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Law Pui-ki. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2005. / Includes bibliographical references (leaves 98-105). / Abstracts in English and Chinese. / Statement --- p.i / Acknowledgments --- p.ii / Abstract --- p.iii / Table of contents --- p.viii / List of figures --- p.xv / List of tables --- p.xvii / Chapter Chapter 1 --- Introduction --- p.1 / Chapter 1.1 --- Schistosomiasis --- p.1 / Chapter 1.1.1 --- Disease burden --- p.1 / Chapter 1.1.2 --- Causative agents --- p.1 / Chapter 1.1.3 --- Transmission --- p.2 / Chapter 1.1.4 --- Pathology of the disease --- p.3 / Chapter 1.1.5 --- Control and therapy --- p.3 / Chapter 1.2 --- Schistosomiasis in China --- p.5 / Chapter 1.3 --- Schistosoma japonicum --- p.6 / Chapter 1.3.1 --- Life cycle of S. japonicum --- p.6 / Chapter 1.3.2 --- Biology of S. japonicum cercaria --- p.9 / Chapter 1.3.3 --- Transformation of cercaria to schistosomulum --- p.10 / Chapter 1.3.4 --- Cercarial stage-specific antigens --- p.12 / Chapter 1.4 --- Aim of study --- p.14 / Chapter Chapter 2 --- Materials and methodology --- p.15 / Chapter 2.1 --- Materials --- p.15 / Chapter 2.1.1 --- Schistosoma japonicum cercaria cDNA library --- p.15 / Chapter 2.1.2 --- Cercarial stage-specific clones --- p.15 / Chapter 2.1.3 --- Adult worm and cercaria RNA --- p.18 / Chapter 2.1.4 --- "Snail intermediate host, Oncomelania hupensis" --- p.18 / Chapter 2.1.5 --- Bacterial strains --- p.18 / Chapter 2.1.6 --- Chemicals --- p.19 / Chapter 2.1.7 --- Kits and reagents --- p.21 / Chapter 2.1.8 --- Nucleic acids --- p.22 / Chapter 2.1.9 --- Solutions --- p.22 / Chapter 2.1.10 --- Enzymes --- p.24 / Chapter 2.1.11 --- Primers --- p.25 / Chapter 2.1.12 --- Antibodies --- p.26 / Chapter 2.2 --- Methodology --- p.27 / Chapter 2.2.1 --- Identification of cercarial stage-specific genes --- p.27 / Chapter 2.2.1.1 --- Sequence analysis of cercarial stage-specific clones --- p.27 / Chapter 2.2.1.2 --- "Confirmation of stage-specific expression of the selected gene, 20H8 and sjCa8, by RNA dot blot" --- p.29 / Chapter 2.2.1.2.1 --- Cloning of S. japonicum glyceraldehyde-3-phosphate dehydrogenase (sjGAPDH) (GenBank accession no. U75571) --- p.29 / Chapter 2.2.1.2.1.1 --- Reverse transcription and PCR amplification of sjGAPDH --- p.29 / Chapter 2.2.1.2.1.2 --- Cloning of sjGADPH in pBluescript II KS (-) --- p.29 / Chapter 2.2.1.2.1.3 --- Sequence verification of cloned sjGAPDH --- p.30 / Chapter 2.2.1.2.2 --- Synthesis of Digoxigenin (DIG)-labeled probe --- p.31 / Chapter 2.2.1.2.2.1 --- Synthesis of DIG-labeled probe by PCR --- p.31 / Chapter 2.2.1.2.2.2 --- Estimation of concentration of DIG-labeled probe --- p.32 / Chapter 2.2.1.2.3 --- RNA dot blot --- p.33 / Chapter 2.2.1.2.3.1 --- Transferring RNA to the membrane using the BIO-RAD blotting manifold --- p.33 / Chapter 2.2.1.2.3.2 --- Hybridization of DIG-labeled probe to detect sjGAPDH --- p.33 / Chapter 2.2.1.2.3.3 --- Detection of the chemiluminescent signal --- p.33 / Chapter 2.2.1.2.3.4 --- Stripping membrane for reprobing --- p.34 / Chapter 2.2.2 --- Characterization of the cercarial stage-specific gene and gene product of 20H8 --- p.35 / Chapter 2.2.2.1 --- Cloning of full-length cDNA of 20H8 --- p.35 / Chapter 2.2.2.1.1 --- 5'RACE of 20H8 --- p.35 / Chapter 2.2.2.1.2 --- Cloning of full-length 20H8 into pBluescript II SK(-) --- p.36 / Chapter 2.2.2.2 --- Analysis of DNA sequence and deduced amino acid sequence of 20H8 --- p.37 / Chapter 2.2.2.3 --- Demonstration of the immunogenicity and antigenicity of 20H8 --- p.38 / Chapter 2.2.2.3.1 --- Expression of 20H8 in E. coli --- p.38 / Chapter 2.2.2.3.1.1 --- "Cloning of 20H8 in an E coli expression vector, pET32a+" --- p.38 / Chapter 2.2.2.3.1.2 --- Expression of recombinant 20H8 protein in E. coli --- p.39 / Chapter 2.2.2.3.2 --- Purification and concentration of recombinant 20H8 protein --- p.40 / Chapter 2.2.2.3.3 --- Production of antiserum --- p.40 / Chapter 2.2.2.3.4 --- Evaluation of immunogenicity of recombinant 20H8 protein --- p.41 / Chapter 2.2.2.3.5 --- Evaluation of antigenicity of recombinant 20H8 protein --- p.42 / Chapter 2.2.2.4 --- Immunolocalization of 20H8 in cercaria --- p.43 / Chapter 2.2.2.4.1 --- Collection of S. japonicum cercaria --- p.43 / Chapter 2.2.2.4.2 --- Immunofluorescence staining of cercaria --- p.43 / Chapter 2.2.3 --- Characterization of the cercarial stage-specific gene and gene product of sjCa8 --- p.45 / Chapter 2.2.3.1 --- Analysis of DNA sequence and deduced amino acid sequence of sjCa8 --- p.45 / Chapter 2.2.3.2 --- Demonstration of the immunogenicity and antigenicity of sjCa8 --- p.46 / Chapter 2.2.3.2.1 --- Expression of sjCa8 in E. coli --- p.46 / Chapter 2.2.3.2.2 --- Purification and concentration of recombinant sjCa8 protein --- p.46 / Chapter 2.2.3.2.3 --- Production of antiserum --- p.46 / Chapter 2.2.3.2.4 --- Evaluation of the immunogenicity of sjCa8 --- p.47 / Chapter 2.2.3.2.5 --- Evaluation of the antigenicity of sjCa8 --- p.47 / Chapter 2.2.3.3 --- Demonstration of calcium-binding property of sjCa8 --- p.48 / Chapter 2.2.3.3.1 --- Calcium-dependent electrophoretic mobility shift --- p.48 / Chapter 2.2.3.3.2 --- Ruthenium red assay --- p.48 / Chapter 2.2.3.4 --- Immunolocalization of sjCa8 in cercaria --- p.49 / Chapter Chapter 3 --- Results --- p.50 / Chapter 3.1. --- Identification of cercarial stage-specific genes --- p.50 / Chapter 3.1.1 --- Identification of cercarial stage-specific genes by microarray --- p.50 / Chapter 3.1.2 --- "Confirmation of stage-specific expression of the selected gene, 20H8 and sjCa8, by RNA dot blot" --- p.54 / Chapter 3.2. --- Characterization of the cercarial stage-specific gene and gene product of 20H8 --- p.57 / Chapter 3.2.1 --- Cloning of full-length cDNA of 20H8 --- p.57 / Chapter 3.2.2 --- Analysis of DNA sequence and deduced amino acid sequence of 20H8 --- p.59 / Chapter 3.2.3 --- Demonstration of the immunogenicity and antigenicity of 20H8 --- p.62 / Chapter 3.2.3.1 --- Expression of 20H8 in E. coli --- p.62 / Chapter 3.2.3.2 --- Purification and concentration of recombinant 20H8 protein --- p.64 / Chapter 3.2.3.3 --- Production of antiserum --- p.65 / Chapter 3.2.3.4 --- Evaluation of immunogenicity of recombinant 20H8 protein --- p.66 / Chapter 3.2.3.5 --- Evaluation of antigenicity of recombinant 20H8 protein --- p.67 / Chapter 3.2.4 --- Immunolocalization of 20H8 in cercaria --- p.68 / Chapter 3.3. --- Characterization of the cercarial stage-specific gene and gene product of sjCa8 --- p.70 / Chapter 3.3.1 --- Analysis of DNA sequence and deduced amino acid sequence of sjCa8 --- p.70 / Chapter 3.3.2 --- Demonstration of the immunogenicity and antigenicity of sjCa8 --- p.75 / Chapter 3.3.2.1 --- Expression of sjCa8 in E. coli --- p.75 / Chapter 3.3.2.2 --- Purification and concentration of recombinant sjCa8 protein --- p.76 / Chapter 3.3.2.3 --- Production of anti-sjCa8 serum --- p.77 / Chapter 3.3.2.4 --- Evaluation of immunogenicity of sjCa8 protein --- p.77 / Chapter 3.3.2.5 --- Evaluation of antigenicity of sjCa8 protein --- p.78 / Chapter 3.3.3 --- Demonstration of calcium-binding property of sjCa8 --- p.79 / Chapter 3.3.3.1 --- Electrophoretic motility shift --- p.80 / Chapter 3.3.3.2 --- Ruthenium red binding assay --- p.80 / Chapter 3.3.4 --- Immunolocalization of sjCa8 in cercaria 81 --- p.81 / Chapter Chapter 4 --- Discussion --- p.83 / Chapter 4.1 --- Identification of cercarial stage-specific genes --- p.83 / Chapter 4.1.1 --- Identification of cercarial stage-specific genes by microarray --- p.83 / Chapter 4.1.2 --- "Confirmation of stage-specific