The vascular endothelium is a single cell layer that lines the lumen of the entire vasculature. It is the site of synthesis of nitric oxide (NO), a vasodilatory compound synthesized by endothelial nitric oxide synthase (eNOS). NO causes intracellular calcium sequestration of the vascular smooth muscle cells, relaxing and dilating the arteries. Age profoundly affects endothelium-dependent vasodilation, leading to specific losses of NO. We sought to determine what causes the age-specific loss of endothelial NO. This was accomplished by investigating whether there are differences in markers of eNOS post-translational regulation elements in the aortic endothelium of young (2-4 months; corresponding to an adolescent human adult) and old (32-34 months; corresponding to a 65-75 year-old human). F 344 x Brown Norway hybrid rats. Results show that maximal eNOS activity significantly declines with age (n=4;p���0.05) though there was no change in eNOS protein levels in the aortic endothelium. Endothelial NOS exists in two distinct subcellular fractions. No alterations were detected in the soluble, inactive fraction while significantly less eNOS protein is detected in the active, plasma membrane fraction of the endothelium (n=4;p���0.02). Endothelial NOS activation is also controlled by its phosphorylation state. In this work we demonstrate that free ceramides and ceramide-activated phosphatase (PP2A) activity are significantly elevated with age in the endothelium and correlate with specific alterations in eNOS phosphorylation status consistent with its inactivation. These changes were concomittent with an age-associated decline in endothelial glutathione (GSH) and increased sphingomyelinase activity which liberates ceramides from membrane sphingolipids. In previously published reports we demonstrated that the dithiol compound R-��-lipoic acid (LA) increased maximal NO synthesis in cultured endothelial cells and that LA improved age-associated loss of eNOS stimulatory phosphorylation in rats. Therefore, we administered pharmacologic doses of LA (40 mg/kg, i.p. over 24 h) to old rats to determine whether it restored NO-dependent vasomotor function. Results show that LA significantly increased endothelial GSH (p���0.05 compared to saline controls), decreased sphingomyelinase activity and reversed the age-related increase in ceramide (p���0.01) in old animals. Finally, LA significantly improved endothelium-dependent vasodilation, suggesting that it might be a good therapeutic agent for age-related vascular endothelial dysfunction. / Graduation date: 2005

Importância. Anticoncepcionais hormonais combinados orais (AHCO) podem levar ao aumento do risco da doença cardiovascular, que pode estar associado a alterações na pressão arterial e na função endotelial. Objetivos. O objetivo deste estudo foi avaliar o impacto do uso de AHCO contendo 20 mg de etinilestradiol (EE) e 3 mg de drospirenona (DRSP) por mulheres jovens normotensas sobre a função endotelial arterial, pressão arterial sistólica (PAS, mmHg) e diástólica (PAD, mmHg), frequência cardíaca (FC, bpm), débito cardíaco (DC, L/min), e sobre a resistência periférica total (RPT, NU). Métodos. Setenta e uma mulheres jovens saudáveis com idade média de 29 ± 1 ano foram avaliadas. Quarenta e três foram analisadas antes da introdução do AHCO e ao final de 6 meses de uso (grupo-caso) e vinte e oito, não usuárias de nenhum método hormonal de contracepção, foram avaliadas quanto aos mesmos parâmetros no mesmo intervalo de tempo (grupo-controle). Resultados. Não se observaram mudanças significantes na função endotélio-dependente (VMF%) e independente (VIE%) e nas medidas de PAS, PAD, FC, DC e da RPT com o uso do AHCO (p> 0,05 para todas as variáveis). Não houve variações significantes nestes parâmetros no grupo-controle. Conclusão: O uso desta formulação de AHCO não causou alterações deletérias na reatividade vascular, e nas variáveis hemodinâmicas em mulheres jovens normotensas. / Background: Combined oral contraceptives (COCs) may lead to a rise in cardiovascular disease risk, possibly associated with changes in blood pressure and endothelial function. Objective: The objective was to evaluate the impact of COC containing 20 mcg of ethinyl estradiol (EE) and 3 mg of drospirenone (DRSP) on the arterial endothelial function, systolic and diastolic blood pressure (SBP and DBP), heart rate (HR), cardiac output (CO), and total peripheral resistance (TPR) of normotensive young women. Methods: Of the 71 women in the study, 43 were evaluated before the introduction of COC and after 6 months of its use (case group) and 28, COC nonusers, were assessed for the same parameters at the same time interval (control group). Results: No significant changes in endothelium-dependent and -independent functions or in measures of SBP, DBP, HR, CO, and TPR caused by COC use were observed in the case group (P> 0.05 for all variables) or in the control group. Conclusion: These data suggest COC with 20 mcg EE and 3 mg DRSP does not alter arterial endothelial function or hemodynamic parameters in normotensive young women

