This thesis focused 011 the role of voltage-gated K+ channels encoded by KCNQ genes (Kv 7 channell) in vascular smooth muscle. An array of techniques were employed including isometric tension recordings, the Langendorff isolated heart, quantitative polymerase chain reaction and Western blotting to investigate Kv 7 function and expression in normotensive and hypertensive rats and mice. With t.he aid of structurally different Kv7 activators and Kv7 blockers, this thesis elucidated an important role for Kv 7 channels in the regulation of vascular reactivity. Increasing concentrations of structurally disparate K" 7 activators (S- l , retigabine, lCA-27243 and BMS-204352) caused concentration-dependent relaxations in segments of aorta, mesenteric artery, and renal artery from normotensive rats and mice. In the Landendorff isolated-heart, the Ky 7 activators dose-dependently increased coronary perfusion with varying efficacies. These data suggest that KCNQ-encoded channels together with KCNE-encoded subunits, contribute La setting the resting membrane potential in vascular smooth muscle, and in doing so can prevent vasoconstriction. In blood vessels taken from hypertensive rats and hypertensive mice, the vasorelaxant effects of the Ky7 activators were impaired significantly, which was associated wil.h reduced Ky 7.4 protein expression in all vessels tested. The attenuation in Ky 7 function and decrease in Ky 7.4 expression across different vascular beds provides important impetus to our understanding of the pathology of hypertension. In contrast, Ky 7 function and Ky 7.4 expression was unaffected in the hypertensive rat cerebral arteries compared to the normotensive rat cerebral arteries. This thesis has identified K" 7 channels as important regulators of the smooth muscle tone in the vasculature and discovered a vessel-specific role for K" 7 channels in hypertension.

Pressure and wire myography are established techniques for the study of small artery structure and function. The series of studies in this thesis have evaluated these techniques in a variety of novel clinical settings in which microvascular dysfunction has not been evaluated previously. In health, normal small artery function is likely to serve a number of different important functions. Changes to basal tone will affect peripheral resistance and thus blood pressure. Furthermore, by regulation of downstream nutritive flow the arterial tone will also influence glycaemia. The myogenic capacity ofthe artery, by constricting and remodeling against higher intraluminal pressure, serves to protect vulnerable downstream capillary beds from elevated central pressures in hypertension. We have shown for the first time that healthy perivascular adipose tissue exerts an anticontractile effect on the adjacent small artery by release of adiponectin. In patients with obesity and metabolic syndrome, this capacity is lost due to inflammation in the adipose tissue. However, we have shown that improvements to metabolic control in patients with Type 1 Diabetes over a 10 year period can reverse the characteristic abnormal hypertrophic remodeling caused by hypertension in diabetes and instead enable eutrophic remodeling which is more commonly seen in patients without diabetes. In the final clinical study, we show that patients with late-life depression, a model of cerebral microvascular target organ damage, despite being matched with control participants for traditional risk factors (diabetes, blood pressure, cholesterol), show impaired small artery endothelial function and abnormal hypertrophic growth of the vessel wall. Lastly, in an animal model with controlled genetic mutations in the cytoskeleton protein adducin, we have shown that deficiencies in the beta subunit affect renal artery myogenic tone which is associated with the development of proteinuria.

Title from spine / Includes bibliographies / 1 v. (various pagings) : / Title page, contents and abstract only. The complete thesis in print form is available from the University Library. / Thesis (D. Sc.)--University of Adelaide, 1988