expression of the selected genes, 20H8 and sjCa8" --- p.85 / Chapter 4.2 --- Characterization of the cercarial stage-specific gene and gene product of 20H8 --- p.86 / Chapter 4.2.2 --- Analysis of DNA sequence and deduced amino acid sequence of 20H8 --- p.86 / Chapter 4.2.3 --- Demonstration of the immunogenicity and antigenicity of 20H8 --- p.88 / Chapter 4.2.4 --- Immunolocalization of 20H8 in cercaria --- p.90 / Chapter 4.3 --- Characterization of the cercarial stage-specific gene and gene product of sjCa8 --- p.91 / Chapter 4.3.1 --- Analysis of DNA sequence and deduced amino acid sequence of sjCa8 --- p.91 / Chapter 4.3.2 --- Demonstration of the immunogenicity and antigenicity of sjCa8 --- p.93 / Chapter 4.3.3 --- Demonstration of calcium-binding property of sjCa8 --- p.94 / Chapter 4.3.4 --- Immunolocalization of sjCa8 in cercaria --- p.94 / Chapter 4.4 --- Conclusions --- p.95 / References --- p.96

Xie Tong. / "May 2004." / Thesis (Ph.D.)--Chinese University of Hong Kong, 2004. / Includes bibliographical references (p. 113-126). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Mode of access: World Wide Web. / Abstracts in English and Chinese.

by Li Ho Kin. / Thesis (M.Phil.)--Chinese University of Hong Kong, 1997. / Includes bibliographical references (leaves 213-245). / ACKNOWLEDGEMENTS --- p.i / ABBREVIATIONS --- p.iii / ABSTRACT --- p.viii / TABLE OF CONTENTS --- p.xi / Chapter CHAPTER 1: --- GENERAL INTRODUCTION --- p.1 / Chapter 1.1 --- Introduction --- p.2 / Chapter 1.2 --- Effector Cells Mediating Anti-Tumour Immunity --- p.5 / Chapter 1.2.1 --- Cytotoxic T Lymphocytes --- p.6 / Chapter 1.2.2 --- Natural Killer Cells --- p.7 / Chapter 1.2.3 --- Lymphokine-Activated Killer Cells --- p.9 / Chapter 1.2.4 --- Macrophages --- p.10 / Chapter 1.2.5 --- Tumour-Infiltrating Lymphocytes --- p.12 / Chapter 1.3 --- Current Modalities for Cancer Treatment --- p.13 / Chapter 1.3.1 --- Surgery --- p.13 / Chapter 1.3.2 --- Radiation Therapy --- p.14 / Chapter 1.3.3 --- Chemotherapy --- p.14 / Chapter 1.3.4 --- Biologic Therapy --- p.15 / Chapter 1.4 --- Biological Response Modifiers as Cancer Therapeutics --- p.17 / Chapter 1.4.1 --- Monoclonal Antibodies --- p.20 / Chapter 1.4.2 --- Differentiating Agents --- p.21 / Chapter 1.4.3 --- Cytokines --- p.24 / Chapter 1.4.4 --- Bioactive Plant Polysaccharides --- p.30 / Chapter 1.5 --- Bacterial Polysaccharides as Potential Immunomodulators and Anti-Tumour Agents --- p.36 / Chapter 1.5.1 --- General Properties of Bacterial Polysaccharides --- p.36 / Chapter 1.5.2 --- Structure and Function of Bacterial Polysaccharides --- p.37 / Chapter 1.5.3 --- Immunomodulatory and Anti-Tumour Activities of Bacterial Polysaccharides --- p.39 / Chapter 1.5.4 --- General Properties of Klebsiella K7 Capsular Antigen --- p.41 / Chapter 1.6 --- Aims and Scopes of the Investigation --- p.44 / Chapter CHAPTER 2: --- MATERIALS AND METHODS --- p.46 / Chapter 2.1 --- Materials --- p.47 / Chapter 2.1.1 --- Animals --- p.47 / Chapter 2.1.2 --- Klebsiella pneumoniae Serotype7 --- p.47 / Chapter 2.1.3 --- Agar Medium for Cultivation of Klebsiella pneumoniae Serotype7 --- p.47 / Chapter 2.1.4 --- Cell Lines --- p.47 / Chapter 2.1.5 --- "Buffers, Culture Medium and Other Reagents" --- p.49 / Chapter 2.1.6 --- Antibodies --- p.54 / Chapter 2.1.7 --- Radioisotopes --- p.55 / Chapter 2.1.8 --- Recombinant Cytokines --- p.55 / Chapter 2.1.9 --- Oligonucleotide Primers and Internal Probes --- p.56 / Chapter 2.1.10 --- Reagents and Solutions for Gene Expression Analysis --- p.59 / Chapter 2.2 --- Methods --- p.64 / Chapter 2.2.1 --- "Extraction, Purification and Characterization of Klebsiella K7 Capsular Antigen" --- p.54 / Chapter 2.2.1.1 --- Extraction and Purification of K7 Capsular Antigen --- p.64 / Chapter 2.2.1.2 --- Gel Filtration of K7 