Based on the results aforementioned, I went on in the second part of the study to examine the impact of aging on EDCF-mediated contractions - the alterations of COX-2-mediated endothelium-dependent contractions and the associated release of prostaglandin(s) in the aortae from aged (>18 month-old) hamsters. Endothelium-dependent contractions in the presence of NG-nitro-L-arginine methyl ester (L-NAME) were significantly greater in the aortae from aged hamsters and contractions could also be observed without L-NAME, which were sensitive to COX-2 inhibitors and TP receptor antagonists. The levels of COX-2 expression, the release of PGF2alpha and vascular sensitivity to PGF 2alpha were augmented in aortae of aged hamsters. The present results indicate a positive impact of aging on COX-2-derived PGF2alpha-mediated endothelium-dependent contractions. / In the first part of the study, I investigated whether COX-2 participated in the occurrence of endothelium-dependent contractions in the aortae from young (-3 month-old) hamsters and identified the most possible EDCF. Endothelium-dependent contractions were elicited by acetylcholine and abolished by COX-2 inhibitors (NS-398, DuP-697 and celecoxib) and thromboxane-prostanoid (TP) receptor antagonists (S 18886, L-655,240 and GR 32191), but not by COX-1 inhibitors (valeryl salicylate and sc 560). RT-PCR and Western blot analysis using aortae with and without endothelium revealed that the COX-2 expression was localized mainly in the endothelium. Levels of prostangladin F2alpha (PGF2alpha ) and prostacyclin (PGI2) increased in response to acetylcholine and the release of both prostaglandins was inhibited by COX-2 but not COX-1 inhibitors. Exogenous PGF2alpha but not PGI2 caused contractions at a concentration that corresponded to the amount released endogenously. The release of PGF2alpha was not affected by the presence of nitric oxide (NO). The results of the present study suggest that a novel constitutive role of COX-2 in endothelium-dependent contractions, with its metabolites PGF2alpha acting as a physiological EDCF in the young hamster aortae. / In the third part of the study, I investigated the relationship and the intracellular signaling cascades linking two pro-inflammatory factors Ang II and COX-2, and tested whether COX-2 mediated the Ang II-induced vascular pathogenesis. Eight hour-incubation with 100 nmol/L Ang II resulted in maximal COX-2 expression in primary rat endothelial cells and it was inhibited by losartan and RNA synthesis inhibitor (actinomycin-D). Inhibitors of either p38 MAPK or ERK1/2 (respectively SB 202190 and PD 98059) decreased the COX-2 expression, and co-treatment with both inhibitors caused an additive effect, suggesting a joint mediation through both kinases. Protein kinase C (PKC) inhibitor (GF109203X), and particularly, the specific PKCdelta inhibitor (rottlerin), prevented Ang II-induced phosphorylation of ERK1/2 and COX-2 expression, indicating an upstream regulation of ERK1/2 by PKC delta. A pivotal role of PKCdelta in Ang II-induced COX-2 expression was further supported by a similar stimulatory effect of PKC activator, signified by the Ang II-stimulated translocation of PKCdelta to the membrane and confirmed by its phosphorylation (Tyr311). Small interfering RNA targeting PKCdelta (siPKCdelta) diminished COX-2 expression, which was abrogated in siPKCdelta-treated cells treated with SB 202190, confirming the parallel pathways of PKC delta-ERK1/2 and p38 MAPK. Aortae and renal arteries from Ang II-infused rats exhibited an increased endothelial COX-2 expression and impaired acetylcholine-induced relaxation that was normalized by celecoxib. Human mesenteric arteries incubated with Ang II demonstrated elevated endothelial COX-2 and MCP-1 expressions, of which the former was inhibited by SB 202190 plus rottlerin and the latter prevented by COX-2 inhibitor celecoxib. Renal arteries from hypertensive or diabetic patients revealed an exaggerated expression of COX-2 and MCP-1 in the endothelium. The present novel findings indicate that the activation of PKCdelta-ERK1/2 and p38 MAPK is critical in Ang II-induced COX-2 up-regulation in endothelial cells, and identify a COX-2-dependent pro-atherosclerotic cytokine MCP-1. (Abstract shortened by UMI.) / Wong, Siu Ling. / Adviser: Huang Yu. / Source: Dissertation Abstracts International, Volume: 73-02, Section: B, page: . / Thesis (Ph.D.)--Chinese University of Hong Kong, 2010. / Includes bibliographical references (leaves 192-228). / 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, [201-] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstract also in Chinese.

