Wai Kei Chan. / Thesis (M.Phil.)--Chinese University of Hong Kong, 1999. / Includes bibliographical references (leaves 155-164). / Abstracts in English and Chinese. / Chapter Chapter 1 --- Introduction / Chapter 1.1. --- Sulfonylureas --- p.1 / Chapter 1.2. --- Biological action of sulfonylurea drugs --- p.2 / Chapter 1.2.1 --- Effects on pancreatic β cells --- p.5 / Chapter 1.2.2. --- Effects on cardiac myocytes --- p.7 / Chapter 1.2.3. --- Effects on smooth muscle cells --- p.11 / Chapter 1.2.4. --- Effects on endothelial cells --- p.14 / Chapter 1.3. --- Side effects and toxicity --- p.15 / Chapter 1.4. --- Objectives of the present study --- p.17 / Chapter Chapter 2 --- Methods and Marterials / Chapter 2.1. --- Tissue and Cell Preparation --- p.20 / Chapter 2.1.1. --- Preparation of the isolated rat aorta and mesenteric artery --- p.20 / Chapter 2.1.2. --- Removal of the functional endothelium --- p.20 / Chapter 2.1.3. --- Cell culture --- p.21 / Chapter 2.1.3.1. --- Materials --- p.21 / Chapter 2.1.3.2. --- Aortic smooth muscle cells in primary culture --- p.21 / Chapter 2.1.3.3. --- Aortic endothelial cells in primary culture --- p.23 / Chapter 2.1.3.4. --- Cultured rat aortic smooth muscle cell line (A7r5) --- p.23 / Chapter 2.1.3.5. --- Cultured human umbilical vein endothelial cell line (ECV-304) --- p.24 / Chapter 2.1.3.6. --- Cell subculture --- p.24 / Chapter 2.1.3.7. --- Immunostaining of endothelial cells in primary culture --- p.24 / Chapter 2.2. --- Organ Bath Set-up --- p.25 / Chapter 2.3. --- Force Measurement --- p.28 / Chapter 2.3.1. --- Vascular action of glibenclamide --- p.28 / Chapter 2.3.1.1. --- Antagonistic effect of glibenclamide on relaxation induced by K+ channel openers --- p.28 / Chapter 2.3.1.2. --- Relaxant response of glibenclamide --- p.29 / Chapter 2.3.1.3. --- Role of endothelium-derived vasoactive factors in glibenclamide induced relaxation --- p.29 / Chapter 2.3.1.4. --- Effect of endothelial prostanoids in glibenclamide-induced relaxation --- p.30 / Chapter 2.3.1.5. --- Effects of putative K+ channel blockers on glibenclamide-induced relaxation --- p.30 / Chapter 2.3.1.6. --- Effect of glibenclamide on high K+- and CaCl2-induced contraction --- p.31 / Chapter 2.3.1.7. --- Effect of glibenclamide on prostaglandin F2α-induced contraction --- p.32 / Chapter 2.3.1.8. --- Effect of glibenclamide on protein kinase C-mediated contraction --- p.32 / Chapter 2.3.2. --- Vascular action of glipizide --- p.33 / Chapter 2.3.3. --- Vascular action of tolbutamide --- p.33 / Chapter 2.3.3.1. --- Contractile response of tolbutamide --- p.33 / Chapter 2.3.3.2. --- Effects of inhibitors of endothelium-derived factors --- p.33 / Chapter 2.3.3.3. --- Effects of inhibitors of Ca2+ influx --- p.34 / Chapter 2.3.3.4. --- Effect of protein kinase C inhibitor --- p.34 / Chapter 2.3.3.5. --- Effects of neural factors --- p.34 / Chapter 2.4. --- Cyclic GMP measurement --- p.35 / Chapter 2.4.1. --- Material --- p.35 / Chapter 2.4.2. --- Methods --- p.35 / Chapter 2.4.2.1. --- Tissue preparation --- p.35 / Chapter 2.4.2.2. --- Plasma and tissue according to protocols provided by Amersham --- p.35 / Chapter 2.4.2.3. --- Cyclic GMP content measurement --- p.36 / Chapter 2.4.2.4. --- Protein content measurement --- p.39 / Chapter 2.4.2.5. --- Cyclic GMP measurement protocol --- p.40 / Chapter 2.5. --- Ca2+ measurement --- p.40 / Chapter 2.5.1. --- Materials --- p.40 / Chapter 2.5.1.1. --- PTI RatioMaster Fluorescence System --- p.40 / Chapter 2.5.1.2. --- Confocal Imaging System --- p.42 / Chapter 2.5.2. --- Method --- p.42 / Chapter 2.5.3. --- Protocols for Ca2+ measurement --- p.45 / Chapter 2.5.3.1. --- Effect of glibenclamide in endothelial cells --- p.45 / Chapter 2.5.3.2. --- Effect of glibenclamide in vascular smooth muscle cells --- p.45 / Chapter 2.5.3.3. --- Effect of tolbutamide in vascular smooth muscle cells --- p.46 / Chapter 2.6. --- Cell proliferation --- p.45 / Chapter 2.6.1. --- Materials --- p.45 / Chapter 2.6.2. --- Method --- p.46 / Chapter 2.6.3. --- Protocols for cell proliferation --- p.47 / Chapter 2.6.3.1. --- Effect of glibenclamide on endothelial cell proliferation --- p.47 / Chapter 2.6.3.2. --- Effect of glibenclamide on aortic smooth muscle cell proliferation --- p.47 / Chapter 2.7. --- Chemicals and solutions --- p.48 / Chapter 2.8. --- Statistical analysis --- p.50 / Chapter Chapter 3 --- Results / Chapter 3.1. --- Glibenclamide --- p.51 / Chapter 3.1.1. --- Effect of glibenclamide on the K+ channel activity --- p.51 / Chapter 3.1.2. --- Relaxant response of glibenclamide --- p.55 / Chapter 3.1.3. --- Effects of inhibitors of nitric oxide activity on glibenclamide- induced relaxation --- p.57 / Chapter 3.1.4. --- Role of endothelial relaxing prostanoids in glibenclamide-induced relaxation --- p.69 / Chapter 3.1.5. --- Effect of putative K+ channel blockers on glibenclamide-induced relaxation --- p.73 / Chapter 3.1.6. --- Effect of glibenclamide on high K+-induced arterial contraction --- p.75 / Chapter 3.1.7. --- Effect of glibenclamide on protein kinase C-mediated contraction --- p.83 / Chapter 3.1.8. --- Effect of glibenclamide on prostaglandin F2α-induced contraction --- p.83 / Chapter 3.2 --- Glipizide --- p.85 / Chapter 3.2.1. --- Relaxant response of glipizide --- p.85 / Chapter 3.3. --- Tolbutamide --- p.91 / Chapter 3.3.1. --- Contractile response to tolbutamide --- p.91 / Chapter 3.3.2. --- Effects of endothelium-derived factors --- p.94 / Chapter 3.3.3. --- Effects of inhibitors of Ca2+ influx on tolbutamide-induced contraction --- p.98 / Chapter 3.3.4. --- "Effects of forskolin, sodium nitroprusside, staurosporine on tolbutamide-induced contraction" --- p.102 / Chapter 3.3.5. --- Effect of neural factors --- p.106 / Chapter 3.4. --- Effect of glibenclamide on cGMP levels --- p.112 / Chapter 3.5. --- Effect of glibenclamide on intracellular[Ca2+ ] in cultured endothelial cells --- p.112 / Chapter 3.6. --- Effect of glibenclamide on intracellular [Ca2+] in cultured aortic smooth muscle cells --- p.115 / Chapter 3.7. --- Effect of tolbutamide on intracellular [Ca2+] in cultured aortic smooth muscle cells --- p.121 / Chapter 3.8. --- Effect of glibenclamide on proliferation of cultured endothelial cells --- p.121 / Chapter 3.9. --- Effect of glibenclamide on proliferation of cultured aortic smooth muscle cells --- p.123 / Chapter Chapter 4 --- Discussion / Chapter 4.1. --- Effect of glibenclamide --- p.133 / Chapter 4.2. --- Effect of glipizide --- p.143 / Chapter 4.3. --- Effect of tolbutamide --- p.144 / Chapter 4.4. --- Conclusion --- p.152 / References --- p.155 / Publications --- p.163

The place of estrogen in women's health remains controversial. Premenopausal women have a lower prevalence of cardiovascular disease (CVD) than men and in observational studies hormone replacement therapy (HRT) decreases CVD in postmenopausal women. However, prospective randomized trials of secondary and primary prevention have failed to substantiate an overall protective effect from HRT and have even shown some harm. To explain this paradox it is necessary to better understand the effects of estrogen on the vascular wall. Estrogen rapidly mediates the activation of eNOS and increases the production of nitric oxide (NO), an important factor for endothelial health. In ovariectomized rats estrogen reduces production of superoxide (O2-) by NAD(P)H oxidase. The decreased function is associated with a decrease in the p47phox component of NAD(P)H oxidase and its interaction with the multicomponent enzyme. In these rats estrogen did not alter eNOS expression and bioavailability of NO, which is in contrast to its acute effects. This highlights the difference between chronic and acute studies. The decrease in O2-production suggests the intracellular signaling. / Estrogen has antiapoptotic effects. Oxidized low-density lipoprotein (oxLDL) and the inflammatory cytokine TNFalpha increased apoptosis which is associated with atherosclerosis. In human umbilical vein endothelial cells (HUVEC), estrogen decreased the extent of TNFalpha and oxLDL induced apoptosis as indicated by the expression of cleaved caspase-3 and FACS assay. Estrogen also preserves the antiapoptotic mitochondrial Bcl-2 and Bcl-xL proteins. / Estrogen has angiogenic properties that can help a healthy endothelium respond to injury. However, estrogen increases the angiogenesis caused by TNFalpha and this could lead to revascularization in the plaques of women with advanced disease. / Overall the balance between the positive and negative aspects of the effects of estrogen on the vascular wall could explain the paradoxical response in older women.

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