The receptor for advanced glycation endproducts (RAGE) is a 35-kDa polypeptide of the immunogloblin superfamily that has been implicated as a mediator of both acute and chronic vascular inflammation. RAGE has also recently been implicated in the pathology of pulmonary hypertension (PH): a rare, progressive disease of the small pulmonary arteries characterised by pulmonary vascular remodelling, thrombosis, vasoconstriction and increased pulmonary vascular resistance. A ligand for RAGE, the calcium binding protein MTS1/S100A4, is expressed in occlusive vascular lesions of patients with advanced PH. MTS1/S100A4 is upregulated and secreted by pulmonary arterial smooth muscle cells (PASMCs) in vitro on activation of the 5HT1b receptor and 5HT transporter (5HTT). Additionally, the proliferative effect of 5HT on these cells, which is mediated by 5HT1b and 5HTT, may be inhibited by antagonism of RAGE or reduced bioavailability of MTS1/S100A4. These data suggest that MTS1/S100A4, through its action at RAGE, is a key mediator of 5HT-induced hPASMC proliferation. Transgenic mice overexpressing MTS1/S100A4 are observed to develop obliterative pulmonary vascular disease and possess increased right ventricular pressure at baseline and after hypoxia when compared to wildtype mice (WT). These increases occur in the absence of an increase in pulmonary vascular remodelling suggesting that MTS1/S100A4 overexpression is associated with some other structural or functional change in the pulmonary circulation. We sought to further our understanding of the role of RAGE in pulmonary hypertension through treatment with a small molecule inhibitor of monocyte chemoattractant protein 1(MCP-1), a marker of downstream of RAGE rage activation; through further characterisation of the MTS1/S100A4 mouse in a chronic hypoxic model of PAH; and through treatment with soluble RAGE (sRAGE) to reduce RAGE ligand bioavailability in vivo. In each case systolic right ventricular pressure (sRVP), right ventricular hypertrophy (RVH) and pulmonary vascular remodelling were measured in normoxic conditions or after a two week chronic hypoxia challenge to induce PH. These in vivo experiments were supplemented with functional studies in isolated intrapulmonary arteries to assess vascular reactivity and vascular elastance as well as studies of pulmonary fibroblast proliferation in vitro. Treatment with the MCP-1 synthesis inhibitor Bindarit produced no detectable effects upon the pulmonary response of mice to chronic hypoxia, though this study may have been hampered by difficulties with the methylcellulose vehicle. MCP-1 produced no degree of proliferation in pulmonary fibroblasts and neither augmented nor inhibited proliferation induced by 5HT. We found little evidence for the exacerbation of PH in MTS1/S100A4 mice in normoxia, hypoxia or after 4 weeks of normoxic recovery. Mean RVP was elevated above that in WT mice exposed to hypoxia. However, MTS1/S100A4 mice appeared protected against hypoxia-induced vascular remodelling and decreases in vascular elastance. No other significant differences in sRVP, RVH or remodelling were observed between strains. Vessels isolated from MTS1/S100A4 mice tended towards an enhanced contractile response to 5HT in normoxia compared with vessels in WT mice but were also more sensitive to the nitric oxide donor SNP. These differences in vasoreactivity were largely abolished by exposure to hypoxia. Treatment with soluble RAGE (sRAGE) to reduce RAGE ligand bioavailability produced a significant reduction in sRVP after hypoxia in comparison to vehicle-dosed mice -possibly associated with the prevention of a hypoxia-induced decrease in proximal vascular elastance. However, no benefit upon the development of remodelling or the extent of RVH was observed. Vessels isolated from mice treated with sRAGE and challenged with hypoxia showed a marked increase in contractility. Further work demonstrated that sRAGE produces a small, slowly developing contraction in isolated vessels and that the maximal force of contraction to 5HT was markedly augmented in the presence of sRAGE. Finally, treatment with sRAGE did not inhibit fibroblast proliferation in vitro as induced by 5HT but was observed to cause a small degree of proliferation alone and to augment hypoxia-induced proliferation. In summary, we have reported a number of seemingly contradictory findings associated with RAGE in pulmonary hypertension. Treatment with sRAGE produced a beneficial reduction in hypoxia-induced PH associated with protection against decreased proximal vascular elastance but produced no change in hypoxia-induced RVH or remodelling as well as greatly increasing vascular contractility. MTS1/S100A4 mice show some evidence of deleterious changes to the pulmonary circulation, but these may be offset by beneficial compensatory mechanisms such as increased sensitivity to nitric oxide and protection against vascular remodelling. MTS1/S100A4 stimulates smooth muscle cell proliferation suggesting that it may involved pulmonary vascular remodelling. However, inhibition of RAGE was observed to enhance fibroblast proliferation in response to hypoxia here. Fibroblasts are important regulators of SMC proliferation in vivo. These findings therefore suggest a more complicated relationship between RAGE, its ligands and the remodelling process. Since both MTS1/S100A4 overexpression and sRAGE treatment in vivo produced findings which are difficult to reconcile using the currently employed techniques, it is clear that furthering our understanding of RAGE will require study with greater focus upon the interaction of different cell types in the pulmonary vasculature and the manner in which the disturbance of this may lead to alterations in the physical and physiological properties of the pulmonary circulation.

