- Other Chemicals and Reagents --- p.53 / Chapter 2.2 --- Methods / Chapter 2.2.1 --- Preparation of Primary Cultured Rat Astrocytes --- p.55 / Chapter 2.2.2 --- Measuring Cell Viability: MTT Assay --- p.56 / Chapter 2.2.3 --- Measuring Cell Proliferation: 3H Thymidine Incorporation Assay --- p.57 / Chapter 2.2.4 --- Measuring Glucose Uptake: Zero-trans Glucose Uptake Assay --- p.58 / Chapter 2.2.5 --- Measuring Ascorbic Acid Accumulation --- p.60 / Chapter 2.2.6 --- Total Protein Extraction --- p.61 / Chapter 2.2.7 --- Western Blotting --- p.62 / Chapter 2.2.8 --- Immunostaining --- p.64 / Chapter 2.2.9 --- Isolation of RNA --- p.64 / Chapter 2.2.10 --- Measurement of RNA Yield --- p.65 / Chapter 2.2.11 --- RNA Gel Electrophoresis --- p.66 / Chapter 2.2.12 --- Reverse Transcription (RT) --- p.66 / Chapter 2.2.13 --- Polymerase Chain Reaction (PCR) --- p.67 / Chapter 2.2.14 --- Separation of PCR Products by Agarose Gel Electrophoresis --- p.67 / Chapter 2.2.15 --- Quantization of PCR Products and Western Blotting --- p.68 / Chapter 2.2.16 --- Real-time PCR --- p.68 / Chapter Chapter 3 --- Results / Chapter 3.1 --- Role of Calcium Ions (Ca2+) in TNF-α-induced Astrocyte Proliferation --- p.70 / Chapter 3.1.1 --- Effects of Changes of Extracellular Ca2+ on Astrocyte Viability --- p.72 / Chapter 3.1.2 --- Effects of Other Divalent Ions on Astrocyte Viability --- p.74 / Chapter 3.1.3 --- Effects of Changes of Intracellular Ca2+ on Astrocyte Viability --- p.78 / Chapter 3.1.4 --- Role of Ca2+ on TNF-α-mduced Proliferation in Astrocytes --- p.85 / Chapter 3.1.5 --- Role of Other Divalent Ions on tnf-α-mduced Proliferation in Astrocytes --- p.90 / Chapter 3.2 --- Effect of Cytokines on Glucose Uptake in Rat Astrocytes --- p.95 / Chapter 3.2.1 --- Basal level of Glucose Uptake in Astrocytes and Effects of Cytokines on Glucose Uptake in Astrocytes --- p.95 / Chapter 3.2.2 --- Signaling Cascade of LPS- and TNF-α-induced Glucose Uptake in Astrocytes --- p.120 / Chapter (A) --- TNFR Subtypes Mediating TNF-a-induced Glucose Uptake --- p.121 / Chapter (B) --- MAPK --- p.125 / Chapter (C) --- PKA --- p.133 / Chapter (D) --- NFkB --- p.139 / Chapter (E) --- Other Mechanisms / Signalling molecules --- p.150 / Chapter (1) --- Interaction with β-Adrenegic Mechanism / Chapter (2) --- Role of cGMP --- p.154 / Chapter (3) --- Effect of Mg2+ on LPS- / TNF-α- induced Glucose Uptake in Astrocytes --- p.156 / Chapter (4) --- Possible Involvement of IGF-1 System --- p.160 / Chapter 3.2.3 --- Summary --- p.163 / Chapter 3.3 --- Effects of LPS and Cytokines on AA Accumulation in Astrocytes --- p.164 / Chapter Chapter 4 --- Discussion / Chapter 4.1 --- Role of Calcium ions (Ca2+) in TNF-α-induced Astrocyte Proliferation --- p.177 / Chapter 4.1.1 --- Drastic Changes in Extracellular Ca2+ Caused Astrocyte Death --- p.178 / Chapter 4.1.2 --- Extraordinary Role of Ca2+ in Astrocytes Survival --- p.178 / Chapter 4.1.3 --- Elevation of [Ca2+]i Reduced Astrocyte Viability --- p.180 / Chapter 4.1.4 --- Failure of Verapamil to Block TNF-α-induced Astrocyte Proliferation --- p.182 / Chapter 4.2 --- Hypothesis for the Relationship between Cytokines and Energy Metabolism --- p.185 / Chapter 4.2.1 --- Mechanism and Signaling Cascade of the Elevated Glucose Uptake --- p.186 / Chapter 4.2.2 --- Increased Glucose Uptake by Cytokines: Friend or Foe? --- p.191 / Chapter 4.2.3 --- Depletion of AA Pool by LPS --- p.194 / Chapter 4.2.4 --- Possible Bedside Application of the Findings --- p.195 / Chapter 4.3 --- Prospects of This Study and Concluding Remarks --- p.197 / Appendix --- p.201 / References --- p.202

by Lam Fung Chun. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2005. / Includes bibliographical references (leaves 187-202). / Abstracts in English and Chinese. / Abstract --- p.i / Abstract in Chinese --- p.iii / Acknowledgements --- p.v / Table of Contents --- p.vi / List of Abbreviations --- p.xiii / List of Tables --- p.xvii / List of Figures --- p.xviii / Publication --- p.xx / Chapter Chapter 1 --- Introduction --- p.1 / Chapter 1.1 --- Epidemiology of Diabetes Mellitus --- p.1 / Chapter 1.2 --- Definition of Diabetes Mellitus --- p.1 / Chapter 1.3 --- Glucose Homeostasis and Diabetes Mellitus --- p.2 / Chapter 1.4 --- Classification of Diabetes Mellitus --- p.6 / Chapter 1.4.1 --- Type 1 Diabetes Mellitus --- p.6 / Chapter 1.4.2 --- Type 2 Diabetes Mellitus --- p.7 / Chapter 1.4.3 --- Gestational Diabetes Mellitus --- p.8 / Chapter 1.4.4 --- Other specific types --- p.8 / Chapter 1.5 --- Diagnostic Criteria of Diabetes Mellitus --- p.9 / Chapter 1.6 --- Complications of Diabetes Mellitus --- p.11 / Chapter 1.7 --- Pharmacological Treatment of Diabetes --- p.12 / Chapter 1.7.1 --- Treatment for type 1 diabetes mellitus --- p.12 / Chapter 1.7.2 --- Treatment for Type 2 diabetes mellitus --- p.13 / Chapter 1.7.2.1 --- Sulfonylureas --- p.14 / Chapter 1.7.1.2 --- Meglitinides --- p.15 / Chapter 1.7.1.3 --- Biguanides --- p.15 / Chapter 1.7.1.4 --- Thazolidinediones --- p.16 / Chapter 1.7.1.5 --- α-Glucosidase inhibitor --- p.16 / Chapter 1.8 --- Diabetes and Traditional Chinese Medicine --- p.17 / Chapter 1.9 --- Objective of this project --- p.18 / Chapter Chapter 2 --- "Botanical, Preparation and Authentication of Traditional Chinese Herbs" --- p.22 / Chapter 2.1 --- Introduction --- p.22 / Chapter 2.2 --- Herbal Materials --- p.22 / Chapter 2.3 --- Authentication of Herbal Material --- p.30 / Chapter 2.4 --- Extraction Method --- p.32 / Chapter 2.4.1 --- Material and Methods --- p.32 / Chapter 2.4.2 --- Results --- p.32 / Chapter 2.4 --- Discussion --- p.32 / Chapter Chapter 3 --- In vitro Studies on Selected Traditional Chinese Herbs --- p.35 / Chapter 3.1. --- Introduction --- p.35 / Chapter 3.2 --- Hepatic Gluconeogenesis Studies --- p.36 / Chapter 3.2.1 --- Introduction --- p.36 / Chapter 3.2.2 --- Material and Methods --- p.41 / Chapter 3.2.2.1 --- Cell Culture of H4IIE --- p.41 / Chapter 3.2.2.2 --- Glucose Production Assay --- p.42 / Chapter 3.2.2.3 --- Bicinchoninic Acid (BCA) Protein Assay --- p.43 / Chapter 3.2.3 --- Results --- p.44 / Chapter 3.3 --- Intestinal Glucose Absorption Studies --- p.46 / Chapter 3.3.1 --- Introduction --- p.46 / Chapter 3.3.2 --- Material and Methods --- p.48 / Chapter 3.3.2.1 --- Preparation of BBMV --- p.48 / Chapter 3.3.2.1.1 --- Chemicals --- p.48 / Chapter 3.3.2.1.2 --- Method --- p.48 / Chapter 3.3.2.2 --- Preparation of Herbal Extracts --- p.50 / Chapter 3.3.2.3 --- BBMV Glucose Uptake Assay --- p.51 / Chapter 3.3.2.4 --- Bicinchoninic Acid (BCA) Protein Assay --- p.54 / Chapter 3.3.3 --- Results --- p.54 / Chapter 3.4 --- Fibroblast Glucose Uptake Studies --- p.57 / Chapter 3.4.1 --- Introduction --- p.57 / Chapter 3.4.2 --- Material and Methods --- p.58 / Chapter 3.4.2.1 --- Cell Culture of Hs68 --- p.58 / Chapter 3.4.2.2 --- 2-Deoxy-D-glucose Uptake Assay --- p.59 / Chapter 3.4.2.3 --- Bicinchoninic Acid (BCA) Protein Assay --- p.60 / Chapter 3.4.3 --- Results --- p.60 / Chapter 3.5 --- Adipocyte Glucose Uptake Studies --- p.63 / Chapter 3.5.1 --- Introduction --- p.63 / Chapter 3.5.2 --- Material and Methods --- p.65 / Chapter 3.5.2.1 --- Cell Culture of 3T3-L1 --- p.65 / Chapter 3.5.2.2 --- Differentiation of 3T3-L1 --- p.65 / Chapter 3.5.2.3 --- 2-Deoxy-D-glucose Uptake Assay --- p.66 / Chapter 3.5.2.4 --- Bicinchoninic Acid (BCA) Protein Assay --- p.68 / Chapter 3.5.3 --- Results --- p.69 / Chapter 3.6 --- Glucose Transporter Type 4 (GLUT4) Expression Studies --- p.71 / Chapter 3.6.1 --- Introduction --- p.71 / Chapter 3.6.2 --- Material and Methods --- p.48 / Chapter 3.6.2.1 --- Cell Culture of 3T3-L1 --- p.71 / Chapter 3.6.2.2 --- Differentiation of 3T3-L1 --- p.71 / Chapter 3.6.2.3 --- GLUT4 Expression Assay --- p.72 / Chapter 3.6.2.4 --- Preparation of RNA --- p.72 / Chapter 3.6.2.5 --- RT-PCR --- p.73 / Chapter 3.6.2.6 --- PCR Analysis on GLUT4 Expression --- p.74 / Chapter 3.6.2.7 --- Real-time PCR --- p.75 / Chapter 3.6.3 --- Results --- p.77 / Chapter 3.7 --- Discussion --- p.81 / Chapter 3.7.1 --- Discussion of Hepatic Gluconeogenesis Studies --- p.81 / Chapter 3.7.2 --- Discussion of Intestinal Glucose Absorption Studies --- p.82 / Chapter 3.7.3 --- Discussion of Fibroblast Glucose Uptake Studies --- p.83 / Chapter 3.7.4 --- Discussion of Adipocyte Glucose Uptake Studies --- p.84 / Chapter 3.7.5 --- Discussion of Glucose Transporter Type 4 (GLUT4) Expression Studies --- p.86 / Chapter 3.7.6 --- Conclusion --- p.87 / Chapter Chapter 4 --- Purification of Cortex Moutan --- p.90 / Chapter 4.1 --- Introduction --- p.90 / Chapter 4.1.1 --- Phytochemical Studies of Cortex Moutan --- p.90 / Chapter 4.2 --- Organic Extraction of Cortex Moutan --- p.93 / Chapter 4.2.1 --- Extraction Material and Methods --- p.93 / Chapter 4.2.2. --- Results --- p.93 / Chapter 4.3 --- BBMV Glucose Uptake Assay with Cortex Moutan Organic Extract (CM-C and CM-D) --- p.96 / Chapter 4.3.1 --- Material and Methods --- p.48 / Chapter 4.3.2 --- Results --- p.96 / Chapter 4.4 --- Fractionation of CM-C and CM-D --- p.98 / Chapter 4.4.1 --- Material and Methods --- p.98 / Chapter 4.4.1.1 --- Chemicals --- p.98 / Chapter 4.4.1.2 --- Methods --- p.98 / Chapter 4.4.2 --- Results --- p.100 / Chapter 4.5 --- BBMV Glucose Uptake Assay of CM-C and CM-D Sub-fractions --- p.105 / Chapter 4.5.1 --- Results --- p.105 / Chapter 4.6 --- Sulfonylation of CM-D1 --- p.107 / Chapter 4.6.1 --- Material and Methods --- p.107 / Chapter 4.6.1.1 --- Chemicals --- p.107 / Chapter 4.6.1.2 --- Methods --- p.107 / Chapter 4.6.2 --- Structure Elucidation of CM-D1s --- p.108 / Chapter 4.6.2.1 --- 1H-NMR Analysis --- p.108 / Chapter 4.6.3 --- BBMV Glucose Uptake Assay of CM-D1s --- p.108 / Chapter 4.6.4 --- Results --- p.108 / Chapter 4.7 --- "Structural Elucidation of CM-D3, CM-D4 and CM-D5" --- p.112 / Chapter 4.7.1 --- Material and Methods --- p.112 / Chapter 4.7.1.1 --- Mass Spectrometry --- p.112 / Chapter 4.7.1.2 --- 1H-NMR Analysis --- p.112 / Chapter 4.7.2 --- Results --- p.113 / Chapter 4.8 --- "BBMV Glucose Uptake Assay of Acetovallione, CM-D3，CM-D4 and CM-D5" --- p.116 / Chapter 4.8.1 --- Results --- p.116 / Chapter 4.9 --- Synthesis of CM-D3s --- p.118 / Chapter 4.9.1 --- Material and Methods --- p.118 / Chapter 4.9.1.1 --- Chemicals --- p.118 / Chapter 4.9.1.2 --- Methods --- p.118 / Chapter 4.9.2 --- Structure Elucidation of synthesized product --- p.119 / Chapter 4.9.3 --- Results --- p.119 / Chapter 4.10 --- Discussion --- p.121 / Chapter Chapter 5 --- In vivo Studies on Selected Herbs --- p.123 / Chapter 5.1 --- Introduction --- p.123 / Chapter 5.1.1 --- Diabetic Animal Models --- p.123 / Chapter 5.1.2 --- Neonatal Streptozotocin-induced Diabetic Rat Model --- p.125 / Chapter 5.2 --- Oral Glucose Tolerance Test (OGTT) --- p.126 / Chapter 5.2.1 --- Animal --- p.126 / Chapter 5.2.2 --- Rat Induction Material and Methods --- p.126 / Chapter 5.2.3 --- Testing Method for diabetic condition of rats --- p.127 / Chapter 5.3.4 --- Results --- p.128 / Chapter 5.3 --- Basal Glycaemia Test --- p.138 / Chapter 5.3.1 --- Animal --- p.138 / Chapter 5.3.2 --- Rat Induction Material and Methods --- p.138 / Chapter 5.3.3 --- Testing Method --- p.138 / Chapter 5.3.4 --- Results --- p.140 / Chapter 5.4 --- Discussion --- p.143 / Chapter Chapter 6 --- General Discussion --- p.147 / Chapter 6.1 --- Introduction --- p.147 / Chapter 6.2 --- Summary of Research Findings --- p.151 / Chapter 6.3 --- Result Interpretation --- p.152 / Chapter 6.3.1 --- Result Interpretation of In Vitro Studies --- p.152 / Chapter 6.3.2 --- Result Interpretation of Cortex Moutan Purification --- p.154 / Chapter 6.3.3 --- Result Interpretation of In Vivo Studies --- p.157 / Chapter 6.4 --- Limitations and Improvements --- p.161 / Chapter 6.5 --- Future Directions --- p.163 / Chapter 6.6 --- Conclusions --- p.169 / Appendices --- p.170 / References --- p.187

Lui Oi Lan. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2005. / Includes bibliographical references (leaves [216]-240). / Abstracts in English and Chinese. / ACKNOWLEDGEMENTS --- p.i / ABBREVIATIONS --- p.ii / ABSTRACT --- p.vii / 撮要 --- p.x / PUBLICATIONS --- p.xiii / TABLE OF CONTENTS --- p.xiv / Chapter CHAPTER 1: --- GENERAL INTRODUCTION / Chapter 1.1 --- Hematopoiesis and Leukemia --- p.1 / Chapter 1.1.1 --- An Overview on Hematopoietic Development --- p.1 / Chapter 1.1.2 --- Leukemia --- p.8 / Chapter 1.1.2.1 --- General Diagnostic Tests for Leukemia --- p.9 / Chapter 1.1.2.2 --- Classification and Epidemiology of Leukemia --- p.10 / Chapter 1.1.2.3 --- Conventional Approaches to Leukemia Therapy --- p.17 / Chapter 1.1.2.4 --- Novel Approaches to Leukemia Therapy --- p.20 / Chapter 1.2 --- Conjugated Linoleic Acid --- p.23 / Chapter 1.2.1 --- Introduction: Historical Development and Occurrence of Conjugated Linoleic Acid --- p.23 / Chapter 1.2.2 --- Phytochemistry and Metabolism of Conjugated Linoleic Acid --- p.24 / Chapter 1.2.2.1 --- Chemical Structures of Conjugated Linoleic Acid Isomers --- p.24 / Chapter 1.2.2.2 --- Biosynthesis of Conjugated Linoleic Acid --- p.26 / Chapter 1.2.2.3 --- Metabolism of Conjugated Linoleic Acid --- p.30 / Chapter 1.2.2.4 --- Mode of Entry and Tissue Incorporation of Conjugated Linoleic Acid --- p.33 / Chapter 1.2.2.5 --- Toxicology of Conjugated Linoleic Acid --- p.33 / Chapter 1.2.3 --- Physiological Activities of Conjugated Linoleic Acid: Reported Health Benefits --- p.35 / Chapter 1.2.3.1 --- Anti-adipogenesis / Chapter 1.2.3.2 --- Anti-diabetogenesis --- p.36 / Chapter 1.2.3.3 --- Anti-atherosclerosis --- p.38 / Chapter 1.2.3.4 --- Anti-carcinogenesis --- p.39 / Chapter 1.2.3.5 --- Anti-tumor Activity --- p.40 / Chapter 1.2.3.6 --- Effects of Conjugated Linoleic Acid on Lipid Metabolism --- p.44 / Chapter 1.2.3.6.1 --- Actions on Phospholipids by Conjugated Linoleic Acid --- p.45 / Chapter 1.2.3.6.2 --- Conjugated Linoleic Acid as a Ligand for the PPAR System --- p.47 / Chapter 1.2.3.7 --- Immunomodulation --- p.47 / Chapter 1.3 --- Aims and Scopes of This Investigation --- p.50 / Chapter CHAPTER 2: --- MATERIALS AND METHODS / Chapter 2.1 --- Materials / Chapter 2.1.1 --- Animals --- p.52 / Chapter 2.1.2 --- Cell Lines --- p.52 / Chapter 2.1.3 --- "Cell Culture Medium, Buffers and Other Reagents" --- p.52 / Chapter 2.1.4 --- Reagents for 3H-Thymidine Incorporation Assay --- p.54 / Chapter 2.1.5 --- Reagents and Buffers for Flow Cytometry --- p.58 / Chapter 2.1.6 --- Reagents for DNA Extraction --- p.59 / Chapter 2.1.7 --- Cell Death Detection ELISAPLUS Kit --- p.63 / Chapter 2.1.8 --- Reagents for Measuring Caspase Activity --- p.65 / Chapter 2.1.9 --- Reagents for Total RNA Isolation --- p.66 / Chapter 2.1.10 --- Reagents and Buffers for RT-PCR --- p.69 / Chapter 2.1.11 --- Reagents and Buffers for Gel Electrophoresis of Nucleic Acids --- p.74 / Chapter 2.1.12 --- "Reagents, Buffers and