Capsular Antigen --- p.65 / Chapter 2.2.1.3 --- Characterization of K7 Capsular Antigen --- p.67 / Chapter 2.2.1.4 --- Determination of the Bio-Toxicity of K7 Capsular Antigen --- p.67 / Chapter 2.2.1.5 --- In Vitro Cytotoxicity of K7 Capsular Antigen on Splenocytes --- p.53 / Chapter 2.2.2 --- Assay for the Hematopoietic and Mitogenic Activities of Klebsiella K7 Capsular Antigen --- p.69 / Chapter 2.2.2.1 --- Isolation and Preparation of Cells --- p.69 / Chapter 2.2.2.2 --- In Vitro Lymphocyte Transformation Assay --- p.70 / Chapter 2.2.2.3 --- In Vitro Assay of Thymocyte Proliferation --- p.70 / Chapter 2.2.2.4 --- Depleting Mouse T Cells by Anti-Thy-1.2 Antibody Plus Complement Treatment --- p.71 / Chapter 2.2.2.5 --- Depleting Mouse B Cells by Anti-mouse B Cell Antibody Plus Complement Treatment --- p.71 / Chapter 2.2.2.6 --- Depleting Macrophages from Spleen Cell Suspension --- p.71 / Chapter 2.2.2.7 --- Flow Cytometric Analysis of Different Cell Populations from Splenocytes --- p.72 / Chapter 2.2.2.8 --- In Vitro Assay of rmIL-3-Stimulated Proliferation of Murine Bone Marrow Cells --- p.73 / Chapter 2.2.2.9 --- In Vitro Cytotoxicity of K7 Capsular Antigen on Bone Marrow Cells --- p.73 / Chapter 2.2.2.10 --- Colony Assay of Murine Bone Marrow Cells --- p.73 / Chapter 2.2.2.11 --- Assay of Differentiation of Murine Bone Marrow Cells --- p.74 / Chapter 2.2.3 --- Assay for Macrophage Activating Activities of Klebsiella K7 Capsular Antigen --- p.75 / Chapter 2.2.3.1 --- Preparation of Murine Peritoneal Exudate Cells (PEC) --- p.75 / Chapter 2.2.3.2 --- Assay of Phagocytic Activity of Peritoneal Macrophages --- p.75 / Chapter 2.2.3.3 --- In Vitro Macrophage-Mediated Cytostatic Activity --- p.76 / Chapter 2.2.3.4 --- Nitric Oxide (NO) Production of Peritoneal Macrophages --- p.76 / Chapter 2.2.3.5 --- In Vivo Migration of Macrophages --- p.77 / Chapter 2.2.3.6 --- Tumour Necrosis Factor (TNF) Production by Peritoneal Macrophages --- p.77 / Chapter 2.2.3.7 --- TNF Bioassay --- p.78 / Chapter 2.2.3.8 --- Gene Expression Analysis in Peritoneal Macrophages --- p.78 / Chapter 2.2.3.8.1 --- Preparation of Cell Lysate --- p.78 / Chapter 2.2.3.8.2 --- RNA Isolation --- p.79 / Chapter 2.2.3.8.3 --- Reverse Transcription of RNA --- p.80 / Chapter 2.2.3.8.4 --- Polymerase Chain Reaction (PCR) --- p.80 / Chapter 2.2.3.8.5 --- Agarose Gel Electrophoresis --- p.82 / Chapter 2.2.3.8.6 --- 3' End Labelling of Oligonucleotide Probes --- p.82 / Chapter 2.2.3.8.7 --- Dot Blot Hybridization --- p.83 / Chapter 2.2.3.8.8 --- DIG Chemiluminescent Detection --- p.83 / Chapter 2.2.4 --- Assay for Anti-Tumour Activities of Klebsiella K7 Capsular Antigen --- p.85 / Chapter 2.2.4.1 --- Assay of Tumour Cell Proliferation --- p.85 / Chapter 2.2.4.2 --- Assay of Anti-Tumour Activity In Vivo --- p.85 / Chapter 2.2.4.3 --- Assay of Lymphokine-Activated Killer Cell Activity --- p.85 / Chapter 2.2.4.4 --- Assay of Natural Killer Cell Activity --- p.87 / Chapter 2.2.4.5 --- Assay for Differentiation-Associated Characteristics of Myeloid Leukemia Cells --- p.88 / Chapter 2.2.4.5.1 --- Determination of the Viability of Myeloid Leukemia JCS cells --- p.88 / Chapter 2.2.4.5.2 --- Assessment of Cell Morphology --- p.88 / Chapter 2.2.4.5.3 --- Surface Antigen Immunophenotyping --- p.88 / Chapter 2.2.4.5.4 --- Assay of Non-specific Esterase Activity --- p.89 / Chapter 2.2.4.5.5 --- Assay of Phagocytosis --- p.90 / Chapter 2.2.5 --- Statistical Analysis --- p.90 / Chapter CHAPTER 3: --- "EXTRACTION, PURIFICATION & CHARACTERIZATION OF KLEBSIELLA K7 CAPSULAR ANTIGEN" --- p.91 / Chapter 3.1 --- Introduction --- p.92 / Chapter 3.2 --- Results --- p.94 / Chapter 3.2.1 --- Extraction and Purification of K7 Capsular Antigen from Klebsiella pneumoniae Serotype7 --- p.94 / Chapter 3.2.2 --- Gel Filtration Chromatography