This thesis is based on a worldwide overview (meta-analysis) of prospective observational studies of blood pressure and cholesterol, involving a centralised collection of data on over one million individuals from 59 studies, which I have co- ordinated since its inception. Analytically, the aim has been to develop and to use appropriate statistical techniques to assess the age- and sex-specific associations of usual blood pressure and of usual cholesterol with cause-specific mortality. Since the data set is uniquely large, and because appropriate methods of analysis (with full account taken of the time-dependent nature of the regression dilution bias) have been developed and used, these associations have been established more reliably. An integral part of the methodological element of the thesis has been to investigate the systematic underestimation of associations between risk factor and disease that are obtained when only a single baseline measurement is used to assess levels of such risk factors (the regression dilution bias). The extent of this bias has been investigated in each study that had repeat measurements of risk factors during follow-up. One particularly novel aspect has been the emphasis on, and methods developed to account for, the regression dilution bias in several studies simultaneously and in an appropriately time-dependent way. This thesis illustrates the extent to which random error and inappropriate statistical analysis lead to misleading conclusions concerning the importance of blood pressure and blood cholesterol, particularly in premature death. Only by studying adequate numbers of deaths (136,000 deaths among 1 million adults during 13 million person- years of follow-up) and by using appropriate statistical techniques - taking proper account of (a) the regression dilution bias; (b) the full range of blood pressure and cholesterol; (c) the opposing effects of HDL.and the remaining non-HDL cholesterol; and (d) age at death - did it become possible to provide reliable results on the true relationships between blood pressure, cholesterol fractions and vascular and other causes of death. These analyses have demonstrated reliably that, as causes of IHD death in early middle age, blood pressure and blood lipids are three to five times more important than suggested by inappropriate analyses, with no clinically relevant inverse associations with cancer or other non-vascular mortality (except, surprisingly, COPD).

Στη διπλωματική εργασία παρουσιάζεται ένα ευφυές σύστημα αποφάσεων για τη διάγνωση και τη θεραπεία της υπέρτασης που ονομάζεται Piesys. Η δημιουργία του ευφυούς συστήματος που βασίζεται στο Piesys έγινε με τη βοήθεια δύο εργαλείων του Clips και του Fuzzytech. / In this project there is a presentation of an expert system called Piesys which is used for the diagnosis and therapy of hypertantion. The creation of the expert system became with the assistance of two expert system tools Clips and Fuzzytech.

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L'hypertension artérielle est un trait polygénique. Les variants génétiques découverts n'avaient qu'un faible effet et n'expliquaient qu'une infime partie (1%) de sa variabilité phénotypique. Ce résultat était à l'origine du concept d'héritabilité manquante. Nous pensons que l'héritabilité manquante pourrait s'expliquer par des interactions gène-gène, gène-environnement et la méthylation d'ADN. Une fois complétées par des études transcriptomiques, nous pourrions révéler les voies moléculaires impliquées. Ainsi, nous avons identifié une nouvelle interaction épistasique fonctionnelle entre le rs5355C>T du gène SELE et le rs6046G>A du gène F7 associée à la pression artérielle systolique. Dans cette même direction, nous avons rapporté deux autres interactions gène-gène. De plus, nous avons trouvé que le SNP intergénique rs7556897C>T dans le locus SLC19A3-CCL20 interagissait avec l'obésité pour influencer la pression artérielle diastolique. En parallèle aux études précédentes, nous avons montré que le rs5030878T>C du gène FPR1 et l'allèle KL-VS du gène Klotho pourraient être impliqués dans la régulation de la pression artérielle via leurs interactions avec l'âge et le traitement antihypertenseur respectivement. Nous avons aussi participé à l'identification d'un nouveau SNP, le rs2000999G>A, expliquant quasiment la moitié de l'héritabilité de l'haptoglobine dont le rôle antihypertenseur est en cours d'investigation. Finalement, nous avons proposé une nouvelle catégorie de SNPs les eMethSNPs. En conclusion, les interactions identifiées dites "fonctionnelles" montrent que l'inflammation de faible niveau est impliquée dans la régulation de la pression artérielle / Essential hypertension is a polygenic trait. Discovered genetic variants were shown to have small effects, consequently explaining a tiny fraction (1%) of its phenotypic variation and resulting to a missing heritability. The missing heritability could be explained by gene-gene, gene-environment interactions and DNA methylation. Once conducted, these approaches should be further complemented by transcriptomic analyses, and thereby to reveal the involved molecular pathways. Therefore, we have shown that rs6046G>A in F7 interacted with rs5355C>T in SELE in order to influence systolic blood pressure levels. In the same direction, we have reported two other gene-gene interactions. We have also found the intergenic SNP rs7556897T>C, located between SLC19A3 and CCL20 locus, interacting with obesity in order to influence diastolic blood pressure. In parallel to those studies, we have shown that rs5030878T>C in FPR1 and KL-VS allele in Klotho may be implicated in BP regulation through their interaction with age and antihypertensive drugs respectively. We have also participated to the identification of a novel SNP, the rs2000999G>A explaining up to the half of haptoglobin?s heritablilty, a protein currently under investigation for its antihypertensive role. Finally, we proposed a new category of functional genetic-epigenetic biomarkers, the eMethSNPs. In conclusion, the identified 'functional' interactions show that low-level inflammation is involved in blood pressure regulation