Materials for Western Blot Analysis" --- p.75 / Chapter 2.2 --- Methods / Chapter 2.2.1 --- Culture of the Tumor Cell Lines --- p.80 / Chapter 2.2.2 --- "Isolation, Preparation and Culture of Mouse Peritoneal Macrophages" --- p.81 / Chapter 2.2.3 --- Determination of Cell Viability --- p.82 / Chapter 2.2.4 --- Determination of Cell Proliferation by [3H]-TdR Incorporation Assay --- p.83 / Chapter 2.2.5 --- In Vivo Tumorigenicity Study --- p.83 / Chapter 2.2.6 --- Analysis of Cell Cycle Profile / DNA Content by Flow Cytometry --- p.83 / Chapter 2.2.7 --- Measurement of Apoptosis --- p.84 / Chapter 2.2.8 --- Determination of the Mitochondrial Membrane Potential --- p.86 / Chapter 2.2.9 --- Measurement of Caspase Activity --- p.87 / Chapter 2.2.10 --- Study of Intracellular Accumulation of Reactive Oxygen Species (ROS) --- p.88 / Chapter 2.2.11 --- Study of the Scavenging Activity of Antioxidants --- p.88 / Chapter 2.2.12 --- Gene Expression Study --- p.89 / Chapter 2.2.13 --- Protein Expression Study --- p.92 / Chapter 2.2.14 --- Measurement of Cell Differentiation --- p.95 / Chapter 2.2.15 --- Statistical Analysis --- p.98 / Chapter CHAPTER 3: --- STUDIES ON THE ANTI-TUMOR ACTICITY OF CONJUGATED LINOLEIC ACID ON MYELOID LEUKEMIA CELLS / Chapter 3.1 --- Introduction / Chapter 3.2 --- Results --- p.99 / Chapter 3.2.1 --- Anti-proliferative Activity of CLA-mix and CLA Isomers on Various Myeloid Leukemia Cell Lines In Vitro --- p.101 / Chapter 3.2.2 --- Cytotoxic Effect of CLA-mix on the WEHI-3B JCS Cells In Vitro --- p.109 / Chapter 3.2.3 --- Cytotoxic Effect of CLA-mix on Primary Murine Myeloid Cells In Vitro --- p.111 / Chapter 3.2.4 --- Kinetic and Reversibility Studies of the Anti-proliferative Activity of CLA-mix on the WEHI-3B JCS Cells --- p.113 / Chapter 3.2.5 --- Effect of CLA-mix and its isomers on the Cell Cycle Profiles of the WEHI-3B JCS Cells In Vitro --- p.116 / Chapter 3.2.6 --- Effect of CLA-mix and its isomer on the Expression of Cell Cycle-regulatory Genes in the WEHI-3B JCS Cells --- p.123 / Chapter 3.2.7 --- Effect of CLA-mix and its isomer on the In V Tumorigenicity of the WEHI-3B JCS Cells ivo --- p.128 / Chapter 3.3 --- Discussion --- p.131 / Chapter CHAPTER 4: --- STUDIES ON THE APOPTOSIS-INDUCING ACTIVITY OF CONJUGATED LINOLEIC ACID ON MYELOID LEUKEMIA CELLS / Chapter 4.1 --- Introduction --- p.141 / Chapter 4.2 --- Results --- p.141 / Chapter 4.2.1 --- Induction of Apoptosis in Both Murine and Human Myeloid Leukemia Cells by CLA --- p.144 / Chapter 4.2.2 --- Effect of CLA and its Isomer on the Mitochondrial Membrane Potential of the WEHI-3B JCS Cells --- p.151 / Chapter 4.2.3 --- Effect of CLA-mix and its Isomer on the Expression of Apoptosis-regulatory Genes of the Bcl-2 Family in the WEHI-3B JCS Cells --- p.154 / Chapter 4.2.4 --- Effect of CLA-mix and its Isomer on the Expression of Apoptosis-regulatory Proteins in the WEHI-3B JCS Cells --- p.158 / Chapter 4.2.5 --- Effect of CLA-mix and its Isomer on the Induction of Caspase Activity in the WEHI-3B JCS Cells --- p.161 / Chapter 4.2.6 --- Effect of CLA-mix and its Isomer on the Induction of ROS in the WEHI-3B JCS Cells --- p.170 / Chapter 4.2.7 --- Effect of Antioxidants on the Induction of ROS by CLA-mix and its Isomer in the WEHI-3B JCS Cells --- p.173 / Chapter 4.2.8 --- Effect of Antioxidants on the Induction of Apoptosis by CLA-mix and its Isomer in the WEHI-3B JCS Cells --- p.176 / Chapter 4.2 --- Discussion / Chapter CHAPTER 5: --- STUDIES ON THE DIFFERENTIATION-INDUCING ACTIVITY OF CONJUGATED LINOLEIC ACID ON MYELOID LEUKEMIA CELLS / Chapter 5.1 --- Introduction --- p.187 / Chapter 5.2 --- Results --- p.190 / Chapter 5.2.1 --- Morphological Alterations in CLA-mix- and CLA isomer-treated WEHI-3B JCS Cells --- p.190 / Chapter 5.2.2 --- Effects of CLA-mix on the Cell Size and Granularity of WEHI-3B JCS Cells --- p.196 / Chapter 5.2.3 --- Studies of the Surface Phenotypic Changes in the CLA-mix-treated WEHI-3B JCS cells --- p.198 / Chapter 5.2.4 --- Studies on the Induction of Monocytic Serine Esterase (MSE) Activity in the CLA-mix-treated WEHI-3B JCS Cells --- p.200 / Chapter 5.2.5 --- Studies on the Induction of Endocytic Activity in the CLA-mix-treated WEHI-3B JCS Cells --- p.201 / Chapter 5.2.6 --- Studies on the Expression of the Differentiation-regulatory Cytokine Genes in the CLA-mix-treated WEHI-3B JCS Cells --- p.202 / Chapter 5.3 --- Discussion --- p.204 / Chapter CHAPTER 6: --- CONCLUSIONS AND FUTURE PERSPECTIVES REFERENCES --- p.208 / REFERENCES --- p.217