of K7 Capsular Antigen --- p.94 / Chapter 3.2.3 --- Characterization of K7 Capsular Antigen --- p.97 / Chapter 3.2.4 --- Determination of Bio-toxicity and Cellular Toxicity of K7 Capsular Antigen --- p.97 / Chapter 3.3 --- Discussion --- p.101 / Chapter CHAPTER 4: --- HEMATOPOIETIC AND MITOGENIC ACTIVITIES OF KLEBSIELLA K7 CAPSULAR ANTIGEN --- p.103 / Chapter 4.1 --- Introduction --- p.104 / Chapter 4.2 --- Results --- p.107 / Chapter 4.2.1 --- Effect of K7 Capsular Antigen on the In Vitro Proliferation of Murine Bone Marrow Cells --- p.107 / Chapter 4.2.2 --- Effect of K7 Capsular Antigen on rmIL-3-Stimulated Proliferation of Murine Bone Marrow Cells --- p.107 / Chapter 4.2.3 --- Effect of K7 Capsular Antigen on the In Vitro Differentiation of Murine Bone Marrow Cells --- p.112 / Chapter 4.2.4 --- Effect of K7 Capsular Antigen on the In Vitro Murine Bone Marrow Colony Formation --- p.112 / Chapter 4.2.5 --- Effect of K7 Capsular Antigen on the Viability of Splenocytes In Vitro --- p.117 / Chapter 4.2.6 --- In Vitro Mitogenic Effect of K7 Capsular Antigen on Splenocytes and Thymocytes --- p.117 / Chapter 4.2.7 --- Characterization of the Lymphocyte Population(s) Responding to K7 Capsular Antigen --- p.121 / Chapter 4.2.8 --- Effect of Polymyxin B Sulphate on the Mitogenic Activity of K7Capsular Antigen --- p.125 / Chapter 4.2.9 --- In Vivo Mitogenic Activity of K7 Capsular Antigen --- p.125 / Chapter 4.2.10 --- Flow Cytometric Analysis of Splenocytes from K7 Capsular Antigen-Treated Mice --- p.128 / Chapter 4.2.11 --- In Vitro Co-mitogenic Activity of K7 Capsular Antigen --- p.128 / Chapter 4.3 --- Discussion --- p.132 / Chapter CHAPTER 5: --- ACTIVATION OF MACROPHAGES BY KLEBSIELLA K7CAPSULAR ANTIGEN --- p.136 / Chapter 5.1 --- Introduction --- p.137 / Chapter 5.2 --- Results --- p.140 / Chapter 5.2.1 --- Effect of K7 Capsular Antigen on the Phagocytic Activity of Macrophages In Vitro --- p.140 / Chapter 5.2.2 --- Effect of K7 Capsular Antigen on the In Vivo Migration of Macrophages --- p.140 / Chapter 5.2.3 --- Effect of K7 Capsular Antigen on the In Vitro Cytostatic Activity of Picolinic Acid (PLA)-Activated Macrophages --- p.144 / Chapter 5.2.4 --- Effect of K7 Capsular Antigen on Nitric Oxide (NO) Production by Macrophages In Vitro --- p.144 / Chapter 5.2.5 --- Effect of K7 Capsular Antigen on the In Vitro Production of Tumour Necrosis Factor (TNF) by Macrophages --- p.148 / Chapter 5.2.6 --- Effect of K7 Capsular Antigen on the In Vitro Induction of Gene Expression in Macrophages --- p.148 / Chapter 5.3 --- Discussion --- p.166 / Chapter CHAPTER 6: --- ANTI-TUMOUR ACTIVITIES OF KLEBSIELLA K7 CAPSULAR ANTIGEN --- p.171 / Chapter 6.1 --- Introduction --- p.172 / Chapter 6.2 --- Results --- p.174 / Chapter 6.2.1 --- Effect of K7 Capsular Antigen In Vitro Growth of Various Tumour Cell Lines --- p.174 / Chapter 6.2.2 --- Effect of K7 Capsular Antigen on the In Vitro Growth of EAT Cells --- p.174 / Chapter 6.2.3 --- Effect of K7 Capsular Antigen on the Activation of LAK cells --- p.185 / Chapter 6.2.4 --- Effect of K7 Capsular Antigen on the Activation of NK Cells --- p.185 / Chapter 6.2.5 --- Effect of K7 Capsular Antigen on the Induction of Monocytic Differentiation of the Murine Myeloid Leukemia JCS Cells --- p.185 / Chapter 6.2.5.1 --- Effect of K7 Capsular Antigen on the Induction of Morphological Changes of JCS Cells --- p.188 / Chapter 6.2.5.2 --- Effect of K7 Capsular Antigen on the Expression of Macrophage Differentiation Antigen on Leukemia JCS Cells --- p.192 / Chapter 6.2.5.3 --- Effect of K7 Capsular Antigen on the Induction of Non-specific Esterase Activity in Leukemia JCS Cells --- p.192 / Chapter 6.2.5.4 --- Effect of K7 Capsular Antigen on the Stimulation of Phagocytic Activity in Leukemia JCS Cells --- p.192 / Chapter 6.3 --- Discussion --- p.198 / Chapter CHAPTER 7: --- CONCLUSIONS & FUTURE PERSPECTIVES --- p.202 / REFERENCES --- p.213

