Biological Activities of Novel Synthetic Tri-peptide Conjugated Chitosan Oligosaccharides

목차 (Table of Contents)

• Abstract i
• Table of Contents iv
• List of Figures ix
• List of Tables xii
• List of Abbreviation xiii
• Chapter I. Peptide-Chitooligosasccharide Conjugation and Screening of Bioactivities
• 1. Introduction 2
• 1.1. Chitooligosaccharides and their bioavailability 2
• 1.2. Chitooligosaccharide derivatization 6
• 1.3. Potential usage of peptide as bioactivity enhancer 7
• 1.4. Conjugation of amino acids and peptides 8
• 2. Materials and Methods 10
• 2.1. Materials and chemicals for synthesis 10
• 2.2. Cell lines and chemicals for screening methods 10
• 2.3. Synthesis of peptide conjugated COSs 11
• 2.4. Instrumental analysis 13
• 2.5. Determination of toxicity of the samples 16
• 2.6. Anti-inflammatory effect screening 17
• 2.6.1. Nitric oxide production assay 17
• 2.7. Anti-cancer effect screening 17
• 2.7.1. Anti-proliferative effect 17
• 2.8. Anti-HIV effect screening 18
• 2.8.1. Syncytia formation assay 18
• 2.8.2. Cell viability assay 18
• 2.9. Screening anti-diabetes activity 19
• 2.9.1. Glycerol assay 19
• 2.10. Statistical analysis 20
• 3. Results and Discussion 21
• 3.1. Synthesis of amino-acid conjugated COSs 23
• 3.2. Determination of the toxicity of the samples 32
• 3.3. Anti-inflammation activity 32
• 3.4. Anti-cancer activity 37
• 3.5. Anti-HIV activity 37
• 3.6. Anti-diabetes activity 43
• 4. Conclusion 45
• References 46
• Chapter II. Investigation Anti-Diabetes and Anti-Obesity Mechanism
• 1. Introduction 54
• 1.1. Enzyme inhibitors 58
• 1.2. Protein glycation and diabetes 59
• 1.3. Role of obesity in type 2 diabetes (insulin resistance) 61
• 1.4. Obesity 62
• 1.5. Chitooligosaccharides and peptides 64
• 2. Materials and Methods 65
• 2.1. Materials and chemicals 65
• 2.2. Cell culture 65
• 2.3. Differentiation of adipocytes 65
• 2.4. Oil red O staining 66
• 2.5. Triglyceride assay 67
• 2.6. RNA extraction 68
• 2.7. Reverse transcription polymerase chain reaction (RT-PCR) 68
• 2.8. Western blot 71
• 2.9. α -glucosidase enzyme assay 71
• 2.10. α -amylase enzyme assay 72
• 2.11. Protein glycation assay 73
• 2.12. Glucose uptake assay 73
• 2.13. Statistical analysis 74
• 3. Results and Discussion 75
• 3.1. Differentiation of adipocytes 75
• 3.2. Effect of COS and its derivative on lipid accumulation of adipocyte cells 75
• 3.3. Inhibition of adipogenic transciption factor expression 83
• 3.4. Effect of QWM-COS and MWQ-COS on protein glycation 86
• 3.5. Effect of WQM-COS and MQW-COS on activity of α-glucosidase and
• α-amylase 88
• 3.6. Effect of QWM-COS and MWQ-COS on glucose uptake 90
• 4. Conclusion 94
• References 95
• Chapter III. Elucidation of the action mechanism against HIV-1 infection
• 1. Introduction 101
• 1.1. Acquired immunodeficiency syndrome (AIDS) and HIV 101
• 1.2. Course of infection 103
• 1.3. Human immunodeficiency virus type 1 106
• 1.4. Genes and structure 107
• 1.5. HIV-1 replication cycle 109
• 1.6. Viral entry 109
• 1.7. Reverse transcription, nuclear import, and integration 110
• 1.8. Gene expression 111
• 1.9. Budding and maturation 112
• 1.10. Anti-HIV therapy 114
• 1.11. Antiretroviral drugs 116
• 1.11.1. Protease inhibitors 116
• 1.11.2. Reverse transcriptase inhibitors 118
• 1.11.3. Integrase inhibitors 119
• 1.11.4. Entry inhibitors 120
• 1.11.5. Fusion inhibitors 120
• 1.11.6. Coreceptor antagonists 121
• 1.12. Current status of HIV/AIDS in Korea 122
• 2. Materials and Methods 126
• 2.1. Materials and chemicals 126
• 2.2. Cells and viruses 126
• 2.3. Cell Viability assay 127
• 2.4. Inhibition of syncytia formation 127
• 2.5. Inhibition of cytopathic effect of HIV-1 128
• 2.6. Western blot analysis 128
• 2.7. p24 antigen capture ELISA 129
• 2.8. Delayed addition of pep-COSs to HIV-1IIIB infected cells 130
• 2.9. Effect of pep-COSs on cell-cell fusion 130
• 2.10. Luciferase gene reporter assay 130
• 2.11. CD4-gp41 ELISA 131
• 2.12. X-Gal staining 132
• 2.13. Polymerase chain reaction 132
• 2.14. Statistical analysis 133
• 3. Results and Discussion 134
• 3.1. Cell viability assay 134
• 3.2. Inhibition of syncytia formation 134
• 3.3. Inhibition of cytopathic effect of HIV-1 137
• 3.4. p24 antigen capture ELISA 139
• 3.5. Luciferase gene reporter assay 141
• 3.6. Delayed addition of pep-COSs to HIV-1IIIB infected cells 141
• 3.7. Effect of pep-COSs on cell-cell fusion 143
• 3.8. CD4-gp41 ELISA 144
• 3.9. X-gal staining 149
• 3.10. Polymerase chain reaction 149
• 4. Conclusion 152
• References 143
• Acknowledgement 162