Wu Min. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2006. / Includes bibliographical references (leaves 87-114). / Abstracts in English and Chinese. / Declaration --- p.i / Acknowledgement --- p.ii / Publication list --- p.iii / Abstract (English) --- p.xi / Abstract (Chinese) --- p.xiv / Abbreviations --- p.xvi / List of figures / tables --- p.xviii / Chapter Chapter 1. --- General Introduction --- p.1 / Chapter 1.1 --- Endothelial function in the regulation of vascular tone --- p.1 / Chapter 1.1.1 --- NO --- p.2 / Chapter 1.1.2 --- PGI2 --- p.5 / Chapter 1.1.3 --- EDHF --- p.6 / Chapter 1.2 --- Alteration of endothelial functions after preservation with cardioplegia /organ preservation solutions in the coronary and pulmonary microcirculations --- p.18 / Chapter 1.2.1 --- Cardioplegia/organ preservation solutions --- p.21 / Chapter 1.2.2 --- Effect of Cardioplegia/organ preservation solutions on endothelial function --- p.22 / Chapter 1.2.2.1 --- Effect of K+ on endothelial function --- p.23 / Chapter 1.2.2.2 --- Effect of other components on endothelial function --- p.24 / Chapter Chapter 2. --- Materials and Methods --- p.26 / Chapter 2.1 --- Isometric force study in coronary/pulmonary resistance arteries --- p.26 / Chapter 2.1.1 --- Preparation of vessels --- p.26 / Chapter 2.1.1.1 --- Preparation of porcine coronary small arteries --- p.26 / Chapter 2.1.1.2 --- Preparation of porcine pulmonary small arteries --- p.26 / Chapter 2.1.2 --- Technique of setting up --- p.29 / Chapter 2.1.2.1 --- Mounting of small vessels --- p.29 / Chapter 2.1.2.2 --- Normalization procedure for small vessels --- p.29 / Chapter 2.1.3 --- EDHF-mediated vasorelaxation --- p.30 / Chapter 2.1.3.1 --- Precontraction and stimuli of EDHF --- p.30 / Chapter 2.1.3.2 --- """True"" response of EDHF" --- p.31 / Chapter 2.1.4 --- Data acquisition and analysis --- p.32 / Chapter 2.2 --- Electrophysiological study --- p.32 / Chapter 2.2.1 --- Preparation of small porcine coronary/pulmonary arteries --- p.32 / Chapter 2.2.2 --- Preparation of microelectrode --- p.32 / Chapter 2.2.3 --- Impaling of microelectrode --- p.33 / Chapter 2.2.4 --- Recording of membrane potential --- p.33 / Chapter 2.3 --- Statistical analysis --- p.34 / Chapter 2.4 --- Chemicals --- p.34 / Chapter Chapter 3. --- Effects of Celsior Solution on Endothelial Function in Resistance Coronary Arteries Compared to St. Thomas' Hospital Solution --- p.37 / Chapter 3.1 --- Abstract --- p.37 / Chapter 3.2 --- Introduction --- p.38 / Chapter 3.3 --- Experimental design and analysis --- p.40 / Chapter 3.3.1 --- Vessel preparation --- p.40 / Chapter 3.3.2 --- Normalization --- p.40 / Chapter 3:3.3 --- "Relaxation study: BK-induced, EDHF-mediated relaxation" --- p.41 / Chapter 3.3.4 --- Cellular electrophysiological study: EDHF-mediated cellular hyperpolarization and associated relaxation --- p.41 / Chapter 3.3.5 --- Data analysis --- p.42 / Chapter 3.4 --- Results --- p.43 / Chapter 3.4.1 --- Relaxation study --- p.43 / Chapter 3.4.1.1 --- Resting force --- p.43 / Chapter 3.4.1.2 --- U46619-induced precontraction --- p.43 / Chapter 3.4.1.3 --- EDHF-mediated relaxation --- p.43 / Chapter 3.4.2 --- Electrophysiological studies --- p.44 / Chapter 3.4.2.1 --- Resting membrane potential --- p.44 / Chapter 3.4.2.2 --- EDHF-mediated cellular hyperpolarization --- p.45 / Chapter 3.4.2.3 --- Cellular hyperpolarization-associated relaxation --- p.45 / Chapter 3.5 --- Discussion --- p.46 / Chapter 3.5.1 --- Effects of Celsior solution on endothelial function --- p.47 / Chapter 3.5.2 --- Effects of ST solution on EDHF-mediated function --- p.48 / Chapter 3.5.3 --- Comparison between Celsior and ST solutions on EDHF-mediated function --- p.48 / Chapter 3.5.4 --- Clinical implications --- p.49 / Chapter Chapter 4. --- Effects of Perfadex and Celsior Solution on Endothelial Function in Resistance Pulmonary Arteries --- p.57 / Chapter 4.1 --- Abstract --- p.57 / Chapter 4.2 --- Introduction --- p.58 / Chapter 4.3 --- Experimental design and analysis --- p.59 / Chapter 4.3.1 --- Vessel Preparation --- p.59 / Chapter 4.3.2 --- Normalization --- p.60 / Chapter 4.3.3 --- Isometric force study --- p.60 / Chapter 4.3.4 --- Electrophysiological studies --- p.61 / Chapter 4.3.5 --- Data analysis --- p.61 / Chapter 4.4 --- Results --- p.62 / Chapter 4.4.1 --- Relaxation study: EDHF-mediated relaxation --- p.62 / Chapter 4.4.1.1 --- Resting force --- p.62 / Chapter 4.4.1.2 --- U46619-induced precontraction --- p.62 / Chapter 4.4.1.3 --- EDHF-mediated relaxation --- p.62 / Chapter 4.4.2 --- Electrophysiological studies --- p.63 / Chapter 4.4.2.1 --- Resting membrane potential --- p.63 / Chapter 4.4.2.2 --- EDHF-mediated cellular hyperpolarization --- p.64 / Chapter 4.4.2.3 --- Cellular hyperpolarization-associated relaxation --- p.64 / Chapter 4.5 --- Discussion --- p.65 / Chapter 4.5.1 --- Effects of Celsior solution on endothelial function during cardiopulmonary surgery --- p.65 / Chapter 4.5.2 --- Effects of Perfadex solution on EDHF-mediated endothelial function --- p.66 / Chapter 4.5.3 --- Comparison between Celsior and Perfadex solutions on EDHF-mediated function --- p.66 / Chapter 4.5.4 --- Clinical implications --- p.67 / Chapter Chapter 5. --- Exploration of the Nature of EDHF - the Effect of H2O2 on the Membrane Potential in the Rat Small Mesenteric Arteries --- p.73 / Chapter Chapter 6. --- General Discussion --- p.75 / Chapter 6.1 --- EDHF-mediated endothelial function in porcine coronary and pulmonary circulation --- p.75 / Chapter 6.1.1 --- Role of EDHF in the regulation of porcine coronary arterial tone --- p.75 / Chapter 6.1.2 --- Role of EDHF in the regulation of porcine pulmonary arterial tone --- p.76 / Chapter 6.2 --- Alteration of EDHF-mediated endothelial functions after exposure to organ preservation solutions --- p.77 / Chapter 6.2.1 --- Effects of hyperkalemic solution on EDHF-mediated endothelial function in coronary and pulmonary circulation --- p.78 / Chapter 6.2.2 --- Effects of low-potassium-based preservation solution on EDHF-mediated endothelial function in pulmonary circulation --- p.79 / Chapter 6.2.3 --- Comparison between hyperkalemic solution and low-potassium-based preservation solution on EDHF-mediated endothelial function --- p.80 / Chapter 6.2.4 --- Effects of other component of organ preservation solutions on EDHF-mediated endothelial function --- p.81 / Chapter 6.3 --- Clinical implications --- p.82 / Chapter 6.4 --- The effect of H202 on the membrane potential in rat small mesenteric arteries --- p.83 / Chapter 6.5 --- Limitation of the study --- p.84 / Chapter 6.6 --- Future investigations --- p.85 / Chapter 6.7 --- Conclusions --- p.85 / References --- p.87

