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Hyperglycemic impairment of CGRP-induced cAMP responses in vascular smooth muscle cells (VSMCs) and the role of cGMP/protein kinase G pathway in regulating apoptosis and proliferation of VSMCs and bone marrow stromal stem cells.January 2006 (has links)
Wong Cheuk Ying. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2006. / Includes bibliographical references (leaves 101-124). / Abstracts in English and Chinese. / Abstract --- p.i / 摘要 --- p.iv / Acknowledgement --- p.vi / List of Abbreviations --- p.vii / Chapter Chapter 1. --- General Introduction --- p.1 / Chapter Chapter 2. --- Methods --- p.4 / Chapter 2.1 --- Measurement of cAMP and cGMP in VSMCs --- p.4 / Chapter 2.1.1 --- Cell culture --- p.4 / Chapter 2.1.2 --- Enzyme-immunoassay colorimetric measurement for cAMP and cGMP --- p.5 / Chapter 2.1.3 --- Statistical analysis --- p.6 / Chapter 2.2 --- Measurement of apoptosis in VSMCs and bone marrow-derived stem cells --- p.6 / Chapter 2.2.1 --- Cell culture --- p.6 / Chapter 2.2.2 --- Hoechst33258 --- p.7 / Chapter 2.2.3 --- Cell Death ELISA plus --- p.7 / Chapter 2.2.4 --- Protein extraction and Western blot analysis of PKG expression --- p.8 / Chapter 2.2.5 --- Statistical analysis --- p.9 / Chapter 2.3 --- Measurement of cell proliferation in VSMCs and bone marrow-derived stem cells --- p.9 / Chapter 2.3.1 --- Cell culture --- p.9 / Chapter 2.3.2 --- Cell count --- p.10 / Chapter 2.3.3 --- MTT assay --- p.11 / Chapter 2.3.4 --- BrdU-(5`Bromo-2-deoxyuridine) ELISA colorimetric assay --- p.11 / Chapter 2.3.5 --- Statistical analysis --- p.12 / Chapter Chapter 3. --- Effects of hyperglycemia on CGRP-induced cAMP response in VSMCs / Chapter 3.1 --- Introduction --- p.13 / Chapter 3.2 --- Results --- p.18 / Chapter 3.3 --- Discussion --- p.22 / Chapter Chapter 4. --- Role of cGMP and protein kinase G in regulation of apoptosis in VSMCs / Chapter 4.1 --- Introduction --- p.26 / Chapter 4.2 --- Results --- p.30 / Chapter 4.3 --- Discussion --- p.44 / Chapter Chapter 5. --- Role of protein kinase G in regulation of proliferation in VSMCs / Chapter 5.1 --- Introduction --- p.55 / Chapter 5.2 --- Results --- p.58 / Chapter 5.3 --- Discussion --- p.67 / Chapter Chapter 6. --- Effects of aging and eNOS- and iNOS-gene deletion (using eNOS- and iNOS-knockout mice) on apoptosis of VSMCs / Chapter 6.1 --- Introduction --- p.73 / Chapter 6.2 --- Results --- p.76 / Chapter 6.3 --- Discussion --- p.79 / Chapter Chapter 7. --- Role of protein kinase G in regulation of apoptosis and proliferation of bone marrow stromal stem cells / Chapter 7.1 --- Introduction --- p.81 / Chapter 7.2 --- Results --- p.84 / Chapter 7.3 --- Discussion --- p.92 / Chapter Chapter 8. --- Overall discussion --- p.95 / Chapter Chapter 9. --- References --- p.101
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Regulation of apoptosis in uterine epithelial cells and ovarian cancer cells by the cGMP/protein kinase G signaling pathway.January 2003 (has links)
Chan Siu Lan. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2003. / Includes bibliographical references (leaves 149-181). / Abstracts in English and Chinese. / Abstract --- p.ii / Chinese Abstract (摘要) --- p.v / Acknowledgements --- p.viii / Publications --- p.x / Table of contents --- p.xii / List of Figures --- p.xvi / List of Table and Diagram --- p.xx / Abbreviations --- p.xxi / Chapter Chapter 1: --- Introduction / Chapter 1.1 --- Major objectives and long-term significance --- p.1 / Chapter 1.2 --- Biological significance of apoptosis --- p.1 / Chapter 1.3 --- Importance of apoptosis in the study of the female reproductive system --- p.3 / Chapter 1.4 --- Specific aims of the present project --- p.3 / Chapter 1.5 --- Experimental approaches --- p.8 / Chapter Chapter 2: --- Materials and Methods / Chapter 2.1 --- General experimental methods --- p.11 / Chapter 2.1.1 --- Culture of cells --- p.11 / Chapter 2.1.1.1 --- Culture of rabbit immortalized uterine epithelial cells --- p.11 / Chapter 2.1.1.2 --- Culture of primary mouse uterine epithelial cells --- p.12 / Chapter 2.1.1.3 --- Culture of human ovarian epithelial cancer cells --- p.13 / Chapter 2.1.2 --- Assessment of apoptotic DNA fragmentation --- p.13 / Chapter 2.1.2.1 --- DNA extraction --- p.14 / Chapter 2.1.2.2 --- Assessment of apoptotic DNA --- p.14 / Chapter 2.1.2.3 --- Assessment of apoptotic DNA by CE-LIF --- p.15 / Chapter 2.1.2.4 --- Assessment of apoptosis by Nuclear Hoechst 33248 Staining / Chapter 2.1.3 --- Assessement of protein content --- p.16 / Chapter 2.1.3.1 --- Protein extraction and western blot analysis --- p.16 / Chapter 2.1.4 --- Adenoviral infection of A2780s cells --- p.18 / Chapter 2.2 --- Preparation of solutions --- p.18 / Chapter 2.3 --- Animals and cell lines --- p.25 / Chapter 2.4 --- Statistical analysis --- p.25 / Chapter Chapter 3: --- Literature Review / Chapter 3.1 --- Morphological analysis of physiological cell death --- p.26 / Chapter 3.1.1 --- Characteristics of apoptosis --- p.27 / Chapter 3.2 --- Methods of detecting apoptosis --- p.31 / Chapter 3.3 --- Molecules controlling apoptosis --- p.33 / Chapter 3.3.1 --- Caspases --- p.33 / Chapter 3.3.2 --- The Bcl-2 family proteins --- p.34 / Chapter 3.4 --- Apoptosis signalling --- p.36 / Chapter 3.4.1 --- The death receptor-dependent pathway --- p.36 / Chapter 3.4.2 --- The mitochondria-dependent pathway --- p.38 / Chapter 3.4.3 --- The endoplasmic-reticulum-dependent pathway --- p.39 / Chapter 3.5 --- Importance of apoptosis in the female reproductive system --- p.40 / Chapter 3.5.1 --- Apoptosis in uterus epithelial cells --- p.40 / Chapter 3.5.2 --- Apoptosis in ovarian cancer cells --- p.42 / Chapter 3.6 --- Regulation of apoptosis by nitric oxide/cGMP/protein kinase G --- p.44 / Chapter 3.6.1 --- Regulation of apoptosis by nitric oxide --- p.44 / Chapter 3.6.2 --- Regulation of apoptosis by cGMP --- p.48 / Chapter 3.6.3 --- Regulation of apoptosis by soluble guanyly cyclase activator --- p.50 / Chapter Chapter 4: --- "Apoptotic DNA fragmentation caused by sodium nitroprusside, a nitric oxide donor, in uterine epithelial cells: ultrasensitive quantitation using the new capillary electrophoresis/laser-induced fluorescence (CE-LIF) technology" / Chapter 4.1 --- Abstract --- p.52 / Chapter 4.2 --- Introduction --- p.53 / Chapter 4.3 --- Results --- p.57 / Chapter 4.4 --- Discussion --- p.61 / Chapter 4.5 --- Figures of Chapter 4 --- p.66 / Chapter Chapter 5: --- Guanylyl-cyclase inhibitors NS2028 and ODQ and protein-kinase-G inhibitor KT5823 trigger apoptotic DNA fragmentation in an immortalized uterine epithelial cell line: anti-apoptotic effects of basal cGMP/PKG / Chapter 5.1 --- Abstract --- p.74 / Chapter 5.2 --- Introduction --- p.75 / Chapter 5.3 --- Results --- p.80 / Chapter 5.4 --- Discussion --- p.83 / Chapter 5.5 --- Figures of Chapter 5 --- p.89 / Chapter Chapter 6: --- "Direct, prolonged activation of soluble guanylyl cyclase by YC-1 or protein kinase G by cGMP analogs enhances the level of apoptosis in an immortalized uterine epithelial cell line, HRE-H9 cells" / Chapter 6.1 --- Abstract --- p.100 / Chapter 6.2 --- Introduction --- p.101 / Chapter 6.3 --- Results --- p.105 / Chapter 6.4 --- Discussion --- p.107 / Chapter 6.5 --- Figures of Chapter 6 --- p.114 / Chapter Chapter 7: --- "ODQ,an inhibitor of soluble guanylyl cyclase, down-regulates XIAP expression and induces apoptosis in human ovarian cancer cells" / Chapter 7.1 --- Abstract --- p.124 / Chapter 7.2 --- Introduction --- p.125 / Chapter 7.3 --- Results --- p.129 / Chapter 7.4 --- Discussion --- p.132 / Chapter 7.5 --- Figures of Chapter 7 --- p.138 / Chapter Chapter 8: --- Overall Conclusion --- p.145 / Chapter Chapter 9: --- References --- p.149
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