Keung Hoi Yee. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2005. / Includes bibliographical references (leaves 151-174). / Abstracts in English and Chinese. / Acknowledgement --- p.I / Abstract --- p.II / 摘要 --- p.V / Content --- p.VII / List of tables --- p.XIII / List of figures --- p.XIV / Abbreviations --- p.XVII / Chapter Chapter 1 --- General Introduction --- p.1 / Chapter 1.1 --- Biology of Pleurotus tuber-regiun (Ptr) --- p.1 / Chapter 1.1.1 --- Ptr grown in the wild --- p.1 / Chapter 1.1.2 --- Cultivation of Ptr --- p.2 / Chapter 1.2 --- Functional food and pharmaceutical application of Ptr sclerotium --- p.3 / Chapter 1.2.1 --- Traditional food and medicinal uses of Ptr sclerotium --- p.3 / Chapter 1.2.2 --- Nutritional value and chemical composition --- p.4 / Chapter 1.2.3 --- Anti-tumor activity --- p.7 / Chapter 1.2.4 --- Anti-viral activity --- p.8 / Chapter 1.2.5 --- Immunologic function --- p.8 / Chapter 1.2.6 --- Pharmaceutical application --- p.9 / Chapter Chapter 2 --- Toxicological evaluation on Ptr sclerotium --- p.11 / Chapter 2.1 --- Introduction --- p.11 / Chapter 2.1.1 --- Toxicological concern of Ptr sclerotium --- p.11 / Chapter 2.1.2 --- Toxicological study --- p.12 / Chapter 2.1.3 --- Biochemical methods for toxicological evaluation --- p.14 / Chapter 2.1.3.1 --- Serum enzyme activities --- p.15 / Chapter 2.1.3.2 --- Other serum analytes --- p.17 / Chapter 2.1.4 --- Histopathological study --- p.20 / Chapter 2.1.5 --- Acute toxicity --- p.21 / Chapter 2.1.6 --- Sub-acute and sub-chronic toxicity --- p.23 / Chapter 2.1.7 --- Objectives --- p.26 / Chapter 2.2 --- Materials and Methods --- p.27 / Chapter 2.2.1 --- Sample materials and chemicals --- p.27 / Chapter 2.2.2 --- Acute toxicity test --- p.27 / Chapter 2.2.2.1 --- Diet and animals --- p.27 / Chapter 2.2.2.2 --- Experimental design --- p.28 / Chapter 2.2.2.3 --- Calculation of sclerotium intake dose --- p.29 / Chapter 2.2.2.4 --- Biochemical assays --- p.30 / Chapter 2.2.2.5 --- Histopathological examination --- p.31 / Chapter 2.2.3 --- Sub-acute and sub-chronic toxicity tests --- p.32 / Chapter 2.2.3.1 --- Diet Preparation --- p.32 / Chapter 2.2.3.2 --- Experimental design --- p.32 / Chapter 2.2.3.3 --- Biochemical assays --- p.36 / Chapter 2.2.3.4 --- Organ weight --- p.40 / Chapter 2.2.3.5 --- Histopathological examination --- p.41 / Chapter 2.2.4 --- Statistical analyses --- p.41 / Chapter 2.3 --- Results and Discussion --- p.42 / Chapter 2.3.1 --- Acute toxicity test --- p.42 / Chapter 2.3.1.1 --- Food consumption --- p.43 / Chapter 2.3.1.2 --- Serum transaminase activities --- p.44 / Chapter 2.3.1.3 --- Histopathology --- p.45 / Chapter 2.3.1.4 --- NOAEL --- p.45 / Chapter 2.3.2 --- Sub-acute toxicity test --- p.50 / Chapter 2.3.2.1 --- Body weight gain --- p.50 / Chapter 2.3.2.2 --- Biochemical assays --- p.51 / Chapter 2.3.2.3 --- Organ per body weight and histopathology --- p.52 / Chapter 2.3.2.4 --- Effects of Ptr sclerotial diets --- p.53 / Chapter 2.3.3 --- Sub-chronic toxicity test --- p.59 / Chapter 2.3.3.1 --- Food and energy consumption --- p.59 / Chapter 2.3.3.2 --- Biochemical assays --- p.63 / Chapter 2.3.3.3 --- Organ per body weight --- p.67 / Chapter 2.3.3.4 --- Body weight increase --- p.75 / Chapter 2.3.3.5 --- NOAEL --- p.80 / Chapter 2.4 --- Summary --- p.81 / Chapter Chapter 3 --- Hepatoprotection of Ptr sclerotium --- p.82 / Chapter 3.1 --- Introduction --- p.82 / Chapter 3.1.1 --- Hepatotoxicity --- p.82 / Chapter 3.1.2 --- Potential hepatoprotection effect of Ptr sclerotium --- p.83 / Chapter 3.1.3 --- Toxicity of CC14 --- p.85 / Chapter 3.1.4 --- Toxicity of AFB! --- p.89 / Chapter 3.1.5 --- Bioactivity of chlorophyllin --- p.92 / Chapter 3.1.6 --- Comet assay --- p.93 / Chapter 3.1.7 --- Objectives --- p.98 / Chapter 3.2 --- Materials and Methods --- p.99 / Chapter 3.2.1 --- Sample materials and chemicals --- p.99 / Chapter 3.2.2 --- Curative and preventive tests of Ptr sclerotium against CCl4-induced hepatotoxicity --- p.99 / Chapter 3.2.2.1 --- Animal and diets --- p.99 / Chapter 3.2.2.2 --- Dose-response of CCl4 on rat model --- p.100 / Chapter 3.2.2.3 --- Biochemical assays --- p.100 / Chapter 3.2.2.4 --- Curative hepatoprotection test on Ptr --- p.101 / Chapter 3.2.2.5 --- Preventive hepatoprotection test on Ptr --- p.101 / Chapter 3.2.3 --- Preventive tests of Ptr sclerotium against AFB1-induced hepato- and geno-toxicity --- p.103 / Chapter 3.2.3.1 --- Dose-response of AFB1 on rat model --- p.103 / Chapter 3.2.3.2 --- Preventive test of Ptr against AFB1 --- p.103 / Chapter 3.2.3.3 --- Biochemical assays --- p.105 / Chapter 3.2.3.4 --- Histopathological examination --- p.105 / Chapter 3.2.4 --- Comet assay --- p.106 / Chapter 3.2.4.1 --- Reagent preparations --- p.106 / Chapter 3.2.4.2 --- Procedures --- p.107 / Chapter 3.2.5 --- Statistical analyses --- p.110 / Chapter 3.3 --- Results and Discussion --- p.111 / Chapter 3.3.1 --- Curative and preventive tests of Ptr sclerotium against CCl4-induced hepatotoxicity --- p.112 / Chapter 3.3.1.1 --- Dose-response of CCl4 on rat model --- p.112 / Chapter 3.3.1.2 --- Curative test of Ptr sclerotium against CCl4-induced hepatotoxicity --- p.116 / Chapter 3.3.1.3 --- Preventive test of Ptr sclerotium against CCl4-induced hepatotoxicity --- p.121 / Chapter 3.3.2 --- Preventive tests of Ptr sclerotium against AFB1-induced hepato- and geno-toxicity --- p.126 / Chapter 3.3.2.1 --- Dose-response of AFB1 on rat model --- p.126 / Chapter 3.3.2.2 --- Preventive test of Ptr sclerotium against AFB1-induced geno- and hepatotoxicity --- p.134 / Chapter 3.3.2.3 --- CHL versus 30% Ptr sclerotial diet --- p.137 / Chapter 3.3.3 --- A comparison of the hepatotoxicity of CC14 and AFB1 --- p.142 / Chapter 3.4 --- Summary --- p.147 / Chapter Chapter 4 --- Conclusions and future work --- p.148 / References --- p.151 / Related publication --- p.175