Chronic treatment with enalapril and valsartan significantly improved endothelium-dependent relaxations of aortas from ovariectomized rats. The present results clearly point to that chronic treatment with enalapril or valsartan reduced expression and function of RAS and associated oxidative stress, thereby augmented NO bioavailability and improved endothelium-dependent relaxations. These results provided novel evidence supporting a potential application of ACEI and ARB in the treatment of endothelial dysfunction-associated vascular complications in postmenopausal women. / Functional studies showed that acetylcholine-induced relaxations in isolated aortas were impaired in a time-dependent manner, from the 4th-week to the 12th-week after ovariectomy. The impaired relaxations were partially restored by acute treatment with losartan [angiotensin II type 1 receptor (AT1R) blocker] and apocynin [NAD(P)H oxidase inhibitor]. The present results demonstrate that estrogen deficiency blunted endothelium-dependent relaxations due to impaired the NO bioavailability, which is closely associated with the reduced eNOS activity and elevated RAS expression and associated NAD(P)H oxidase-mediated oxidative stress in the vascular wall. / The present study shows that chronic consumption of cranberry juice restored the endothelium-dependent relaxations in aortas from ovariectomized rats. In ovariectomized rats, the phenylephrine-induced a higher active vascular tension; which was prevented by chronic consumption of cranberry juice. The present data also shows that cranberry juice administration significantly reduces the elevated serum levels of total cholesterol, triglyceride, high density lipoprotein (HDL) cholesterol, non-HDL (nHDL) cholesterol, and nHDL/HDL. The active ingredients in the cranberry juice organic extract accounting for the vascular benefit remain to be further examined even though the extract causes endothelial NO-dependent relaxations in normal rat aortas and contains several bioactive compounds, some of which may protect the vascular function. This study provides the first line of evidence concerning a significant vascular benefit of chronic consumption of cranberry juice during estrogen deficiency. (Abstract shortened by UMI.) / The present study used ovariectomized female rats that mimic the "equivalent" state of menopause in human and investigated whether dysregulation of RAS components contribute to endothelial dysfunction and whether chronic treatment with ACEI (enalapril) or ARB (valsartan) could restore endothelial function in ovariectomized rats. / The second objective of the present study was to investigate whether or not consumption of cranberry juice, a popular drink in Western countries, could restore endothelial function during estrogen deficiency and to elucidate the cellular mechanisms underlying the improved endothelial function. / Yung, Lai Ming. / Adviser: Huana Yu. / Source: Dissertation Abstracts International, Volume: 70-06, Section: B, page: 3252. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2008. / Includes bibliographical references (leaves 148-168). / 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.

Zhang Rongzhen. / "July 2004." / Thesis (Ph.D.)--Chinese University of Hong Kong, 2004. / Includes bibliographical references (p. 130-176). / 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.