by Ho Cheong Yip. / Thesis (M.Phil.)--Chinese University of Hong Kong, 1997. / Includes bibliographical references (leaves 152-164). / ACKNOWLEDGMENTS --- p.I / ABBREVIATIONS --- p.II / ABSTRACT --- p.VI / CHAPTER / Chapter 1 --- GENERAL INTRODUCTION --- p.1 / Chapter 1.1 --- Immunomodulators of biological origin --- p.1 / Chapter 1.2 --- Effector cells involved in anti-tumor immunity --- p.3 / Chapter 1.2.1 --- Cytotoxic T lymphocytes --- p.3 / Chapter 1.2.2 --- Macrophages --- p.4 / Chapter 1.2.3 --- Natural killer cells --- p.5 / Chapter 1.2.4 --- Lymphokine-activated killer cells --- p.7 / Chapter 1.2.5 --- Tumor-infiltrating lymphocytes --- p.9 / Chapter 1.3 --- Cytokines involved in immunomodulation and anti-tumor immunity --- p.11 / Chapter 1.3.1 --- Interleukin-1 --- p.11 / Chapter 1.3.2 --- Interleukin-2 --- p.13 / Chapter 1.3.3 --- Interleukin-3 --- p.14 / Chapter 1.3.4 --- Tumor necrosis factor-alpha --- p.15 / Chapter 1.3.5 --- Interferons --- p.18 / Chapter 1.4 --- Carbohydrates used as potential immunopotentiating agents --- p.19 / Chapter 1.5 --- General properties of K1 capsular antigen --- p.20 / Chapter 2 --- AIM AND SCOPE OF THIS DISSERTATION --- p.22 / Chapter 3 --- MATERIALS AND METHODS --- p.24 / Chapter 3.1 --- Materials --- p.24 / Chapter 3.2 --- Methods --- p.33 / Chapter (I) --- "Extraction, purification and characterization of K1 capsular antigen from Klebsiella pneumoniae" / Chapter 3.2.1 --- Extraction and purification of Kl capsular antigen --- p.33 / Chapter 3.2.2 --- Gel filtration of K1 capsular antigen --- p.35 / Chapter 3.2.3 --- Characterization of K1 capsular antigen --- p.35 / Chapter 3.2.3.1 --- Determination of carbohydrate and protein contents --- p.35 / Chapter 3.2.3.2 --- Determination of uronic acid content --- p.35 / Chapter 3.2.4 --- Determination of bio-toxicity of K1 capsular antigen --- p.35 / Chapter 3.2.5 --- Treatment of K1 capsular antigen with sodium periodate --- p.36 / Chapter 3.2.6 --- Treatment of K1 capsular antigen with sodium hydroxide followed by acetic acid neutralization --- p.36 / Chapter (II) --- Isolation and preparation of cells / Chapter 3.2.7 --- Murine splenocytes --- p.37 / Chapter 3.2.8 --- Removal of red blood cells and dead cells using Ficoll-paque gradient method --- p.37 / Chapter 3.2.9 --- Murine thymocytes --- p.37 / Chapter 3.2.10 --- Murine peritoneal exudate cells (PEC) --- p.38 / Chapter 3.2.11 --- Bone marrow cells --- p.38 / Chapter (III) --- Assays of immunomodulatory activities of K1 capsular antigen on lymphocytes / Chapter 3.2.12 --- In vitro mitogenic assay --- p.39 / Chapter 3.2.13 --- In vivo mitogenic assay --- p.39 / Chapter 3.2.14 --- "In vitro co-mitogenic assay of K1 capsular antigen with Polymyxin B sulfate, LPS or Con A" --- p.40 / Chapter 3.2.15 --- In vitro mitogenic effect of K1 capsular antigen on lymphocyte sub-populations --- p.40 / Chapter 3.2.16 --- Assay of murine interleukin-2 --- p.41 / Chapter 3.2.17 --- Assay of murine interleukin-1 --- p.41 / Chapter (IV) --- Assays of immunomodulatory activities of K1 capsular antigen on macrophages and bone marrow cells / Chapter 3.2.18 --- In vivo migration of macrophages --- p.42 / Chapter 3.2.19 --- Assay of interleukin-1 produced from murine macrophages --- p.42 / Chapter 3.2.20 --- Assay of nitric oxide produced from murine macrophages --- p.43 / Chapter 3.2.21 --- Assay of proliferation of murine bone marrow cells --- p.44 / Chapter 3.2.22 --- Assay of differentiation of murine bone marrow cells --- p.44 / Chapter (V) --- Assays of anti-tumor activities of K1 capsular antigen / Chapter 3.2.23 --- In vitro cytostatic effect of K1 capsular antigen on murine tumor cell lines --- p.45 / Chapter 3.2.24 --- In vivo anti-tumor