肝癌是造成癌症相關死亡的主要原因之一。而常規化療受耐藥性的發展和各種副作用的限制。由於無毒性和鲜明的生物药物能力，從蘑菇提取的代謝物在癌症治療中獲得更多的注意和关注。我們以前的研究已經證明來自平菇香菇多醣蛋白複合物的抗癌作用。本研究的目的是探討一種含有多醣蛋白複合物的秀珍菇（PP）熱水提取物在肝癌細胞中抗癌活性的分子機制。 / 我們的研究結果表明，用PP处理过的肝癌細胞，不僅顯著的显示出降低的體外腫瘤細胞的增殖和侵襲，也增強化療藥物順鉑的藥物敏感性。無論是口服和腹腔注射都顯著抑制移植免疫BALB / c裸小鼠的腫瘤生長。同时，PP也能在體外和體內实验顯著抑制PI3K/Akt信號通路在肝癌細胞。有趣的是，当过表达AKT时，Myr-AKT，PP的這種抑制癌细胞生长的效果有减弱的趋势，同时也反映在PP对癌细胞侵襲抑制的作用上。印跡和酶聯免疫吸附試驗結果表明，在PP处理过的肝癌細胞中，血管內皮生長因子（VEGF）的表達和分泌減少了。此外， rhVEGF的加入减弱了 PP对PI3K/Akt通路和肝癌细胞表型的抑製作用。 / 我們的研究結果表明，PP能在體外和體內试验中抑制肝癌細胞增殖，侵襲和耐藥性，通过抑制分泌血管內皮生長因子誘導PI3K/Akt的信號通路。這項研究表明了PP的潛在治療肝癌的治療意義。 / Liver cancer or hepatocellular carcinoma is one of the leading causes of cancer-related deaths. Conventional chemotherapies are limited by the development of drug resistance and various side effects. Because of its non-toxicity and potent biopharmacological activity, metabolites derived from mushrooms have received more attention in cancer therapy. Our previous studies have demonstrated the anti-cancer effects of polysaccharide-protein complexes derived from the Pleurotus mushrooms. The aim of this study was to investigate the underlying molecular mechanism of the anti-cancer activity of a hot water extract containing a polysaccharide-protein complex isolated from Pleurotus pulmonarius (PP) in liver cancer cells. / Our results indicated that exposure of liver cancer cells to PP not only significantly reduced the in vitro cancer cell proliferation and invasion but also enhanced the drug-sensitivity to the chemotherapeutic drug Cisplatin. Both oral administration and intraperitoneal injection of PP significantly inhibited the tumor growth in xenograft BALB/c nude mice. PP triggered a marked suppression of the PI3K/AKT signaling pathway in liver cancer cells in vitro and in vivo, and overexpression of the constitutively active form of AKT, Myr-AKT, abrogated this effect and the inhibited proliferation and invasion by PP. Both western blot and ELISA results showed that PP-treated liver cancer cells had reduced expression and secretion of vascular endothelial growth factor (VEGF). Addition of recombinant human VEGF attenuated the inhibitory effects of PP on PI3K/AKT pathway and the cancer phenotypes. / Our results demonstrated that PP suppressed the proliferation, invasion, and drug-resistance of liver cancer cells in vitro and in vivo, mediated by the inhibition of autocrine VEGF-induced PI3K/AKT signaling pathway. All these results suggest the potential therapeutic implication of PP in the treatment of human liver cancer. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Xu, Wenwen. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2012. / Includes bibliographical references (leaves 83-99). / Abstracts also in Chinese. / Thesis Committee --- p.i / English Abstract --- p.ii / Chinese Abstract --- p.iv / Acknowledgements --- p.v / List of Tables --- p.vi / List of Figures --- p.vii / Abbreviations --- p.x / Content page --- p.xiv / Chapter Chapter 1 --- Literature Review --- p.1 / Chapter 1.1 --- Mushroom as functional foods --- p.1 / Chapter 1.1.1 --- Introduction of functional food --- p.1 / Chapter 1.1.2 --- Functional food and cancer --- p.1 / Chapter 1.1.3 --- Edible Mushroom as functional food --- p.4 / Chapter 1.1.4 --- Pleurotus pulmonarius and its function --- p.7 / Chapter 1.2 --- Hepatocellular carcinoma --- p.9 / Chapter 1.2.1 --- Liver and hepatocellular carcinoma --- p.9 / Chapter 1.2.2 --- Carcinogenesis of liver cancer --- p.12 / Chapter 1.2.2.1 --- Hallmarks of cancer --- p.12 / Chapter 1.2.2.2 --- Cell cycle --- p.13 / Chapter 1.2.2.3 --- Apoptosis --- p.15 / Chapter 1.2.2.4 --- Angiogenesis --- p.17 / Chapter 1.2.2.5 --- Invasion and metastasis --- p.19 / Chapter 1.2.2.6 --- Drug resistance --- p.21 / Chapter 1.2.3 --- The role of PI3K/AKT pathway --- p.23 / Chapter 1.2.4 --- The role of growth factor Vascular endothelial growth factor (VEGF) in HCC --- p.25 / Chapter 1.3 --- Research objectives --- p.27 / Chapter 1.3.1 --- Hypothesis and objectives --- p.27 / Chapter 1.3.2 --- Experimental design --- p.28 / Chapter Chaper 2 --- Materials and Methods --- p.29 / Chapter 2.1 --- Materials --- p.29 / Chapter 2.1.1 --- Mushroom Pleurotus pulmonarius --- p.29 / Chapter 2.1.2 --- Drugs and cell lines --- p.29 / Chapter 2.1.3 --- Antibodies list --- p.30 / Chapter 2.1.4 --- Animal models --- p.32 / Chapter 2.2 --- Sample preparation and structure investigation --- p.32 / Chapter 2.2.1 --- Polysaccharide extraction from mushroom --- p.32 / Chapter 