L'hypothèse que l'hypertension peut conférer un état d'hypercoagulabilité découle des principales complications liées à l'hypertension, l'infarctus du myocarde et accident vasculaire cérébral. Notre objectif était de déterminer si l'hypertension spontanée confère des changements dans les protéines de la coagulation et de la capacité de production de thrombine dans le sang et la paroi vasculaire. Nous avons utilisé le modèle de rats spontanément hypertendus (SHR) par rapport aux rats Wistar. La génération de thrombine est plus faible dans le plasma riche en plaquettes et plasma sans plaquettes de SHR par rapport à Wistar. Ceci est lié aux faibles concentrations en facteur tissulaire (TF) et en prothrombine, ainsi qu?un taux plus élevé de TFPI dans le plasma des SHR. En revanche, l'ajout d'anneaux d'aorte thoracique de SHR à un pool de plasma Wistar a donné lieu à une augmentation plus importante dans la génération de thrombine par rapport à l'ajout d'anneaux équivalent du Wistar. Alors qu'aucune différence n'a été observée pour les cellules endothéliales, la formation de thrombine était plus élevée à la surface de culture CML aortiques des SHR que des Wistar. L'exposition des phospholipides chargés négativement est plus élevée sur les anneaux et les CML des SHR que les Wistar. Les activités TF et TFPI étaient plus élevés chez les CML des SHR. Ces résultats montrent une opposition de génération de thrombine à la surface de la paroi artérielle et dans le plasma des rats SHR par rapport aux rats Wistar. Le phénotype thrombotique plus élevé de la paroi vasculaire SHR est dû à la capacité du CML à soutenir la génération de thrombine. Ces résultats suggèrent que le remodelage de la membrane phospholipidique et la synthèse de molécules pro-coagulantes induites par hypertension dans les CML sont des substrats pour une formation de thrombine accrue dans la paroi vasculaire / The hypothesis that hypertension may confer a hypercoagulable state arises from the main complications associated with hypertension, stroke and myocardial infarction. Our objective was to determine whether spontaneous hypertension confers changes in the coagulation proteins and the thrombin generating capacity in blood and the vascular wall. We used the model of spontaneously hypertensive rats (SHR) compared with Wistar rats. Thrombin generation was lower in platelet-rich plasma and platelet-free plasma from SHR compared to Wistar. This was related to lower tissue factor (TF) and prothrombin as well as higher TFPI levels in SHR plasma. In contrast, the addition of thoracic aorta rings of SHR to a Wistar plasma pool resulted in a higher increase in thrombin generation compared to the addition of equivalent rings from Wistar. Whereas no difference was observed for endothelial cells, thrombin formation was higher at the surface of cultured SHR aortic SMCs than from Wistar. Exposure of negatively-charged phospholipids was higher on SHR than on Wistar rings as well as on SMCs. TF and TFPI activities were higher in SHR SMCs. These results show opposite thrombin generating capacity of plasma and vessel walls in SHR compared to Wistar. The higher prothrombotic phenotype of the SHR vessel wall was due to the ability of SMCs to support thrombin generation. These findings suggest that the hypertension-induced membrane phospholipid reorganization and synthesis of procoagulant molecules in SMCs provide substrates for increased thrombin formation within the vessel wall

Includes 4 published papers by the author as part of appendix 9 / Bibliography: leaves 1-19 (second sequence) / System requirements for accompanying computer disk: IBM-compatible computer. Other requirements: Dbase. / 1 v. (various pagings) ; / Title page, contents and abstract only. The complete thesis in print form is available from the University Library. / Summary: describes and evaluates an economical method of collecting a large amount of data on thousands of patients suffering from essential hypertension and establishes the reliability of the data collected in this way. Also provides information on the tolerability and effectiveness of nifedipine / Thesis (M.D)--Dept. of Clinical and Experimental Pharmacology, University of Adelaide, 1993