activities of K1 capsular antigen --- p.45 / Chapter 3.2.25 --- In vitro stimulation of TNF-α-like factor release from thioglycollate-elicited murine peritoneal macrophages by K1 capsular antigen --- p.46 / Chapter 3.2.26 --- Effects of K1 capsular antigen on TNF-α production in normal mice in vivo --- p.47 / Chapter 3.2.27 --- Effects of K1 capsular antigen on TNF-α production as well as EAT growth in vivo --- p.48 / Chapter 3.2.28 --- In vitro induction of macrophage-mediated cytostasis on tumor cells --- p.48 / Chapter 3.2.29 --- In vitro induction of macrophage-mediated cytolysis on tumor cells --- p.49 / Chapter 3.2.30 --- In vivo induction of alloreactive cytotoxic T cells --- p.49 / Chapter 3.2.31 --- In vitro induction of lymphokine-activated killer cell activity --- p.50 / Chapter 3.2.32 --- In vivo induction of lymphokine-activated killer cell activity --- p.51 / Chapter 3.2.33 --- In vivo induction of lymphokine-activated killer cell activity in tumor-bearing mice --- p.52 / Chapter 3.2.34 --- In vivo induction of lymphokine-activated killer cell activity in tumor-bearing mice with in vitro culture --- p.52 / Chapter 3.2.35 --- Phenotypic characterization of LAK cells by flow cytometry --- p.53 / Chapter 3.2.36 --- Winn-type tumor-inhibition of LAK cells --- p.54 / Chapter 3.2.37 --- Adoptive transfer of LAK cells to tumor-bearing mice --- p.54 / Chapter 3.2.38 --- In vivo stimulation of tumor-infiltrating lymphocytes --- p.55 / Chapter 3.2.39 --- Statistical analysis --- p.57 / Chapter 4 --- "EXTRACTION, PURIFICATION AND CHARACTERIZATION OF K1 CAPSULAR ANTIGEN FROM KLEBSIELLA PNEUMONIAE" / Introduction --- p.58 / Results --- p.59 / Chapter 4.1 --- Extraction and purification of Kl capsular antigen from Klebsiella pneumoniae --- p.59 / Chapter 4.2 --- Gel filtration of Kl capsular antigen --- p.59 / Chapter 4.3 --- Characterization of Kl capsular antigen --- p.62 / Chapter 4.4 --- Determination of toxicity (LC50) of Kl capsular antigen by brine shrimp assay --- p.62 / Discussion --- p.62 / Chapter 5 --- IMMUNOMODULATORY ACTIVITIES OF K1 CAPSULAR POLYSACCHARIDE ANTIGEN FROM KLEBSIELLA PNEUMONIAE / Introduction --- p.67 / Results --- p.69 / Chapter 5.1 --- Mitogenic activity of Kl capsular antigen on murine lymphocytes in vitro --- p.69 / Chapter 5.2 --- Mitogenic activity of K1 capsular antigen on murine lymphocytes in vivo --- p.69 / Chapter 5.3 --- Evidences to support the mitogenic activity of K1 capsular antigen is due to its polysaccharide structure rather than due to the contamination by LPS --- p.74 / Chapter 5.4 --- Effect of Kl capsular antigen on IL-2 production from murine lymphocytes in vitro --- p.79 / Chapter 5.5 --- Effect of K1 capsular antigen on IL-1 -like factor production from murine thymocytes in vitro --- p.79 / Chapter 5.6 --- Immunopotentiating activities of Kl capsular antigen on macrophages --- p.85 / Chapter 5.6.1 --- In vivo migration of macrophages in K1-treated mice --- p.85 / Chapter 5.6.2 --- Effect ofKl capsular antigen on the macrophage EL-1-like factor production in vitro --- p.85 / Chapter 5.6.3 --- Effect ofKl capsular antigen on the macrophage nitric oxide (NO) production in vitro --- p.85 / Chapter 5.7 --- Effects of Kl capsular antigen on the proliferation and differentiation of murine bone marrow cells --- p.89 / Discussion ´ب --- p.94 / Chapter 6 --- ANTI-TUMOR ACTIVITIES OF K1 CAPSULAR POLYSACCHARIDE ANTIGEN FROM KLEBSIELLA PNEUMONIAE / Introduction --- p.100 / Results --- p.102 / Chapter 6.1 --- In vitro cytostatic effects of Kl capsular antigen on murine tumor cell lines --- p.102 / Chapter 6.2 --- In vivo anti-tumor activities of Kl capsular antigen --- p.102 / Chapter 6.3 --- Effects of Kl capsular antigen on TNF-α production and on EAT growth in vivo --- p.110 / Chapter 6.4 --- In vitro induction of