2.2.2 --- Endotoxin test --- p.32 / Chapter 2.2.3 --- Determination of monosaccharide profile by gas chromatography and mass spectrometry (GC/MS) --- p.33 / Chapter 2.2.3.1 --- Sample preparation for gas chromatography analysis --- p.33 / Chapter 2.2.3.1.1 --- Acid depolymerisation --- p.33 / Chapter 2.2.3.1.2 --- Neutral sugar derivatization --- p.33 / Chapter 2.2.3.1.3 --- External monosaccharide standard preparation --- p.34 / Chapter 2.2.3.2 --- Gas chromatography-mass spectrometry (GC/MS) --- p.34 / Chapter 2.2.4 --- Determination of total sugar by phenol-sulfuric acid method (Dubois, 1956) --- p.36 / Chapter 2.2.5 --- Determination of protein content by Lowry-Folin method (Lowry et al.,1951) --- p.37 / Chapter 2.3 --- Biological assays --- p.38 / Chapter 2.3.1 --- In vitro assays --- p.38 / Chapter 2.3.1.1 --- MTT assay --- p.38 / Chapter 2.3.1.2 --- Colony formation assay --- p.38 / Chapter 2.3.1.3 --- Plasmid transfection --- p.39 / Chapter 2.3.1.4 --- In vitro cell invasion assay --- p.39 / Chapter 2.3.1.5 --- Cell cycle analysis --- p.39 / Chapter 2.3.1.6 --- Western blot analysis --- p.40 / Chapter 2.3.1.7 --- VEGF ELISA Kit --- p.42 / Chapter 2.3.2 --- In vivo assays --- p.43 / Chapter 2.3.2.1 --- Tumor xenograft nude mouse model --- p.43 / Chapter 2.3.2.2 --- Immunohistochemistry --- p.45 / Chapter 2.3.2.3 --- H&Estaining --- p.45 / Chapter 2.3.3 --- Statistical analysis --- p.45 / Chapter Chaper 3 --- Results and discussion --- p.46 / Chapter 3.1 --- The yield and chemical characteristic of PP --- p.46 / Chapter 3.1.1 --- The yield of PP from mushroom Pleurotus pulmonarius --- p.46 / Chapter 3.1.2 --- Total carbohydrate and protein content --- p.47 / Chapter 3.1.3 --- Monosaccharide composition by GC-MS --- p.48 / Chapter 3.2 --- Toxicity of the PP water by Limulus amebocyte lysate (LAL) test --- p.48 / Chapter 3.2.1 --- Limulus amebocyte lysate (LAL) test --- p.48 / Chapter 3.3 --- Effects of PP on the proliferation of liver cancer cell lines --- p.50 / Chapter 3.3.1 --- MTT assay --- p.50 / Chapter 3.3.2 --- Colony-formation assay --- p.51 / Chapter 3.3.3 --- Cytotoxic effects of PP against normal liver cell --- p.52 / Chapter 3.3.4 --- The anti-proliferative effect of PP on other cancer types --- p.53 / Chapter 3.3.5 --- Cell cycle analysis by flow cytometry of PP treated liver cancer cells --- p.54 / Chapter 3.3.6 --- Protein expression by western blot analysis of P treated liver cancer cells --- p.56 / Chapter 3.4 --- Anti-cancer effect of PP on liver cancer cells through inactivation of PI3K/AKT signaling pathway --- p.57 / Chapter 3.4.1 --- Effect of PP on inactivation of PI3K/AKT pathway --- p.57 / Chapter 3.4.2 --- The abrogated inhibitory effect of PP on Huh7 with overexpression of AKT. --- p.59 / Chapter 3.4.3 --- The abrogated inhibitory effect of PP on PI3K/AKT signal pathway with overexpression of the constitutively active form of AKT, Myr-AKT --- p.60 / Chapter 3.5 --- Inhibition of VEGF expression and secretion by PP --- p.62 / Chapter 3.5.1 --- ELISA result of PP on VEGF secretion --- p.62 / Chapter 3.5.2 --- The attenuated inhibitory effect of PP on cell proliferation with addition of rhVEGF --- p.63 / Chapter 3.5.3 --- The attenuated inhibitory effect of PP on PI3K/AKT signal pathway with addition of rhVEGF --- p.64 / Chapter 3.6 --- Effect of PP on enhancing the chemosensitivity of liver cancer cells to Cisplatin --- p.66 / Chapter 3.6.1 --- Synergistic effect of PP with cisplatin (DDP) in liver cancer cells --- p.66 / Chapter 3.6.2 --- The abrogated drug-resistant effect by PP by overexpression of the constitutively active form of AKT, Myr-AKT --- p.67 / Chapter 3.6.3 --- The abrogated drug-resistant effect of PP with addition of rhVEGF --- p.68 / Chapter 3.7 --- The anti-invasive potential of PP on liver cancer cells. --- p.69 / Chapter 3.7.1 --- Boyden chamber assay --- p.69 / Chapter 3.7.2 --- The attenuated anti-invasive effect of PP on liver cancer cells with overexpression of constitutively activated AKT --- p.71 / Chapter 3.7.3 --- The attenuated anti-invasive effect of PP on liver cancer cells with addition of rhVEGF --- p.72 / Chapter 3.8 --- The anti-tumor effect of PP in vivo --- p.73 / Chapter 3.8.1 --- The anti-tumor effect of PP by using tumor xenograft model --- p.73 / Chapter 3.8.2 --- Body weight of nude mice treated with PP --- p.75 / Chapter 3.8.3 --- Harmful effect of PP on nude mice --- p.76 / Chapter 3.8.4 --- Immunohistochemist analysis of mice tumor xenograft treated with PP --- p.77 / Chapter 3.8.5 --- Western blot anaylysis using the tumor tissues harvested from mice xenograftes treated with PP --- p.78 / Chapter Chapter 4 --- Conclusion and future Plan --- p.81 / Reference --- p.83 / Related Publication List --- p.100

Yu, Le. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2009. / Includes bibliographical references (leaves 171-198). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstracts in English and Chinese.