macrophage-mediated cytostatic effect on tumor cells by K1 capsular antigen --- p.110 / Chapter 6.5 --- In vitro induction of macrophage-mediated cytolytic effect on tumor cells by K1 capsular antigen --- p.115 / Chapter 6.6 --- Effect of Kl capsular antigen on the activation of alloreactive cytotoxic T cells --- p.115 / Chapter 6.7 --- Effect of Kl capsular antigen on the activation of lymphokine- activated killer (LAK) cells in vitro and in vivo --- p.115 / Chapter 6.8 --- Phenotypic characterization of LAK cells by flow cytometry --- p.123 / Chapter 6.9 --- Winn-type tumor inhibition assay of LAK cells --- p.126 / Chapter 6.10 --- Adoptive transfer of LAK cells to tumor-bearing mice --- p.132 / Chapter 6.11 --- Effect ofKl capsular antigen on the activation of tumor- infiltrating lymphocytes (TILs) in tumor-bearing mice --- p.132 / Discussion --- p.136 / Chapter 7 --- CONCLUSIONS AND FUTURE PERSPECTIVES --- p.146 / BIBLIOGRAPHY --- p.152

<p>Chronic lymphocytic leukemia (CLL) cells are considered to be derived from antigen-exposed B cells. To further explore the antigen-driven selection behind the leukemogenesis of CLL, we performed immunoglobulin (Ig) specificity screening of 7 CLL cell lines and 23 primary CLL clones from patient peripheral blood. We also included a recombinant monovalent monoclonal antibody (mAb) belonging to a subset of CLL cases with identical or semiidentical heavy chain complementarity determining region 3 (HCDR3) of the IGHV3-21 gene rearrangement. We found CLL mAb specificities against vimentin, filamin B, cofilin-1, proline-rich acidic protein 1, cardiolipin, oxidized low density lipoprotein and Streptococcus pneumoniae polysaccarides. These molecules are functionally associated with microbial infection and/or apoptotic cell removal. An antigen-driven selection would therefore imply that CLL B cell precursors are involved in the elimination and scavenging of pathogens and apoptotic cells, which could trigger the development of the disease.</p><p>The limited in vitro survival of CLL cells makes Epstein-Barr virus (EBV) immortalization of CLL cells a useful experimental model for studies on antibody-specificity screening. Considering the intricate procedure of EBV transformation of CLL cells and the many false cell lines used worldwide, we also wanted to characterize and evaluate the authentic origin of several previously established CLL cell lines and their normal lymphoblastoid counterparts. Three of the CLL cell lines tested were truly authentic (I83-E95, CLL-HG3 and CII), two had features of a biclonal Ig expression (232B4 and WaC3CD5+), one was only tentatively verified (PGA-1), whereas one cell line could not be verified (EHEB) due to lack of original patient cells for comparison. Two of the presumed normal lymphoblastoid cell lines tested were shown to be a neoplastic CLL clone. This study emphasizes the importance of proper cell line authentication and we will continue to verify additional cell lines not yet proven authentic.</p><p>In conclusion, we provide evidence for natural Ab production by CLL cells and suggest that these cells might be derived from B cell precursors involved in the innate immunity and, thus, providing a first-line-defence against pathogens and in elimination of apoptotic cells.</p>

Thesis (Ph. D.)--University of Chicago, Dept. of Pathology, December 2001. / Includes bibliographical references. Also available on the Internet.

Thesis (Ph. D.)--University of Washington, 2006. / Vita. Includes bibliographical references (leaves 130-150).

Thesis (Ph. D.)--Ohio State University, 2006. / Title from first page of PDF file. Includes bibliographical references (p. 255-300).

Thesis (Ph. D.)--University of Chicago, Committy on immunology, June 2001. / Includes bibliographical references. Also available on the Internet.