An alternative complementary approach was used to verify the antinociceptive effect of 4 CMMs aqueous extracts in a Drosophila adult model. Drosophila adults were subjected to CMM treatments and then placed on an in-house-designed heating device for noxious heat stimulation. Their behavioral outputs were quantified and expressed as heat avoidance index (AI) for revealing the degree of antinociceptive effect of CMMs. By comparing the AI value of non-CMM treated control group with CMM-treated groups at temperature challenge 32°C, it was found that an AI value of 0.2 was obtained for non-CMM-treated control group whereas CMMs-treated groups showed AI values ranged from 0.33 to 0.4. The increase of AI value in those CMM-treated groups means that Drosophila adults became more susceptible to noxious heat stimulation. This indicates that those identified CMMs by the larvae model possess strong and versatile antinoceiceptive activities in Drosophila adults. / In addition, reverse transcription PCR (RT PCR) analysis was performed to study the effects of CMMs on the mRNA expression of three nociceptive-related genes painless, nompC and CG4536. These three genes all belong to the Transient Receptor Potential (TRP) families and have been shown to be involved in heat response. The results indicate that the gene expression level for nompC was significantly down-regulated with fold changes ranging from 0.2 to 0.7 upon 2 hrs treatment of three aqueous CMM extracts Citrus aurantium, Angelica dahurica and Vitex trifolia. However, there is no significant difference in gene expression level for painless and CG4536. / In this study, it has been demonstrated that Drosophila are feasible to use for screening CMMs with antinociceptive activity. While the data of the relative gene expression level for those target genes observed in this study may also serve as biomarkers for providing more evidence to investigate drugs have antinociceptive effects. In the future, such information paves the way for further development in the study of antinociceptive drugs. / Nociception is the reception of signals in the central nervous system (CNS) triggered by specialized sensory receptors which received stimuli such as electrical, thermal, mechanical, or chemical and response to escape from danger. Similar to humans, the fruitfly Drosophila display evolutionarily conserved nociceptive response that makes it suitable for in vivo nociceptive study. In this study, Drosophila larvae were used as initial screening model to investigate the antinociceptive effect that was caused by 61 randomly selected Chinese Medicinal Materials (CMMs). Upon noxious heat stimulation, 73% of larvae in the control group produced a stereotypical rolling behavior within 1 s. Among those tested CMMs, the results indicated that 4 aqueous CMMs extracts from Citrus aurantium L. (family: Rutaceae), Angelica dahurica (Fish. ex Hoffm.) Benth. et Hook (family: Umbelliferae), Vitex trifolia L. var. simplicifolia Cham. (family: Verbenaceae) and Panax notoginseng (Burk.) F. H. Chen (family: Araliaceae) were found to have strong antinociceptive effect on Drosophila larvae since less than 40% of the larvae have produced stereotypical rolling behavior within 1 s upon noxious heat stimulation. / "September 2007." / Advisers: Ming Liang Song; Ho Yin Edwin Chan. / Source: Dissertation Abstracts International, Volume: 69-08, Section: B, page: 4768. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2007. / Includes bibliographical references (p. 134-139). / 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.

Dysmenorrhoea, defined as cramping pain in the lower abdomen occurring during menstruation, is known to affect up to 90% of women of childbearing age to varying degrees. The underlying causes of this condition are believed to be due to a number of factors, but are mainly attributed to increased myometrial activity, increased prostaglandin production and hormonal influences. Although there are pharmaceutical treatments available, they mainly concentrate on symptomatic relief, with the main treatment being the use of non-steroidal anti-inflammatory drugs (NSAIDs) to directly relieve the pain. Other treatments include the use of the combined oral contraceptives which are believed to influence myometrial contractility via regulation of hormonal activity. However due to the gastric and contraceptive side effects of these treatments respectively, other alternative treatments are becoming increasingly popular. One such treatment is the use of Bak foong pills (BFP), a traditional Chinese medicine used in China for the treatment of various gynaecological disorders including primary dysmenorrhoea. The aims of the current project were therefore to highlight the major beneficial effects of BFP and attempt to elucidate its major mechanisms of action in treating dysmenorrhoea. / The study demonstrated that BFP's anti-dysmenorrhoeal properties were due to a combination of hormonal, myometrial relaxant and analgesic effects. Treatment of mice with BFP caused an estrogen-like effects as demonstrated with increased cystic fibrosis transmembrane conductance regulator (CFTR) mRNA expression. Furthermore, serum estrogen and progesterone levels were also elevated in BFP treated rats. BFP was also able to significantly reduce myometrial contractions, indicating that BFP's anti-dysmenorrhoeal effect may be aided by reduced contractility of the myometrium following treatment. The uterine relaxation caused by BFP was not dependant on increases in nitric oxide or cAMP, but appeared to affect calcium mobilization. Investigation of the analgesic effect of BFP, assessed using a visceral pain model in mice showed that following sub-chronic (72 hour) treatment with BFP, there was a significant reduction in pain response, demonstrating that BFP had direct analgesic effect. (Abstract shortened by UMI.) / Rowlands Dewi Kenneth. / "July 2005." / Adviser: Hsiao Chang Chan. / Source: Dissertation Abstracts International, Volume: 67-07, Section: B, page: 3533. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2005. / Includes bibliographical references (p. 148-165). / 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. / School code: 1307.